VX- ?

CAS  2446817-72-5

HYDRATE 2446818-26-2

Acetic acid, 1-​methylethyl ester 2446818-27-3

C₂₁ H₂₀ F N₃ O_{3, 381.4}

1H-Indole-3-propanamide, 2-(4-fluorophenyl)-N-[(3S,4R)-4-hydroxy-2-oxo-3-pyrrolidinyl]-

3-[2-(4-fluorophenyl)-lH-indol-3-yl]-N-[(3S,4R)-4-hydroxy-2- oxo-pyrrolidin-3-yl ] propanamide

use in treating focal segmental glomerulosclerosis (FSGS) and/or non-diabetic kidney disease (NDKD).

NEW DRUG APPROVALS

one time

$10.00

Click here to purchase.

PATENT

WO/2021/158666

SOLID FORMS OF APOL1 INHIBITOR AND METHODS OF USING SAME

Compound I is disclosed as Compound 87 in U.S. Provisional Application No.62/780,667 filed on December 17, 2018, U.S. Application No. 16/717,099 filed onDecember 17, 2019, and PCT International Application No. PCT/US2019/066746 filed on December 17, 2019, the entire contents of each of which are incorporated herein by reference.

Compound I, which can be employed in the treatment of diseases mediated by APOLl, such as FSGS and NDKD

Example 1. Synthesis of Compound

Preparation of Compound I and Forms Thereof

Compound I Compound I /^‘– PrOAc solvate Form A

n-pentanol/

n-heptane

Compound I

Form B

Step 1. Synthesis of 3-[2-(4-fluorophenyl)-lH-indol-3-yl]propanoic acid (C101)

[00156] To a mixture of C104 (100.0 g, 1.0 equiv) and phenyl hydrazine hydrochloride (72.2 g, 1.05 eqiv) was charged AcOH (800 mL, 8 vol). The mixture was agitated and heated to 85 °C for 16 hours. The batch was cooled to 22 °C. A vacuum was applied and the batch distill at <70 °C to ~3 total volumes. The batch was cooled to 19- 25 °C. The reactor was charged with iPrOAc (800 mL, 8 vol) and then charged with water (800 mL, 8 vol). The internal temperature was adjusted to 20 – 25 °C and the biphasic mixture was stirred for no less than 0.5 h. Stirring was stopped and the phases allowed to separate for no less than 0.5 h. The lower aqueous layer was removed. 1 N HC1 (500 mL, 5 vol) was charged to the reactor. The internal temperature was adjusted to 20 – 25 °C, and the

biphasic mixture was stirred for no less than 0.5 h. Stirring was stopped and phases were allowed to separate for no less than 0.5 h. The lower aqueous layer was removed. The reactor was charged with 1 N HC1 (500 mL, 5 vol). The internal temperature was adjusted to 20 – 25 °C, and the biphasic mixture was stirred for no less than 0.5 h. Stirring was stopped and phases were allowed to separate for no less than 0.5 h. The lower aqueous layer was removed. Water (500 mL, 5 vol) was charged to the reactor. The internal temperature was adjusted to 20 – 25 °C, and the biphasic mixture was stirred for no less than 0.5 h. Stirring was stopped and phases were allowed to separate for no less than 0.5 h. The lower aqueous layer was removed. Water (500 mL, 5 vol) was charged to the reactor. The internal temperature was adjusted to 20 – 25 °C, and the biphasic mixture was stirred for no less than 0.5 h. Stirring was stopped and phases were allowed to separate for no less than 0.5 h. The lower aqueous layer was removed. The organic phase was distilled under vacuum at <75 °C to 3 total volumes. The reactor was charged with toluene (1000 mL, 10 vol). The organic phase was distilled under vacuum at <75 °C to 5 total volumes. The reactor was charged with toluene (1000 mL, 10 vol). The organic phase was distilled under vacuum at <75 °C to 5 total volumes. The resulting slurry was heated to an internal temperature of 85 °C until complete dissolution of solids was achieved. The mixture was allowed to stir for 0.5 h at 85 °C and then cooled to an internal temperature of 19 – 25 °C over 5 h. The mixture was allowed to stir at 25 °C for no less than 2 h. The slurry was filtered. The filter cake was washed with toluene (1 x 2 vol (200 mL) and 1 x 1.5 vol (150 mL)). The solids were dried under vacuum with nitrogen bleed at 60 °C to afford product C101 (95.03 g, 70%).

Step 2. Synthesis of Compound I

[00157] A mixture of 3-[2-(4-fluorophenyl)-lH-indol-3-yl]propanoic acid C101 (50 g, 1.0 equiv), S2 hydrochloride (28.3 g, 1.05 equiv), and CDMT (34.1 g, 1.1 equiv) was charged with 2-MeTHF (200 mL, 4 vol) and DMF (50 mL, 1 vol) and the mixture was agitated. The internal temperature adjusted to <13 °C. The reactor was charged with NMM (64.5 g, 3.5 equiv) over 1 h, while maintaining internal temperature <20 °C. The internal temperature was adjusted to 25 °C and the batch was stirred at that temperature for 14 h. The batch was cooled to 10 °C and charged with water (250 mL, 5 vol) while keeping the internal temperature <20 °C. The batch was then warmed to 20 – 25 °C. Stirring was stopped, and the phases allowed to separate for 10 min. The lower aqueous phase was removed. The aqueous layer was back extracted with 2-MeTHF (2 x 200 mL, 2 x 4 vol) at

20 – 25 °C. The combined organic phases were washed with 1 N HC1 (500 mL, 10 vol) at 20 – 25 °C by mixing for 10 min and settling for 10 min. The lower aqueous phase was removed. The organic phases were washed with 0.25 N HC1 (2 x 250 mL, 2 x 5 vol) at 20 – 25 °C by mixing for 10 min and settling for 10 min for each wash. Lower aqueous phases were removed after each wash. The organic phase was washed with water (250 mL, 5 vol) at 20 – 25 °C by mixing for 10 min and settling for 10 min. The reactor was charged with 20 wt % Nuchar RGC® and stirred for 4 h. The reaction mixture was filtered through a pad of celite®. The reactor and celite® pad were rinsed with 2-MeTHF. The combined organics were distilled under vacuum at <50 °C to 5 total volumes. The reactor was charged with iPrOAc (500 mL, 10 vol). The organic phase was distilled under vacuum at <50 °C to 5 total volumes. The mixture was charged with additional iPrOAc (400 mL, 8 vol) and distillation under vacuum was repeated. The mixture was charged with additional iPrOAc (250 mL, 5 vol), heated to an internal temperature of 75 °C and stirred for 5 h. The slurry was cooled to 25 °C, over 5 h and stirred for no less than 12 h. The slurry was filtered and the filter cake washed with iPrOAc (2 x 50 mL, 2 x 1 vol). The solids were dried under vacuum with nitrogen bleed at 55 – 60 °C to afford Compound I as an iPrOAc solvate (60.38 g including 9.9% w/w iPrOAc, 80.8% yield).

Recrystallization to Form A of Compound I

[00158] Compound I as an iPrOAc solvate (17.16 g after correction for iPrOAc content, 1.0 equiv) was charged to a reactor. A mixture of IP A (77 mL, 4.5 vol) and water (137 mL, 8 vol) were charged to the reactor. The slurry was heated to an internal temperature of 75 °C. The batch was cooled to an internal temperature of 25 °C over 10 h and then stirred at 25 °C for at least 12 h. The slurry was filtered. The filter cake was washed with 36/64 IP A/water (2 x 52 mL, 2 x 3 vol). The solids were dried under vacuum with nitrogen bleed at 60 °C to afford Compound I as a neat, crystalline form (Form A, 15.35 g, 89%).

[00159] The X-ray powder diffractogram of Compound I Form A (FIG. 50) was acquired at room temperature using a PANalytical Empyrean diffractometer equipped with PIXcel ID detector. The peaks are listed in Table A below.

Table A. XRPD of Form A of Compound I

|

I

PATENT

• WO2020131807

Alternative Preparation I of Compound 87 (Indole preparation route C)

Step 1. Synthesis of 2-(4-fluorophenyl)-lH-indole (C98)

[00401] To a stirred suspension of indole (5 g, 42.7 mmol) and (4- fluorophenyl)boronic acid (8.96 g, 64.0 mmol) in AcOH (200 mL) was

added Pd(OAc)2.Trimer (1.44 g, 6.4 mmol) and the mixture stirred at room temperature for 16 h under 02-balloon pressure. Then the reaction mixture was filtered through a Celite® pad, washed with EtOAc (500 mL). The filtrates were washed with water, sat. NaHC03 solution, brine solution, then dried over Na2S04 and concentrated under reduced pressure. Purification by silica gel chromatography (Gradient: 0-10 % EtOAc in heptane) yielded the product afforded 2-(4-fluorophenyl)-lH-indole (5.5 g, 61 %). ‘H NMR (300 MHz, DMSO-de) 5 11.51 (s, 1H), 7.9 (t, J = 5.4 Hz, 2H), 7.52 (d, J = 7.8 Hz, 1H), 7.39 (d, J = 8.1 Hz, 1H), 7.30 (t, J = 8.7 Hz, 2H), 7.09 (t, J = 12 Hz, 1H), 6.99 (t, J = 7.5 Hz, 1H), 6.86 (s, 1H). LCMS m/z 212.4 [M+H]⁺.

Step 2. Synthesis of methyl (E)-3-[2-(4-fluorophenyl)-lH-indol-3-yl]prop-2-enoate (C99)

[00402] 2-(4-fluorophenyl)-lH-indole (1.0 g, 4.76 mmol) and methyl 3,3-dimethoxypropanoate (0.81 mL, 5.7 mmol) were suspended in dichloromethane (15 mL). Trifluoroacetic acid (2.00 mL, 26 mmol) was added rapidly via syringe, resulting in a clear brown solution. The reaction mixture was heated to 40 °C for three hours. The reaction was diluted with dichloromethane (15 mL) to give an amber solution which was washed with saturated aqueous NaHCCh (25 mL) to yield a bright yellow/light amber biphasic mixture. The phases were separated and the organic layer was washed with saturated NaHCCh (30 mL), then dried (MgSCh) and filtered. The mixture was concentrated under a nitrogen stream overnight. The crude product was obtained as a yellow powder. The product was dissolved in minimum 2-MeTHF and pentane added until the suspension became lightly cloudy. The suspension was allowed to stand overnight, and the precipitate was filtered off. The filter cake was washed with heptane (2 x 15 mL), and dried in vacuo at 40 °C to afford the product as a yellow powder. Methyl (E)-3-[2-(4-fluorophenyl)-lH-indol-3-yl]prop-2-enoate (1.30 g, 86 %). ¾ NMR (300 MHz, Chloroform -if) d 8.41 (s, 1H), 8.01 – 7.95 (m, 1H), 7.92 (d, J = 16.0 Hz,

1H), 7.58 – 7.50 (m, 2H), 7.46 – 7.41 (m, 1H), 7.33 – 7.27 (m, 2H), 7.22 (t, J = 8.6 Hz, 2H), 6.59 (d, J = 16.0 Hz, 1H), 3.79 (s, 3H). LCMS m/z 295.97 [M+H]⁺.

Step 3. Synthesis of methyl 3-[2-(4-fluorophenyl)-lH-indol-3-yl]propanoate (CIOO)

[00403] To a solution of methyl (E)-3-[2-(4-fluorophenyl)-lH-indol-3-yl]prop-2-enoate (7 g, 0.02 mol) in EtOAc (350 mL) was added Palladium on carbon (4 g, 10 %w/w, 0.004 mol) and stirred at room temperature for 2 h under an atmosphere of H2 (bladder pressure). The reaction mixture was filtered through a pad of Celite® and washed with EtOAc (400 mL). The filtrates was concentrated to afford methyl 3-[2-(4-fluorophenyl)-lH-indol-3-yl]propanoate (7.1 g, 100 %). ¹H MR (300 MHz, DMSO-<fc) 5 11.2 (s, 1H), 7.65 (q, J = 5.4 Hz, 2H), 7.54 (d, J = 8.1 Hz, 1H), 7.36 (t, J = 9.0 Hz, 3H), 7.10 (t, J = 8.1 Hz, 1H), 7.02 (t, J = 7.8 Hz, 1H), 3.53 (s, 3H), 3.10 (t, J = 15.9 Hz, 2H), 2.63 (t, J = 15.9 Hz, 2H). LCMS m/z 298.21 [M+H]⁺. The product was used directly in the subsequent step without further purification.

Step 4. Synthesis of 3-[2-(4-fluorophenyl)-lH-indol-3-yl]propanoic acid (C101)

[00404] To stirred solution of methyl 3-[2-(4-fluorophenyl)-lH-indol-3-yl]propanoate (14.4 g, 0.05mol) in THF (300 mL), MeOH (300 mL) and H2O (250 mL) was cooled to -10°C. LiOH.H20 (10.1 g, 0.24 mol) was slowly added in a portion-wise manner. The reaction mixture was allowed to stir at room temperature for 16 h. The mixture was

evaporated and ice cold water (200 mL) was added, pH was adjusted to pH- 2 with 1M HC1 (400 mL, Cold solution). The mixture was stirred for 10 minutes, filtered and dried to afford 3-[2-(4-fhiorophenyl)-lH-indol-3-yl]propanoic acid (12.9 g, 94 %). ‘H NMR (400 MHz, DMSCMJ) 5 12.11 (s, 1H), 11.18 (s, 1H), 7.65 (q, J = 5.2 Hz, 2H), 7.56 (d, J = 7.6 Hz, 1H), 7.36 (t, J = 8.8 Hz, 3H), 7.10 (t, J = 8 Hz, 1H), 7.01 (t, J = 8 Hz, 1H), 3.06 (t, J = 16.4 Hz, 2H), 2.55 (t, J = 16 Hz, 2H). LCMS m/z 284.21 [M+H]⁺.

Step 5. Synthesis of 3-[2-(4-fluorophenyl)-lH-indol-3-yl]-N-[(3S,4R)-4-hydroxy-2- oxo-pyrrolidin-3-yl ] propanamide (87)

[00405] A mixture of 3-[2-(4-fluorophenyl)-lH-indol-3-yl]propanoic acid C101 (40 g, 120.0 mmol) and (3S,4R)-3-amino-4-hydroxy-pyrrolidin-2-one (Hydrochloride salt) S2 (23.8 g, 156.0 mmol) in DMF (270 mL) was stirred at room temperature for 5 minutes. CDMT (27.2 g, 154.9 mmol) and NMM (53 mL, 482.1 mmol) were added and the mixture was stirred at room temperature for 2 h. The mixture was poured into water (140 mL) and then stirred for 1 h at room temperature, then filtered and washing the solids with water (50 mL). The solids were dissolved in 1 : 1 IP A/water (-400 mL, until all solids dissolved) with heating (reflux) and stirring. The mixture was allowed to cool slowly to room temperature overnight. The mixture was cooled to 0 oC and stirred to break up crystals for filtration. The crystals were then filtered off, rinsed with cold 1 : 1 IP A/water to afford a tan solid (45 g). The solid was dissolved in IPA (200 mL) and heated to 80 °C to dissolve the solid. Activated charcoal (10 g) was added and the mixture was heated with stirring for 30 minutes. The mixture was filtered through Celite ® and solvent removed under reduced pressure. A mixture of 40:60 IP A/water (350 mL) was added to the solid and the mixture was heated until all solids dissolved. The mixture was cooled to room temperature over 5 h. Solids precipitated within the mixture. The mixture was then cooled to 0 °C and stirred for 1 h. The solids were filtered off and air dried on funnel for 1 h, then in a vacuum at 55 °C overnight to afford the product. 3-[2-(4-fluorophenyl)-lH-indol-3-yl]-N-[(3S,4R)-4-hydroxy-2-oxo-pyrrolidin-3-yl]propanamide (36.6 g, 79 %). ¾ NMR (300 MHz, Methanol-i¾) d 7.63 (ddt, J= 8.6, 5.1, 2.7 Hz, 3H), 7.35 (dt, J= 8.1, 1.0 Hz, 1H), 7.25 – 7.16 (m, 2H), 7.11 (ddd, J= 8.1, 7.0, 1.3 Hz, 1H), 7.03 (ddd, J = 8.0, 7.0, 1.2 Hz, 1H), 4.34 (td, J= 7.6, 6.8 Hz, 1H), 4.22 (d, J= 7.7 Hz, 1H), 3.55 (dd, J= 9.9, 7.5 Hz, 1H), 3.26 – 3.18 (m, 2H), 3.10 (dd, J= 9.9, 6.8 Hz, 1H), 2.69 – 2.59 (m, 2H). LCMS m/z 382.05 [M+H]⁺. The

product contained 0.23 % IPA by weight by NMR (1439 ppm IPA by residual solvent analysis). Purity is 99.5 % by (qNMR).

Alternative Preparation II of Compound 87 ( Indole Preparation route D)

Step 1. Synthesis of 5-(4-fluorophenyl)-5-oxo-pentanoic acid (Cl 04)

[00406] To a stirred suspension of AlCb(13.9 g, 0.10 mol) in dichloromethane (50 mL) was added a solution of tetrahydropyran-2,6-dione (5.93 g, 0.05

mol) in dichloromethane (100 mL) at 0 °C over a period of 15 minutes and stirred for 30 min. Then to the reaction mixture was added fluorobenzene (5 g, 0.05 mol) at 0 °C over a period of 15 min, gradually allowed to room temperature and stirred for 16 h. Then the reaction mixture was added to ice water (50 mL) under stirring. The resulting solid was filtered to afford a light yellow solid. The solid was diluted with 3 % NaOH solution (50 mL) and dichloromethane (50 mL). The aqueous layer was separated and acidified with IN HC1 at 0 °C. The mixture was then extracted with EtOAc (100 mL), dried over Na2SC>4, and concentrated under reduced pressure. The solid was then washed with pentane and dried to afford 5-(4-fluorophenyl)-5-oxo-pentanoic acid as an off white solid. (6 g, 53 %). ¾ NMR (300 MHz, DMSO-^) d 12.07 (s, 1H), 8.06 (d, J = 6 Hz, 1H), 8.02 (d, J = 5.4 Hz, 1H), 7.36 (t, J = 8.7 Hz, 2H), 3.06 (t, J = 12 Hz,

2H), 2.31 (t, J = 7.2 Hz, 2H), 1.86-1.78 (m, 2H). LCMS m/z 211.18 [M+H]⁺.

Step 2. Synthesis of 3-[2-(4-fluorophenyl)-lH-indol-3-yl]propanoic acid (Cl 01) [00407] Phenylhydrazine (Hydrochloride salt) (375.7 g, 2.6 mol) was combined with the 5-(4-fluorophenyl)-5-oxo-pentanoic acid (507.7 g, 2.4 mol) in a 12 L three-necked round-bottomed flask equipped with an overhead stirrer, temperature probe, and reflux condenser. AcOH (5 L) was added. The stirring was initiated and ZnCk (605 g, 4.44 mol) was added. The white suspension rapidly thickened after a few minutes (due to formation of the hydrazine intermediate). Approx. 500 mL of extra AcOH was added to aid stirring. The reaction was then heated to 100 °C for three hours. The reaction was cooled to room temperature and poured into water (approx. 6 L). The mixture was extracted with EtOAc (approx 8 L). The extract was washed with water, dried

(MgS04), filtered, and evaporated in vacuo to afford a golden yellow solid. The solid was triturated with approx. 4 L of 10 % EtOAc/DCM and filtered. The filter cake was washed with 50 % dichloromethane/heptane (approx 1 L). The filter cake was dissolved in 40 % EtOAc/dichloromethane (approx. 2L) and filtered over a plug of silica gel. The plug was eluted with 40 % EtOAc/ dichloromethane until the product had been eluted (checked by TLC (25 % EtOAc/ dichloromethane)). The filtrate was evaporated in vacuo to afford 382.6 g of an off-white solid (Crop 1). All filtrates were combined and evaporated in vacuo. The remaining solid was dissolved in 10 %

EtOAc/dichloromethane (approx. 1 L) and chromatographed on a 3 kg silica gel cartridge on the ISCO Torrent (isocratic gradient of 10 % EtOAc/dichloromethane). Product fractions were combined and evaporated in vacuo to afford a yellow solid that was slurried with dichloromethane, cooled under a stream of nitrogen, and filtered. The filter cake was washed with 50 % dichloromethane/heptane and dried in vacuo to afford 244.2 g of product (Crop 2). Altogether, both crops afforded 3-[2-(4-fluorophenyl)-lH-indol-3-yl]propanoic acid (626.8 g, 93 %). ¾ NMR (300 MHz, DMSO-i/e) d 12.15 (s, 1H), 11.20 (s, 1H), 7.74 – 7.62 (m, 2H), 7.57 (d, J = 7.8 Hz, 1H), 7.47 – 7.28 (m, 3H), 7.11 (ddd, J = 8.1, 7.0, 1.2 Hz, 1H), 7.02 (ddd, J = 7.9, 7.0, 1.1 Hz, 1H), 3.17 – 2.85 (m, 2H), 2.61 – 2.52 (m, 2H) ppm. ¹⁹F NMR (282 MHz, DMSO-i/e) d -114.53 ppm. LCMS m/z 284.15 [M+H]⁺.

Step 3. Synthesis of 3-[2-(4-fluorophenyl)-lH-indol-3-yl]-N-[(3S,4R)-4-hydroxy-2- oxo-pyrrolidin-3-yl ] propanamide (87)

[00408] A 3-L three neck RBF under nitrogen was equipped with a 150 mL addition funnel and thermocouple, then loaded with 3-[2-(4-fluorophenyl)-lH-indol-3-yl]propanoic acid (77.2 g, 228.6 mmol), (3S,4R)-3-amino-4-hydroxy-pyrrolidin-2-one

(Hydrochloride salt) (36.6 g, 239.9 mmol) and CDMT (44.2 g, 251.7 mmol). DMF (320 mL) was added and the orange slurry was cooled to -5 °C (acetone/brine/dry ice). NMM (88 mL, 800.4 mmol) was added via a funnel over 75 minutes to keep the internal temp <0 °C. The slurry was stirred at between -10 and 0 °C for 1 hour, then allowed to warm to ambient temperature progressively over 2 hours. Additional reagents were added (10 % of the initial quantities), and the mixture was stirred overnight at ambient temperature. Water (850 mL) was added over 60 minutes, maintaining the internal temperature at <25 °C (ice bath). This slow water addition allows for complete dissolution of any visible salt before precipitation of the product. The resulting thick slurry was stirred at ambient temperature overnight. The solid was recovered by filtration and washed with water (3 x 500 mL). The solid was dried under a stream of air at ambient temperature, then purified by crystallization.

Crystallization of 3- [2-( 4-fluorophenyl)-lH-indol-3-yl ]-N-[ ( 3S, 4R)-4-hydroxy-2-oxo- pyrrolidin-3-yl ] propanamide (87)

[00409] Under nitrogen atmosphere, a 2-L, 3 -neck flask equipped with addition funnel and thermocouple was charged with a light brown suspension of the crude 3-[2-(4-fluorophenyl)-lH-indol-3-yl]-N-[(3S,4R)-4-hydroxy-2-oxo-pyrrolidin-3-yljpropanamide (89.5 g) in IPA (225 mL, 2.5 vol). The slurry was heated to 50 °C and water (675 mL, 7.5 vol) was added until near-complete dissolution of solid was observed. The temperature was adjusted to 70 °C-to achieve full dissolution, yielding a clear amber solution. After 30 minutes, the heat source was removed and the mixture was cooled to ambient temperature over the weekend, stirring gently while maintaining the nitrogen atmosphere. The solid was recovered by filtration, washed with IPA:H20 = 1 :2 (2 x 300 mL, 2 x 3.3 vol) dried under a stream of air overnight to afford the product. 3-[2-(4-fluorophenyl)-lH-indol-3-yl]-N-[(3S,4R)-4-hydroxy-2-oxo-pyrrolidin-3-yl]propanamide (84.8 g, 92 %). ¾ NMR (300 MHz, DMSO-^) d 11.19 (s, 1H), 8.23 (d, J= 7.5 Hz, 1H), 7.77 (s, 1H), 7.72 – 7.63 (m, 2H), 7.60 (d, J= 7.8 Hz, 1H), 7.41 -7.31 (m, 3H), 7.12 (ddd, J= 8.1, 7.0, 1.2 Hz, 1H), 7.03 (ddd, J= 8.0, 7.0, 1.1 Hz, 1H), 5.49 (d, J= 5.0 Hz, 1H), 4.20 – 4.06 (m, 2H), 3.38 (s, 1H), 3.11 – 3.00 (m, 2H), 2.92 (dd, J= 9.4, 6.6 Hz, 1H). LCMS m/z 382.15 [M+H]⁺.

Crystallization of 3- [2-( 4-fluorophenyl)-lH-indol-3-yl J-N-[ ( 3S, 4R)-4-hydroxy-2-oxo- pyrrolidin-3-yl ] propanamide (87)

[00410] A 2-L, 3-neck flask equipped with addition funnel and thermocouple was charged with a light brown suspension of the crude 3-[2-(4-fluorophenyl)-lH-indol-3- yl]-N-[(3S,4R)-4-hydroxy-2-oxo-pyrrolidin-3-yl]propanamide in IPA (225 mL, 1 vol). The slurry was heated to 50 °C and water (675 mL, 3 vol) was added until near- complete dissolution of solid observed (mL). Temperature was increased to 70 °C under nitrogen (full dissolution, yielding a clear amber solution). After 30 minutes, the heat was removed and the mixture cooled to ambient temperature over the weekend, stirring gently under nitrogen atmosphere. The solid was recovered by filtration and washed with IPAiLLO = 1 :2 (2 x 300 mL).The solid was dried under a stream of air overnight to afford the product. 3-[2-(4-fluorophenyl)-lH-indol-3-yl]-N-[(3S,4R)-4-hydroxy-2-oxo- pyrrolidin-3-yl]propanamide (84.8 g, 92 %). ¾ NMR (300 MHz, DMSO-i/e) d 11.19 (s, 1H), 8.23 (d, J= 7.5 Hz, 1H), 7.77 (s, 1H), 7.72 – 7.63 (m, 2H), 7.60 (d, J= 7.8 Hz,

1H), 7.41 – 7.31 (m, 3H), 7.12 (ddd, J= 8.1, 7.0, 1.2 Hz, 1H), 7.03 (ddd, 7= 8.0, 7.0,

1.1 Hz, 1H), 5.49 (d, J= 5.0 Hz, 1H), 4.20 – 4.06 (m, 2H), 3.38 (s, 1H), 3.11 – 3.00 (m, 2H), 2.92 (dd, J= 9.4, 6.6 Hz, 1H). LCMS m/z 382.15 [M+H]⁺.

Large Scale Preparation of Compound 87

/- PrOAc solvate Form A

Step 1. Synthesis of 3-[2-(4-fluorophenyl)-lH-indol-3-yl]propanoic acid (C101)

[00411] To a mixture of C104 (100.0 g, 1.0 equiv) and phenyl hydrazine hydrochloride (72.2 g, 1.05 eqiv) was charged AcOH (800 mL, 8 vol). The mixture was agitated and heated to 85 °C for 16 hours. The batch was cooled to 22 °C. A vacuum was applied and the batch distill at <70°C to ~3 total volumes. The batch was cooled to 19- 25 °C. The reactor was charged with iPrOAc (800 mL, 8 vol) and then charged with water (800 mL, 8 vol). The internal temperature was adjusted to 20 – 25 °C and the biphasic mixture was stirred for no less than 0.5 h. Stirring was stopped and the phases allowed to separate for no less than 0.5 h. The lower aqueous layer was removed. 1 N HC1 (500 mL, 5 vol) was charged to the reactor. The internal temperature was adjusted to 20 – 25 °C, and the biphasic mixture was stirred for no less than 0.5 h. Stirring was stopped and phases were allowed to separate for no less than 0.5 h. The lower aqueous layer was removed. The reactor was charged with 1 N HC1 (500 mL, 5 vol). The internal temperature was adjusted to 20 – 25 °C, and the biphasic mixture was stirred for no less than 0.5 h.

Stirring was stopped and phases were allowed to separate for no less than 0.5 h. The lower aqueous layer was removed. Water (500 mL, 5 vol) was charged to the reactor.

The internal temperature was adjusted to 20 – 25 °C, and the biphasic mixture was stirred for no less than 0.5 h. Stirring was stopped and phases were allowed to separate for no less than 0.5 h. The lower aqueous layer was removed. Water (500 mL, 5 vol) was charged to the reactor. The internal temperature was adjusted to 20 – 25 °C, and the biphasic mixture was stirred for no less than 0.5 h. Stirring was stopped and phases were allowed to separate for no less than 0.5 h. The lower aqueous layer was removed. The organic phase was distilled under vacuum at <75 °C to 3 total volumes. The reactor was charged with toluene (1000 mL, 10 vol). The organic phase was distilled under vacuum at <75 °C to 5 total volumes. The reactor was charged with toluene (1000 mL, 10 vol). The organic phase was distilled under vacuum at <75 °C to 5 total volumes. The resulting slurry was heated to an internal temperature of 85 °C until complete dissolution of solids was achieved. The mixture was allowed to stir for 0.5 h at 85 °C and then cooled to an internal temperature of 19 – 25 °C over 5 h. The mixture was allowed to stir at 25 °C for no less than 2 h. The slurry was filtered. The filter cake was washed with toluene (1 x 2 vol (200 mL) and 1 x 1.5 vol (150 mL)). The solids were dried under vacuum with nitrogen bleed at 60 °C to afford product C101 (95.03 g, 70%).

Purification of Compound 87 by Recrystallization to Form A

[00412] Compound 87 as an iPrOAc solvate (17.16 g after correction for iPrOAc content, 1.0 equiv) was charged to a reactor. A mixture of IP A (77 mL, 4.5 vol) and water (137 mL, 8 vol) were charged to the reactor. The slurry was heated to an internal temperature of 75 °C. The batch was cooled to an internal temperature of 25 °C over 10 h and then stirred at 25 °C for at least 12 h. The slurry was filtered. The filter cake was washed with 36/64 IP A/water (2 x 52 mL, 2 x 3 vol). The solids were dried under vacuum with nitrogen bleed at 60 °C to afford Compound 87 as a neat, crystalline form (Form A, 15.35 g, 89%).

Synthetic Procedure

[00413] A mixture of 3-[2-(4-fluorophenyl)-lH-indol-3-yl]propanoic acid C101 (50 g, 1.0 equiv), S2 hydrochloride (28.3 g, 1.05 equiv), and CDMT (34.1 g, 1.1 equiv) was charged with 2-MeTHF (200 mL, 4 vol) and DMF (50 mL, 1 vol) and the mixture was agitated. The internal temperature adjusted to <13 °C. The reactor was charged with NMM (64.5 g, 3.5 equiv) over 1 h, while maintaining internal temperature <20 °C. The internal temperature was adjusted to 25 °C and the batch was stirred at that temperature for 14 h. The batch was cooled to 10 °C and charged with water (250 mL, 5 vol) while keeping the internal temperature <20 °C. The batch was then warmed to 20 – 25 °C. Stirring was stopped, and the phases allowed to separate for 10 min. The lower aqueous phase was removed. The aqueous layer was back extracted with 2-MeTHF (2 x 200 mL, 2 x 4 vol) at 20 – 25 °C. The combined organic phases were washed with 1 N HC1 (500 mL, 10 vol) at 20 – 25 °C by mixing for 10 min and settling for 10 min. The lower aqueous phase was removed. The organic phases were washed with 0.25 N HC1 (2 x 250 mL, 2 x 5 vol) at 20 – 25 °C by mixing for 10 min and settling for 10 min for each wash. Lower aqueous phases were removed after each wash. The organic phase was washed with water (250 mL, 5 vol) at 20 – 25 °C by mixing for 10 min and settling for 10 min. The reactor was charged with 20 wt % Nuchar RGC® and stirred for 4 h. The reaction mixture was filtered through a pad of celite®. The reactor and celite® pad were rinsed with 2-MeTHF. The combined organics were distilled under vacuum at <50 °C to 5 total volumes. The reactor was charged with iPrOAc (500 mL, 10 vol). The organic phase was distilled under vacuum at <50 °C to 5 total volumes. The mixture was charged with additional iPrOAc (400 mL, 8 vol) and distillation under vacuum was repeated. The mixture was charged with additional iPrOAc (250 mL, 5 vol), heated to an internal

temperature of 75 °C and stirred for 5 h. The slurry was cooled to 25 °C, over 5 h and stirred for no less than 12 h. The slurry was filtered and the filter cake washed with iPrOAc (2 x 50 mL, 2 x 1 vol). The solids were dried under vacuum with nitrogen bleed at 55 – 60 °C to afford Compound 87 as an iPrOAc solvate (60.38 g including 9.9% w/w iPrOAc, 80.8% yield).

Form A of Compound 87

[00414] Compound 87 hydrate form was converted to the dehydrated, neat crystalline form (Form A) after drying.

Hydrate Form A of Compound 87

[00415] A mixture of IP A (4.5 vol) and water (8 vol) was added to compound 87

(iPrOAc solvate containing ~2.5 – 11 wt% iPrOAc, 1.0 equiv). The slurry was heated to an internal temperature of 75 °C and filtered hot. The filtrate was cooled to 25 °C for at least 12 h. The slurry was filtered. The filter cake was washed with 36/64 IP A/water (2 x 3 vol). The solids were dried under vacuum with nitrogen bleed at 55 – 60 °C. The product was isolated as Hydrate form.

IPAC Solvate of Compound 87:

[00416] The large scale synthesis described above provided an iPrOAc solvate containing ~2.5 – 11 wt% iPrOAc after drying.

Amorphous Form of Compound 87

[00417] ~lg of compound 87 was dissolved in 22mL of acetone. The solution was evaporated using a Genevac. The resulted solid was dried at 60C under vacuum overnight. The dried solid was amorphous form.

///////////

O=C(N[C@@H]1C(=O)NC[C@H]1O)CCc1c2ccccc2[NH]c1c1ccc(F)cc1

SIMILAR

predicted

VX 147

cas 2446816-88-0 predicted

O=C(N[C@@H]1C(=O)NC[C@H]1O)CCc1c2cc(F)cc(F)c2[NH]c1c1ccc(F)cc1

• OriginatorVertex Pharmaceuticals
• ClassSmall molecules; Urologics
• Mechanism of ActionApolipoprotein L1 inhibitors

• Orphan Drug StatusNo
• New Molecular EntityYes

Highest Development Phases

• Phase IIFocal segmental glomerulosclerosis
• Phase IKidney disorders

Most Recent Events

• 14 Apr 2020Phase-II clinical trials in Focal segmental glomerulosclerosis in USA (PO) (EudraCT2020-000185-42) (NCT04340362)
• 31 Dec 2019Vertex Pharmaceuticals completes phase I clinical trial in Focal segmental glomerulosclerosis and Kidney disorders (In volunteers) in USA (PO)
• 05 Aug 2019Vertex Pharmaceuticals plans a phase II proof-of-concept trial for focal segmental glomerulosclerosis in 2020

VX 148

297730-05-3

Name: VX-148
CAS#: 297730-05-3
Chemical Formula: C23H25N5O4
Exact Mass: 435.19065
Molecular Weight: 435.48
Elemental Analysis: C, 63.44; H, 5.79; N, 16.08; O, 14.70

• OriginatorVertex Pharmaceuticals
• ClassAntipsoriatics
• Mechanism of ActionInosine monophosphate dehydrogenase inhibitors
• DiscontinuedPsoriasis; Transplant rejection; Viral infections

• 13 Nov 2003Interim data from a media release have been added to the adverse events and Skin Disorders therapeutic trials sections
• 23 May 2003Vertex Pharmaceuticals has completed enrolment in a phase IIa trial for Psoriasis in Iceland
• 24 Dec 2002Phase-II clinical trials in Psoriasis in Iceland (unspecified route)

VX-148 is a second-generation, orally administered inhibitor of inosine monophosphate dehydrogenase (IMPDH). The IMPDH enzyme plays a key role in regulating immune response and proliferation of specific cell types, including lymphocytes. VX-148 is a developed for the treatment of autoimmune diseases.

Investigated for use/treatment in autoimmune diseases, psoriasis and psoriatic disorders, and viral infection.

VX-148 is a novel, uncompetitive IMPDH inhibitor with a K(i) value of 6 nM against IMPDH type II enzyme. VX-148 is slightly more potent than mycophenolic acid and VX-497 in inhibiting the proliferation of mitogen-stimulated primary human lymphocytes (IC(50) value of ~80 nM). The inhibitory activity of VX-148 is alleviated in the presence of exogenous guanosine. VX-148 does not inhibit proliferation of nonlymphoid cell types such as fibroblasts, indicating selectivity for inhibition of IMPDH activity. VX-148 is orally bioavailable in rats and mice; oral administration of VX-148 inhibits primary antibody response in mice in a dose-dependent manner with an ED(50) value of 38 mg/kg b.i.d. VX-148 significantly prolongs skin graft survival at 100 mg/kg b.i.d. in mice.

SYN

WO 0056331

The intermediate carbamate (V) has been obtained as follows. The reaction of 4-bromo-3-methoxynitrobenzene (I) with CuCN in NMP at 150 C gives 2-methoxy-4-nitrobenzonitrile (II), which is reduced with H2 over Pd/C in ethyl acetate to yield 4-amino-2-methoxybenzonitrile (III). Finally, this compound is condensed with phenyl carbamate (IV) by means of NaHCO3 in ethyl acetate to afford the desired carbamate intermediate (V).

SYN

The reduction of 3-nitroacetophenone (VI) by means of NaBH4 in ethanol gives 1-(3-nitrophenyl)ethanol (VII), which is treated with DPPA and DBU in hot toluene to yield the azido derivative (VIII). The reduction of (VIII) with PPh3 in THF/water affords 1-(3-nitrophenyl)ethylamine (IX) as a racemic mixture that is submitted to optical resolution with L-(+)-tartaric acid to provide the desired (S)-isomer (X). The reduction of the nitro group of (X) by means of H2 over Pd/C in methanol gives 1(S)-(3-aminophenyl)ethylamine (XI), which is condensed with 2(R)-hydroxypentanenitrile (XII) and CDI to yield the carbamate (XIII). Finally, this compound is condensed with intermediate carbamate (V) by means of TEA in hot ethyl acetate to afford the target urea.

1. Jain J, Almquist SJ, Heiser AD, Shlyakhter D, Leon E, Memmott C, Moody CS, Nimmesgern E, Decker C: Characterization of pharmacological efficacy of VX-148, a new, potent immunosuppressive inosine 5′-monophosphate dehydrogenase inhibitor. J Pharmacol Exp Ther. 2002 Sep;302(3):1272-7. [Article]

////////////VX 148, phase 2

O=C(O[C@H](CC)CC#N)N[C@H](C1=CC=CC(NC(NC2=CC=C(C#N)C(OC)=C2)=O)=C1)C

NEW DRUG APPROVALS

one time

$10.00

Click here to purchase.

Pralnacasan

VX 740

cas 192755-52-5

(4S,7S)-N-[(2R,3S)-2-ethoxy-5-oxooxolan-3-yl]-7-(isoquinoline-1-carbonylamino)-6,10-dioxo-2,3,4,7,8,9-hexahydro-1H-pyridazino[1,2-a]diazepine-4-carboxamide

N-[(4S,7S)-4-{[(2R,3S)-2-ethoxy-5-oxooxolan-3-yl]carbamoyl}-6,10-dioxo-octahydro-1H-pyridazino[1,2-a][1,2]diazepin-7-yl]isoquinoline-1-carboxamide

 (1S,9S)-N-((2R,3S)-2-Ethoxy-5-oxotetrahydrofuran-3-yl)-9-((isoquinolin-1-ylcarbonyl)amino)-6,10-dioxooctahydro-6-H-pyridazino(1,2-a)(1,2)diazepine-1-carboxamide

6H-Pyridazino(1,2-a)(1,2)diazepine-1-carboxamide, N-((2R,3S)-2-ethoxytetrahydro-5-oxo-3-furanyl)octahydro-9-((1-isoquinolinylcarbonyl)amino)-6,10-dioxo-, (1S,9S)-

• HMR 3480
• HMR3480
• HMR3480/VX-740
• Pralnacasan
• UNII-N986NI319S
• VX 470
• VX-740

C26H29N5O7, 523.543

NSAID, ICE inhibitor & metastasis inhibitor.пралнаказан [Russian] [INN]برالناكاسان [Arabic] [INN]普那卡生 [Chinese] [INN]

Pralnacasan is an orally bioavailable pro-drug of a potent, non-peptide inhibitor of interleukin-1beta converting enzyme (ICE).Pralnacasan is a potent, non-peptide inhibitor of interleukin-1beta converting enzyme (ICE, aka Caspase-1). It was originally discovered by Vertex Pharmaceuticals and licensed for development to Aventis Pharma. In 2003 Aventis and Vertex Pharmaceuticals agreed to voluntarily discontinue development based on results from a 9-month animal toxicity trial that showed liver abnormalities due to chronic high doses of pralnacasan. Pralnacasan has also been investigated for the treatment of Partial Epilepsy; advancing to Phase II clinical trials.Pralnacasan is a potent, non-peptide inhibitor of interleukin-1beta converting enzyme (ICE). Pralnacasan is an oral, anti-cytokine drug candidate licensed for development by Aventis Pharma from Vertex Pharmaceuticals. In November 2003, Aventis and Vertex Pharmaceuticals announced that they had voluntarily suspended the phase II clinical trials of pralnacasan due to results from an animal toxicity study that demonstrated liver abnormalities after a nine-month exposure to pralnacasan at high doses. While no similar liver toxicity has been seen to date in human trials, the companies will evaluate the animal toxicity results before proceeding with the phase II clinical program.Pralnacasan inhibits interleukin-1beta converting enzyme (ICE), an enzyme that regulates the production of IL-1 and IFN gamma – intercellular mediators that initiate and sustain the process of inflammation. Inhibiting ICE may be an effective strategy for curtailing damaging inflammatory processes common to a number of acute and chronic conditions, such as rheumatoid arthritis (RA) and osteoarthritis. 
PAPERhttps://pubs.rsc.org/en/content/articlelanding/2017/ob/c7ob01403a/unauth
IDrugs (2003), 6(2), 154-158. 
Chemistry (Weinheim an der Bergstrasse, Germany) (2017), 23(2), 360-369PAPER 
Bioorganic & Medicinal Chemistry Letters (2006), 16(16), 4233-4236.https://www.sciencedirect.com/science/article/abs/pii/S0960894X06006184?

Abstract

Novel 1-(2-acylhydrazinocarbonyl)cycloalkyl carboxamides were designed as peptidomimetic inhibitors of interleukin-1β converting enzyme (ICE). A short synthesis was developed and moderately potent ICE inhibitors were identified (IC₅₀ values <100 nM). Most of the synthesized examples were selective for ICE versus the related cysteine proteases caspase-3 and caspase-8, although several dual-acting inhibitors of ICE and caspase-8 were identified. Several of the more potent ICE inhibitors were also shown to inhibit IL-1β production in a whole cell assay (IC₅₀ < 500 nM).

Graphical abstract

Novel 1-(2-acylhydrazinocarbonyl)cycloalkyl carboxamides were designed and synthesized as selective peptidomimetic inhibitors of interleukin-1β converting enzyme (ICE IC₅₀ values <100 nM).

PAPEROrganic letters (2014), 16(13), 3488-91.https://pubs.acs.org/doi/10.1021/ol501425b

Abstract

Peptides containing N2-acyl piperazic or 1,6-dehydropiperazic acids can be formed efficiently via a novel multicomponent reaction of 1,4,5,6-tetrahydropyridazines, isocyanides, and carboxylic acids. Remarkably, the reaction’s induced intramolecularity can enable the regiospecific formation of products with N2-acyl piperazic acid, which counters the intrinsic and troublesome propensity for piperazic acids to react at N1 in acylations. The utility of the methodology is demonstrated in the synthesis of the bicyclic core of the interleukin-1β converting enzyme inhibitor, Pralnacasan.
PatentWO 9722619WO 9903852WO 9952935
PATENTWO 2000042061https://patents.google.com/patent/WO2000042061A1/enThe invention particularly relates to the process as defined above in which the compound of formula (I) is 9- (1, 3-dihydro-1,3, dioxo-2H-isoindol-2-yl) -3 ,, 7, 8, 9, 10-hexahydro-6, 10-dioxo-6H-pyridazino- [1,2- a] [1, 2] ethyl diazepine-1-carboxylate:

The invention particularly relates to the process as defined above in which the compound of formula (Iopt) is- (lS-cis) -9- (1, 3-dihydro-l, 3-dioxo-2H-isoindol-2-yl) – 3,4,7,8,9, 10-hexahydro-β, 10-dioxo -6H-pyridazino- [1,2- a] [1, 2] ethyl diazeρine-1-carboxylate:

The compounds of formula (I) can be generally used for the synthesis of medicaments as indicated in patent EP 94095. The compounds of formulas (II) and (III) and (F) are known and can be prepared according to the experimental method described below.The invention also relates to the application of the process as defined above as an intermediate step for the preparation of a compound of formula (V)

via the compound of formula (Iopt) as defined above, characterized in that this process comprises the steps of the process for the preparation of the compounds of formula (Iopt) from the compounds of formula (II) as defined above.The subject of the invention is also the application as defined above, characterized in that the compound of formula (Iopt) is (lS-cis) -9- (1, 3-dihydro-l, 3-dioxo -2H- isoindol-2-yl) -3,4,7,8,9, 10-hexahydro-6, 10-dioxo-6H- pyridazino- [1,2-a] [1, 2] diazepine-1- ethyl carboxylate

The subject of the invention is also the application of the process as defined above as an intermediate step in the overall process for preparing the compounds of formula (I) and (Iopt) as defined above. Finally, the subject of the invention is, as intermediate compound, the compound of formula (IA) as defined above.Preparation 1 Preparation of bis (phenylmethyl) 1,2-hydrazinecarboxylate1.5 liters of methanol and 25 g of 80% hydrazine monohydrate are placed under nitrogen. Cooled to 0 ° C and then introduced 75 g of benzyl chloroformate and a solution of 93 g of sodium carbonate in 1100 ml of demineralized water. Maintaining the reaction mixture for 1 hour at 0 ° C, drained and washed by displacement with a mixture of 100 ml of methanol and 100 ml of water, then washed by displacement with 500 ml of water at 0 C °. Dried and obtained 107.6 g of the desired product. Preparation 2Preparation of N-phthaloyl-L-glutamic anhydride D (+) 2-tetrahydro-2,6,6-dioxo-2H-pyran-3-yl-1H-isoindole-1,3 (2H) – dione (R)Stage a: N-phthaloyl-L-glutamic acid2- (1, 3-dihydro-1,3, dioxo-2H-isoindole-2-yl) acid – pentanedioic (2S)To a solution of 14.4 g of sodium carbonate in 180 ml of water is added 10 g of L-glutamic acid then 16 g of N-carbethoxyphthalimide (nefkens reagent, commercial). The mixture is stirred at ambient temperature for 2 hours and then extracted with ethyl acetate. The organic phase is evaporated under reduced pressure until a dry extract is obtained and 2.74 g of crude product is obtained. Washing is carried out with sodium bicarbonate, then after return to the acid and extraction with ethyl acetate, 370 mg of expected product and H ₂ N-C0 ₂ Et are isolated. Furthermore, the aqueous phase is brought to pH = 2 with 36% hydrochloric acid at a temperature below 5 ° C and then extracted with ethyl acetate, washed with a saturated chloride solution. sodium, dry, filter and concentrate under reduced pressure until 22.7 g of expected product is obtained in the form of an oil.Mass spectrum (MH) ^“ = 276 ^“ Infrared (Nujol):1775 cm ^“1 (m), 1720 cm ^{” 1} (F, complex): CO 1611 cm ^“1 : Aromatic Stage b:To the product obtained in stage a), 160 ml of tetrahydrofuran are added and 18.6 g of DCC (1, 3-Dicyclohexyl-carbodiimide) dissolved in 55 ml of tetrahydrofuran are added dropwise over 30 minutes. Stirred for 1 hour at 15-17 ° C, then filtered, rinsed with tetrahydrofuran, evaporated under reduced pressure until a dry extract is obtained which is taken up in isopropyl ether. After 30 minutes of stirring, the filter is washed and dried. 14.98 g of expected product are obtained. α _D = -52.63 ^λ H NMR (DMSO) 2.12 (m, 1H); 2.61 (m, 1H); 2.98 (dm, 1H); 3.16 (ddd, 1H); 5.48 (dd, 1H); 7.82 (m,> 4H)Example 1: (IS-cis) -9- (1, 3-dihydro-1,3, dioxo-2H-isoindol-2-yl) -3,4,7,8,9,10-hexahydro-6,10 -dioxo-6H-pyridazino- [1,2- a] [1,2] diazepine-1-ethyl carboxylate. Stage a: Preparation of 2,5-dibromopentanoic acid 39 ml of bromine are added to a mixture of 106 g of 5-bromopentanoic acid and 1 ml of phosphorus tribromide. The reaction mixture is brought to 70-80 ° C for 16 h 30. The reaction medium is brought to 100 ° C for 15 minutes and allowed to return to room temperature. 147 g of sought product is obtained.Stage b: Preparation of ethyl 2,5-dibromopentanoate24.37 g of oxalyl chloride are added to a mixture containing 50 g of the acid prepared in the preceding stage, 15 drops of dimethylformamide and 300 ml of dichloromethane. The reaction mixture is kept under stirring at at room temperature, until the reaction is complete. The reaction mixture is cooled to 10 ° C and 50 ml of ethyl alcohol are added. Stirred for 30 minutes at 10 ° C, allowed to return to room temperature and stirred for 3 hours at room temperature. It is brought to dryness and the desired product is obtained. Stage c: CyclizationPreparation of (S) -tetrahydro-1,2,3-pyridazinetricarboxylate of 3-ethyl-1,2-bis (phenylmethyl) and (R) -tetrahydro-1,2,3-pyridazinetricarboxylate of 1,2 -bis (phenylmethyl). A suspension of 12.1 g of ethyl 2,5-dibromopentanoate (stage b) in 50 ml of diglyme is introduced at 20-25 ° C. in a suspension containing 10.42 g of 1,2-hydrazine carboxylate of bis (phenylmethyl) (preparation 1), 65 ml of diglyme and 8.26 g of potassium carbonate. The suspension obtained is heated to 90 ° C. and stirring is continued for 48 hours. Cooled to 20 ° C, poured into a solution containing 50 ml of 2N hydrochloric acid and 150 ml of a mixture of water and ice. Extraction is carried out with ethyl acetate, washing with water and drying. It is filtered, rinsed with ethyl acetate and dried. Finally, the crude product is purified by chromatography on silica, eluting with a heptane / ethyl acetate mixture 40/20 and 10.71 g of sought product is obtained. Stage d: Acylation and hydrogenolysisPreparation of α, (IS) – [3-oxo-3- (tetrahydro-3-ethoxycarbonyl-1 (2H) -pyridazinyl) propyl] -1,3-dihydro-1,3-dioxo-2H-isoindole acid -2-aceticThe mixture consisting of 15g of tetrahydro-1,2,3-pyridazinetricarboxylate of 3-ethyl-1,2-bis (phenylmethyl) is placed under hydrogen pressure (1.3 bar) for 24 hours. R + S mixture as prepared in stage c 150 ml of tetrahydrofuran, 2.5 g of palladium on carbon (10%) and 9.08 g of phthaloylglutamic acid anhydride as prepared according to preparation 2. After filtration, we evaporated under reduced pressure until a dry extract is obtained which is taken up in 100 ml of ethyl acetate and 150 ml of a saturated solution of sodium bicarbonate. It is extracted 3 times and the bicarbonate solution is acidified to pH = 3 with 36% hydrochloric acid. It is extracted 3 times with dichloromethane and washed with water. 13.16 g of crude product are obtained, which product is purified by chromatography on silica, eluting with a toluene / ethyl acetate / acetic acid 20/80 / 1.5 mixture to obtain 12.7 g of the expected product.NMR (250MHz, CDC1 ₃ ): 1.24 (d, 3H, OCH ₂ CH ₃ ); 4.12 (q, 2H, OCH ₂ CH ₃ ); 4.36-4.40 (m, 1H, Hl in alpha or beta position); 4.69-4.92 (m, 1H, H9 in the alpha position); 7.70 – 7.86 H aromatic. Stage el: cyclization with POCl ₃– (lS-cis) -9- (1, 3-dihydro-l, 3-dioxo-2H-isoindol-2-yl) – 3,4,7,8,9, 10-hexahydro-6, 10-dioxo -6H-pyridazino- [1,2- a] [1, 2] ethyl diazepine-1-carboxylate. – (lR-trans) -9- (1, 3-dihydro-1,3, dioxo-2H-isoindol-2-yl) – 3,4,7,8, 9, 10-hexahydro-6,10-dioxo -6H-pyridazino- [1,2-a] [1,2] diazepine-1-ethyl carboxylate.To a solution of 20 ml of dichloroethane heated beforehand to 75 ° C., the following solutions A and B are added over 3 hours: A: 417 mg of the ester prepared in stage d in 4 ml of dichloroethane to which 1 ml of a solution of 1.2 ml of 2,6-lutidine in 5 ml of dichloroethane. B: 1 ml of a solution of 1.9 ml of P0Cl ₃ in 10 ml of dichloroethane, then the mixture is stirred for 1 hour at this temperature. Cool to 10 ° C., add demineralized water, extract with dichloromethane and evaporate under reduced pressure to obtain a crude product (0.415 g) which is purified by chromatography on silica eluting with the heptane / dichloromethane mixture. / ethyl acetate 1/1/1. 161.8 mg of the SS diastereoisomer, 126.7 mg of the SR diastereoisomer and 5.8 mg of the SS + SR mixture are isolated. Stage e2: cyclization with POBr ₃– (lS-cis) -9- (1, 3-dihydro-l, 3-dioxo-2H-isoindol-2-yl) – 3,4,7, 8, 9, 10-hexahydro-6, 10-dioxo -6H-pyridazino- [1, 2- a] [1, 2] ethyl diazepine-1-carboxylate. – (lR-trans) -9- (1, 3-dihydro-l, 3-dioxo-2H-isoindol-2-yl) – 3,4,7, 8, 9, 10-hexahydro-6, 10-dioxo -6H-pyridazino- [1, 2- a] [1, 2] ethyl diazepine-1-carboxylate.To a solution of 20 ml of dichloroethane heated beforehand to 80 ° C., the following solutions A and B are added over 3 hours:A: 417 mg of the ester prepared in stage d in 4 ml of dichloroethane to which 1 ml of a solution of 2.4 ml of 2,6-lutidine in 10 ml of dichloroethane was added. B: 1 ml of a solution of 5.85 g of POBr ₃ in 10 ml of dichloroethane, then the mixture is stirred for 1 hour at this temperature. Cool to 10 ° C, add demineralized water, extract with dichloromethane and evaporate under reduced pressure to obtain a crude product (0.419 g) which is purified by chromatography on silica eluting with the heptane / dichloromethane / mixture 1/1/1 ethyl acetate. 163 mg of the SS diastereoisomer, 143 mg of the SR diastereoisomer and 6.2 mg of the SS + SR mixture are isolated.Stage f: deracemization / epimerization – (lS-cis) -9- (1, 3-dihydro-l, 3-dioxo-2H-isoindol-2-yl) – 3,4,7,8, 9, 10-hexahydro -6,10-dioxo-6H-pyridazino- [1, 2- a] [1, 2] ethyl diazepine-1-carboxylate.Is introduced at a temperature of -45 / -48 ° C in one hour 30 minutes, a solution containing 0.029 g of potassium terbutylate and 0.3 ml of dimethylformamide in a mixture containing 0.194 g of the mixture SS + SR prepared in stage d , 1.5 ml of dimethylformamide and 0.75 ml of terbutanol. The mixture is kept stirring for 1 hour and, after cooling to -50 ° C., 0.4 g of powdered ammonium chloride is introduced. Stirred 10 minutes at -45 ° C, add 1 ml of ammonium chloride at 20 ° C and stirred again 10 minutes. 2 ml of water are added after 5 minutes demineralized. Extracted with ethyl acetate, washed with demineralized water, decanted, concentrated and dried. 0.166 g of expected SS diastereoisomer is obtained. ” _D = -75.3 ° (1% in methanol) NMR (250MHz, CDC1 ₃ ): 1.73 (m, 3H, H-2alpha H-3alpha H-3beta; 1.24 (d, 3H, OCH ₂ CH ₃ ); 2.38 (m, 3H, H2beta, H7alpha, H8 alpha); 2.92 (m, 1H, H4alpha); 3.39 – 3.44 (m, 1H, H8beta); 3.62 (m, 1H, H7beta); 4.23 (m, 2H, OCH ₂ CH ₃ ); 4.66-4.71 (m, 1H, H4 in beta position); 5.26-5.41 (m, 2H, Hl and H9 in the alpha position); 7.72 – 7.88 H aromatics. 
PATENT 
WO 2000010979https://patents.google.com/patent/WO2000010979A1/en

 formula II, said compound has the structure:

In the synthesis of these inhibitors, the terminal carbon of Ri adjacent the -COOH moiety contains a protecting substituent. Preferably that protecting

substituent is

The synthesis steps from compound H to the inhibitors set forth above involve removal of the protecting substituent on R_x; coupling of the R₅-NH- or R_5′-NH- moiety in its place; hydrolysis of the R₂ group;N .(CJ₂)_m.—Tand coupling of the amine ( ^(Ch,2)a ^Rs or -NH-Z)in its place. The removal of the protecting substituent on Ri is typically carried out with hydrazine. The subsequent coupling of the R₅-NH- or R_5′-NH- moiety is achieved with standard coupling reagents, such as EDC, DCC or acid chloride . Depending upon the nature of R₂, its hydrolysis may be achieved with an acid (when R₂ is t-butyl), a hydroxide (when R₂ is any other alkyl, alkenyl or alkynyl or Ar) or hydrogenolysis (when R₂ is an Ar-substituted alkyl, alkenyl or alkynyl) . This produces the corresponding acid from the ester.The acid is then coupled to the amine with standard coupling reagents, such as EDC, DCC or acid chloride .In order that this invention be more fully understood, the following examples are set forth. These examples are for the purpose of illustration only and are not to be construed as limiting the scope of the invention in any way. EXAMPLE 1Synthesis of a 7,6 Scaffold for a Caspase InhibitorA.

Compound A’ was dissolved m 5 equivalents of S0C1₂ and then heated to 80°C for 1 hour. The solution was then cooled to 50°C and 2 equivalents of bromine were added. The solution was incubated at 50°C for an additional 12 hours until the red color disappeared. We then cooled the solution to 10°C and added 4 volumes of water. The solution was then re-heated to 50°C for another hour. We then separated the organic and aqueous layer, washed the organic layer consecutively with water, Na₂S0 and then brme, removing the aqueous layer after each washing. The final organic layer was then isolated, dried over Na₂S0 and concentrated to produce compound B’ as an amber oil.B.

Compound B’ was treated with 1 equivalent of tert-butanol and 0.1 equivalents of 4- (dimethylammo) – pyπdme a solution of and the resulting solution cooled to 7°C. We then added a solution of 1 equivalent of DCC m toluene while maintaining reaction temperature at less than 22°C. The cooling bath was removed and the reaction was stirred at ambient temperature under a nitrogen atmosphere for 16 hours. The reaction mixture was then diluted with hexane and cooled to 9°C . The resulting solids were removed by filtration. The filtrate was washed consecutively with 0. IN HC1, water, and then sodium bicarbonate. The filtrate was then dried over sodium sulfate and concentrated in vacuo to afford compound C as a yellow oil.C.

Compound D’ was combined with 1.2 equivalents of compound C and dissolved in DMF at ambient temperature under nitrogen atmosphere. We then added granular sodium sulfate, 2.5 equivalents of LiOH monohydrate, and then 0.1 equivalents Bu₄NI to the resulting solution. The reaction temperature was maintained at between 20°C and 30°C and allowed to stir for 16 hours. The reaction mixture was then diluted with ethyl acetate and water and the layers separated. The organic layer was washed with water and then brine, dried over sodium sulfate and concentrated in vacuo to produce an amber oil. This oil was then dissolved in 5 volumes of ethanol at ambient temperature. We then added 2.5 volumes of water. The resulting mixture was allowed to stir until a white solid formed (approximately 5 hours) . The crystallized product was isolated via filtration then dried in vacuo to afford compound E’ as a white solid.D.

We dissolved compound E’ in THF. We then added, at ambient temperature under a nitrogen atmosphere, 0.02 equivalents of triethylamine and 0.01 equivalents of Pd(OAc)₂. A solution of 2.5 equivalents of triethylsilane (Et₃SiH) in THF was then added and the resulting black solution was allowed to stir for 16 hours to complete the reaction. We then added a saturated, aqueous solution of sodium bicarbonate followed by a solution of compound F’ in THF. After 30 minutes, the layers were separated and the aqueous layer acidified to pH 4.5 with aqueous citric acid. The product in the aqueous layer was then extracted into ethyl acetate. The organic layer was isolated, washed with brine, dried over sodium sulfate and concentrated in vacuo to produce a white foam. This crude product was then recrystallized from MTBE to afford compound G’ as a white powder. E.

Method #1:To a suspension of compound G’ and 0.1 equivalents of DMF m dichloroethane, at 70°C we added 5 equivalents of 2, 6-lutιdme simultaneously with 2.5 equivalents of S0C1₂ over a period of 3 hours. The reaction was then diluted with toluene and washed consecutively with NaHC0₃ and br e. The solution was then dried over Na₂S0₄ and concentrated in vacuo to afford compound H’ as a yellow solid.Method #2:To a suspension of compound G’ m dichloroethane, at 70°C, we added 4 equivalents of 2,6- lutid e followed by 2 equivalents of methanesulfonyl chloride. The resulting solution was stirred at 70°C for 12 hours. The reaction was then diluted with toluene and washed consecutively with NaHC0₃ and brme. The solution was then dried over Na₂S0₄ and concentrated in vacuo to afford compound H’ as a white solid. Method #2 produced a significantly higher yield of H’ as compared to Method #1. EXAMPLE 2 Use of Intermediate H’ to Produce an Inhibitor of ICE A.

t-Butyl-9-amino-6 , 10-dioxo-l ,2,3,4,7,8,9, 10-octahydro-6- H-pyridazino [1 ,2-a] [1 ,2] diazepine-1-carboxylate (GB2,128,984) To a suspension of H’ (107 g, 0.25 mol) in ethanol (900 iriL) was added hydrazine (27 L, 0.55 mol) and the resulting mixture was allowed to stir at ambient temperature. After 4 hours, the reaction was concentrated in vacuo and the resulting white solid was suspended in acetic acid (IL of 2N) and allowed to stir at ambient temperature for 16 hours. The resulting white solid was filtered off and washed with water. The filtrate was made basic by the addition of solid sodium carbonate and the product extracted with dichloromethane. The organic layer was washed with brine, dried over magnesium sulfate and concentrated in vacuo to afford 79 mg of compound I’ as a yellow viscous oil.B.

t-Butyl-9- (isoquinolin-1-oylamino) -6, 10-dioxo- 1,2,3,4,7,8,9, 10-octahydro-6-H-pyridazino [ 1 , 2-a] [1,2] diazepine-1-carboxylate To a solution of the amine I’ (79 g, 0.265 mol) and isoquinolin-1-carboxylic acid (56g, 0.32 mol) in dichloromethane : DMF (400mL: 400mL) was added hydroxybenztriazole (54 g, 0.4 mol) and l-(3- dimethylaminopropyl) -3-ethylcarbodiimide hydrochloride (74 g, 0.39 mol) and the resulting mixture was allowed to stir at ambient temperature for 16 hours. The reaction mixture was poured into water and extracted with ethyl acetate. The ethyl acetate layer was washed with 0.5N sodium bisulfate, water, sodium bicarbonate, brine, dried over sodium sulfate and concentrated in vacuo to afford 122 g of compound J’ as an orange solid-foam.C.

9- (isoquinolin-1-oylamino) -6, 10-dioxo-l ,2 ,3 ,4 , 7 , 8 , 9 , 10- octahydro-6-H-pyridazino [1 ,2-a] [1,2] diazepine-1- carboxylate A solution of the ester J’ (122 g) in dichloromethane and trifluoroacetic acid (200 mL) was allowed to stir at ambient temperature for 16 hours. The reaction mixture was concentrated to a black oil which was then triturated with acetonitrile and ether to afford 98 g of compound K’ as a pale yellow solid. D .

K'[IS, 9S (2RS, 3S) ] N-(2-benzyloxγ-5-oxotetrahydrofuran-3- yl) -6 , 10-dιoxo-9- (ιsoquιnolιn-1-oγlamιno) -1,2,3,4,7,8,9, 10-octahydro-6-H-pyrιdazιno [ 1 , 2-a] [1,2] dιazepιne-l-carboxamιde To a solution of (3S, 2RS) 3- allyloxycarbonylammo-2- (4-chlorobenzyl) oxy-5- oxotetrahydrofuran [Bioorg. & Med. Chem. Lett., 2, pp. 615-618 (1992)] (4.4 g, 15.1 mmol) in dichloromethane was added N, N-dimethylbarbituric acid (5.9g, 3.8 mmol) then tetrakispalladium ( 0) tπphenyl phosphme (1.7 g, 1.5 mmol) and the resulting mixture was allowed to stir at ambient temperature for 15 minutes. To the resulting mixture was added the acid, compound K’ (5.0 g, 12.6 mmol), hydroxybenztπazole (2.0 g, 14.8 mmol) then and 1- (3-dιmethylammopropyl) -3-ethylcarbodιιmιde hydrochloride (2.7g, 14 mmol) and the reaction was allowed to stir for 3 hours at ambient temperature. The reaction mixture was then poured into water and extracted with ethyl acetate. The organics were washed with 0.5M sodium bisulfate, water, sodium bicarbonate, br e, dried over magnesium sulfate and concentrated m vacuo to afford 2.6 g of the crude product as a yellow foam. The crude material was purified by column chromatography (Sι0₂, dichloromethane : acetone 9:1 – 3:1) to afford 1.2 g of the compound L’ . Compound L’ and related compounds that may be synthesized using the method of this invention as an intermediate step are described in WO 97/22619, the disclosure of which is herein incorporated by reference. Those related compounds may be synthesized from the product of the method of this invention, H or H’ , through modifications of the procedure set forth in Example 2. Such modifications are well known in the art. 
PATENTWO 2001083458https://patents.google.com/patent/WO2001083458A2/enScheme IV

C 2 5,> R’==OH (S)-VI-a *^•* 6 6., R R”==<CI

Example 1

(S) -t-butyl- bis- (1,2-benzyloxycarbonyl) – hexahydropyridazine-3-carboxylate (>90% ee) : To a solution of bis-Cbz hydrazine and (R) -t-butyl-2, 5- dimesylvalerate (from the diol prepared by the method of Schmidt et al., Synthesis, p. 223 (1996)) in DMF was added Na₂S0₄ then TBAF (2.5 equivalents). The resulting reaction mixture was allowed to stir at room temperature for 24 hrs. The reaction was then diluted with ethyl acetate. The organic layer was washed sequentially with 10% citric acid and brine, dried over anhydrous Na₂S0₄ and concentrated in vacuo to afford the title compound. The optical purity of the title compound was greater than 90% ee as determined by HPLC using a ChiralPak^® AD column and eluting with ethanol at 0.7 ml per minute.Example 2

(S) -t-butyl-bis- (1 ,2-benzyloxycarbonyl) – hexahydropyridazine-3-carboxylate (40% ee) : To a solution of bis-Cbz hydrazine and (R) -t-butyl-2, 5-dimesylvalerate(96.5% ee) in DMF was added Na₂S0₄ then K₂C0₃ (5 equivalents) and TBAI (0.1 equivalents). The resulting reaction mixture was heated at 80°C for 24 hrs. The reaction was allowed to cool and diluted with ethyl acetate. The organic layer was washed sequentially with 10% citric acid and brine, dried over anhydrous Na₂S0₄ and concentrated in vacuo to afford the title compound as a 70:30 mixture of the S:R enantiomers (40% ee, as determined by HPLC using a ChiralPak^® AD column, eluting with ethanol at 0.7 ml/min) .Example 3

Racemic t-butyl- bis- (1 ,2-benzyloxycarbonyl) – hexahydropyridazine-3-carboxylate: To a solution of bis- Cbz hydrazine and (R) -t-butyl-2, 5-dimesylvalerate (96.5% ee) in THF was added NaH (2 equivalents) . The resulting reaction mixture was stirred at room temperature. The reaction was quenched then diluted with ethyl acetate. The organic layer was washed sequentially with 10% citric acid and brine, dried over anhydrous Na₂S0₄ and concentrated in vacuo to afford the title compound as a racemic mixture.Example 4 A. Deprotection and salt formation

Hexahydro-pyridazine-3-carboxylic acid tert-butyl ester , L-tartaric acid salt (B) : Compound A was combined with 10% Pd/C (10% w/w) in tetrahydrofuran. The resulting suspension was stirred at 60 °C under a hydrogen atmosphere until deprotection complete. The catalyst was removed via filtration, to the filtrate was added L- tartaric acid (1 equivalent) and the resulting solution concentrated in vacuo.B. Enantiomeric Enrichment

Hθ

The concentrate (B) was taken up in n-butanol(10 volumes), heated to reflux, then allowed to slowly cool to ambient temperature while stirring. The resulting solids were collected via filtration to afford(S) -piperazic acid, t-butyl ester as the tartrate salt (C) in 33% yield.C. Chiral AnalysisCompound (C) was suspended in water and DCM and cooled. We then added NaOH to basify the aqueous layer. The layers were then separated and to the organic layer we added two equivalents of benzyl chloroformate andNaOH. After stirring for 1 hour, the layers were again separated and the organic layer was washed with water.The organic layer was then dried over MgS0₄ and then concentrated in vacuo to produce the bis-Cbz piperazic acid, t-butyl ester for chiral HPLC analysis. The bis-Cbz piperazic acid, t-butyl ester was applied to a Chiralpak AD HPLC column (Chiral Technologies, Exton, PA) and eluted with ethanol at 0.8 ml/minute. Fractions from the column were quantitate by absorption at 210 nm. The results demonstrated that (S)- piperazic acid, t-butyl ester accounted for 94.5% of the piperazic acid, t-butyl ester present in the preparation.

Example 5 Conversion of Intermediate IV to Intermediate Vl-a ^Cbzy

IV’ C0₂t-Bu yi-a C0₂t-Bu Tetrahydro-pyridazine-l,3-dicarboxylic acid 1-benzyl ester 3-tert-butyl ester (Vl-a) : Compound IV (1 mmol) is combined with toluene and sodium hydroxide (aqueous, 2M, 3 equivalents) and the resulting mixture cooled to 1 °C. A solution of benzylchloroformate (1.05 equivalents) in toluene is added while maintaining the reaction pH at 10 or higher by the addition of sodium hydroxide, as needed. After stirring an additional 1 hour, allow the mixture to warm to room temperature then extract with ethyl acetate. The organic layer is washed with brine, dried over sodium sulfate and concentrated to afford Vl-a.Example 6 Conversion of Intermediate X to an Inhibitor of ICE

A. Phthalimide removal to form IX-b

X IX-b t-Butyl-9-amino-6 , 10-dioxo-l ,2,3,4,7,8,9, 10-octa ydro-6-H-pyridazino[l,2-a] [1,2] diazepine-1-carboxylate (GB 2,128,984): To a suspension of X (107 g, 0.25 mol) in ethanol (900 mL) was added hydrazine (27 mL, 0.55 mol) and the resulting mixture was allowed to stir at ambient temperature. After 4 hours, the reaction was concentrated in va cuo and the resulting white solid was suspended in acetic acid (1L of 2N) and allowed to stir at ambient temperature for 16 hours. The resulting white solid was filtered off and washed with water. The filtrate was made basic by the addition of solid sodium carbonate and the product extracted with dichloromethane. The organic layer was washed with brine, dried over magnesium sulfate and concentrated in va cuo to afford 79g of compound IX-b as a yellow viscous oil.B. Formation of compound XII

IX-b XII t-Butyl-9- (isoquinolin-1-oylamino) -6 , 10-dioxo- 1,2,3,4,7,8,9, 10-octahydro-6-H-pyridazino [1 , 2-a] [1,2] diazepine-1-carboxylate (XII) : To a solution of IX-b (79 g, 0.265 mol) and isoquinolin-1-carboxylic acid (56g, 0.32 mol) in dichloromethane and DMF (400mL: 00mL) was added hydroxybenzotriazole (54 g, 0.4 mol) and l-(3- dimethylaminopropyl) -3-ethylcarbodiimide hydrochloride (74 g, 0.39 mol) and the resulting mixture was allowed to stir at ambient temperature for 16 hours. The reaction mixture was poured into water and extracted with ethyl acetate. The ethyl acetate layer was washed with 0.5N sodium bisulfate, water, sodium bicarbonate, brine, dried over sodium sulfate and concentrated in vacuo to afford 122 g of compound XII as an orange solid-foam.t-Butyl ester hydrolysis to form compound XIII

XIII 9- (isoquinolin-1-oylamino) -6 , 10-dioxo-l ,2,3,4,7,8,9, 10- octahydro-6-H-pyridazino [1 , 2-a] [1 , 2] diazepine-1- carboxylate (XIII) : A solution of the ester XII (from step B) (122 g) in dichloromethane and trifluoroacetic acid (200 mL) was allowed to stir at ambient temperature for 16 hours. The reaction mixture was concentrated to a black oil which was then triturated with acetonitrile and ether to afford 98 g of compound XIII as a pale yellow solid.D. Formation of compound 4-b

[1S, 9S (2RS,3S) ]N- (2-benzyloxy-5-oxotetrahydrofuran-3- yl) -6,10-dioxo-9- (isoquinolin-1-oylamino) – 1,2,3,4,7,8,9, 10-octahydro-6-H-pyridazino [1 , 2-a] [1,2] diazepine-1-carboxamide (4-b) : To a solution of (3S, 2RS) 3-allyloxycarbonylamino-2-benzyloxy-5-oxotetrahydrofuran [Bioorq. & Med. Chem. Lett., 2, pp. 615-618 (1992)] (4.4 g, 15.1 mmol) in dichloromethane was added N,N- dimethylbarbituric acid (5.9g, 3.8 mmol) then tetrakispalladium(O) triphenyl phosphine (1.7 g, 1.5 mmol) and the resulting mixture was allowed to stir at ambient temperature for 15 minutes. To the resulting mixture was added the acid, compound XIII (from step C) (5.0 g, 12.6 mmol), hydroxybenzotriazole (2.0 g, 14.8 mmol), then 1- (3-dimethylaminopropyl) -3-ethylcarbodiimide hydrochloride (2.7g, 14 mmol) and the reaction was allowed to stir for 3 hours at ambient temperature. The reaction mixture was then poured into water and extracted with ethyl acetate. The organics were washed with 0.5M sodium bisulfate, water, sodium bicarbonate, brine, dried over magnesium sulfate and concentrated in vacuo to afford 2.6 g of the crude product as a yellow foam. The crude material was purified by column chromatography (Si0₂, dichloromethane: acetone 9:1 – 3:1) to afford 1.2 g of the compound 4-b. Compounds of formulae VII and VIII, and related compounds, that may be synthesized using the method of this invention as an intermediate step are described in WO 97/22619 and United States Patent 6,204,261 the disclosure of which is herein incorporated by reference. Those related compounds may be synthesized from the product of the method of this invention, I, IV, or V, through modifications of the procedure set forth in Examples 4 through 6. Such modifications are well known in the art.PATENTUS 6559304https://patents.google.com/patent/US6559304B1PATENTWO 2008074816https://patents.google.com/patent/WO2008074816A1/en

Patent 

Publication numberPriority datePublication dateAssigneeTitleEP0094095A2 *1982-05-121983-11-16F. Hoffmann-La Roche AgBicyclic carboxylic acids and their alkyl and aralkyl estersUS4692438A *1984-08-241987-09-08Hoffmann-La Roche Inc.Pyridazo-diazepines, diazocines, and -triazepines having anti-hypertensive activityWO1993023403A1 *1992-05-151993-11-25Merrell Dow Pharmaceuticals Inc.NOVEL MERCAPTOACETYLAMIDO PYRIDAZO[1,2]PYRIDAZINE, PYRAZOLO[1,2]PYRIDAZINE, PYRIDAZO[1,2-a][1,2]DIAZEPINE AND PYRAZOLO[1,2-a][1,2]DIAZEPINE DERIVATIVES USEFUL AS INHIBITORS OF ENKEPHALINASE AND ACEWO1994011353A1 *1992-11-121994-05-26University College LondonProcess for the preparation of (3r)- and (3s)-piperazic acid derivativesWO1995035308A1 *1994-06-171995-12-28Vertex Pharmaceuticals IncorporatedINHIBITORS OF INTERLEUKIN-1β CONVERTING ENZYMEFamily To Family CitationsUS6204261B11995-12-202001-03-20Vertex Pharmaceuticals IncorporatedInhibitors of interleukin-1β Converting enzyme inhibitorsFR2777888B11998-04-272004-07-16Hoechst Marion Roussel IncNOVEL DERIVATIVES OF ACID (3,4,7,8,9,10-HEXAHYDRO-6,10- DIOXO-6H-PYRIDAZINO [1,2-A] [1,2] DIAZEPINE-1-CARBOXYLIC, THEIR PROCESS OF PREPARATION AND THEIR APPLICATION TO THE PREPARATION OF MEDICINESFR2777889B11998-04-272004-07-09Hoechst Marion Roussel IncNOVEL DERIVATIVES OF OCTAHYDRO-6,10-DIOXO-6H- PYRIDAZINO [1,2-A] [1,2] DIAZEPINE-1-CARBOXYLIC, THEIR PREPARATION PROCESS AND THEIR APPLICATION TO THE PREPARATION OF THERAPEUTICALLY ACTIVE COMPOUNDS 

////////////////Pralnacasan, VX 740, VX 470, HMR 3480, пралнаказан , برالناكاسان , 普那卡生 , 

CCOC1C(CC(=O)O1)NC(=O)C2CCCN3N2C(=O)C(CCC3=O)NC(=O)C4=NC=CC5=CC=CC=C54

NEW DRUG APPROVALS

ONE TIME

$10.00

Click here to purchase.

Naftopidil

1-[4-(2-methoxyphenyl)piperazin-1-yl]-3-naphthalen-1-yloxypropan-2-ol

C24H28N2O3, 392.49

CAS 57149-07-2

KT-611, Flivas, Avishot

1-(4-(2-methoxyphenyl)piperazin-1-yl)-3-(naphthalen-1-yloxy)propan-2-ol

Naftopidil (Flivas), BM-15275NaftopidilCAS Registry Number: 57149-07-2 
CAS Name: 4-(2-Methoxyphenyl)-a-[(1-naphthalenyloxy)methyl]-1-piperazineethanolAdditional Names:RS-1-[4-(2-methoxyphenyl)-1-piperazinyl]-3-(1-naphthoxy)-2-propanol; 1-(2-methoxyphenyl)-4-[3-(naphth-1-yloxy)-2-hydroxypropyl]-piperazine 
Manufacturers’ Codes: KT-611Trademarks: Avishot (Kanebo); Flivas (Asahi)Molecular Formula: C24H28N2O3Molecular Weight: 392.49Percent Composition: C 73.44%, H 7.19%, N 7.14%, O 12.23%Literature References: a1-Adrenergic blocker and serotonin (5HT1A) receptor agonist. Prepn: E. C. Witte et al.,DE2408804; eidem,US3997666 (1975, 1976 both to Boehringer Mann.). Clinical pharmacodynamics: R. Kirsten et al.,Eur. J. Clin. Pharmacol.46, 271 (1994). Clinical pharmacokinetics: M. J. G. Farthing et al.,Postgrad. Med. J.70, 363 (1994). HPLC determn in human plasma: G. Niebch et al.,J. Chromatogr.534, 247 (1990). Clinical evaluation in BPH: K. Yasuda et al.,Prostate25, 46 (1994). Review of pharmacology and clinical experience: H. M. Himmel, Cardiovasc. Drug Rev.12, 32-47 (1994). 
Properties: Crystals from isopropanol, mp 125-126°; also reported as colorless crystals, mp 125-129°. Insol in water. Partition coefficient (octanol/water): 75. LD50 in mice, rats (g/kg): 1.3, 6.4 orally (Himmel).Melting point: mp 125-126°; mp 125-129°Log P: Partition coefficient (octanol/water): 75Toxicity data: LD50 in mice, rats (g/kg): 1.3, 6.4 orally (Himmel) 
Derivative Type: DihydrochlorideCAS Registry Number: 57149-08-3Molecular Formula: C24H28N2O3.2HClMolecular Weight: 465.41Percent Composition: C 61.94%, H 6.50%, N 6.02%, O 10.31%, Cl 15.24%Properties: Crystals from methanol/ethanol (1:2), mp 212-213°.Melting point: mp 212-213° 
Therap-Cat: Antihypertensive; a-blocker in treatment of symptomatic benign prostate hypertrophy.Keywords: a-Adrenergic Blocker; Antihypertensive.

Naftopidil (INN, marketed under the brand name Flivas) is a drug used in benign prostatic hypertrophy which acts as a selective α₁-adrenergic receptor antagonist or alpha blocker.^[1]

PATENT

DE 2408804

CN 101671317

CN 102816136

JP 2013023467

JP 2014118360

IN 2011CH00466

US 20150353473

CN 104744405

IN 2013CH06042

IN 2012DE02071

JP 2016044182

PAPER

ChemMedChem (2009), 4(3), 393-9.

The Journal of organic chemistry (2013), 78(18), 9076-84.

e-EROS Encyclopedia of Reagents for Organic Synthesis (2014), 1-5

European journal of medicinal chemistry (2015), 96, 83-91.

Bioorganic & medicinal chemistry letters (2018), 28(9), 1534-1539.

ChemistrySelect (2019), 4(26), 7745-7750.

Green Chemistry (2019), 21(16), 4422-4433.  |

PAPER

https://www.scielo.br/j/jbchs/a/q5qDxfT9mSwtL9hhQYxyhgs/?lang=en#

(S)-1-(4-(2-Methoxyphenyl)piperazin-1-yl)-3-(naphthalene1-yloxy)propan-2-ol (2b) To a solution of epoxide 8b (0.1 g, 0.5 mmol) in anhydrous 2-propanol (10 mL) was added 1-(2-methoxyphenyl) piperazine (0.096 g, 0.5 mmol) and the reaction mixture was refluxed for 32 h. After completion of reaction, the solvent was removed under reduced pressure and purification was carried out by flash column chromatography (230-400 mesh silica). The EtOAc:petroleum ether (60:40) was used as solvent system for elution, it afforded the (S)-(+)-naftopidil 2b as a yellow solid (0.156 g, 80%); mp 126-127°C; [α]D 25 +4.3o (c 1.55, MeOH);3 [α]D 25 +4.5o (c 1.5, MeOH); IR (CHCl3) νmax/cm-1 3403, 3031, 2977, 2907, 1261, 1225; 1 H NMR (300 MHz, CDCl3) d 2.58-2.70 (m, 4H, N-CH2), 2.80-2.85 (m, 2H, CH2N), 3.03-3.51 (m, 4H, NCH2), 3.51 (bs, 1H, OH), 3.75 (s, 3H, OCH3), 4.02-4.10 (m, 2H, OCH2), 4.19-4.23 (m, 1H, CH), 6.72-6.85 (m, 2H, Ar-H), 6.83-6.85 (d, 2H, J 3.9 Hz, Ar-H), 6.87-6.95 (1H, m, Ar-H), 7.14-7.29 (1H, m, Ar-H), 7.33-7.42 (3H, m, Ar-H), 7.69-7.72 (m, 1H, Ar-H), 8.19-8.22 (m, 1H, Ar-H); 13C NMR (75 MHz, CDCl3) d 50.44 (NCH2), 53.43 (NCH2), 55.17 (OCH3), 60.85 (CH2N), 65.47 (CH), 70.36 (OCH2), 104.73 (Ar), 111.03 (Ar), 118.05 (Ar), 120.39 (Ar), 120.83 (Ar), 121.78 (Ar), 122.91 (Ar), 125.07 (Ar), 125.41 (Ar), 125.67 (Ar), 126.26 (Ar), 127.32 (Ar), 134.31 (Ar), 140.87 (Ar), 152.04 (Ar), 154.21 (Ar); LC-MS m/z 393.36 (M+ + 1), 415.36 (M+ + Na); For compound 2a: [α]D 25 -10.6o (c 1, MeOH,);6 [α]D 25 -11.7o (c 1, MeOH).

PATENT

CN 1473820

PATENT

WO 2018026371

https://patents.google.com/patent/WO2018026371A1/en

PATENT

JP-2021104982

Naftopidil monohydrochloride dihydrate and its use for the preparation of naftopidil , which is known as an ameliorating agent for dysuria associated with benign prostatic hyperplasia.Naftopidil is known as an ameliorating agent for dysuria associated with benign prostatic hyperplasia. Since naftopidil is administered as a free form, there is a need for a method for preparing the free form that can be obtained efficiently and with high purity. 
Japanese Unexamined Patent Publication No. 50-12186 (Patent Document 1) discloses a method for preparing naftopidil, and states that naftopidil was obtained in a yield of 29% to 79% in the examples thereof. In particular, in Example 3, naftopidil is obtained via naftopidil hydrochloride anhydride, but the yield is 49%, and the purity is not described. 
Japanese Patent Application Laid-Open No. 2013-23467 (Patent Document 2) reacts 1- (2-methoxyphenyl) piperazin with 2-[(1-naphthyloxy) methyl] oxylane to obtain crude naftopidil, which is obtained as toluene. Discloses a method for obtaining purified naftopidil from water and water, as well as a mixed solvent of toluene and methanol. In this method, the yield of crude naftopidil did not reach 80%, and the purity after two purification operations using toluene and water, and then toluene and methanol was said to be 99.62% at the highest. ing. In this method, crude naftopidil is not chlorinated with hydrochloric acid. 
In Indian patent application 466 / CHE / 2011 (Patent Document 3), crude naftopidil was recrystallized from ethyl acetate to obtain naftopidil in a yield of 79% and a purity of 99.90%, and further recrystallized from methanol to obtain purity. It discloses a method of obtaining 99.99% naftopidil. Even with this method, crude naftopidil is not chlorinated with hydrochloric acid. 
Indian Patent Application 2071 / DEL / 2012 (Patent Document 4) discloses a method for producing green chemical naftopidil using metal nanoparticles. Here, naftopidil is purified by column chromatography using silica gel to obtain naftopidil in a yield of 63%, but the purity is not disclosed.patcit 1: Japanese Patent Application
Laid-Open No. 50-12186 patcit 2: Japanese Patent Application Laid-Open No.
2013-23467 patcit 3: Indian Patent Application 466 / CHE / 2011
patcit 4: Indian Patent Application 2071 / DEL / 2012Production
of Naftopidil Monohydrochloride Dihydrate The naftopidil monohydrochloride dihydrate according to the present invention is preferably prepared according to the following scheme.
[Chem. 2]

That is, it can be obtained by reacting 2-[(1-naphthyloxy) methyl] oxylane with 1- (2-methoxyphenyl) piperazine by adding a solvent such as toluene, and then adding / presenting hydrochloric acid. ..The present invention will be described in more detail with reference to the following examples. The reactions in the examples below, and the numbers given to the compounds, are as shown in the scheme below.
[Chem. 3]

Example 1
100 g of 1 -naphthol [1] was dissolved in chloromethyloxylan [2], and a sodium methoxide methanol solution was added dropwise. After completion of the reaction, the reaction was washed with water and the organic layer was concentrated to obtain 2-[(1-naphthyloxy) methyl] oxylan [3] (yield 89%). 
Example 2
A toluene solution of 1- (2-methoxyphenyl) piperazin [4] was added dropwise to a toluene solution of 5.0 g of 2-[(1-naphthyloxy) methyl] oxylan [3]. After completion of the reaction, the mixture was washed with water and cooled by adding hydrochloric acid. After the suspension is filtered off, it is dried and (2RS) -1- [4- (2-methoxyphenyl) piperazin-1-yl] -3- (naphthalene-1-yloxy) propan-2-ol. Hydrochloride dihydrate [5] was obtained (yield 95%). 
Example 3
A toluene solution of 1- (2-methoxyphenyl) piperazin [4] was added dropwise to a toluene solution of 5.0 g of 2-[(1-naphthyloxy) methyl] oxylan [3]. After completion of the reaction, the mixture was washed with water, methanol and hydrochloric acid were added to separate the liquids, and the mixture was cooled. After the suspension is filtered off, it is dried and (2RS) -1- [4- (2-methoxyphenyl) piperazin-1-yl] -3- (naphthalene-1-yloxy) propan-2-ol. Hydrochloride dihydrate [5] was obtained (yield 81%). 
Example 4
A toluene solution of 1- (2-methoxyphenyl) piperazin [4] was added dropwise to a toluene solution of 5.0 g of 2-[(1-naphthyloxy) methyl] oxylan [3]. After completion of the reaction, the mixture was washed with water, methanol and hydrochloric acid were added, and the mixture was cooled. After the suspension is filtered off, it is dried and (2RS) -1- [4- (2-methoxyphenyl) piperazin-1-yl] -3- (naphthalene-1-yloxy) propan-2-ol. Hydrochloride dihydrate [5] was obtained (yield 86%). 
Example 5
A toluene solution of 1- (2-methoxyphenyl) piperazin [4] was added dropwise to a toluene solution of 100 g of 2-[(1-naphthyloxy) methyl] oxylan [3]. After completion of the reaction, the mixture was washed with water, methanol and hydrochloric acid were added, and the mixture was cooled. After the suspension is filtered off, it is dried and (2RS) -1- [4- (2-methoxyphenyl) piperazin-1-yl] -3- (naphthalene-1-yloxy) propan-2-ol. Hydrochloride dihydrate [5] was obtained (yield 92%). 
Example 6
(2RS) -1- [4- (2-methoxyphenyl) piperazin-1-yl] -3- (naphthalene-1-yloxy) propan-2-ol monohydrochloride dihydrate [5 ] Toluene and sodium hydroxide aqueous solution were added to 7.0 g. The organic layer was washed with water and concentrated, and then metall and acetonitrile were added and cooled. After the suspension is filtered off, it is dried and (2RS) -1- [4- (2-methoxyphenyl) piperazin-1-yl] -3- (naphthalene-1-yloxy) propan-2-ol [ 6] was obtained (yield 82%, chemical purity 99.98%). 
Example 7
(2RS) -1- [4- (2-methoxyphenyl) piperazin-1-yl] -3- (naphthalene-1-yloxy) propan-2-ol monohydrochloride dihydrate [5 ] Toluene and an aqueous sodium hydroxide solution were added to 12.0 g. The organic layer was washed with water and concentrated, then metall was added and cooled. After the suspension is filtered off, it is dried and (2RS) -1- [4- (2-methoxyphenyl) piperazin-1-yl] -3- (naphthalene-1-yloxy) propan-2-ol [ 6] was obtained (yield 90%, chemical purity 99.99%). 
Example 8
(2RS) -1- [4- (2-Methoxyphenyl) piperazin-1-yl] -3- (naphthalene-1-yloxy) propan-2-ol monohydrochloride dihydrate [5 ] Toluene, methanol, and potassium hydroxide aqueous solution were added to 116 g. The organic layer was washed with water and concentrated, then 2-propanol was added and cooled. After the suspension is filtered off, it is dried and (2RS) -1- [4- (2-methoxyphenyl) piperazin-1-yl] -3- (naphthalene-1-yloxy) propan-2-ol [ 6] was obtained (yield 90%, chemical purity 99.98%). 
Comparative Example 1
A toluene solution of 1- (2-methoxyphenyl) piperazin [4] was added dropwise to a 10.0 g toluene solution of 2-[(1-naphthyloxy) methyl] oxylan [3]. After completion of the reaction, the mixture was washed with water and cooled. After the suspension is filtered off, it is dried and (2RS) -1- [4- (2-methoxyphenyl) piperazin-1-yl] -3- (naphthalene-1-yloxy) propan-2-ol [ 6] Crude crystals were obtained (yield 89%). 
Comparative Example 2
(2RS) -1- [4- (2-methoxyphenyl) piperazin-1-yl] -3- (naphthalene-1-yloxy) propan-2-ol [6] obtained in Comparative Example 1. Methoxyol and acetonitrile were added to 6.0 g of the crude crystals of the above, and the mixture was cooled. After the suspension is filtered off, it is dried and (2RS) -1- [4- (2-methoxyphenyl) piperazin-1-yl] -3- (naphthalene-1-yloxy) propan-2-ol [ 6] was obtained (yield 85%, chemical purity 99.96%). 
Naftopidil one identification hydrochloride dihydrate
(1) water and HCl content
mosquito – Le Fischer – water content value measured by the law was 7.3% to 7.5%. The amount of HCl measured by neutralization titration was 8.0% to 8.1%. Determined from these naftopidil: HCl: _{H 2} When calculating these molar ratios from O weight ratio of approximately 1: 1: 2. From this, it was judged that naftopidil monohydrochloride dihydrate was obtained.
(2) Powder X-ray Diffraction
The chart of the results of powder X-ray diffraction (Cu-Kα) of naftopidil monohydrochloride dihydrate was as shown in FIG. For reference, a chart of naftopidil is shown as FIG.
(3) Differential Thermal Analysis / Thermogravimetric Analysis
(TG / DTA) The chart of the results of differential thermal analysis / thermogravimetric analysis (TG / DTA) of naphthopidyl monohydrochloride dihydrate is as shown in FIG. rice field. Here, the measurement conditions were such that the heating rate was 5 ° C./min. For reference, a chart of naftopidil is shown as FIG. 
PAPERShivani; Journal of Organic Chemistry 2007, V72(10), P3713-3722 https://pubs.acs.org/doi/10.1021/jo062674j

References

1. ^ Sakai H, Igawa T, Onita T, Furukawa M, Hakariya T, Hayashi M, Matsuya F, Shida Y, Nishimura N, Yogi Y, Tsurusaki T, Takehara K, Nomata K, Shiraishi K, Shono T, Aoki D, Kanetake H (2011). “Efficacy of naftopidil in patients with overactive bladder associated with benign prostatic hyperplasia: prospective randomized controlled study to compare differences in efficacy between morning and evening medication”. Hinyokika Kiyo. 57 (1): 7–13. PMID 21304253.

/////////////////Naftopidil, KT 611, a-Adrenergic Blocker, Antihypertensive.

COC1=CC=CC=C1N2CCN(CC2)CC(COC3=CC=CC4=CC=CC=C43)O

NEW DRUG APPROVALS

ONE TIME

$10.00

Click here to purchase.

Belzutifan

CAS 1672668-24-4

383.34 g·mol^{−1  C₁₇H₁₂F₃NO₄S}

3-[[(1S,2S,3R)-2,3-difluoro-1-hydroxy-7-methylsulfonyl-2,3-dihydro-1H-inden-4-yl]oxy]-5-fluorobenzonitrile

MK-6482, PT-2977, UNII-7K28NB895L, 7K28NB895L

3-[(1S,2S,3R)-2,3-Difluoro-1-hydroxy-7-methylsulfonylindan-4-yl]oxy-5-fluorobenzonitrile

3-{[(1s,2s,3r)-2,3-Difluoro-1-Hydroxy-7-(Methylsulfonyl)-2,3-Dihydro-1h-Inden-4-Yl]oxy}-5-Fluorobenzonitrile

GTPL11251, PT 2977 [WHO-DD], BDBM373040

FDA APPROVED 8/13/2021, Welireg

To treat von Hippel-Lindau disease under certain conditions

EMA Drug Information

FDA approves belzutifan for cancers associated with von Hippel-Lindau disease

On August 13, 2021, the Food and Drug Administration approved belzutifan (Welireg, Merck), a hypoxia-inducible factor inhibitor for adult patients with von Hippel-Lindau (VHL) disease who require therapy for associated renal cell carcinoma (RCC), central nervous system (CNS) hemangioblastomas, or pancreatic neuroendocrine tumors (pNET), not requiring immediate surgery.

Belzutifan was investigated in the ongoing Study 004 (NCT03401788), an open-label clinical trial in 61 patients with VHL-associated RCC (VHL-RCC) diagnosed based on a VHL germline alteration and with at least one measurable solid tumor localized to the kidney. Enrolled patients had other VHL-associated tumors, including CNS hemangioblastomas and pNET. Patients received belzutifan 120 mg once daily until disease progression or unacceptable toxicity.

The primary efficacy endpoint was overall response rate (ORR) measured by radiology assessment, as assessed by an independent review committee using RECIST v1.1. Additional efficacy endpoints included duration of response (DoR), and time- to- response (TTR). An ORR of 49% (95% CI:36, 62) was reported in patients with VHL-associated RCC. All patients with VHL-RCC with a response were followed for a minimum of 18 months from the start of treatment. The median DoR was not reached; 56% of responders had DoR ≥ 12 months and a median TTR of 8 months. In patients with other VHL-associated non-RCC tumors, 24 patients with measurable CNS hemangioblastomas had an ORR of 63% and 12 patients with measurable pNET had an ORR of 83%. Median DoR was not reached, with 73% and 50% of patients having response durations ≥ 12 months for CNS hemangioblastomas and pNET, respectively.

The most common adverse reactions, including laboratory abnormalities, reported in ≥ 20% of patients who received belzutifan were decreased hemoglobin, anemia, fatigue, increased creatinine, headache, dizziness, increased glucose, and nausea. Anemia and hypoxia from belzutifan use can be severe. In Study 004, anemia occurred in 90% of patients and 7% had Grade 3 anemia. Patients should be transfused as clinically indicated. The use of erythropoiesis stimulating agents for treatment of anemia is not recommended in patients treated with belzutifan. In Study 004, hypoxia occurred in 1.6% of patients. Belzutifan can render some hormonal contraceptives ineffective, and belzutifan exposure during pregnancy can cause embryo-fetal harm.

The recommended belzutifan dosage is 120 mg administered orally once daily with or without food.
View full prescribing information for Welireg.

This review was conducted under Project Orbis, an initiative of the FDA Oncology Center of Excellence. Project Orbis provides a framework for concurrent submission and review of oncology drugs among international partners. For this review, FDA collaborated with the Australian Therapeutic Goods Administration (TGA), Health Canada, and the Medicines and Healthcare products Regulatory Agency (MHRA) of the United Kingdom. The application reviews are ongoing at the other regulatory agencies.

This review used the Real-Time Oncology Review (RTOR) pilot program, which streamlined data submission prior to the filing of the entire clinical application, as well as the Assessment Aid and the Product Quality Assessment Aid, voluntary submissions from the applicant to facilitate the FDA’s assessment. The FDA approved this application approximately 1 month ahead of the FDA goal date.

This application was granted priority review for this indication. A description of FDA expedited programs is in the Guidance for Industry: Expedited Programs for Serious Conditions-Drugs and Biologics.

Belzutifan, sold under the brand name Welireg, is a medication used for the treatment of von Hippel–Lindau disease-associated renal cell carcinoma.^{[1][2][3][4][5][6]} It is taken by mouth.^[1]

The most common side effects include decreased hemoglobin, anemia, fatigue, increased creatinine, headache, dizziness, increased glucose, and nausea.^[2]

Belzutifan is an hypoxia-inducible factor-2 alpha (HIF-2α) inhibitor.^[1][2][7]

Belzutifan is the first drug to be awarded an “innovation passport” from the UK Medicines and Healthcare products Regulatory Agency (MHRA).^[8][4] Belzutifan was approved for medical use in the United States in August 2021.^[2][9] Belzutifan is the first hypoxia-inducible factor-2 alpha inhibitor therapy approved in the U.S.^[9]

Medical uses

Belzutifan is indicated for treatment of adults with von Hippel-Lindau (VHL) disease who require therapy for associated renal cell carcinoma (RCC), central nervous system (CNS) hemangioblastomas, or pancreatic neuroendocrine tumors (pNET), not requiring immediate surgery.^[2]

PATENT

WO  2019191227

https://patents.google.com/patent/WO2019191227A1/en

PATENT

WO 2015035223

https://patents.google.com/patent/WO2015035223A1/enScheme 9

[01237] 3-r(15,25.3 ?^‘)-2.3-difluoro-l-hvdroxy-7-methylsulfonyl-indan-4- νΠο^χν-5-fluoro-benzonitrile (Compound 289)[01238] Step A: r(15.2/?V4- -cvano-5-fluoro-phenoxy)-2-fluoro-7- methylsulfonyl-indan-l -vH acetate: To a stirred solution of 3-fluoro-5-[(15,27?)-2-fluoro-l – hydroxy-7-methylsulfonyl-indan-4-yl]oxy-benzonitrile (2.00 g, 5.47 mmol) in DCM (27 mL) was added 4-(dimethylamino)pyridine (0.2 g, 1.64 mmol) and triethylamine (1.53 mL, 10.9 mmol). Acetic anhydride (1.00 mL, 10.9 mmol) was added dropwise at 0 °C under nitrogen. The reaction mixture was stirred at ambient temperature overnight. The reaction mixture was diluted with DCM, washed with saturated aqueous NaHC0₃ and brine, dried andconcentrated. The residue was purified by flash chromatography on silica gel (20-40% EtOAc/hexane) to give [(lS,2/?)-4-(3-cyano-5-fluoro-phenoxy)-2-fluoro-7-methylsulfonyl- indan-l-yl] acetate (1.95 g, 87%). LCMS ESI (+) m/z 408 (M+H).[01239] Step B: Γ( 1 .25.35)-3-bromo-4-(3-cvano-5-fluoro-Dhenoxy)-2-fluoro- 7-methylsulfonyl-indan-l-yll acetate and f(15.25,3/?)-3-bromo-4-(3-cyano-5-fluoro- phenoxy)-2-fluoro-7-methylsulfonyl-indan-l -yl1 acetate: To a stirred solution of [(15,2/?)-4- (3-cyano-5-fluoro-phenoxy)-2-fluoro-7-methylsulfonyl-indan-] -yl] acetate (1.95 g, 4.79 mmol) in 1 ,2-dichloroethane (24 mL) was added N-bromosuccinimide (0.94 g, 5.27 mmol) and 2,2′-azobisisobutyronitrile (8 mg, 0.05 mmol). The reaction mixture was heated at 80 °C for 3 hours. After cooling, the reaction mixture was diluted with DCM, washed with saturated aqueous NaHC0₃ and brine, dried and concentrated. The residue was purified by column chromatography on silica gel (20-30% EtOAc hexane) to give [(lS,2S,3S)-3-bromo- 4-(3-cyano-5-fluoro-phenoxy)-2-fluoro-7-methylsulfonyl-indan-l-yl] acetate (1 .52 g, 65%). LCMS ESI (+) m/z 486, 488 (M+H). Further elution with 30-50% EtOAc/hexane gave the more polar product [(lS,2S,3/?)-3-bromo-4-(3-cyano-5-fluoro-phenoxy)-2-fluoro-7- methylsulfonyl-indan-l -yl] acetate (0.583 g, 25%). LCMS ESI (+) m/z 486, 488 (M+H). [01240] Step C: rd5.2^.3 V4-(3-cvano-5-fluoro-phenoxy)-2-fluoro-3- hvdroxy-7-methylsulfonyl-indan- 1 -yll acetate: To a combined mixture of [(1 ,25,35)-3- bromo-4-(3-cyano-5-fluoro-phenoxy)-2-fluoro-7-methylsulfonyl-indan-l -yl] acetate and [( 15,2S,3/?)-3-bromo-4-(3-cyano-5-fluoro-phenoxy)-2-fluoro-7-methylsulfonyl-indan- 1 -yl] acetate prepared in Step B (2.05 g, 4.22 mmol) were added 1 ,2-dimethoxyethane (28 mL) and water (0.050 mL) followed by silver perchlorate hydrate (1.42 g, 6.32 mmol). The reaction mixture was heated at 70 °C for 2 hours. After cooling, the reaction mixture was diluted with EtOAc and filtered through Celite. The filtrate was washed with water and brine, dried and concentrated. The residue was purified by flash chromatography on silica gel (20-50%) to give [(15,2/?,35)-4-(3-cyano-5-fluoro-phenoxy)-2-fluoro-3-hydroxy-7-methylsulfonyl-indan- 1 -yl] acetate (0.416 g, 23%) as the less polar product. LCMS ESI (+) m/z 441 (M+NH₄⁺). Further elution with 60% EtOAc/hexane gave [(15,2/?,3R)-4-(3-cyano-5-fluoro-phenoxy)-2- fluoro-3-hydroxy-7-methylsulfonyl-indan-l-yl] acetate (0.58 g, 32 %). LCMS ESI (+) m/z 441 (M+NH₄⁺).[01241] Step D: r(15.25.3/? -4-(3-cvano-5-fluoro-phenoxyV2.3-difluoro-7- methylsulfonyl-indan-l-vH acetate: To a stirred solution of [(15,2/?,35)-4-(3-cyano-5-fluoro- phenoxy)-2-fluoro-3-hydroxy-7-methylsulfonyl-indan-l-yl] acetate (416 mg, 0.98 mmol) in DCM (10 mL) was added (diethylamino)sulfur trifluoride (DAST) (0.26 mL, 2.0 mmol) at – 78 °C under nitrogen. The reaction mixture was allowed to warm to 0 °C and stirred for 15 minutes. The reaction was quenched by saturated aqueous NaHC0₃. The mixture was partitioned between EtOAc and water. The aqueous layer was extracted with EtOAc. The combined organic layers were washed with brine, dried and concentrated. The residue was purified by flash chromatography on silica gel (20-40% EtOAc/hexane) to give [(15,25,3/?)- 4-(3-cyano-5-fluoro-phenoxy)-2,3-difluoro-7-methylsulfonyl-indan-l -yl] acetate (310 mg, 74%). LCMS ESI (+) m/z 426 (M+H).[01242] Step E: 3-r(15.25.3^)-2.3-difluoro-l-hvdroxy-7-methylsulfonyl-indan-4-vnoxy-5-fluoro-benzonitrile (Compound 289): Prepared as described in Example 288 Step F substituting [(l ?)-4-(3-cyano-5-fluoro-phenoxy)-3,3-difluoro-7-methylsulfonyl-indan- 1-yl] acetate with [(15,25,3/?)-4-(3-cyano-5-fluoro-phenoxy)-2,3-difluoro-7-methylsulfonyl- indan-l-yl] acetate. LCMS ESI (+) m/z 384 (M+H); Ή NMR (400 MHz, CDC1₃): δ 8.13 (d, 1H), 7.31-7.25 (m, 1 H), 7.23-7.19 (m, 1 H), 7.14-7.09 (m, 1H), 7.04 (d, 1H), 6.09-5.91 (m, 1 H), 5.87-5.80 (m, 1 H), 5.25-5.05 (m, 1H), 3.32 (s, 3H), 2.95 (d, 1H). 
PatentWO 2016145032https://patentscope.wipo.int/search/en/detail.jsf?docId=WO2016145032&tab=PCTDESCRIPTIONCOMPD 289

PATENTWO 2016145045WO 2016168510WO 2016057242WO 2019191227 

Merck Team Wins 2021 Pete Dunn Award

‎05-17-2021 10:52 AM

The ACS Green Chemistry Institute (GCI) Pharmaceutical Roundtable honors the work of Stephen Dalby, François Lévesque, Cecilia Bottecchia and Jonathan McMullen at Merck with the 2021 Peter J. Dunn Award for Green Chemistry & Engineering Impact in the Pharmaceutical Industry. The team’s innovation is titled, “Greener Manufacturing of Belzutifan (MK-6482) Featuring a Photo-Flow Bromination.”

Belzutifan is an important new drug used in the treatment of cancer and other non-oncology diseases. Acquired by Merck in 2019 through the purchase of Peloton Therapeutics, a new, greener manufacturing process for its synthesis was needed. Over the next 18 months, the team developed a more direct route from commodity chemical to API, employed new reaction conditions, particularly in the oxidation sequence, and incorporated new technology, photo-flow.

Despite this accelerated timeline, the team achieved a five-fold improvement in overall yield with a commensurate 73% reduction in process mass intensity (PMI) compared to the original route. Notably, the Merck team also developed a visible light-initiated radical bromination performed in flow. According to the L.-C. Campeau, Executive Director and Head of Process Chemistry and Discovery Process Chemistry at Merck, this is the “first example of a photo-flow reaction run on manufacturing scale at Merck and represents the linchpin of the synthesis.”

The improved process for Belzutifan, which is expected to launch this year, will reduce the waste associated with its manufacture and is aligned with Merck’s corporate sustainability goals.

“The Merck team delivered an excellent example of the application of innovative technologies to develop a more sustainable synthesis of the pharmaceutically-active compound, Belzutifan,” comments Paul Richardson, Director of Oncology and Chemical Synthesis at Pfizer and Co-Chair of the ACS GCI Pharmaceutical Roundtable. “Using the guiding principles of green chemistry, for example, in the use of catalysis and a relatively benign reaction media, further illustrate the Merck team’s work as worthy of recognition for the 2021 Peter Dunn Award.”

The award will be presented at the June 11 GC&E Friday, part of the 25th Annual Green Chemistry & Engineering Conference. During this session from 10 a.m. – 1 p.m., Stephen Dalby & Jon MacMullen will be discussing the details of this innovative process.

References

1. ^ Jump up to:^a b c d https://www.accessdata.fda.gov/drugsatfda_docs/label/2021/215383s000lbl.pdf
2. ^ Jump up to:^{a b c d e f} “FDA approves belzutifan for cancers associated with von Hippel-Lindau”. U.S. Food and Drug Administration (FDA). 13 August 2021. Retrieved 13 August 2021.  This article incorporates text from this source, which is in the public domain.
3. ^ “Belzutifan”. SPS – Specialist Pharmacy Service. 18 March 2021. Retrieved 25 April 2021.
4. ^ Jump up to:^a b “MHRA awards first ‘innovation passport’ under new pathway”. RAPS (Press release). Retrieved 25 April 2021.
5. ^ “Merck Receives Priority Review From FDA for New Drug Application for HIF-2α Inhibitor Belzutifan (MK-6482)” (Press release). Merck. 16 March 2016. Retrieved 25 April 2021 – via Business Wire.
6. ^ “FDA Grants Priority Review to Belzutifan for von Hippel-Lindau Disease–Associated RCC”. Cancer Network. Retrieved 26 April 2021.
7. ^ {{cite journal |vauthors=Choueiri TK, Bauer TM, Papadopoulos KP, Plimack ER, Merchan JR, McDermott DF, Michaelson MD, Appleman LJ, Thamake S, Perini RF, Zojwalla NJ, Jonasch E | display-authors=6 |title=Inhibition of hypoxia-inducible factor-2α in renal cell carcinoma with belzutifan: a phase 1 trial and biomarker analysis |journal=Nat Med |volume= |issue= |pages= |date=April 2021 |pmid=33888901 |doi=10.1038/s41591-021-01324-7 }
8. ^ “First Innovation Passport awarded to help support development and access to cutting-edge medicines”. Medicines and Healthcare products Regulatory Agency (MHRA) (Press release). 26 February 2021. Retrieved 14 August 2021.
9. ^ Jump up to:^a b “FDA Approves Merck’s Hypoxia-Inducible Factor-2 Alpha (HIF-2α) Inhibitor Welireg (belzutifan) for the Treatment of Patients With Certain Types of Von Hippel-Lindau (VHL) Disease-Associated Tumors” (Press release). Merck. 13 August 2021. Retrieved 13 August 2021 – via Business Wire.

External links

• “Belzutifan”. Drug Information Portal. U.S. National Library of Medicine.
• Clinical trial number NCT04195750 for “A Study of Belzutifan (MK-6482) Versus Everolimus in Participants With Advanced Renal Cell Carcinoma (MK-6482-005)” at ClinicalTrials.gov
• Clinical trial number NCT03401788 for “A Phase 2 Study of Belzutifan (PT2977, MK-6482) for the Treatment of Von Hippel Lindau (VHL) Disease-Associated Renal Cell Carcinoma (RCC) (MK-6482-004)” at ClinicalTrials.gov

/////////Belzutifan, Welireg, FDA 2021, APPROVALS 2021, MK 6482, PT 977, Antineoplastic

CS(=O)(=O)C1=C2C(C(C(C2=C(C=C1)OC3=CC(=CC(=C3)C#N)F)F)F)O

NEW DRUG APPROVALS

ONE TIME

$10.00

Click here to purchase.

AVASOPASEM

Average: 518.83
Monoisotopic: 517.134397

Chemical FormulaC₂₁H₃₅Cl₃MnN₅

manganese(2+);(4S,9S,14S,19S)-3,10,13,20,26-pentazatetracyclo[20.3.1.0^4,9.0^14,19]hexacosa-1(26),22,24-triene;dichloride

• Manganese, dichloro((4aS,13aS,17aS,21aS)-1,2,3,4,4a,5,6,12,13,13a,14,15,16,17,17a,18,19,20,21,21a-eicosahydro-7,11-nitrilo-7H-dibenzo(b,H)-5,13,18,21-tetraazacycloheptadecine-kappaN5,kappaN13,kappaN18,kappaN21,kappaN22)-, (pb-7-11-2344’3′)-

CAS 435327-40-5

• A superoxide dismutase mimetic.

• GC 4419
• M-40419
• SC-72325A
• For the Reduction of The Severity and Incidence of Radiation and Chemotherapy-Induced Oral Mucositis

Avasopasem manganese, also known as GC4419, is a highly-selective small molecule mimetic of superoxide dismutase (SOD) being investigated for the reduction of radiation-induced severe oral mucositis.^1,2 This drug has potential application for radiation-induced esophagitis and oral mucositis, in addition to being currently tested against COVID-19.

Avasopasem manganese is a superoxide dismutase mimetic that rapidly and selectively converts superoxide to hydrogen peroxide and oxygen in order to protect normal tissue from radiation therapy-induced damage.¹ This drug is currently being investigated against oral mucositis, esophagitis, and COVID-19.

PATENT

https://patentscope.wipo.int/search/en/detail.jsf?docId=WO2018152353

Transition metal pentaaza 15-membered macrocyclic ring complexes having the macrocyclic ring system corresponding to Formula A have been shown to be effective in a number of animal and cell models of human disease, as well as in treatment of conditions afflicting human patients.

For example, in a rodent model of colitis, one such compound, GC4403, has been reported when administered by intraperitoneal (ip) injection to significantly reduce the injury to the colon of rats subjected to an experimental model of colitis (see Cuzzocrea et al., Europ. J. Pharmacol., 432, 79-89 (2001)).

GC4403 administered ip has also been reported to attenuate the radiation damage arising both in a clinically relevant hamster model of acute, radiation-induced oral mucositis (Murphy et al., Clin. Can. Res., 74(13), 4292 (2008)), and lethal total body irradiation of adult mice (Thompson et al., Free Radical Res., 44(5), 529-40 (2010)).

Similarly, another such compound, GC4419, administered ip has been shown to attenuate VEGFr inhibitor-induced pulmonary disease in a rat model (Tuder, et al., Am. J. Respir. Cell Mol. Biol., 29, 88-97 (2003)), and to increase the anti-tumor activity of anti-metabolite and anti-mitotic agents in mouse cancer models (see, e.g., WO2009/143454). In other studies, GC4419 and GC4403 have been shown to be similarly potent in various animal models of disease. Additionally, another such compound, GC4401, administered ip has been shown to provide protective effects in animal models of septic shock (S. Cuzzocrea, et. al., Crit. Care Med., 32(1 ), 157 (2004)) and pancreatitis (S. Cuzzocrea, et. al., Shock, 22(3), 254-61 (2004)).

[0003] Certain of these compounds have also been shown to possess potent anti-inflammatory activity and prevent oxidative damage in vivo. For example, GC4403 administered ip has been reported to inhibit inflammation in a rat model of inflammation (Salvemini, et.al., Science, 286, 304 (1999)), and prevent joint disease in a rat model of collagen-induced arthritis (Salvemini et al., Arthritis & Rheumatism, 44(12), 2009-2021 (2001)). In addition, these compounds have been reported to possess analgesic activity and to reduce inflammation and edema by systemic administration in the rat-paw carrageenan hyperalgesia model, see, e.g., U.S. Pat. No. 6,180,620.

[0004] Compounds of the class comprising GC4419 have also been shown to be safe and effective in the prevention and treatment of disease in human subjects. For example, GC4419 administered by intravenous (iV) infusion has been shown to reduce oral mucositis in head-and-neck cancer patients undergoing chemoradiation therapy (Anderson, C, Phase 1 Trial of Superoxide Dismutase (SOD) Mimetic GC4419 to Reduce Chemoradiotherapy (CRT)-lnduced Mucositis (OM) in Patients (pts) with Mouth or Oropharyngeal Carcinoma (OCC), Oral Mucositis Research Workshop,

MASCC/ISOO Annual Meeting on Supportive Care in Cancer, Copenhagen, Denmark (June 25, 2015)).

[0005] However, the administered dose when delivered systemically, for example by a parenteral route, can be limited in animal models and particularly in humans by systemic exposure and resulting toxicity that appears to be similar in nature among the pentaaza 15-membered macrocyclic ring dismutase mimetics of Formula A, particularly GC4403, GC4419, GC4401 and related compounds sharing the dicyclohexyl and pyridine motif in the macrocycle ring (e.g., compounds sharing the dicyclohexyl and pyridine motif generally include compounds according to Formula (I) below herein having W as an unsubstituted pyridine moiety, and wherein U and V are transcyclohexanyl fused rings) . For example, the maximum tolerated dose of GC4403 delivered as a 30-minute iv infusion in humans is 25 mg, or roughly 0.35 mg/kg in a 70-kg subject, and similar limitations exist for animal parenteral dosing. Thus, the efficacy of treatment of conditions such as local inflammatory disease or tissue damage of the alimentary canal may be limited when using systemic delivery of GC4403 and similar compounds.

[0006] In each of these compounds comprising the pentaaza 15-membered macrocyclic ring of Formula A, the five nitrogens contained in the macrocyclic ring each form a coordinate covalent bond with the manganese (or other transition metal coordinated by the macrocycle) at the center of the molecule. Additionally, manganese (or other appropriate transition metal coordinated with the macrocycle) forms coordinate covalent bonds with “axial ligands” in positions perpendicular to the roughly planar macrocycle. Such coordinate covalent bonds are characterized by an available “free” electron pair on a ligand forming a bond to a transition metal via donation and sharing of the electron pair thus forming a two-electron bond between the metal and the donor atom of the ligand (Cotton, F.A. & G. Wilkinson, Advanced Inorganic Chemistry, Chapter 5, “Coordination Compounds”, 2^nd revised edn., Interscience Publishers, p.139 (1966); lUPAC Gold Book, online version http://goldbook.iupac.org/C01329.html). The coordinate covalent nature of the bonds between manganese (or other such appropriate transition metal) and the five macrocyclic ring nitrogens and between manganese (or other such transition metal) and each of the two chloro axial ligands is evidenced, for example, by the “single crystal” X-ray crystal structure of GC4403 (Fig. 11 ) and GC4419 (Fig. 12).

[0007] Coordination compounds contrast with ionic compounds, for example, salts, where in the solid state the forces between anions and cations are strictly coulombic electrostatic forces of attraction between ions of opposite charge. Thus, in salts, discrete cations and anions provide the force to maintain the solid state structure; e.g., such as the chloride ion and the sodium ion in a typical salt such as sodium chloride (Cotton, F.A. & G. Wilkinson, Advanced Inorganic Chemistry, Chapter 5, “The Nature of Ionic Substances”, 2^nd revised edn., Interscience Publishers, pp. 35-36, 45-49 (1966).

[0008] Although pentaaza 15-membered macrocyclic ring complexes have been disclosed in the literature for a number of anti-inflammatory indications, the representative disclosures discussed above illustrate that such compounds are generally administered by intraperitoneal (ip) or intravenous (iv) injection to potentiate systemic bioavailability. Local (e.g. topical) administration has been reported as ineffective in animal models of inflammatory disease, particularly when measured against the efficacy of systemic administration methods (Murphy et al., Clin. Can. Res., 74(13), 4292 (2008); WO 2008/045559). One research group has reported inhibition of colonic tissue injury and neutrophil accumulation by intracolonic administration of a prototype pentaaza macrocycle superoxide dismutase mimetic (MnPAM) (having a different structure from GC4403), though that disclosure neither addresses systemic bioavailability of the compounds described therein, nor explore limitations resulting from systemic bioavailability impacting safety and/or efficacy of that specific compound (Weiss et al., J. Biol. Chem., 271(42): 26149-26156 (1996); Weiss, R. and Riley, D., Drugs Future, 21 (4): 383-389 (1996)).

[0009] Aspects of the present disclosure provide for formulations of pentaaza macrocyclic ring complexes of the class comprising GC4419, GC4403, and GC4401 that exhibit limited systemic bioavailability when administered orally (e.g. less than 20%, less than 15%, and even less than 10% bioavailability when dosed in appropriate oil-based formulations; see Table 1 and when combined with other formulations even less than 5%, and even less than 1%; see Example 28). In general, drug absorption from the gastrointestinal tract occurs via passive uptake so that absorption is favored when the drug is in a non-ionized (neutral) and lipophilic form. See, e.g., Goodman & Gilman’s: The Pharmacological Basis of Therapeutics, Ninth Edition, p. 5-9 (1996). Without wishing to be limited to any particular theory, this is also believed to be the case for this class of compounds, as exemplified by GC4403, where the axial ligands are both chloro moieties forming a coordinate covalent bond to the manganese and a neutral complex results:

The Mn(ll) pentaaza macrocyclic ring dichloro complexes, such as GC4419, GC4401, GC4444, and GC4403 (structures shown below) were synthesized using literature procedures. For GC4403 the chiral R,R-diaminocyclohexane is utilized as starting material,² whereas for GC4419, the mirror-image enantiomer of GC4403, the chiral S,S-diaminocyclohexane is utilized instead.^3,4 The remainder of the synthesis of GC4419 can be identical in all respects to the method published for GC4403.² The synthesis of the GC4401 complex was reported previously in reference 5.

[00213] The synthesis of GC4444 which contains the additional 11-R-Methyl substituent generating a fifth chiral center on carbon (and is also derived from R,R-diaminocyclohexane) is made from the corresponding chiral tetraamine whose synthesis was published in reference 6 as Example 5C.

Syntheses of Axial Ligand Derivatives

[00214] The same Mn(II) pentaaza macrocyclic ring dichloro complexes (GC4419, GC4403, GC4444 and GC4401 ) were also used as the starting material precursors for the syntheses of other axial ligand bound derivatives using a generic synthesis scheme in which a large excess of a salt of an anion is used to displace the chloro ligand thereby generating the new compound.

EXAMPLE 2

[00215] Synthesis of Manganese(ll)bis-acetato[(4aS,13aS,17aS,21aS)-1,2,3,4,48,5,6,12,13,13a,14,15,16,17, 17a,18,19,20,21,21a- Eicosahydro-11,7-nitrilo-7H-dibenzo[b,h][1,4,7,10] tetraazacycloheptadecine-KN5, κΝ13, κΝ18, κΝ21, κΝ22]-, [bis-Acetato (GC4419)]. GC4701

[00216] Using a 500-mL Erlenmeyer, 100 mL of deionized (“DI”) water was added to 5.3 g of GC4419; the mixture was stirred vigorously for 15-20 min, then sonicated for 5 min. The resulting light brownish suspension was filtered through a 10-20 μ fritted funnel (ca. 0.3 g of solid material remained in the funnel). The resulting clear solution was added into a sodium acetate solution (ca. 429 mmol, 21 equiv in 100 mL DI water) as a stream in one portion. No solid separated and the yellowish solution was stirred for 5 additional min. The solution was transferred to a separatory funnel and extracted (3 X 50 mL) with dichloromethane. The organic layers were separated, combined, and transferred back into a separatory funnel. The dichloromethane solution was back-extracted (2 X 50 mL) with aqueous sodium acetate (32 g/100 mL). The dichloromethane layer was dried over MgSO₄ (ca. 10 g) for 30 min (w/stirring), filtered using a 10-20 μ fritted funnel, and the solution taken to dryness using a rotavap. To the yellow oily solid resulting from taking the solution to dryness was added methanol (50 mL). This solution was then again taken to dryness on the rotovap to yield a light yellow foam/glass. This material was dried in vacuo at room temperature for two days.

[00217] The isolated yellowish brittle (4.11 g, 75% yield based on GC4419) was analyzed by HPLC and showed a purity of 99.7% and elemental analysis showed 0.98 wt. % residual chlorine. The elemental analysis is consistent with the expected bis-(acetato) structure C₂₅H₄₁MnN₅O₄●2H₂O. Anal Cal’d: C, 53.00% ; H, 8.01 %; N, 12.36%, and Mn, 9.70%. Anal Found: C, 53.10% ; H, 8.34% ; Mn, 9.86%, N, 12.56%, and CI (as total halogen content), 0.98 wt. %.

Patent

WO 2002071054

https://patents.google.com/patent/WO2002071054A1/enSuperoxide dismutase (SOD) enzymes are enzymes that catalyze the dismutation of the free radical superoxide, the one-electron reduction product of molecular oxygen. The dismutation of the free radical superoxide involves the conversion of this one-electron reduction product of molecular oxygen to the nonradical molecular oxygen. Superoxide dismutase enzymes are a class of oxidoreductases which contain either Cu/Zn, Fe, or Mn at the active site. Superoxide dismutase (SOD) mimetic compounds are low molecular weight catalysts which mimic the natural enzyme function of the superoxide dismutase enzymes. Thus, superoxide dismutase mimetic compounds also catalyze the conversion of superoxide into oxygen and hydrogen peroxide, rapidly eliminating the harmful biologically generated superoxide species that are believed to contribute to tissue pathology in a number of diseases and disorders. These diseases and disorders include reperfusion diseases, such as those following myocardial infarct or stroke, inflammatory disorders such as arthritis, and neurological disorders such as Parkinson’s disease. Chem Reviews, 1999 vol 99, No. 9, 2573-2587.Superoxide dismutase mimetic compounds possess several advantages over the superoxide dismutase enzymes themselves in that their chemical properties can be altered to enhance stability, activity and biodistribution while still possessing the ability to dismutase the harmful superoxide. Superoxide dismutase mimetic compounds have generated intense interest and have been the focus of considerable efforts to develop them as a therapeutic agent for the treatment of a wide range of diseases and disorders, including reperfusion injury, ischemic myocardium post-ischemic neuropathies, inflammation, organ transplantation and radiation induced injury. Most of the superoxide dismutase mimics currently being developed as therapeutic agents are synthetic low molecular weight manganese-based superoxide dismutase mimetic compounds. Chem Reviews, 2576. Superoxide dismutase mimetic compounds are metal complexes in which the metal can coordinate axial ligands. Examples of such metal complexes include, but are not limited to, complexes of the metals Mn and Fe. Many of the complexes of the metals Mn and Fe do not possess superoxide dismutase activity but possess properties that enable them to be put to other therapeutic and diagnostic uses. These therapeutic and diagnostic uses include MRI imaging enhancement agents, peroxynitrite decomposition catalysts, and catalase mimics. These metal complexes, however, share the structural similarity of possessing a metal that can coordinate exchangeable ligands. These metal complexes exist in water as a mixture of species in which various ligands are possible. An illustration of such a mixture is provided by M40403 , a Mn(π) complex of a nitrogen-containing fifteen membered macrocyclic ligand, shown in Scheme 1. One of the forms for this metal complex is the dichloro complex, which when dissolved in water another form is generated where one of the chloride anions immediately dissociates from the metal generating the [Mn(Cl)(aquo)]+ complex. The problem in aqueous solvent systems or any solvent which has a potential donor atom is that there are a variety of potential ligands available to coordinate axially to the Mn(π) ion of the complex, hi conducting an analysis of a sample containing a metal complex by high performance liquid chromatography (HPLC) the chromatogram tends to be very broad and unresolved due to the presence of the various species of complexes, as shown in Scheme 1. This phenomena makes the identification and quantification of metal complexes by standard HPLC techniques quite difficult. Therefore, in light of the developing roles of metal complexes as therapeutics in the treatment of various disorders and diagnostic agents, a substantial need exists for an effective and workable high performance liquid chromatography method for analyzing metal complexes.

Scheme 1An additional complication which exists is the issue of the acid stability of the metal complex. As the pH decreases, the rate at which the complex becomes protonated and experiences instability increases. This presents particular problems for the use of HPLC as a method of detection and quantification of the metal complexes because the mobile phase used for reverse phase HPLC frequently contains mixtures of organic solvents and water in various combinations with trifluoroacetic acid. The trifluoroacetic acid is commonly present between about 0.1 to about 0.5% by weight. The presence of the trifluoroacetic acid causes the complex to dissociate. This dissociation destroys the potential of any such method to be used for release testing for purity. Furthermore, the trifluoroacetate anion causes the formation of some of the trifluoroacetato complex which could possess a different retention time from the chloro complexes thus, confusing the chromatography. Thus, the phenomenon of ligand exchange, coupled with the acid instability of the metal complexes, provides considerable challenges to the effort to detect and quantify metal complexes using HPLC. These challenges and needs have surprisingly been met by the invention described below.Analytical HPLC is a powerful method to obtain information about a sample compound including information regarding identification, quantification and resolution of a compound. HPLC has been used particularly for the analysis of larger compounds and for the analysis of inorganic ions for which liquid chromatography is unsuitable. Skoog, D.A., West, M.A., Analytical Chemistry, 1986, p. 520. As an analytical tool HPLC takes advantage of the differences in affinity that a particular compound of interest has for the stationary phase and the mobile phase (the solvent being continuously applied to the column). Those compounds having stronger interactions with the mobile phase than with the stationary phase will elute from the column faster and thus have a shorter retention time. The mobile phase can be altered in order to manipulate the interactions of the target compound and the stationary phase. In normal-phase HPLC the stationary phase is polar, such as silica, and the mobile phase is a nonpolar solvent such as hexane or isopropyl ether. In reversed- phase HPLC the stationary phase is non-polar, often a hydrocarbon, and the mobile phase is a relatively polar solvent. Since 1974 when reversed-phase packing materials became commercially available, the number of applications for reversed- phase HPLC has grown, and reversed- phase HPLC is now the most widely used type of HPLC. Reversed-phase HPLC’s popularity can be attributed to its ability to separate a wide variety of organic compounds. Reversed-phase chromatography is especially useful in separating the related components of reaction mixtures, and therefore is a useful analytical tool for determining the various compounds produced by reactions. To create a non-polar stationary phase silica or synthetic polymer based adsorbents are modified with hydrocarbons. The most popular bonded phases are Cl, C4, C8 and C18. Silica based adsorbents modified with trimethylchlorosilane (Cl) and butyldimethylchlorosilane (C4) have a few applications in HPLC, mainly for protein separation or purification. These adsorbents show significant polar interactions. Octyl (C8) and octadecyl (C18) modified adsorbents are the most widely used silica based adsorbents, with almost 80% of all HPLC separations being developed with these adsorbents.The most important parameter in reversed-phase HPLC is the mobile phase. The type of mobile phase employed in the HPLC will have a significant effect on the retention of the analytes in the sample, and varying the composition of the mobile phase allows the chromatographer to adjust the retention times of target components in the mixture to desired values. This ability provides the HPLC method with flexibility. The mobile phase in reversed-phase chromatography has to be polar and it also has to provide reasonable competition for the adsorption sites for the analyte molecules. Solvents that are commonly employed as eluent components in reversed-phase HPLC are acetonitrile, dioxane, ethanol, methanol, isopropanol, tetrahydrofuran, and water. In reversed phase HPLC of high molecular weight biological compounds, the solvents acetonitrile, isopropanol or propanol are most frequently used. Popular additives to the mobile phase for the improvement of resolution include mixtures of phosphoric acid and amines and periϊuorinated carboxylic acids, especially trifluoroacetic acid (TFA). HPLC exploits the differences in affinity that a particular compound of interest has for the stationary phase and the mobile phase. This phenomenon can be utilized to separate compounds based on the differences in their physical properties. Thus, HPLC can be used to separate stereoisomers, diastereomers, enantiomers, mirror image stereoisomers, and impurities. Stereoisomers are those molecules which differ from each other only in the way their atoms are oriented in space. The particular arrangement of atoms that characterize a particular stereoisomer is known as its optical configuration, specified by known sequencing rules as, for example, either + or – (also D or L) and/or R or S. Stereoisomers are generally classified as two types, enantiomers or diastereomers. Enantiomers are stereoisomers which are mirror-images of each other. Enantiomers can be further classified as mirror-image stereoisomers that cannot be superimposed on each other and mirror-image stereoisomers that can be superimposed on each other. Mirror- image stereoisomers that can be superimposed on each other are known as meso compounds. Diastereomers are stereoisomers that are not mirror images of each other. Diastereomers have different physical properties such as melting points, boiling points, solubilities in a given solvent, densities, refractive indices, etc. Diastereomers can usually be readily separated from each other by conventional methods, such as fractional distillation, fractional crystallization, or chromatography, including HPLC.Enantiomers, however, present special challenges because their physical properties are identical. They generally cannot be separated by conventional methods, especially if they are in the form of a racemic mixture. Thus, they cannot be separated by fractional distillation because their boiling points are identical and they cannot be separated by fractional crystallization because their solubilites are identical (unless the solvent is optically active). They also cannot be separated by conventional chromatography such as HPLC because (unless the adsorbent is optically active) they are held equally onto the adsorbent. HPLC methods employing chiral stationary phases are a very common approach to the separation of enantiomers. To be able to separate racemic mixtures of stereoisomers, the chiral phase has to form a diastereomeric complex with one of the isomers, or has to have some other type of stereospecific interaction. The exact mechanism of chiral recognition is not yet completely understood. In reversed-phaseHPLC a common type of chiral bonded phase is chiral cavity phases.The ability to be able to separate diastereomers and enantiomers by HPLC is a useful ability in evaluating the success of synthetic schemes. It is often desirable to separate stereoisomers as a means of evaluating the enantiomeric purity of production samples. All references listed herein are hereby incorporated by reference in their entiretyExamples 1 (traditional mobile phase) and 2 (mobile phase containing excess of salt of a coordinating anion).

+X^“

Scheme 2 Any metal complex possessing a metal that is capable of coordinating a monodentate ligand can be used in the present invention. Examples of such metal complexes include, but are not limited to, complexes of the metals Mn and Fe. The metal complexes of the invention preferably have therapeutic and diagnostic utilities. These therapeutic and diagnostic utilities include, but are not limited to, use as superoxide dismutase mimetic compounds, MRI imaging enhancement agents, peroxynitrite decomposition catalysts, and catalase mimics. The preferred metal complexes for use in the invention are superoxide dismutase mimetic compounds. Examples of such superoxide dismutase mimetic compounds include, but are not limited to, the following complexes of the metals Mn and Fe. Iron based superoxide dismutase mimetics include, but are not limited to, Fe^ra(salen) complexes, Fe^ra(l,4,7,10,13-pentaazacyclopentadecane) derivatives and Fe^ffl(porphyrinato) complexes. Manganese based superoxide dismutase mimetic compounds include, but are not limited to, metal complexes containing manganese(π) or manganese(m). Examples of manganese based superoxide dismutase mimetic compounds include Mn^m(porphyrinato) complexes, Mn^ffl(salen) complexes, and Mn^π(l ,4,7, 10, 13-pentaazacyclopentadecane) derivatives. Mn^π(l ,4,7, 10,13- pentaazacyclopentadecane) derivatives are more preferred for use in the invention. Examples of Mn^π(l,4,7,10,13-pentaazacyclopentadecane) derivatives preferred for use in the invention include, but are not limited to, M40403 and M40401, as shown in Scheme 3 below.Furthermore, stereoisomers of all of the above metal complexes can be used in the process of the present invention. Diastereomers of the same metal complexes can also be detected and separated by the method of the present invention. As it is often desirable to separate stereoisomers as a means of evaluating the chemical and optical purity of production samples, the metal complexes can also comprise products of a reaction stream. Enantiomers of any of the metal complexes referenced above can be used in the chiral HPLC method of the invention for the separation of enantiomers of a metal complex.

M40403 M40401

M40484Scheme 3The ligand is a coordinating anion that binds to the metal cation of the metal complex. The coordinating anion can serve as an axial ligand for a superoxide dismutase mimetic compound. Examples of such anions include, but are not limited to, chloride anions, thiocyanate anions, stearate anions, acetate anions, trifluoroacetate anions, carboxylate anions, formate anions, or azide anions. Preferred anions include chloride anions, thiocyanate anions, and formate anions. More preferred anions are chloride anions. The more preferred anions in the chiral HPLC embodiment of the invention are thiocyanate anions. When present in an excess, the thiocyanate anions bind to the coordinating metal of the complexes preferentially to the chloride anions. An excess of thiocyanate anions will produce the bis(thiocyanato) complexes of M40403 and M40419 as shown in Scheme 4.

M40403 M40403-(SCN)₂

M40419 M40419-(SCN)₂Scheme 4An example of the use of the acetate anion as the coordinating anion with M40403 is shown in Scheme 5 below. Scheme 6 illustrates the use of the formate anion as the coordinating anion with M40403.

M40403 M40403-(OAc)₂Scheme 5

M40403 M40403-(Formate)₂Scheme 6The coordinating anion is supplied by a salt of the coordinating anion. Salts of the chloride anion include, but are not limited to, sodium chloride, lithium chloride, potassium chloride, ammonium chloride, or tetraalkylammonium chloride. Preferred salts of the chloride anion include sodium chloride, lithium chloride and tetrabutylammonium chloride. Salts of the thiocyanate anion include, but are not limited to, sodium thiocyanate, potassium thiocyanate, ammonium thiocyanate, or lithium thiocyanate. Preferred salts of the thiocyanate anion include sodium thiocyanate and potassium thiocyanate. Salts of the acetate anion include, but are not limited to, potassium acetate, sodium acetate, ammonium acetate, ammonium trifluoroacetate and lithium acetate. Preferred salts of the acetate anion include ammonium acetate. Salts of the formate anion include, but are not limited to, potassium formate, sodium formate, ammonium formate and lithium formate. Preferred salts of the formate anion include ammonium formate. Salts of the cyanate anion include but are not limited to, sodium cyanate, potassium cyanate, or ammonium cyanate. Salts of the carboxylate anion include, but are not limited to, potassium carboxylate, ammonium carboxylate and sodium carboxylate. Salts of the stearate anion include, but are not limited to, lithium stearate and sodium stearate. Salts of the azide anion include, but are not limited to, sodium azide, potassium azide, and lithium azide. The salt added to the mobile phase can also be a mixture of any of these salts. Examples include a mixture of tetrabutylammonium chloride and lithium chloride.EXAMPLESExperimental For Examples 1-8 Chemicals, Solvents and MaterialsAll solvents used in the study were HPLC grade or equivalent. All chemicals were ACS reagent grade or equivalent.HPLC System and Data AnalysisThe HPLC chromatography was performed using a Gilson system (Model 306 pump, Model 155 UN-V detector, Model 215 liquid handler, Unipoint Software,Win98), a Narian system (Model 310 pump, Model 340 UN-N detector, Model 410 autosampler Star Workstation, Win98) or SSI system (Acuflow Series IN pump, Acutect 500 UV-N detector, Alcott Model 718 autosampler, HP Model 3395 integrator).Example 1HPLC Analysis of M40403 using Method 1

M40403 Method 1: Analytical Column: Waters YMC ODS-AQ S5 120A (4.6 x 50 mm); System A: 0.1% trifluoroacetic acid in H₂O; System B: 0.08% trifluoroacetic acid in acetonitrile; Gradient: 10-50% system B over 10 min; Flow rate: 3ml/min; Detector wavelength: 265. Injected 20 μl of stock solution of M40403 prepared by dissolving 1 mg in 1 ml of water and diluting with 1 ml of system A. The HPLC chromatogram of M40403 using method 1 is shown in Figure 1. Example 2 HPLC Analysis of M40403 using Method 2Method 2: Analytical Column: Waters YMC 9DS-AQ S5 12θΛ (4.6 x 50 MM); System A: 0.5 N aqueous NaCl; System B: 1 :4 water/CH₃CN; Gradient: 10-50% system B over 9 min; Flow rate: 3mL/min; Detector wavelength: 265 nm. Injected 20 μl of stock solution of M40403 prepared by dissolving 1 mg in 1 ml of system A. The HPLC chromatogram of M40403 using method 2 is shown in Figure 2.Example 3 HPLC Analysis of M40403 using Method 3Method 3: Analytical Column: Waters Symmetry Shield RP18, 5 μm, 250 x 4.6 mm;Mobile Phase: Acetonitrile: 0.125 M Tetrabutylammonium Chloride in water (pH 6.5), 5%: 95% H₂0(v/v); Flow rate: 1 mL/min; Detection wavelength: 265nm. Injected 20 μl of stock solution of M40403 prepared by dissolving 1 mg in 1 ml of mobile phase. The HPLC chromatogram of M40403 using method 3 is shown in Figure 3.The HPLC chromatogram of M40403 and related compounds using method 3 is shown in Figure 3a. Method 3 allows a separation of M40402 (bisimine of M40403), M40414 (monoimine of M40403) and M40475 (free ligand of M40403) (see chromatogram in Figure 3a).Example 4HPLC Analysis of M40403 using Method 4Method 4: Analytical Column: Waters Symmetry Shield RP18, 5 μm, 250 x 4.6 mm; Mobile Phase: Acetonitrile: 0.125 M Tetrabutylammonium Chloride and 0.5 M LiCl in water (pH 6.5), 5%: 95% H₂0 (v/v); Flow rate: lmL/min; Detection wavelength: 265 nm. Injected 20 μl of stock solution of M40403 prepared by dissolving 1 mg in 1 ml of system A. The HPLC chromatogram of M40403 using method 4 is shown in Figure 4.The HPLC chromatogram of M40403 and related compounds using method 4 is shown in Figure 4a. Method 4 allows a separation of M40402 (bisimine of M40403), M40414 (monoimine of M40403) and M40475 (free ligand of M40403) and all diastereomers of M40403 (see chromatogram in Figure 4a).Example 5 HPLC Analysis of M40401 using Method 1

M40401 Method 1: Analytical Column: Waters YMC ODS-AQ S5 120A (4.6 x 50 mm); System A: 0.1 % trifluoroacetic acid in H₂O; System B: 0.08% trifluoroacetic acid in acetonitrile; Gradient: 10-50% system B over 10 min; Flow rate: 3ml/min; Detector wavelength: 265. Injected 20 μl of stock solution of M40401 prepared by dissolving 1 mg in 1 ml of water and diluting with 1 ml of system A. The HPLC chromatogram of M40401 using method 1 is shown in Figure 5.Example 6 HPLC with various NaCl concentrations:An HPLC was taken of M40401 with various concentrations of NaCl.Analytical Column: Waters YMC 9DS-AQ S5 120 A (4.6 x 50 mm);System A: (A) H₂O (no NaCl) ; (B) 0.01 M NaCl in water; (C) 0.5 M NaCl in water;System B: acetonitrile; Gradient: 0-100% system B over 10 min; Flow: 3 ml/min;Detector wavelength: 265 nm. Injected 20 μl of stock solution of M40401 prepared by dissolving 1 mg in 1 ml of system A. The HPLC chromatogram of M40401 using various NaCl concentrations is shown in Figure 6. Example 7 HPLC Analysis of M40401 using Method 2Method 2: Analytical Column: Waters YMC ODS-AQ S5 12θΛ (4.6 x 50 MM); System A: 0.5 N aqueous NaCl; System B: 1 :4 water/CH₃CN; Gradient 1 : 10-50% system B over 9 min; Flow rate: 3 mL/min; Detector wavelength: 265 nm. Injected 20 μl of stock solution of M40403 prepared by dissolving 1 mg in 1 ml of system A.The HPLC chromatogram of M40401 using method 2 is shown in Figure 7. Method 2 allows a separation of M40472 (bisimine of M40401), M40473 (monoimine of M40401), free ligand of M40403 and two isomers of M40401 (M40406, M40404).Example 8HPLC Analysis of M40401 using Method 3Method 3: Analytical Column: Waters Symmetry Shield RP18, 5 m, 250 4.6 mm; Mobile Phase: Acetonitrile: 0.125 M Tetrabutylammom^‘um Chloride in H₂0 (pH 6.5), 5: 95%) H₂0 (v/v); Flow rate: lmL/min; Detection wavelength: 265 nm. The HPLC chromatogram of M40401 using method 3 is shown in Figure 8.Method 3 allows a separation of M40472 (bisimine of M40401), M40473 (monoimine of M40401), free ligand of M40403 and two isomers of M40401 (M40406, M40404).Example 9 HPLC Analysis of M40401 using Method 4Method 4: Analytical Column: Waters Symmetry Shield RP18, 5 μm, 250 x 4.6 mm;Mobile Phase: Acetonitrile: 0.125 M Tetrabutylammonium Chloride and 0.5 M LiCl in water (pH 6.5), 5: 95%> H₂O (v/v); Flow rate: 1 mL/min; Detection wavelength: 265 nm; Injected 20 μl of stock solution of M40401 prepared by dissolving 1 mg in 1 ml of a mobile phase. The HPLC chromatogram of M40401 using method 4 is shown in Figure 9.The HPLC chromatogram of M40401 and related compounds using method 4 is shown in Figure 9a. Method 4 allows a separation of M40472 (bisimine of M40401), M40473 (monoimine of M40401), free ligand of M40403 and two isomers of M40401 (M40406, M40404). Example 10HPLC of M40403-(HCOO^“)₂ Using Formate AnionAn HPLC of M40403 employing the formate anion was taken. Analytical Column: Waters YMC 9DS-AQ S5 120 A (4.6 x 50 mm); System A: 0.025 M ammonium formate in water; System B: 1 : 4 = 0.125 M ammonium formate in water/ acetonitrile; Gradient: 0-100% system B over 10 min; Flow: 3 ml/min;Detector wavelength: 265 nm. Injected 20 μl of stock solution of M40403-(Formate)₂ prepared by dissolving 1 mg in 1 ml of system A. The HPLC chromatogram of M40403-(HCOO^“)₂ is shown in Figure 10.Example 11 HPLC of M40403-(OAc)₂ Using Acetate AnionAn HPLC of M40403 employing the acetate anion was taken.Analytical Column: Waters YMC 9DS-AQ S5 120 A (4.6 x 50 mm); System A: 0.025 M ammonium acetate in water; System B: 1: 4 = 0.125 M ammonium acetate in water/ acetonitrile; Gradient: 0-100% system B over 10 min; Flow: 3 ml/min;Detector wavelength: 265 nm. Injected 20 μl of stock solution of M40403-(OAc)₂ prepared by dissolving 1 mg in 1 ml of system A. The HPLC chromatogram of M40403 -(OAc)₂ is shown in Figure 11.Example 12An HPLC method to separate the diastereomers of superoxide dismutase mimetic compound M40403. Four stereoisomer mixtures were prepared (Part A) as shown in Schemes 5-9 and then separated (Part B) via reversed-phase high performance liquid chromatography. Part A: Synthesis of Stereoisomers Of M40403M40403 is synthesized from its single-isomer, tetra-amine precursor M40400 in the reaction shown in Scheme 7.

M40400 M40402

M40403Scheme 7The various stereoisomers of M40403 are synthesized from the various isomers of 1,2-diaminocyclohexane which provides the chiral carbon centers in M40403. The 1,2-diaminocyclohexane isomers used to prepare the R,R+R,S) M40403 stereoisomer mixture of Set 1 are shown in Scheme 6. Similarly, the 1,2-diaminocyclohexane isomers used to prepare the (R,R+S,S) M40403 stereoisomer mixture of Set 2 are shown in Scheme 7. The 1,2-diaminocyclohexane isomers used to prepare the (R,S+R,S) M40403 stereoisomer mixture of Set 3 are shown in Scheme 8. The 1,2- diaminocyclohexane isomers used to prepare the (S,S+R,S) M40403 stereoisomer mixture of Set 4 are shown in Scheme 9. As shown in Schemes 6-9 the M40403 diastereomers are prepared by template cyclization, followed by reduction with sodium borohydride.

Scheme 8

(S.S.S.S)Scheme 9

(S.R.R.S)Scheme 10

Scheme 11Table 1

Part B: Separation of Stereoisomer MixturesChemicals, Materials, and MethodsTetrabutylammonium chloride hydrate (98%, 34,585-7) was purchased from Aldrich Chemical Company. Sodium chloride (99.6%, S-9888) was purchased from Sigma Chemical Company. All other solvents (HPLC-grade unless otherwise indicated) and reagents were purchased from Fisher Scientific and were of the finest grade available. The SymmetryShield® RP₁₈ column (4.6 mm x 250 mm, 5 μm particle size) and its corresponding guard column were purchased from Waters Corporation. Reversed-Phase HPLC ExperimentsPreparation of Standard SolutionsHPLC Mobile phased was an aqueous solution consisting of 0.125 M tetrabutylammonium chloride (TBAC) and 0.5 M LiCl, prepared by adding tetrabutylammonium chloride hydrate (36.99 g) and solid LiCl (21.2 g) to a 1 L volumetric flask, diluting to volume with Millipore water, and inverting the flask several times to obtain a homogeneous solution. The resulting solution was filtered through a 0.45 μm nylon filter prior to use. Mobile phase B was HPLC-grade acetonitrile. Samples of each diastereoisomer set for HPLC-UN analysis were prepared at concentrations of ~ 3.0 mg/mL in a 50:50 mixture of 0.5 M LiCl in MeOH:

PATENT

WO/2021/163397

SOLID STATE FORMS OF AVASOPASEM MANGANESE AND PROCESS FOR PREPARATION THEREOF

Avasopasem manganese (GC4419), has the following chemical structure:

[0003] Avasopasem manganese is a highly selective small molecule superoxide dismutase (SOD) mimetic which is being developed for the reduction of radiation-induced severe oral mucositis (SOM). The compound is described in U.S. Patent No. 8,263,568.

[0004] Polymorphism, the occurrence of different crystalline forms, is a property of some molecules and molecular complexes. A single molecule may give rise to a variety of polymorphs having distinct crystal structures and physical properties like melting point, thermal behaviors (e.g., measured by thermogravimetric analysis (“TGA”), or differential scanning calorimetry (“DSC”)), X-ray diffraction (XRD) pattern, infrared absorption fingerprint, and solid state (¹³C) NMR spectrum. One or more of these techniques may be used to distinguish different polymorphic forms of a compound.

[0005] Different salts and solid state forms (including solvated forms) of an active pharmaceutical ingredient may possess different properties. Such variations in the properties of different salts and solid state forms and solvates may provide a basis for improving formulation, for example, by facilitating better processing or handling characteristics, changing the

dissolution profile in a favorable direction, or improving stability (polymorph as well as chemical stability) and shelf-life. These variations in the properties of different salts and solid state forms may also offer improvements to the final dosage form, for instance, if they serve to improve bioavailability. Different salts and solid state forms and solvates of an active pharmaceutical ingredient may also give rise to a variety of polymorphs or crystalline forms, which may in turn provide additional opportunities to assess variations in the properties and characteristics of a solid active pharmaceutical ingredient.

[0006] Discovering new solid state forms and solvates of a pharmaceutical product may yield materials having desirable processing properties, such as ease of handling, ease of processing, storage stability, and ease of purification or as desirable intermediate crystal forms that facilitate conversion to other polymorphic forms. New solid state forms of a pharmaceutically useful compound can also provide an opportunity to improve the performance characteristics of a pharmaceutical product. It enlarges the repertoire of materials that a formulation scientist has available for formulation optimization, for example by providing a product with different properties, including a different crystal habit, higher crystallinity, or polymorphic stability, which may offer better processing or handling characteristics, improved dissolution profile, or improved shelf-life (chemical/physical stability). For at least these reasons, there is a need for additional solid state forms (including solvated forms) of Avasopasem manganese.

EXAMPLES

Preparation of starting materials

[00119] Avasopasem manganese can be prepared according to methods known from the literature, for example U.S. Patent No. 8,263,568. Alternatively, Avasopasem manganese can be prepared by the template method reported for the enantiomeric analogue GC4403, which has the formula:

GC4403 is disclosed in International Appl. No. WO 98/58636 (as compound SC-72325) and Riley, D.P, and Schall, O.F., Advances in Inorganic Chemistry (2007), 59, 233-263. Thus, GC4403 can be synthesized via the template route described in the literature using the chiral R,R-l,2-diamminocyclohexane [Salvemini, D., et ah, Science (1999), 286, 304-6 , and Aston, K, et al., Inorg. Chem. (2001), 40(8), 1779-89] Avasopasem manganese (GC4419) can be prepared by the same method except that the chiral R,R-l,2-diamminocyclohexane is replaced with S,S-1 ,2-diamminocyclohexane.

Example 1: Preparation of Avasopasem manganese Form AMI

[00120] Avasopasem manganese (0.1 grams) was dissolved in dichloromethane (0.5 ml) at 25-30°C in a test tube. The solution was filtered through 0.45 micron filter and the clear solution was subjected to slow solvent evaporation at 25°C by covering the tube with paraffin film with a pin hole. After, 2 days, the obtained solid was analyzed by XRD- Form AMI; as shown in Figure 1

1. GlobeNewswire: Galera Therapeutics Announces Avasopasem Manganese Improved Markers of Chronic Kidney Disease in Patients Receiving Cisplatin [Link]
2. Galera Therapeutics: AVASOPASEM (GC4419) [Link]

///////////AVASOPASEM, Avasopasem manganese, GC-4419,  GC4419, GC 4419, M 40419, M40419; M-40419, SC 72325A, SC-72325A, SC72325A,

[Cl-].[Cl-].[Mn++].C1CC[C@@H]2NCC3=CC=CC(CN[C@H]4CCCC[C@@H]4NCCN[C@H]2C1)=N3

NEW DRUG APPROVALS

ONE TIME

$10.00

Click here to purchase.

(Heavy chain)
QVQLVQSGAE VKKPGSSVKV SCKASGYSFS DYYMHWVRQA PGQGLEWMGQ INPTTGGASY
NQKFKGKATI TVDKSTSTAY MELSSLRSED TAVYYCARYY YGRHFDVWGQ GTTVTVSSAS
TKGPSVFPLA PCSRSTSEST AALGCLVKDY FPEPVTVSWN SGALTSGVHT FPAVLQSSGL
YSLSSVVTVP SSSLGTKTYT CNVDHKPSNT KVDKRVESKY GPPCPPCPAP EFLGGPSVFL
FPPKPKDTLM ISRTPEVTCV VVDVSQEDPE VQFNWYVDGV EVHNAKTKPR EEQFNSTYRV
VSVLTVLHQD WLNGKEYKCK VSNKGLPSSI EKTISKAKGQ PREPQVYTLP PSQEEMTKNQ
VSLTCLVKGF YPSDIAVEWE SNGQPENNYK TTPPVLDSDG SFFLYSRLTV DKSRWQEGNV
FSCSVMHEAL HNHYTQKSLS LSLGK
(Light chain)
DIVMTQSPDS LAVSLGERAT INCKASQSVD YDGDSYMNWY QQKPGQPPKL LIYAASNLES
GVPDRFSGSG SGTDFTLTIS SLQAEDVAVY YCQQSNEDPY TFGQGTKLEI KRTVAAPSVF
IFPPSDEQLK SGTASVVCLL NNFYPREAKV QWKVDNALQS GNSQESVTEQ DSKDSTYSLS
STLTLSKADY EKHKVYACEV THQGLSSPVT KSFNRGEC
(Disulfide bridge: H22-H96, H132-L218, H145-H201, H224-H’224, H227-H’227, H259-H319, H365-H423, H’22-H’96, H’132-L’218, H’145-H’201, H’259-H’319, H’365-H’423, L23-L92, L138-L198, L’23-L’92, L’138-L’198)

Pepinemab

VX15/2503

Antineoplastic, Anti-human semaphorin 4D antibody

Monoclonal antibody
Treatment of solid tumors, multiple sclerosis and Huntington’s disease

• Moab VX15/2503
• Pepinemab
• UNII-BPZ4A29SYE
• VX-15
• VX15
• VX15/2503

• OriginatorVaccinex
• DeveloperBristol-Myers Squibb; Children’s Oncology Group; Emory University; Merck KGaA; National Cancer Institute (USA); Teva Pharmaceutical Industries; UCLAs Jonsson Comprehensive Cancer Center; Vaccinex
• ClassAntibodies; Antidementias; Antineoplastics; Immunotherapies; Monoclonal antibodies
• Mechanism of ActionCD100 antigen inhibitors

• Orphan Drug StatusYes – Huntington’s disease
• New Molecular EntityYes

• Phase IIHuntington’s disease
• Phase I/IIAlzheimer’s disease; Non-small cell lung cancer; Osteosarcoma; Solid tumours; Squamous cell cancer
• Phase IColorectal cancer; Malignant melanoma; Pancreatic cancer
• No development reportedMultiple sclerosis

• 22 May 2021Pepinemab is still in phase I trials for Colorectal cancer and Pancreatic cancer in USA (NCT03373188)
• 17 May 2021Phase-I/II clinical trials in Squamous cell cancer (Combination therapy, Late-stage disease, Metastatic disease, Recurrent, Second-line therapy or greater) in USA (IV) (NCT04815720)
• 17 May 2021Vaccinex plans a phase I/II trial for Alzheimer’s disease (In volunteers), in H2 2021

Semaphorin 4D (SEMA4D) plays a role in multiple cellular processes that contribute to the pathophysiology of neuroinflammatory/neurodegenerative diseases. SEMA4D is, therefore, a uniquely promising target for therapeutic development.

Pepinemab is a novel monoclonal antibody that blocks the activity of SEMA4D, and preclinical testing has demonstrated the beneficial effects of anti-SEMA4D treatment in a variety of neurodegenerative disease models. Vaccinex is committed to the development of this potentially important antibody that has the potential to help people with different neurodegenerative disorders that share common mechanisms of pathology.

Note: Pepinemab (VX15/2503) is an investigational drug currently in clinical studies. It has not been demonstrated to be safe and effective for any disease indication. There is no guarantee that pepinemab (VX15/2503) will be approved for the treatment of any disease by the U.S. Food and Drug Administration or by any other health authority worldwide.

////////////////////Pepinemab, VX15/2503, vx 15, Antineoplastic, Anti-human semaphorin 4D antibody, Monoclonal antibody, solid tumors, multiple sclerosis,  Huntington’s disease, PEPTIDES

NEW DRUG APPROVALS

ONE TIME

$10.00

Click here to purchase.

Medigen vaccine

MVC COVID-19 vaccine

• MVC-COV1901

track it https://covid19.trackvaccines.org/vaccines/24/

MVC-COV1901 is a vaccine candidate developed and commercialized by Medigen Vaccine Biologics Corporation. The vaccine candidate contains a perfusion form of the SARS-Cov2 recombinant spike protein. Medigen has combined forces with Dynavax, which offers an advanced adjuvant, CpG 1018 (also known as ISS-1018), for use with its vaccine. As of September 2020, the vaccine candidate is in Phase 1 clinical trials to assess its safety and immunogenicity (NCT04487210).

The MVC COVID-19 vaccine, designated MVC-COV1901 and also known as the Medigen COVID-19 vaccine, is a protein subunit COVID-19 vaccine developed by Medigen Vaccine Biologics Corporation [zh] in Taiwan, American company Dynavax Technologies and the U.S. National Institute of Health.^[1][2]

This vaccine is made by the recombinant S-2P spike protein adjuvanted with CpG 1018 supplied by Dynavax.^[3] Preliminary results from Phase I trials on 77 participants were published in June 2021, indicating what the authors described as “robust” immune system response elicited by the vaccine.^[4]

The study authors have assessed the humoral immune response by measuring quantities of binding IgG to S protein, and also the cellular immune response by measuring the quantities of IFN-γ and IL-4 secreting T cells.^[4]

Taiwan-based Medigen Vaccine Biologics Corporation (MVC) and Dynavax Technologies Corporation, in the US, have announced the rollout of its COVID-19 vaccine, MVC-COV1901. Approximately 600,000 people are anticipated to receive the Medigen vaccine this week.

Ryan Spencer, Chief Executive Officer of Dynavax commented, “We are pleased that Medigen’s vaccine is now available for the people of Taiwan. We are very excited for this first, of hopefully multiple, EUAs and approvals for COVID-19 vaccines that include CpG 1018 adjuvant. Considering the limitations of current vaccines and the global vaccine shortage, we believe adjuvanted vaccines can contribute significantly to current vaccination efforts.”

In July, MVC received Taiwan Emergency Use Authorization and approval for inclusion in Taiwan’s COVID-19 vaccine immunization program, MVC-COV1901.

MVC COVID-19 vaccine is indicated for adults over 20 years old and is administered in two doses 28 days apart for prevention of COVID-19.

The Advisory Committee recommended that MVC should submit safety monitoring report monthly during the declared EUA period and should submit a vaccine effectiveness report within one year after obtaining EUA approval.

(CNN)Taiwan’s President Tsai Ing-wen received her first shot of the island’s homegrown Covid-19 vaccine on Monday, a public show of support for the new drug which is central to plans for inoculation self sufficiency amid low immunization rates and struggles to obtain vaccines from overseas.Monday’s island-wide rollout of the Medigen Covid-19 vaccine, developed by Taipei-based Medigen Vaccine Biologics Corporation, comes after the drug was approved for emergency use last month by Taiwanese authorities for anyone above 20 years old, with at least 28 days between the two doses.The vaccine has yet to complete phase 3 clinical trials and no efficacy data is available.  Paul Torkehagen, Medigen’s director of overseas business development, told CNN in May that the company designed a “very large” phase 2 clinical trial to ensure the vaccine’s safety and effectiveness, with 3,800 participants. Normally, a stage 2 clinical trial only involves several hundred people. Data from the trials showed that 99.8% of participants were able to form antibodies against Covid-19 after taking two doses of the vaccine, Medigen’s CEO Charles Chen said.   
Taiwanese President Tsai Ing-wen, center, receives her first shot of the island’s first domestically developed coronavirus vaccine at the Taiwan University Hospital in Taipei, Taiwan on Monday, August 23.   
Taiwan’s Centers for Disease Control said in a July 19 statement that the vaccine posed no serious health effects. Taiwan has ordered 5 million doses of the vaccine from Medigen and more than 700,000 people have already signed up to receive it, according to Reuters.In a Facebook post after receiving the vaccine at a hospital in Taipei, Tsai said she hadn’t suffered from any post-vaccination pain and thanked the health care workers who had administered the shot.”Taking the vaccine can protect yourself, your family, as well as medical staff,” Tsai wrote. “Let’s do our part in boosting Taiwan’s collective defense against the virus!”With its borders sealed to most travelers and strict measures enacted to contain local outbreaks, Taiwan has so far been largely successful in containing Covid-19, reporting fewer than 16,000 total confirmed infections and 828 deaths. But the island has struggled to vaccinate its more than 23 million population, partly due to difficulties obtaining doses from international suppliers.Taiwan’s government has only managed to import around 10 million Covid-19 vaccines, according to Reuters. In July it ordered another 36 million doses of the Moderna shot.Fewer than 5% of Taiwan’s population has received both doses of their Covid-19 vaccine, according to Reuters, as the island delays second dose vaccinations so more people can receive a first shot.On Monday, Taiwan reported four new Covid-19 cases, according to the Central Epidemic Command Center (CECC). Authorities announced on the weekend they would ease virus prevention measures to allow for larger gatherings and the opening of study centers and indoor amusement parks.But Health and Welfare Minister Chen Shih-chung said current Covid-19 restrictions — which include the closure of bars and nightclubs — would remain in place until at least September 6, with the possibility of an extension if the global outbreak continued to grow.Taiwan could become increasingly isolated if it keeps pursuing its “Covid zero” strategy, with both Australia and New Zealand hinting they might abandon the approach once vaccinations reach a certain level.In an opinion piece published on Sunday, Australian Prime Minister Scott Morrison said that while lockdowns to prevent Covid-19 transmission were “sadly necessary for now,” they may not be once vaccination rates increased to the targets of 70% and 80%.”This is what living with Covid is all about. The case numbers will likely rise when we soon begin to open up. That is inevitable,” he said.In neighboring New Zealand, which has also attempted to eliminate the virus within its borders, Covid-19 response minister Chris Hipkins told local media the highly-contagious Delta variant raised “some pretty big questions about what the long-term future of our plans are.”“At some point we will have to start to be more open in the future,” he said.

History

On 16 February 2020, Medigen Vaccine Biologics Corp. (MVC) signed a collaboration agreement with National Institutes of Health (NIH) for COVID-19 vaccine development. The partnership will allow MVC to obtain NIH’s COVID-19 vaccine and related biological materials to conduct animal studies in Taiwan.^[5]

On 23 July 2020, Medigen Vaccine Biologics (MVC) announced collaboration with Dynavax Technologies to develop COVID-19 vaccine. The COVID-19 candidate vaccine will have the combination of SARS-CoV2 spike protein created by MVC and Dynavax’s vaccine adjuvant CpG 1018, which was used in a previously FDA-approved adult hepatitis B vaccine.^[6][7]

Clinical trials

On 13 October 2020, Medigen Vaccine Biologics received Taiwan’s government subsidies for the initiation of Phase 1 Clinical Trial in Taiwan starting early October. The Phase 1 Clinical Trial was held at National Taiwan University Hospital with 45 participants ranging the age of 20-50.^[8][9]

On 25 January 2021, Medigen Vaccine Biologics initiated Phase 2 Clinical Trial for its COVID-19 vaccine candidate MVC-COV1901 with the first participant being dosed. The Phase 2 Clinical Trial for the MVC COVID-19 vaccine was a randomized, double-blinded, and multi-center clinical trial, planned to enroll 3,700 participants of any age 20 above.^[3][10][11]

On 10 June 2021, Medigen Vaccine Biologics released its COVID-19 vaccine Phase 2 interim analysis results, which demonstrates good safety profile in participants. The Phase 2 Clinical Trial in the end included 3,800 participants with all participants receiving second dose by 28 April 2021. Medigen Vaccine Biologics announced that it will request Emergency Use Authorization (EUA) with the concluding of the Phase 2 Clinical Trial.^[12]

On 20 July 2021, Medigen Vaccine Biologics filed a Phase 3 Clinical Trial IND application with Paraguay’s regulatory authority, which was later approved. The Phase 3 Clinical Trial, however, was different from regular Phase 3 Clinical Trial, which uses immune-bridging trial to compare the performance of MVC COVID-19 vaccine with the Oxford-AstraZeneca COVID-19 vaccine.^[13] The decision was a controversial announcement as immune-bridging trials were not fully approved or widely accepted by health authorities. In addition, the accuracy of immune-bridging trials were also been questioned for years.^{[citation needed]}

Adolescents trial

In July 2021, Medigen commenced phase II trials for adolescents aged 12-18.^[14]

Authorization

See also: List of COVID-19 vaccine authorizations § Medigen

On July 19, 2021, MVC COVID-19 vaccine obtained Emergency Use Authorization (EUA) approval from the Taiwanese government after fulfilling EUA requirements set by Taiwanese authority.^[15] The EUA, however, was met with controversy due to the lack of efficacy data and Phase 3 Clinical Trial. On August 23, 2021, President Tsai Ing-Wen was among the first Taiwanese to receive a dose of the vaccine. ^[16]

References

1. ^ “Dynavax and Medigen Announce Collaboration to Develop a Novel Adjuvanted COVID-19 Vaccine Candidate”. GlobeNewswire. 23 July 2020. Retrieved 7 June 2021.
2. ^ 黃驛淵 (10 June 2021). “【獨家】【國產疫苗解盲1】高端實體疫苗針劑首曝光 「每天9萬劑」生產基地直擊” (in Chinese). Mirror Media.
3. ^ Jump up to:^a b “Medigen Vaccine Biologics COVID-19 Vaccine Adjuvanted with Dynavax’s CpG 1018 Announces First Participant Dosed in Phase 2 Clinical Trial in Taiwan”. http://www.medigenvac.com. Retrieved 7 August 2021.
4. ^ Jump up to:^a b Hsieh SM, Liu WD, Huang YS, Lin YJ, Hsieh EF, Lian WC, Chen C, Janssen R, Shih SR, Huang CG, Tai IC, Chang SC (25 June 2021). “Safety and immunogenicity of a Recombinant Stabilized Prefusion SARS-CoV-2 Spike Protein Vaccine (MVCCOV1901) Adjuvanted with CpG 1018 and Aluminum Hydroxide in healthy adults: A Phase 1, dose-escalation study”. EClinicalMedicine: 100989. doi:10.1016/j.eclinm.2021.100989. ISSN 2589-5370. PMC 8233066. PMID 34222848.
5. ^ “MVC and NIH Collaborate to Develop COVID-19 Vaccine”. http://www.medigenvac.com. Retrieved 7 August 2021.
6. ^ “Medigen Collaborates with Dynavax to Develop Novel Adjuvanted COVID-19 Vaccine Candidate”. http://www.medigenvac.com. Retrieved 7 August 2021.
7. ^ “MVC Signed an License Agreement with NIH on COVID-19 Vaccine”. Medigen. 5 May 2020. Retrieved 27 July 2021.
8. ^ “Medigen’s COVID-19 Vaccine Combined with Dynavax’s CpG 1018 Adjuvant Receives Taiwan Government Subsidy with First Participant Dosed in Early October”. http://www.medigenvac.com. Retrieved 7 August 2021.
9. ^ “A Study to Evaluate MVC-COV1901 Vaccine Against COVID-19 in Adult (COVID-19)”. clinicaltrials.gov. United States National Library of Medicine. Retrieved 11 March 2021.
10. ^ “A Study to Evaluate the Safety and Immunogenicity of MVC-COV1901 Against COVID-19”. clinicaltrials.gov. United States National Library of Medicine. Retrieved 11 March 2021.
11. ^ “A Study to Evaluate MVC-COV1901 Vaccine Against COVID-19 in Elderly Adults”. clinicaltrials.gov. United States National Library of Medicine. 28 March 2021. Retrieved 3 April 2021.
12. ^ “MVC Released COVID-19 Vaccine Phase 2 Interim Analysis Result”. http://www.medigenvac.com. Retrieved 7 August 2021.
13. ^ “MVC Announces Paraguay Approval of IND Application for Phase 3 Clinical Trial”. http://www.medigenvac.com. Retrieved 7 August 2021.
14. ^ “A Study to Evaluate MVC-COV1901 Vaccine Against COVID-19 in Adolescents”. clinicaltrials.gov. United States National Library of Medicine. 6 July 2021. Retrieved 6 July 2021.
15. ^ “MVC COVID-19 Vaccine Obtains Taiwan EUA Approval”. http://www.medigenvac.com. Retrieved 7 August 2021.
16. ^ Taiwan begins contested rollout of new Medigen domestic vaccine, Nikkei Asia, Erin Hale, August 23, 2021

////////Medigen vaccine, MVC COVID-19 vaccine, SARS-CoV-2, covid 19, corona virus, taiwan, approvals 2021, iss 1018, CpG 1018, MVC-COV1901

NEW DRUG APPROVALS

one time

$10.00

Click here to purchase.

FPTIPLSRLF DNAMLRAHRL HQLAFDTYQE FEEAYIPKEQ KYSFLQNPQT SLCFSESIPT
PSNREETQQK SNLELLRISL LLIQSWLEPV QFLRSVFANS LVYGASDSNV YDLLKDLEEG
IQTLMGRLED GSPRTGQIFK QTYSKFDTNS HNDDALLKNY GLLYCFRKDM DKVETFLRIV
QCRSVEGSCG F
(Disulfide bridge: 53-165, 182-189)

Lonapegsomatropin, ロナペグソマトロピン

FDA APPROVED, 25/8/21, Skytrofa, Treatment of growth hormone deficiency

To treat short stature due to inadequate secretion of endogenous growth hormone

1934255-39-6 CAS, UNII: OP35X9610Y

Molecular Formula, C1051-H1627-N269-O317-S9[-C2-H4-O]4n

ACP 001; ACP 011; lonapegsomatropin-tcgd; SKYTROFA; TransCon; TransCon growth hormone; TransCon hGH; TransCon PEG growth hormone; TransCon PEG hGH; TransCon PEG somatropin, 

WHO 10598

PEPTIDE

Biologic License Application (BLA): 761177
Company: ACENDIS PHARMA ENDOCRINOLOGY DIV A/S

SKYTROFA is a human growth hormone indicated for the treatment of pediatric patients 1 year and older who weigh at least 11.5 kg and have growth failure due to inadequate secretion of endogenous growth hormone (GH) (1).

• OriginatorAscendis Pharma
• DeveloperAscendis Pharma; VISEN Pharmaceuticals
• ClassGrowth hormones; Hormonal replacements; Polyethylene glycols
• Mechanism of ActionSomatotropin receptor agonists
• Orphan Drug StatusYes – Somatotropin deficiency
• RegisteredSomatotropin deficiency

• 25 Aug 2021Registered for Somatotropin deficiency (In children, In infants) in USA (SC)
• 27 May 2021Ascendis Pharma expects European Commission decision on the Marketing Authorisation Application (MAA) for Somatotropin deficiency (In children, In infants, In neonates) in fourth quarter of 2021
• 27 May 2021Phase-III clinical trials in Somatotropin deficiency (In children, Treatment-naive) in Japan (SC)

Ascendis Pharma A/S Announces U.S. Food and Drug Administration Approval of SKYTROFA® (lonapegsomatropin-tcgd), the First Once-weekly Treatment for Pediatric Growth Hormone Deficiency

https://www.globenewswire.com/news-release/2021/08/25/2286624/0/en/Ascendis-Pharma-A-S-Announces-U-S-Food-and-Drug-Administration-Approval-of-SKYTROFA-lonapegsomatropin-tcgd-the-First-Once-weekly-Treatment-for-Pediatric-Growth-Hormone-Deficiency.html

SKYTROFA, the first FDA approved treatment utilizing TransCon technology, is a long-acting prodrug of somatropin that releases the same somatropin used in daily therapies –

– Once weekly SKYTROFA demonstrated higher annualized height velocity (AHV) at week 52 compared to a daily growth hormone with similar safety and tolerability –

– Availability in the U.S. expected shortly supported by a full suite of patient support programs –

– Ascendis Pharma to host investor conference call today, Wednesday, August 25 at 4:30 p.m. E.T. –

COPENHAGEN, Denmark, Aug. 25, 2021 (GLOBE NEWSWIRE) — Ascendis Pharma A/S (Nasdaq: ASND), a biopharmaceutical company that utilizes its innovative TransCon technologies to potentially create new treatments that make a meaningful difference in patients’ lives, today announced that the U.S. Food and Drug Administration (FDA) has approved SKYTROFA (lonapegsomatropin-tcgd) for the treatment of pediatric patients one year and older who weigh at least 11.5 kg (25.4 lb) and have growth failure due to inadequate secretion of endogenous growth hormone (GH).

As a once-weekly injection, SKYTROFA is the first FDA approved product that delivers somatropin (growth hormone) by sustained release over one week.

“Today’s approval represents an important new choice for children with GHD and their families, who will now have a once-weekly treatment option. In the pivotal head-to-head clinical trial, once-weekly SKYTROFA demonstrated higher annualized height velocity at week 52 compared to somatropinⁱ,” said Paul Thornton, M.B. B.Ch., MRCPI, a clinical investigator and pediatric endocrinologist in Fort Worth, Texas. “This once-weekly treatment could reduce treatment burden and potentially replace the daily somatropin therapies, which have been the standard of care for over 30 years.”

Growth hormone deficiency is a serious orphan disease characterized by short stature and metabolic complications. In GHD, the pituitary gland does not produce sufficient growth hormone, which is important not only for height but also for a child’s overall endocrine health and development.

The approval includes the new SKYTROFA^® Auto-Injector and cartridges which, after first removed from a refrigerator, allow families to store the medicine at room temperature for up to six months. With a weekly injection, patients switching from injections every day can experience up to 86 percent fewer injection days per year.

“SKYTROFA is the first product using our innovative TransCon technology platform that we have developed from design phase through non-clinical and clinical development, manufacturing and device optimization, and out to the patients. It reflects our commitment and dedication to addressing unmet medical needs by developing a pipeline of highly differentiated proprietary products across multiple therapeutic areas,” said Jan Mikkelsen, Ascendis Pharma’s President and Chief Executive Officer. “We are grateful to the patients, caregivers, clinicians, clinical investigators, and our employees, who have all contributed to bringing this new treatment option to children in the U.S. with GHD.”

In connection with the commercialization of SKYTROFA, the company is committed to offering a full suite of patient support programs, including educating families on proper injection procedures for SKYTROFA as the first once-weekly treatment for children with GHD.

“It is wonderful that patients and their families now have the option of a once-weekly growth hormone therapy,” said Mary Andrews, Chief Executive Officer and co-founder of the MAGIC Foundation, a global leader in endocrine health, advocacy, education, and support. “GHD is often overlooked and undertreated in our children and managing it can be challenging for families. We are excited about this news as treating GHD is important, and children have a short time to grow.”

The FDA approval of SKYTROFA was based on results from the phase 3 heiGHt Trial, a 52-week, global, randomized, open-label, active-controlled, parallel-group trial that compared once-weekly SKYTROFA to daily somatropin (Genotropin^®) in 161 treatment-naïve children with GHDⁱⁱ. The primary endpoint was, AHV at 52 weeks for weekly SKYTROFA and daily hGH treatment groups. Other endpoints included adverse events, injection-site reactions, incidence of anti-hGH antibodies, annualized height velocity, change in height SDS, proportion of subjects with IGF-1 SDS (0.0 to +2.0), PK/PD in subjects < 3 years, and preference for and satisfaction with SKYTROFA.

At week 52, the treatment difference in AHV was 0.9 cm/year (11.2 cm/year for SKYTROFA compared with 10.3 cm/year for daily somatropin) with a 95 percent confidence interval [0.2, 1.5] cm/year. The primary objective of non-inferiority in AHV was met for SKYTROFA in this trial and further demonstrated a higher AHV at week 52 for lonapegsomatropin compared to daily somatropin, with similar safety, in treatment-naïve children with GHD.

No serious adverse events or discontinuations related to SKYTROFA were reported. Most common adverse reactions (≥ 5%) in pediatric patients include: infection, viral (15%), pyrexia (15%), cough (11%), nausea and vomiting (11%), hemorrhage (7%), diarrhea (6%), abdominal pain (6%), and arthralgia and arthritis (6%)ⁱⁱ. In addition, both arms of the study reported low incidences of transient, non-neutralizing anti-hGH binding antibodies and no cases of persistent antibodies.

Conference Call and Webcast Information

A live webcast of the conference call will be available on the Investors and News section of the Ascendis Pharma website at www.ascendispharma.com. A webcast replay will be available on this website shortly after conclusion of the event for 30 days.

The Following Information is Intended for the U.S. Audience Only

INDICATION

SKYTROFA® is a human growth hormone indicated for the treatment of pediatric patients 1 year and older who weigh at least 11.5 kg and have growth failure due to inadequate secretion of endogenous growth hormone (GH).

IMPORTANT SAFETY INFORMATION

• SKYTROFA is contraindicated in patients with:
  • Acute critical illness after open heart surgery, abdominal surgery or multiple accidental trauma, or if you have acute respiratory failure due to the risk of increased mortality with use of pharmacologic doses of somatropin.
  • Hypersensitivity to somatropin or any of the excipients in SKYTROFA. Systemic hypersensitivity reactions have been reported with post-marketing use of somatropin products.
  • Closed epiphyses for growth promotion.
  • Active malignancy.
  • Active proliferative or severe non-proliferative diabetic retinopathy.
  • Prader-Willi syndrome who are severely obese, have a history of upper airway obstruction or sleep apnea or have severe respiratory impairment due to the risk of sudden death.
    
• Increased mortality in patients with acute critical illness due to complications following open heart surgery, abdominal surgery or multiple accidental trauma, or those with acute respiratory failure has been reported after treatment with pharmacologic doses of somatropin. Safety of continuing SKYTROFA treatment in patients receiving replacement doses for the approved indication who concurrently develop these illnesses has not been established.
  
• Serious systemic hypersensitivity reactions including anaphylactic reactions and angioedema have been reported with post-marketing use of somatropin products. Do not use SKYTROFA in patients with known hypersensitivity to somatropin or any of the excipients in SKYTROFA.
  
• There is an increased risk of malignancy progression with somatropin treatment in patients with active malignancy. Preexisting malignancy should be inactive with treatment completed prior to starting SKYTROFA. Discontinue SKYTROFA if there is evidence of recurrent activity.
  
• In childhood cancer survivors who were treated with radiation to the brain/head for their first neoplasm and who developed subsequent growth hormone deficiency (GHD) and were treated with somatropin, an increased risk of a second neoplasm has been reported. Intracranial tumors, in particular meningiomas, were the most common of these second neoplasms. Monitor all patients with a history of GHD secondary to an intracranial neoplasm routinely while on somatropin therapy for progression or recurrence of the tumor.
  
• Because children with certain rare genetic causes of short stature have an increased risk of developing malignancies, practitioners should thoroughly consider the risks and benefits of starting somatropin in these patients. If treatment with somatropin is initiated, carefully monitor these patients for development of neoplasms. Monitor patients on somatropin therapy carefully for increased growth, or potential malignant changes of preexisting nevi. Advise patients/caregivers to report marked changes in behavior, onset of headaches, vision disturbances and/or changes in skin pigmentation or changes in the appearance of preexisting nevi.
  
• Treatment with somatropin may decrease insulin sensitivity, particularly at higher doses. New onset type 2 diabetes mellitus has been reported in patients taking somatropin. Undiagnosed impaired glucose tolerance and overt diabetes mellitus may be unmasked. Monitor glucose levels periodically in all patients receiving SKYTROFA. Adjust the doses of antihyperglycemic drugs as needed when SKYTROFA is initiated in patients.
  
• Intracranial hypertension (IH) with papilledema, visual changes, headache, nausea, and/or vomiting has been reported in a small number of patients treated with somatropin. Symptoms usually occurred within the first 8 weeks after the initiation of somatropin and resolved rapidly after cessation or reduction in dose in all reported cases. Fundoscopic exam should be performed before initiation of therapy and periodically thereafter. If somatropin-induced IH is diagnosed, restart treatment with SKYTROFA at a lower dose after IH-associated signs and symptoms have resolved.
  
• Fluid retention during somatropin therapy may occur and is usually transient and dose dependent.
  
• Patients receiving somatropin therapy who have or are at risk for pituitary hormone deficiency(s) may be at risk for reduced serum cortisol levels and/or unmasking of central (secondary) hypoadrenalism. Patients treated with glucocorticoid replacement for previously diagnosed hypoadrenalism may require an increase in their maintenance or stress doses following initiation of SKYTROFA therapy. Monitor patients for reduced serum cortisol levels and/or need for glucocorticoid dose increases in those with known hypoadrenalism.
  
• Undiagnosed or untreated hypothyroidism may prevent response to SKYTROFA. In patients with GHD, central (secondary) hypothyroidism may first become evident or worsen during SKYTROFA treatment. Perform thyroid function tests periodically and consider thyroid hormone replacement.
  
• Slipped capital femoral epiphysis may occur more frequently in patients undergoing rapid growth. Evaluate pediatric patients with the onset of a limp or complaints of persistent hip or knee pain.
  
• Somatropin increases the growth rate and progression of existing scoliosis can occur in patients who experience rapid growth. Somatropin has not been shown to increase the occurrence of scoliosis. Monitor patients with a history of scoliosis for disease progression.
  
• Cases of pancreatitis have been reported in pediatric patients receiving somatropin. The risk may be greater in pediatric patients compared with adults. Consider pancreatitis in patients who develop persistent severe abdominal pain.
  
• When SKYTROFA is administered subcutaneously at the same site over a long period of time, lipoatrophy may result. Rotate injection sites when administering SKYTROFA to reduce this risk.
  
• There have been reports of fatalities after initiating therapy with somatropin in pediatric patients with Prader-Willi syndrome who had one or more of the following risk factors: severe obesity, history of upper airway obstruction or sleep apnea, or unidentified respiratory infection. Male patients with one or more of these factors may be at greater risk than females. SKYTROFA is not indicated for the treatment of pediatric patients who have growth failure due to genetically confirmed Prader-Willi syndrome.
  
• Serum levels of inorganic phosphorus, alkaline phosphatase, and parathyroid hormone may increase after somatropin treatment.
  
• The most common adverse reactions (≥5%) in patients treated with SKYTROFA were: viral infection (15%), pyrexia (15%), cough (11%), nausea and vomiting (11%), hemorrhage (7%), diarrhea (6%), abdominal pain (6%), and arthralgia and arthritis (6%).
  
• SKYTROFA can interact with the following drugs:
  
  • Glucocorticoids: SKYTROFA may reduce serum cortisol concentrations which may require an increase in the dose of glucocorticoids.
  • Oral Estrogen: Oral estrogens may reduce the response to SKYTROFA. Higher doses of SKYTROFA may be required.
  • Insulin and/or Other Hypoglycemic Agents: SKYTROFA may decrease insulin sensitivity. Patients with diabetes mellitus may require adjustment of insulin or hypoglycemic agents.
  • Cytochrome P450-Metabolized Drugs: Somatropin may increase cytochrome P450 (CYP450)-mediated antipyrine clearance. Carefully monitor patients using drugs metabolized by CYP450 liver enzymes in combination with SKYTROFA.

You are encouraged to report side effects to FDA at (800) FDA-1088 or www.fda.gov/medwatch. You may also report side effects to Ascendis Pharma at 1-844-442-7236.

Please click here for full Prescribing Information for SKYTROFA.

About SKYTROFA^® (lonapegsomatropin-tcgd)

SKYTROFA^® is a once-weekly prodrug designed to deliver somatropin over a one-week period. The released somatropin has the same 191 amino acid sequence as daily somatropin.

SKYTROFA single-use, prefilled cartridges are available in nine dosage strengths, allowing for convenient dosing flexibility. They are designed for use only with the SKYTROFA^® Auto-Injector and may be stored at room temperature for up to six months. The recommended dose of SKYTROFA for treatment-naïve patients and patients switching from daily somatropin is 0.24 mg/kg body weight, administered once weekly. The dose may be adjusted based on the child’s weight and insulin-like growth factor-1 (IGF-1) SDS.

SKYTROFA has been studied in over 300 children with GHD across the Phase 3 program which consists of the heiGHt Trial (for treatment-naïve patients), the fliGHt Trial (for treatment-experienced patients), and the enliGHten Trial (an ongoing long-term extension trial). Patients who completed the heiGHt Trial or the fliGHt Trial were able to continue into the enliGHten Trial and some have been on SKYTROFA for over four years.

SKYTROFA is being evaluated for pediatric GHD in Phase 3 trials in Japan and Greater China, including the People’s Republic of China, Hong Kong, Macau and Taiwan. Ascendis Pharma is also conducting the global Phase 3 foresiGHt Trial in adults with GHD. SKYTROFA has been granted orphan designation for GHD in both the U.S. and Europe.

About TransCon Technologies

TransCon refers to “transient conjugation.” The proprietary TransCon platform is an innovative technology to create new therapies that are designed to potentially optimize therapeutic effect, including efficacy, safety and dosing frequency. TransCon molecules have three components: an unmodified parent drug, an inert carrier that protects it, and a linker that temporarily binds the two. When bound, the carrier inactivates and shields the parent drug from clearance. When injected into the body, physiologic conditions (e.g., pH and temperature) initiate the release of the active, unmodified parent drug in a predictable manner. Because the parent drug is unmodified, its original mode of action is expected to be maintained. TransCon technology can be applied broadly to a protein, peptide or small molecule in multiple therapeutic areas, and can be used systemically or locally.

About Ascendis Pharma A/S

Ascendis Pharma is applying its innovative platform technology to build a leading, fully integrated biopharma company focused on making a meaningful difference in patients’ lives. Guided by its core values of patients, science and passion, the company utilizes its TransCon technologies to create new and potentially best-in-class therapies.

Ascendis Pharma currently has a pipeline of multiple independent endocrinology rare disease and oncology product candidates in development. The company continues to expand into additional therapeutic areas to address unmet patient needs.

Ascendis is headquartered in Copenhagen, Denmark, with additional facilities in Heidelberg and Berlin, Germany, in Palo Alto and Redwood City, California, and in Princeton, New Jersey.

Please visit www.ascendispharma.com (for global information) or www.ascendispharma.us (for U.S. information).

NEW DRUG APPROVALS

ONE TIME

$10.00

Click here to purchase.

///////////Lonapegsomatropin, Skytrofa, APPROVALS 2021, FDA 2021, PEPTIDE, ロナペグソマトロピン , ACP 00, ACP 011,  lonapegsomatropin-tcgd, TransCon, TransCon growth hormone, TransCon hGH, TransCon PEG growth hormone, TransCon PEG hGH, TransCon PEG somatropin, ORPHAN DRUG

Difelikefalin acetate

ジフェリケファリン酢酸塩

CAS 1024829-44-4

D-Phe-D-Phe-D-Leu-D-Lys-[ω(4-aminopiperidine-4- carboxylic acid)]-OH

FDA APPROVED, 2021/8/23, FORSUVA

Analgesic, Antipruritic, Opioid receptor agonist

Treatment of moderate-to-severe pruritus associated with chronic kidney disease in adults undergoing hemodialysis

Difelikefalin, CR-845; MR-13A-9; MR-13A9

4-amino-1- (D-phenylalanyl-D-phenylalanyl-D-leucyl-D-lysyl) piperidine-4-carboxylic acid

C36H53N7O6, 679.40573

Difelikefalin, sold under the brand name Korsuva , is an analgesic opioid peptide used for the treatment of moderate-to-severe pruritus. It acts as a peripherally specific, highly selective agonist of the κ-opioid receptor (KOR).^[3][4][5][6]

Difelikefalin was approved for medical use in the United States in August 2021.^[2][7][8]

Difelikefalin acts as an analgesic by activating KORs on peripheral nerve terminals and KORs expressed by certain immune system cells.^[3] Activation of KORs on peripheral nerve terminals results in the inhibition of ion channels responsible for afferent nerve activity, causing reduced transmission of pain signals, while activation of KORs expressed by immune system cells results in reduced release of proinflammatory, nerve-sensitizing mediators (e.g., prostaglandins).^[3]

NEW DRUG APPROVALS

ONE TIME

$10.00

Click here to purchase.

Research

It is under development by Cara Therapeutics as an intravenous agent for the treatment of postoperative pain.^[3][4][6] An oral formulation has also been developed.^[6] Due to its peripheral selectivity, difelikefalin lacks the central side effects like sedation, dysphoria, and hallucinations of previous KOR-acting analgesics such as pentazocine and phenazocine.^[3][4] In addition to use as an analgesic, difelikefalin is also being investigated for the treatment of pruritus (itching).^[3][4][5] Difelikefalin has completed phase II clinical trials for postoperative pain and has demonstrated significant and “robust” clinical efficacy, along with being safe and well tolerated.^[4][6] It has also completed a phase III clinical trial for uremic pruritus in hemodialysis patients.^[9]Kappa opioid receptors have been suggested as targets for intervention for treatment or prevention of a wide array of diseases and conditions by administration of kappa opioid receptor agonists. See for example, Jolivalt et al., Diabetologia, 49(11):2775-85; Epub Aug. 19, 2006), describing efficacy of asimadoline, a kappa receptor agonist in rodent diabetic neuropathy; and Bileviciute-Ljungar et al., Eur. J. Pharm. 494:139-46 (2004) describing the efficacy of kappa agonist U-50,488 in the rat chronic constriction injury (CCI) model of neuropathic pain and the blocking of its effects by the opioid antagonist, naloxone. These observations support the use of kappa opioid receptor agonists for treatment of diabetic, viral and chemotherapy- induced neuropathic pain. The use of kappa receptor agonists for treatment or prevention of visceral pain including gynecological conditions such as dysmenorrheal cramps and endometriosis has also been reviewed. See for instance, Riviere, Br. J. Pharmacol. 141:1331-4 (2004).[0004] Kappa opioid receptor agonists have also been proposed for the treatment of pain, including hyperalgesia. Hyperalgesia is believed to be caused by changes in the milieu of the peripheral sensory terminal occur secondary to local tissue damage. Tissue damage (e.g., abrasions, burns) and inflammation can produce significant increases in the excitability of polymodal nociceptors (C fibers) and high threshold mechanoreceptors (Handwerker et al. (1991) Proceeding of the VIth World Congress on Pain, Bond et al., eds., Elsevier Science Publishers BV, pp. 59-70; Schaible et al. (1993) Pain 55:5-54). This increased excitability and exaggerated responses of sensory afferents is believed to underlie hyperalgesia, where the pain response is the result of an exaggerated response to a stimulus. The importance of the hyperalgesic state in the post-injury pain state has been repeatedly demonstrated and appears to account for a major proportion of the post-injury/inflammatory pain state. See for example, Woold et al. (1993) Anesthesia and Analgesia 77:362-79; Dubner et al.(1994) In, Textbook of Pain, Melzack et al., eds., Churchill-Livingstone, London, pp. 225-242.[0005] Kappa opioid receptors have been suggested as targets for the prevention and treatment of cardiovascular disease. See for example, Wu et al. “Cardioprotection of Preconditioning by Metabolic Inhibition in the Rat Ventricular Myocyte – Involvement of kappa Opioid Receptor” (1999) Circulation Res vol. 84: pp. 1388-1395. See also Yu et al. “Anti-Arrhythmic Effect of kappa Opioid Receptor Stimulation in the Perfused Rat Heart: Involvement of a cAMP-Dependent Pathway”(1999) JMoI Cell Cardiol, vol. 31(10): pp. 1809-1819.[0006] It has also been found that development or progression of these diseases and conditions involving neurodegeneration or neuronal cell death can be prevented, or at least slowed, by treatment with kappa opioid receptor agonists. This improved outcome is believed to be due to neuroprotection by the kappa opioid receptor agonists. See for instance, Kaushik et al. “Neuroprotection in Glaucoma” (2003) J. Postgraduate Medicine vol. 49 (1): pp. 90-95. [0007] The presence of kappa opioid receptors on immune cells (Bidlak et al.,(2000) Clin. Diag. Lab. Immunol. 7(5):719-723) has been implicated in the inhibitory • action of a kappa opioid receptor agonist, which has been shown to suppress HIV-I expression. See Peterson PK et al, Biochem Pharmacol 2001, 61(19):1145-51. [0008] Walker, Adv. Exp. Med. Biol. 521: 148-60 (2003) appraised the antiinflammatory properties of kappa agonists for treatment of osteoarthritis, rheumatoid arthritis, inflammatory bowel disease and eczema. Bileviciute-Ljungar et al., Rheumatology 45:295-302 (2006) describe the reduction of pain and degeneration in Freund’s adjuvant-induced arthritis by the kappa agonist U-50,488.[0009] Wikstrom et al, J. Am. Soc. Nephrol. 16:3742-7 (2005) describes the use of the kappa agonist, TRK-820 for treatment of uremic and opiate-induced pruritis, and Ko et al., J. Pharmacol. Exp. Ther. 305: 173-9 (2003) describe the efficacy of U- 50,488 in morphine-induced pruritis in the monkey. [0010] Application of peripheral opioids including kappa agonists for treatment of gastrointestinal diseases has also been extensively reviewed. See for example, Lembo, Diges. Dis. 24:91-8 (2006) for a discussion of use of opioids in treatment of digestive disorders, including irritable bowel syndrome (IBS), ileus, and functional dyspepsia.[0011] Ophthalmic disorders, including ocular inflammation and glaucoma have also been shown to be addressable by kappa opioids. See Potter et ah, J. Pharmacol. Exp. Ther. 309:548-53 (2004), describing the role of the potent kappa opioid receptor agonist, bremazocine, in reduction of intraocular pressure and blocking of this effect by norbinaltorphimine (norBNI), the prototypical kappa opioid receptor antagonist; and Dortch-Carnes et al, CNS Drug Rev. 11(2): 195-212 (2005). U.S. Patent 6,191,126 to Gamache discloses the use of kappa opioid agonists to treat ocular pain. Otic pain has also been shown to be treatable by administration of kappa opioid agonists. See U.S. Patent 6,174,878 also to Gamache.[0012] Kappa opioid agonists increase the renal excretion of water and decrease urinary sodium excretion (i.e., produces a selective water diuresis, also referred to as aquaresis). Many, but not all, investigators attribute this effect to a suppression of vasopressin secretion from the pituitary. Studies comparing centrally acting and purportedly peripherally selective kappa opioids have led to the conclusion that kappa opioid receptors within the blood-brain barrier are responsible for mediating this effect. Other investigators have proposed to treat hyponatremia with nociceptin peptides or charged peptide conjugates that act peripherally at the nociceptin receptor, which is related to but distinct from the kappa opioid receptor (D. R. Kapusta, Life ScL, 60: 15-21, 1997) (U.S. Pat. No. 5,840,696). U.S. Pat Appl. 20060052284.
PATENTJpn. Tokkyo Koho, 5807140US 20090156508WO 2008057608

PATENTUS 20100075910https://patents.google.com/patent/US8236766B2/en

Example 2Synthesis of Compound (2): D-Phe-D-Phe-D-Leu-D-Lys-[ω(4-aminopiperidine-4-carboxylic acid)]-OHSee the scheme of FIG. 3 and Biron et al., Optimized selective N-methylation of peptides on solid support. J. Peptide Science 12: 213-219 (2006). The amino acid derivatives used were Boc-D-Phe-OH, Fmoc-D-Phe-OH, Fmoc-D-Leu-OH, Fmoc-D-Lys(Dde)-OH, and N-Boc-amino-(4-N-Fmoc-piperidinyl)carboxylic acid. HPLC and MS analyses were performed as described in the synthesis of compound (1) described above.The fully protected resin-bound peptide was synthesized manually starting from 2-Chlorotrityl chloride resin (1.8 g, 0.9 mmol; Peptide International). Attachment of N-Boc-amino-(4-N-Fmoc-piperidinyl)carboxylic acid followed by peptide chain elongation and deprotection of Dde in D-Lys(Dde) at Xaa₄ was carried out according to the procedure described in the synthesis of compound (1). See above. The resulting peptide resin (0.9 mmol; Boc-D-Phe-D-Phe-D-Leu-D-Lys-(N-Boc-amino-4-piperidinylcarboxylic acid)-[2-Cl-Trt resin]) was split and a portion of 0.3 mmol was used for subsequent cleavage. The peptide resin (0.3 mmol) was then treated with a mixture of TFA/TIS/H₂O (15 ml, v/v/v=95:2.5:2.5) at room temperature for 90 minutes. The resin was then filtered and washed with TFA. The filtrate was evaporated in vacuo and the crude synthetic peptide amide (0.3 mmol; D-Phe-D-Phe-D-Leu-D-Lys-[ω(4-aminopiperidine-4-carboxylic acid)]-OH) was precipitated from diethyl ether.For purification, the crude synthetic peptide amide (0.3 mmol) was dissolved in 2% acetic acid in H₂O (50 ml) and the solution was loaded onto an HPLC column and purified using TEAP buffer system with a pH 5.2 (buffers A=TEAP 5.2 and B=20% TEAP 5.2 in 80% ACN). The compound was eluted with a linear gradient of buffer B, 7% B to 37% B over 60 minutes. Fractions with purity exceeding 95% were pooled and the resulting solution was diluted with two volumes of water. The diluted solution was then loaded onto an HPLC column for salt exchange and further purification with a TFA buffer system (buffers A=0.1% TFA in H₂O and B=0.1% TFA in 80% ACN/20% H₂O) and a linear gradient of buffer B, 2% B to 75% B over 25 minutes. Fractions with purity exceeding 97% were pooled, frozen, and dried on a lyophilizer to yield the purified synthetic peptide amide as white amorphous powder (93 mg). HPLC analysis: t_R=16.43 min, purity 99.2%, gradient 5% B to 25% B over 20 min; MS (MH⁺): expected molecular ion mass 680.4, observed 680.3.Compound (2) was also prepared using a reaction scheme analogous to that shown in FIG. 3 with the following amino acid derivatives: Fmoc-D-Phe-OH, Fmoc-D-Leu-OH, Fmoc-D-Lys(Boc)-OH, and Boc-4-amino-1-Fmoc-(piperidine)-4-carboxylic acid.The fully protected resin-bound peptide was synthesized manually starting from 2-Chlorotrityl chloride resin (PS 1% DVB, 500 g, 1 meq/g). The resin was treated with Boc-4-amino-1-Fmoc-4-(piperidine)-4-carboxylic acid (280 g, 600 mmol) in a mixture of DMF, DCM and DIEA (260 mL of each) was added. The mixture was stirred for 4 hours and then the resin was capped for 1 h by the addition of MeOH (258 mL) and DIEA (258 mL).The resin was isolated and washed with DMF (3×3 L). The resin containing the first amino acid was treated with piperidine in DMF (3×3 L of 35%), washed with DMF (9×3 L) and Fmoc-D-Lys(Boc)-OH (472 g) was coupled using PyBOP (519 g) in the presence of HOBt (153 g) and DIEA (516 mL) and in DCM/DMF (500 mL/500 mL) with stiffing for 2.25 hours. The dipeptide containing resin was isolated and washed with DMF (3×3.6 L). The Fmoc group was removed by treatment with piperidine in DMF(3×3.6 L of 35%) and the resin was washed with DMF (9×3.6 L) and treated with Fmoc-D-Leu-OH (354 g), DIC (157 mL) and HOBt (154 g) in DCM/DMF (500 mL/500 mL) and stirred for 1 hour. Subsequent washing with DMF (3×4.1 L) followed by cleavage of the Fmoc group with piperidine in DMF (3×4.2 L of 35%) and then washing of the resin with DMF (9×4.2 L) provided the resin bound tripeptide. This material was treated with Fmoc-D-Phe-OH (387 g), DIC (157 mL) and HOBt (153 g) in DCM/DMF (500 mL/500 mL) and stirred overnight. The resin was isolated, washed with DMF (3×4.7 L) and then treated with piperidine in DMF (3×4.7 L of 35%) to cleave the Fmoc group and then washed again with DMF (9×4.7 L). The tetrapeptide loaded resin was treated with Fmoc-D-Phe-OH (389 g), DIC (157 mL) and HOBt (154 g) in DCM/DMF (500 mL/500 mL) and stirred for 2.25 hours. The resin was isolated, washed with DMF (3×5.2 L) and then treated piperidine (3×5.2 L of 35%) in DMF. The resin was isolated, and washed sequentially with DMF (9×5.2 L) then DCM (5×5.2 L). It was dried to provide a 90.4% yield of protected peptide bound to the resin. The peptide was cleaved from the resin using TFA/water (4.5 L, 95/5), which also served to remove the Boc protecting groups. The mixture was filtered, concentrated (⅓) and then precipitated by addition to MTBE (42 L). The solid was collected by filtration and dried under reduced pressure to give crude synthetic peptide amide.For purification, the crude synthetic peptide amide was dissolved in 0.1% TFA in H₂O and purified by preparative reverse phase HPLC (C18) using 0.1% TFA/water—ACN gradient as the mobile phase. Fractions with purity exceeding 95% were pooled, concentrated and lyophilized to provide pure synthetic peptide amide (>95.5% pure). Ion exchange was conducted using a Dowex ion exchange resin, eluting with water. The aqueous phase was filtered (0.22 μm filter capsule) and freeze-dried to give the acetate salt of the synthetic peptide amide (2) with overall yield, 71.3%, >99% purity.Hydrochloride, hydrobromide and fumarate counterions were evaluated for their ability to form crystalline salts of synthetic peptide amide (2). Approximately 1 or 2 equivalents (depending on desired stoichiometry) of hydrochloric acid, hydrobromic acid or fumaric acid, as a dilute solution in methanol (0.2-0.3 g) was added to synthetic peptide amide (2) (50-70 mg) dissolved in methanol (0.2-0.3 g). Each individual salt solution was added to isopropyl acetate (3-5 mL) and the resulting amorphous precipitate was collected by filtration and dried at ambient temperature and pressure. Crystallization experiments were carried out by dissolving the 10-20 mg of the specific amorphous salt obtained above in 70:30 ethanol-water mixture (0.1-0.2 g) followed by the addition of ethanol to adjust the ratio to 90:10 (˜0.6-0.8 mL). Each solution was then seeded with solid particles of the respective precipitated salt. Each sample tube was equipped with a magnetic stir bar and the sample was gently stirred at ambient temperature. The samples were periodically examined by plane-polarized light microscopy. Under these conditions, the mono- and di-hydrochloride salts, the di-hydrobromide salt and the mono-fumarate salt crystallized as needles of 20 to 50 μm in length with a thickness of about 1 μm.PATENT

WO 2008057608

https://patents.google.com/patent/WO2008057608A2/en Compound (2): D-Phe-D-Phe-D-Leu-D-Lys-[ω(4-aminopiperidine-4- carboxylic acid)]-OH (SEQ ID NO: 2):

EXAMPLE 2: Synthesis of compound (2)[00288] D-Phe-D-Phe-D-Leu-D-Lys-[ω(4-aminopiperidine-4-carboxylic acid)]-OH (SEQ ID NO: 2):[00289] See the scheme of Figure 2 and B iron et al., Optimized selective N- methylation of peptides on solid support. J. Peptide Science 12: 213-219 (2006). The amino acid derivatives used were Boc-D-Phe-OH, Fmoc-D-Phe-OH, Fmoc-D-Leu- OH, Fmoc-D-Lys(Dde)-OH, and N-Boc-amino-(4-N-Fmoc-piperidinyl) carboxylic acid. HPLC and MS analyses were performed as described in the synthesis of compound (1) described above.[00290] The fully protected resin-bound peptide was synthesized manually starting from 2-Chlorotrityl chloride resin (1.8 g, 0.9 mmol; Peptide International). Attachment of N-Boc-amino-(4-N-Fmoc-piperidinyl) carboxylic acid followed by peptide chain elongation and deprotection of Dde in D-Lys(Dde) at Xa^ was carried out according to the procedure described in the synthesis of compound (1). See above. The resulting peptide resin (0.9 mmol; Boc-D-Phe-D-Phe-D-Leu-D-Lys-(N- Boc-amino-4-piperidinylcarboxylic acid)-[2-Cl-Trt resin]) was split and a portion of 0.3 mmol was used for subsequent cleavage. The peptide resin (0.3 mmol) was then treated with a mixture of TFA/TIS/H₂O (15 ml, v/v/v = 95:2.5:2.5) at room temperature for 90 min. The resin was then filtered and washed with TFA. The filtrate was evaporated in vacuo and the crude peptide (0.3 mmol; D-Phe-D-Phe-D- Leu-D-Lys-[ω(4-aminopiperidine-4-carboxylic acid)]-OH) was precipitated from diethyl ether.[00291] For purification, the crude peptide (0.3 mmol) was dissolved in 2% acetic acid in H₂O (50 ml) and the solution was loaded onto an HPLC column and purified using TEAP buffer system with a pH 5.2 (buffers A = TEAP 5.2 and B = 20% TEAP 5.2 in 80% ACN). The compound was eluted with a linear gradient of buffer B, 7%B to 37%B over 60 min. Fractions with purity exceeding 95% were pooled and the resulting solution was diluted with two volumes of water. The diluted solution was then loaded onto an HPLC column for salt exchange and further purification with a TFA buffer system (buffers A = 0.1% TFA in H₂O and B = 0.1% TFA in 80% ACN/20% H₂O) and a linear gradient of buffer B, 2%B to 75%B over 25 min. Fractions with purity exceeding 97% were pooled, frozen, and dried on a lyophilizer to yield the purified peptide as white amorphous powder (93 mg). HPLC analysis: t_R = 16.43 min, purity 99.2%, gradient 5%B to 25%B over 20 min; MS (M+H⁺): expected molecular ion mass 680.4, observed 680.3.[00292] Compound (2) was also prepared using a reaction scheme analogous to that shown in figure 2 with the following amino acid derivatives: Fmoc-D-Phe-OH, Fmoc-D-Leu-OH, Fmoc-D-Lys(Boc)-OH, and Boc-4-amino-l-Fmoc-(piperidine)-4- carboxylic acid.[00293] The fully protected resin-bound peptide was synthesized manually starting from 2-Chlorotrityl chloride resin (PS 1%DVB, 500 g, 1 meq/g). The resin was treated with Boc-4-amino-l-Fmoc-4-(piperidine)-4-carboxylic acid (280 g, 600 mmol) in a mixture of DMF, DCM and DIEA (260 mL of each) was added. The mixture was stirred for 4 hours and then the resin was capped for Ih by the addition of MeOH (258 mL) and DIEA[00294] (258 mL). The resin was isolated and washed with DMF (3 x 3 L). The resin containing the first amino acid was treated with piperidine in DMF (3 x 3 L of 35%), washed with DMF (9 x 3 L) and Fmoc-D-Lys(Boc)-OH (472 g) was coupled using PyBOP (519 g) in the presence of HOBt (153 g) and DIEA (516 mL) and in DCM/DMF (500 mL/ 500 mL) with stirring for 2.25 hours. The dipeptide containing resin was isolated and washed with DMF (3 x 3.6 L). The Fmoc group was removed by treatment with piperidine in DMF [00295] , (3 x 3.6 L of 35%) and the resin was washed with DMF (9 x 3.6 L) and treated with Fmoc-D-Leu-OH (354 g), DIC (157 mL) and HOBt (154 g) in DCM/DMF (500 mL / 500 mL) and stirred for 1 hour. Subsequent washing with DMF (3 x 4.1 L) followed by cleavage of the Fmoc group with piperidine in DMF (3 x 4.2 L of 35%) and then washing of the resin with DMF (9 x 4.2 L) provided the resin bound tripeptide. This material was treated with Fmoc-D-Phe-OH (387 g), DIC (157 mL) and HOBt (153 g) in DCM/DMF (500 mL / 500 mL) and stirred overnight. The resin was isolated, washed with DMF (3 x 4.7 L) and then treated with piperidine in DMF (3 x 4.7 L of 35%) to cleave the Fmoc group and then washed again with DMF (9 x 4.7 L). The tetrapeptide loaded resin was treated with Fmoc-D-Phe-OH (389 g), DIC (157 mL) and HOBt (154 g) in DCM/DMF (500 mL / 500 mL) and stirred for 2.25 hours. The resin was isolated, washed with DMF (3 x 5.2 L) and then treated piperidine (3 x 5.2 L of 35%) in DMF. The resin was isolated, and washed sequentially with DMF (9 x 5.2 L) then DCM (5 x 5.2 L). It was dried to provide a 90.4% yield of protected peptide bound to the resin. The peptide was cleaved from the resin using TFA/ water (4.5 L, 95/5), which also served to remove the Boc protecting groups. The mixture was filtered, concentrated (1/3) and then precipitated by addition to MTBE (42 L). The solid was collected by filtration and dried under reduced pressure to give crude peptide.[00296] For purification, the crude peptide was dissolved in 0.1% TFA in H₂O and purified by preparative reverse phase HPLC (C 18) using 0.1% TF A/water – ACN gradient as the mobile phase. Fractions with purity exceeding 95% were pooled, concentrated and lyophilized to provide pure peptide (> 95.5% pure). Ion exchange was conducted using a Dowex ion exchange resin, eluting with water. The aqueous phase was filtered (0.22 μm filter capsule) and freeze-dried to give the acetate salt of the peptide (overall yield, 71.3%, >99% pure).

PATENT

WO 2015198505

κ opioid receptor agonists are known to be useful as therapeutic agents for various pain. Among, kappa opioid receptor agonist with high selectivity for peripheral kappa opioid receptors, are expected as a medicament which does not cause the central side effects. Such as peripherally selective κ opioid receptor agonist, a synthetic pentapeptide has been reported (Patent Documents 1 and 2).　The following formula among the synthetic pentapeptide (A)

[Formula 1] Being Represented By Compounds Are Useful As Pain Therapeutics. The Preparation Of This Compound, Solid Phase Peptide Synthesis Methods In Patent Documents 1 And 2 Have Been Described.Document 1 Patent: Kohyo 2010-510966 JP
Patent Document 2: Japanese Unexamined Patent Publication No. 2013-241447　Compound (1) or a salt thereof and compound (A), for example as shown in the following reaction formula, 4-aminopiperidine-4-carboxylic acid, D- lysine (D-Lys), D- leucine (D-Leu) , it can be prepared by D- phenylalanine (D-Phe) and D- phenylalanine (D-Phe) sequentially solution phase peptide synthesis methods condensation.[Of 4]The present invention will next to examples will be described in further detail.Example
1 (1) Synthesis of Cbz-D-Lys (Boc) -α-Boc-Pic-OMe (3)
to the four-necked flask of 2L, α-Boc-Pic- OMe · HCl [α-Boc-4 – aminopiperidine-4-carboxylic acid methyl hydrochloride] were charged (2) 43.7g (148mmol), was suspended in EtOAc 656mL (15v / w). To the suspension of 1-hydroxybenzotriazole (HOBt) 27.2g (178mmol), while cooling with Cbz-D-Lys (Boc) -OH 59.2g (156mmol) was added an ice-bath 1-ethyl -3 – (3-dimethylcarbamoyl amino propyl) was added to the carbodiimide · HCl (EDC · HCl) 34.1g (178mmol). After 20 minutes, stirring was heated 12 hours at room temperature. After completion of the reaction, it was added and the organic layer was 1 N HCl 218 mL of (5.0v / w). NaHCO to the resulting organic layer ₃ Aq. 218ML (5.0V / W), Et ₃ N 33.0 g of (326Mmol) was stirred for 30 minutes, and the mixture was separated. The organic layer HCl 218ML 1N (5.0V / W), NaHCO ₃ Aq. 218mL (5.0v / w), NaClaq . Was washed successively with 218ML (5.0V / W), Na ₂ SO ₄　dried addition of 8.74g (0.2w / w). Subjected to vacuum filtration, was concentrated under reduced pressure resulting filtrate by an evaporator, and pump up in the vacuum pump, the Cbz-D-Lys (Boc) -α-Boc-Pic-OMe (3) 88.9g as a white solid obtained (96.5% yield, HPLC purity 96.5%).[0033](2) D-Lys (Boc) Synthesis Of -Arufa-Boc-Pic-OMe (4)
In An Eggplant-Shaped Flask Of 2L, Cbz-D-Lys (Boc) -Arufa-Boc-Pic-OMe (3) 88.3g (142mmol) were charged, it was added and dissolved 441mL (5.0v / w) the EtOAc. The 5% Pd / C to the reaction solution 17.7g (0.2w / w) was added, After three nitrogen substitution reduced pressure Atmosphere, Was Performed Three Times A Hydrogen Substituent. The Reaction Solution Was 18 Hours With Vigorous Stirring At Room Temperature To Remove The Pd / C And After The Completion Of The Reaction Vacuum Filtration. NaHCO The Resulting Filtrate ₃ Aq. 441ML And (5.0V / W) Were Added For Liquid Separation, And The Organic Layer Was Extracted By The Addition Of EtOAc 200ML (2.3V / W) In The Aqueous Layer. NaHCO The Combined Organic Layer ₃ Aq. 441ML And (5.0V / W) Were Added for liquid separation, and the organic layer was extracted addition of EtOAc 200mL (2.3v / w) in the aqueous layer. NaClaq the combined organic layers. 441mL and (5.0v / w) is added to liquid separation, was extracted by the addition EtOAc 200ML Of (2.3V / W) In The Aqueous Layer. The Combined Organic Layer On The Na ₂ SO ₄　Dried Addition Of 17.7 g of (0.2W / W), Then The Filtrate Was Concentrated Under Reduced Pressure Obtained Subjected To Vacuum Filtration By an evaporator, and pump up in the vacuum pump, D-Lys (Boc) -α-Boc-Pic- OMe (4) to give 62.7g (90.5% yield, HPLC purity 93.6%).(3) Cbz-D-Leu -D-Lys (Boc) -α-Boc-Pic-OMe synthesis of (5)
in the four-necked flask of 2L, D-Lys (Boc) -α-Boc-Pic-OMe (4) was charged 57.7 g (120 mmol), was suspended in EtOAc 576mL (10v / w). HOBt 19.3g (126mmol) to this suspension, was added EDC · HCl 24.2g (126mmol) while cooling in an ice bath added Cbz-D-Leu-OH 33.4g (126mmol). After 20 minutes, after stirring the temperature was raised 5 hours at room temperature, further the EDC · HCl and stirred 1.15 g (6.00 mmol) was added 16 h. After completion of the reaction, it was added liquid separation 1N HCl 576mL (10v / w) . NaHCO to the resulting organic layer ₃ Aq. 576ML (10V / W), Et ₃ N 24.3 g of (240Mmol) was stirred for 30 minutes, and the mixture was separated. The organic layer HCl 576ML 1N (10V / W), NaHCO ₃ Aq. 576mL (10v / w), NaClaq . Was washed successively with 576ML (10V / W), Na ₂ SO ₄　dried addition of 11.5g (0.2w / w). After the filtrate was concentrated under reduced pressure obtained subjected to vacuum filtration by an evaporator, and pump up in the vacuum pump, the Cbz-D-Leu-D- Lys (Boc) -α-Boc-Pic-OMe (5) 85.8g It was obtained as a white solid (98.7% yield, HPLC purity 96.9%).(4) D-Leu-D -Lys (Boc) -α-Boc-Pic-OMe synthesis of (6)
in an eggplant-shaped flask of 1L, Cbz-D-Leu- D-Lys (Boc) -α-Boc-Pic -OMe the (5) 91.9g (125mmol) were charged, was added and dissolved 459mL (5.0v / w) the EtOAc. The 5% Pd / C to the reaction solution 18.4g (0.2w / w) was added, After three nitrogen substitution reduced pressure atmosphere, was performed three times a hydrogen substituent. The reaction solution was subjected to 8 hours with vigorous stirring at room temperature to remove the Pd / C and after the completion of the reaction vacuum filtration. NaHCO the resulting filtrate ₃ Aq. 200mL (2.2v / w) were added to separate liquid, NaHCO to the organic layer ₃ Aq. 200mL (2.2v / w), NaClaq . It was sequentially added washed 200mL (2.2v / w). To the resulting organic layer Na ₂ SO ₄　dried added 18.4g (0.2w / w), to the filtrate concentrated under reduced pressure obtained subjected to vacuum filtration by an evaporator, and a pump-up with a vacuum pump. The resulting amorphous solid was dissolved adding EtOAc 200mL (2.2v / w), was crystallized by the addition of heptane 50mL (1.8v / w). Was filtered off precipitated crystals by vacuum filtration, the crystals were washed with a mixed solvent of EtOAc 120mL (1.3v / w), heptane 50mL (0.3v / w). The resulting crystal 46.1g to added to and dissolved EtOAc 480mL (5.2v / w), was crystallized added to the cyclohexane 660mL (7.2v / w). Was filtered off under reduced pressure filtered to precipitate crystals, cyclohexane 120mL (1.3v / w), and washed with a mixed solvent of EtOAc 20mL (0.2v / w), and 30 ° C. vacuum dried, D-Leu- as a white solid D-Lys (Boc) -α- Boc-Pic-OMe (6) to give 36.6 g (48.7% yield, HPLC purity 99.9%).(5) Synthesis of Cbz-D-Phe-D- Leu-D-Lys (Boc) -α-Boc-Pic-OMe (7)
to the four-necked flask of 1L, D-Leu-D- Lys (Boc) -α-Boc-Pic-OMe with (6) 35.8g (59.6mmol) was charged, it was suspended in EtOAc 358mL (10v / w). To this suspension HOBt 9.59g (62.6mmol), Cbz- D-Phe-OH 18.7g was cooled in an ice bath is added (62.6mmol) while EDC · HCl 12.0g (62.6mmol) It was added. After 20 minutes, a further EDC · HCl After stirring the temperature was raised 16 hours was added 3.09 g (16.1 mmol) to room temperature. After completion of the reaction, it was added and the organic layer was 1N HCl 358mL of (10v / w). NaHCO to the resulting organic layer ₃ Aq. 358ML (10V / W), Et ₃ N 12.1 g of (119Mmol) was stirred for 30 minutes, and the mixture was separated. The organic layer HCl 358ML 1N (10V / W), NaHCO ₃ Aq. 358mL (10v / w), NaClaq . Was washed successively with 358ML (10V / W), Na ₂ SO ₄　dried addition of 7.16g (0.2w / w). After the filtrate was concentrated under reduced pressure obtained subjected to vacuum filtration by an evaporator, and pump up in the vacuum pump, Cbz-D-Phe-D -Leu-D-Lys (Boc) -α-Boc-Pic-OMe (7) was obtained 52.5g as a white solid (yield quant, HPLC purity 97.6%).(6) D-Phe-D -Leu-D-Lys (Boc) synthesis of -α-Boc-Pic-OMe ( 8)
in an eggplant-shaped flask of 2L, Cbz-D-Phe- D-Leu-D-Lys ( Boc) -α-Boc-Pic- OMe (7) the 46.9g (53.3mmol) were charged, the 840ML EtOAc (18V / W), H ₂ added to and dissolved O 93.8mL (2.0v / w) It was. The 5% Pd / C to the reaction mixture 9.38g (0.2w / w) was added, After three nitrogen substitution reduced pressure atmosphere, was performed three times a hydrogen substituent. The reaction solution was subjected to 10 hours with vigorous stirring at room temperature to remove the Pd / C and after the completion of the reaction vacuum filtration. NaHCO the resulting filtrate ₃ Aq. 235mL (5.0v / w) were added to separate liquid, NaHCO to the organic layer ₃ Aq. 235mL (5.0v / w), NaClaq . It was added sequentially cleaning 235mL (5.0v / w). To the resulting organic layer Na ₂ SO ₄　dried addition of 9.38g (0.2w / w), then the filtrate was concentrated under reduced pressure obtained subjected to vacuum filtration by an evaporator, pump up with a vacuum pump to D-Phe -D-Leu-D-Lys ( Boc) -α-Boc-Pic-OMe (7) was obtained 39.7g (yield quant, HPLC purity 97.3%).351mL was suspended in (10v / w). To this suspension HOBt 7.92g (51.7mmol), Boc-D-Phe-OH HCl HCl(8) D-Phe-D -Phe-D-Leu-D-Lys-Pic-OMe Synthesis Of Hydrochloric Acid Salt (1)
In An Eggplant-Shaped Flask Of 20ML Boc-D-Phe-D -Phe-D- Leu-D- lys (Boc) -α -Boc- Pic-OMe (9) and 2.00gg, IPA 3.3mL (1.65v / w), was suspended by addition of PhMe 10mL (5v / w). It was stirred at room temperature for 19 hours by addition of 6N HCl / IPA 6.7mL (3.35v / w). The precipitated solid was filtered off by vacuum filtration and dried under reduced pressure to a white solid of D-Phe-D-Phe- D- Leu-D-Lys-Pic- OMe 1.59ghydrochloride (1) (yield: 99 .0%, HPLC purity 98.2%) was obtained.(9) D-Phe-D -Phe-D-Leu-D-Lys-Pic-OMe Purification Of The Hydrochloric Acid Salt (1)
In An Eggplant-Shaped Flask Of 20ML-D-Phe-D- Phe D-Leu -D-Lys- pic-OMe hydrochloride crude crystals (1) were charged 200mg, EtOH: MeCN = 1: after stirring for 1 hour then heated in a mixed solvent 4.0 mL (20v / w) was added 40 ° C. of 5 , further at room temperature for 2 was time stirring slurry. Was filtered off by vacuum filtration, the resulting solid was dried under reduced pressure a white solid ((1) Purification crystals) was obtained 161 mg (80% yield, HPLC purity 99.2% ).(10) D-Phe-D -Phe-D-Leu-D-Lys-Pic Synthesis (Using Purified
(1)) Of (A) To A Round-Bottomed Flask Of 10ML D-Phe-D-Phe-D- -D-Lys Leu-Pic-OMe Hydrochloride Salt (1) Was Charged With Purified Crystal 38.5Mg (0.0488Mmol), H ₂ Was Added And Dissolved O 0.2ML (5.2V / W). 1.5H Was Stirred Dropwise 1N NaOH 197MyuL (0.197mmol) at room temperature. After completion of the reaction, concentrated under reduced pressure by an evaporator added 1N HCl 48.8μL (0.0488mmol), to obtain a D-Phe-D-Phe- D-Leu-D-Lys- Pic (A) (yield: quant , HPLC purity 99.7%).

D-Phe-D-Phe- D-Leu-D-Lys-Pic-OMe (1) physical properties ¹ H NMR (400 MHz, 1M DCl) [delta] ppm by: 0.85-1.02 (yd,. 6 H), 1.34-1.63 ( m, 5 H), 1.65-2.12 ( m, 5 H), 2.23-2.45 (m, 2 H), 2.96-3.12 (m, 4 H), 3.19 (ddt, J = 5.0 & 5.0 & 10.0 Hz), 3.33-3.62 (m, 1 H), 3.68-3.82 (m, 1 H), 3.82-3.95 (m, 4 H), 3.95-4.18 (m, 1 H), 4.25-4.37 (m, 2 H), 4.61-4.77 (M, 2 H), 7.21-7.44 (M, 10 H) ¹³ C NMR (400MHz, 1M DCl) Deruta Ppm: 21.8, 22.5, 24.8, 27.0, 30.5, 30.8, 31.0, 31.2, 31.7, 37.2 , 37.8, 38.4, 39.0, 39.8, 40.4, 40.6, 41.8, 42.3, 49.8, 50.2, 52.2, 52.6, 54.6, 55.2, 57.7, 57.9, 127.6, 128.4, 129.2, 129.6, 129.7, 129.8 dp 209.5 ℃Example 2
(Trifluoroacetic Acid (TFA)
Use) (1) D-Phe-D-Phe-D-Leu-D-Lys-Pic-OMe TFA Synthesis Of Salt (1)
TFA 18ML Eggplant Flask Of 50ML (18V / W) , 1- Dodecanethiol 1.6ML (1.6V / W), Triisopropylsilane 0.2ML (0.2V / W), H ₂ Sequentially Added Stirring The O 0.2ML (0.2V / W) Did. The Solution To The Boc-D-Phe- D- Phe-D-Leu-D -Lys (Boc) -α-Boc-Pic-OMe the (9) 1.00g (1.01mmol) was added in small portions with a spatula. After completion of the reaction, concentrated under reduced pressure by an evaporator, it was added dropwise the resulting residue in IPE 20mL (20v / w). The precipitated solid was filtered off, the resulting solid was obtained and dried under reduced pressure to D-Phe-D-Phe- D-Leu -D-Lys-Pic-OMe · TFA salt as a white solid (1) (Osamu rate 93.0%, HPLC purity 95.2%).(2) D-Phe-D -Phe-D-Leu-D-Lys-Pic synthesis of (A)
to a round-bottomed flask of 10mL D-Phe-D-Phe -D-Leu-D-Lys-Pic-OMe TFA were charged salt (1) 83mg (0.0843mmol), was added and dissolved H2O 431μL (5.2v / w). Was 12h stirring dropwise 1N NaOH 345μL (0.345mmol) at room temperature. After completion of the reaction, concentrated under reduced pressure by an evaporator added 1N HCl 84.3μL (0.0843mmol), to obtain a D-Phe-D-Phe- D-Leu-D-Lys-Pic (A) ( yield: quant, HPLC purity 95.4%).Example
3 (HCl / EtOAc
Use) (1) In An Eggplant-Shaped Flask Of 30ML Boc-D-Phe-D -Phe-D-Leu-D-Lys (Boc) -Arufa-Boc-Pic-OMe (9) 1. It was charged with 00g (1.01mmol ), was added and dissolved EtOAc7.0mL (7.0v / w). 4N HCl / EtOAc 5.0mL (5.0v / w) was added after 24h stirring at room temperature, the precipitated solid was filtered off by vacuum filtration, washed with EtOAc 2mL (2.0v / w). The resulting solid D-Phe-D-Phe- D-Leu-D-Lys-Pic-OMe hydrochloride (1) was obtained 781mg of a white solid was dried under reduced pressure (the 96.7% yield, HPLC purity 95.4%).(2) D-Phe-D -Phe-D-Leu-D-Lys-Pic (A) Synthesis of
eggplant flask of 10mL D-Phe-D-Phe -D-Leu-D-Lys-Pic-OMe hydrochloride were charged salt (1) 90 mg (0.112 mmol), H ₂ was added and dissolved O 0.47mL (5.2v / w). Was 12h stirring dropwise 1N NaOH 459μL (0.459mmol) at room temperature. After completion of the reaction, concentrated under reduced pressure by an evaporator added 1N HCl 0.112μL (0.112mmol), was obtained D-Phe-D-Phe- D-Leu-D-Lys-Pic (A) ( yield: quant, HPLC purity 93.1%).4 Example
Compound (1) Of The Compound By Hydrolysis Synthesis Of (The A) (Compound (1) Without
Purification) Eggplant Flask 10ML D-Phe-D-Phe -D-Leu-D-Lys-Pic-OMe (1) Charged Hydrochloride Were (Without Pre-Step Purification) 114.5Mg (0.142Mmol), H ₂ Was Added And Dissolved O 595MyuL (5.2V / W). Was 14H Stirring Dropwise 1N NaOH 586MyuL (0.586Mmol) At Room Temperature. After Completion Of the reaction, concentrated under reduced pressure by an evaporator added 1N HCl 0.15μL (0.150mmol), was obtained D-Phe-D-Phe- D-Leu-D-Lys-Pic (A) (yield: quant, HPLC purity 95.2 %).Example 1 Comparative
Path Not Via The Compound (1) (Using Whole Guard Boc-D-Phe-D-Phe-D-Leu-D-Lys (Boc) -Alpha-Boc-Pic-OMe
(A)) (1) D–Boc Phe- D-Phe-D-Leu-D-Lys (Boc) -Arufa-Boc-Pic-OH Synthesis Of
Eggplant Flask Of 30ML Boc-D-Phe-D -Phe-D-Leu-D- Lys (Boc) -α- Boc-Pic -OMe (9) were charged 1.00g (1.00mmol), was added and dissolved MeOH 5.0mL (5.0v / w). After stirring for four days by the addition of 1N NaOH 1.1 mL (1.10mmol) at room temperature, further MeOH 5.0mL (5.0v / w), 1N NaOH 2.0mL the (2.0mmol) at 35 ℃ in addition 3h and the mixture was stirred. After completion of the reaction, 1 N HCl 6.1 mL was added, After distilling off the solvent was concentrated under reduced pressure was separated and the organic layer was added EtOAc 5.0mL (5.0mL) .NaClaq. 5.0mL (5.0v / w) Wash the organic layer was added, the organic layer as a white solid was concentrated under reduced pressure to Boc-D-Phe-D- Phe-D-Leu-D-Lys (Boc) – α-Boc-Pic-OH 975.1mg (99.3% yield, HPLC purity 80.8% )(2) D-Phe-D -Phe-D-Leu-D-Lys-Pic synthesis of (A)
to a round-bottomed flask of 20mL Boc-D-Phe-D -Phe-D-Leu-D-Lys (Boc) It was charged -α-Boc-Pic-OH ( 10) 959mg (0.978mmol), was added and dissolved EtOAc 4.9mL (5.0v / w). And 4h stirring at room temperature was added dropwise 4N HCl / EtOAc 4.9mL (5.0mL) at room temperature. After completion of the reaction, it was filtered under reduced pressure, a white solid as to give D-Phe-D-Phe- D-Leu-D-Lys-Pic the (A) (96.4% yield, HPLC purity 79.2%) .　If not via the compound of the present invention (1), the purity of the compound obtained (A) was less than 80%. 

PATENThttp://www.google.com/patents/US20110212882

References

1. ^ Janecka A, Perlikowska R, Gach K, Wyrebska A, Fichna J (2010). “Development of opioid peptide analogs for pain relief”. Curr. Pharm. Des. 16 (9): 1126–35. doi:10.2174/138161210790963869. PMID 20030621.
2. ^ Jump up to:^a b https://www.accessdata.fda.gov/drugsatfda_docs/label/2021/214916s000lbl.pdf
3. ^ Jump up to:^{a b c d e f g h i j} Raymond S. Sinatra; Jonathan S. Jahr; J. Michael Watkins-Pitchford (14 October 2010). The Essence of Analgesia and Analgesics. Cambridge University Press. pp. 490–491. ISBN 978-1-139-49198-3.
4. ^ Jump up to:^{a b c d e} Jeffrey Apfelbaum (8 September 2014). Ambulatory Anesthesia, An Issue of Anesthesiology Clinics. Elsevier Health Sciences. pp. 190–. ISBN 978-0-323-29934-3.
5. ^ Jump up to:^a b Alan Cowan; Gil Yosipovitch (10 April 2015). Pharmacology of Itch. Springer. pp. 307–. ISBN 978-3-662-44605-8.
6. ^ Jump up to:^a b c d Charlotte Allerton (2013). Pain Therapeutics: Current and Future Treatment Paradigms. Royal Society of Chemistry. pp. 56–. ISBN 978-1-84973-645-9.
7. ^ “Korsuva: FDA-Approved Drugs”. U.S. Food and Drug Administration. Retrieved 24 August 2021.
8. ^ “Vifor Pharma and Cara Therapeutics announce U.S. FDA approval of Korsuva injection for the treatment of moderate-to-severe pruritus in hemodialysis patients” (Press release). Vifor Pharma. 24 August 2021. Retrieved 24 August 2021 – via Business Wire.
9. ^ Fishbane S, Jamal A, Munera C, Wen W, Menzaghi F (2020). “A phase 3 trial of difelikefalin in hemodialysis patients with pruritus”. N Engl J Med. 382 (3): 222–232. doi:10.1056/NEJMoa1912770. PMID 31702883.

External links

• “Difelikefalin”. Drug Information Portal. U.S. National Library of Medicine.
• Clinical trial number NCT03422653 for “A Study to Evaluate the Safety and Efficacy of CR845 in Hemodialysis Patients With Moderate-to-Severe Pruritus (KALM-1)” at ClinicalTrials.gov
• Clinical trial number NCT03636269 for “CR845-CLIN3103: A Global Study to Evaluate the Safety and Efficacy of CR845 in Hemodialysis Patients With Moderate-to-Severe Pruritus (KALM-2)” at ClinicalTrials.gov

//////////Difelikefalin acetate, FDA 2021,  APPROVALS 2021, FORSUVA, ジフェリケファリン酢酸塩 , Difelikefalin, CR 845,  MR 13A-9, MR-13A9, PEPTIDE

Heavy chain)
EVQLVESGGG LVQPGGSLRL SCAASGFTFS DYNMAWVRQA PGKGLEWVAT ITYEGRNTYY
RDSVKGRFTI SRDNAKNSLY LQMNSLRAED TAVYYCASPP QYYEGSIYRL WFAHWGQGTL
VTVSSASTKG PSVFPLAPSS KSTSGGTAAL GCLVKDYFPE PVTVSWNSGA LTSGVHTFPA
VLQSSGLYSL SSVVTVPSSS LGTQTYICNV NHKPSNTKVD KKVEPKSCDK THTCPPCPAP
ELLGGPSVFL FPPKPKDTLM ISRTPEVTCV VVDVSHEDPE VKFNWYVDGV EVHNAKTKPR
EEQYNSTYRV VSVLTVLHQD WLNGKEYKCK VSNKALPAPI EKTISKAKGQ PREPQVYTLP
PSRDELTKNQ VSLTCLVKGF YPSDIAVEWE SNGQPENNYK TTPPVLDSDG SFFLYSKLTV
DKSRWQQGNV FSCSVMHEAL HNHYTQKSLS LSPGK
(Light chain)
AIQLTQSPSS LSASVGDRVT ITCRADESVR TLMHWYQQKP GKAPKLLIYL VSNSEIGVPD
RFSGSGSGTD FRLTISSLQP EDFATYYCQQ TWSDPWTFGQ GTKVEIKRTV AAPSVFIFPP
SDEQLKSGTA SVVCLLNNFY PREAKVQWKV DNALQSGNSQ ESVTEQDSKD STYSLSSTLT
LSKADYEKHK VYACEVTHQG LSSPVTKSFN RGEC
(Disulfide bridge: H22-H96, H152-H208, H228-L214, H234-H’234, H237-H’237, H269-H329, H375-H433, H’22-H’96, H’152-H’208, H’228-L’214, H’269-H’329, H’375-H’433, L23-L88, L134-L194, L’23-L’88, L’134-L’194)

Bimekizumab

ビメキズマブ (遺伝子組換え)

UCB 4940

EU APPROVED, 2021/8/20, Bimzelx

Immunoglobulin G1, anti-​(human interleukin 17A​/interleukin 17F) (human-​Rattus norvegicus monoclonal UCB4940 heavy chain)​, disulfide with human-​Rattus norvegicus monoclonal UCB4940 light chain, dimer

Protein Sequence

Sequence Length: 1338, 455, 455, 214, 214multichain; modified (modifications unspecified)

Bimzelx 160 mg solution for injection in pre-filled syringe Bimzelx 160 mg solution for injection in pre-filled pen

The active substance in Bimzelx, bimekizumab, is a monoclonal antibody, a protein designed to attach to interleukins IL-17A, IL-17F and IL-17AF, which are messenger molecules in the body’s immune system (the body’s natural defences). High levels of these interleukins have been shown to be involved in developing inflammatory diseases caused by the immune system, such as plaque psoriasis. By attaching to these interleukins, bimekizumab prevents them from interacting with their receptors (targets) on the surface of the epidermis (outer layer of the skin), which reduces inflammation and improves the symptoms related to plaque psoriasis.,,, https://www.ema.europa.eu/en/documents/overview/bimzelx-epar-medicine-overview_en.pdf

Antipsoriatic, Anti-IL-17A/IL-17F antibody, Monoclonal antibody
Treatment of moderate to severe plaque psoriasis

Bimekizumab, sold under the brand name Bimzelx, is a humanized anti-IL17A, anti-IL-17F, and anti-IL17AF monoclonal antibody^[1][2] that is used to treat plaque psoriasis.^[1]

The most common side effects include upper respiratory tract infections (nose and throat infection) and oral candidiasis (thrush, a fungal infection in the mouth or throat).^[1]

Bimekizumab was approved for medical use in the European Union in August 2021.^[1][3]

Drug: bimekizumab
Company: UCB
Used for: psoriasis
Est. 2026 sales: $1.63 billion

Monoclonal antibody treatments for psoriasis are stacking up—but UCB hopes to muscle into the market with bimekizumab this year. The anti-IL-17A and IL-17F injection showed up both Johnson & Johnson’s Stelara and Novartis blockbuster Cosentyx in trials.

UCB’s Stelara head-to-head, the Be Vivid study presented in June at the American Academy of Dermatology and later published in The Lancet,  found 85% of bimekizumab patients had a 90% or greater reduction in the area and severity of their psoriasis symptoms at 16 weeks. Complete skin clearance, indicated by a score of PASI 100, happened in 59% of patients.

Stelara, for its part, helped just half of patients reach PASI 90 and 21% achieve complete skin clearance over the same time period.

That Be Vivid readout raised expectations of a potentially favorable outcome in UCB’s head-to-head study with Novartis blockbuster Cosentyx (secukinumab), called Be Radiant.

RELATED: UCB’s bimekizumab blows J&J’s Stelara away in phase 3, raising expectations for Cosentyx showdown

In July, UCB announced that in that phase 3 study, its candidate had “demonstrate(d) superiority to secukinumab for complete skin clearance at both weeks 16 and 48.” The full study results will be presented “in due course,” UCB promised.

The data from the Cosentyx trial could be worth a lot to UCB, Evaluate wrote in June, adding that Jefferies analysts at the time expected annual sales of bimekizumab to top out around $1.5 billion. If bimekizumab beats Cosentyx, the sales forecast could rise to above $2 billion, it said at the time.

Without specific Cosentyx-topping data from the Be Radiant study in hand, Evaluate pegs consensus sales estimates at $1.63 billion in 2026.

One concern for UCB is whether the smaller pharma will be able to compete with the big marketing budgets in psoriasis. AbbVie’s Skyrizi and Humira, Novartis’ Cosentyx, Eli Lilly’s Taltz and Amgen’s Otezla are just a handful of the psoriasis drugs that have spent millions on mainstream TV ads to build brand names.

RELATED: DiCE scores $80M to roll oral IL-17 psoriasis med into the clinic

In September, the FDA and EMA accepted UCB’s biologics license application (BLA) for bimekizumab for adults with moderate to severe plaque psoriasis, the company reported. Ongoing phase 3 trials are evaluating the drug to treat a variety of other conditions, including psoriatic arthritis, ankylosing spondylitis, non-radiographic axial spondyloarthritis and hidradenitis suppurativa.

In the meantime, more competition is on the way. South San Francisco biotech DiCE Molecules, for its part, last month nabbed new funding to the tune of $80 million to roll its oral small molecule IL-17 program into a clinical trial in psoriasis and build out preclinical programs.

In addition to IL-17 rivals, others are also looking to get in on the action—particularly, several TYK2 inhibitors. Bristol Myers Squibb’s deucravacitinib recently bested Otezla in a study, while both Pfizer and Nimbus Therapeutics are in phase 2 studies with prospects of their own.

Psoriatic arthritis (PsA) is a complex and heterogeneous inflammatory disease that affects 20% to 30% of patients with psoriasis and is associated with substantial disability, impaired quality of life (QoL), and several comorbidities.^1–3 It involves diverse clinical domains that extend beyond musculoskeletal manifestations (peripheral and axial arthritis, enthesitis and dactylitis): eg, nails, gut, and eyes, in addition to latent or manifest psoriasis.

Although there is still a huge gap in knowledge on the pathophysiology of PsA, what is known has fortunately turned into new treatment approaches that have improved symptoms and outcomes for PsA patients over the last two decades. Pro-inflammatory cytokines have been recognized as potential treatment targets in inflammatory diseases and have led to the creation of a number of anti-cytokine monoclonal antibodies that have revolutionized its treatment, such as TNFα and IL-12/23 inhibitors.⁴ More recently, the IL-17 pathway has been shown to play an important role in the pathophysiology of psoriatic disease and its blockage has shown to be clinically beneficial, as demonstrated with IL-17A inhibitors secukinumab and ixekizumab.⁴ Some patients, however, still do not respond, stop responding over time or suffer from side effects, leading to drug discontinuation, and other times combination strategies are required to control all PsA’s disease domains. Thus, there is still a great need for novel therapeutic options.⁵

Dual inhibitor antibodies target two different cytokines simultaneously potentially offering a better disease control. Interleukin (IL)-17A and IL-17F share structural homology and have a similar biologic function. IL-17A is classically considered to be the most biologically active, but recent studies have shown that IL-17F is also increased in psoriatic skin and synovial cell in psoriatic arthritis, supporting the rationale for targeting both IL-17A and IL-17F in psoriatic disease. Bimekizumab is the first-in-class monoclonal antibody designed to simultaneously target IL-17A and IL-17F.

Medical uses

Bimekizumab is indicated for the treatment of moderate to severe plaque psoriasis in adults who are candidates for systemic therapy.^[1]

History

This drug is being developed by Belgian pharmaceutical UCB. Phase III trials have demonstrated that bimekizumab is superior to not only adalimumab^[4] but also secukinumab^[5] for the treatment of plaque psoriasis.

Names

Bimekizumab is the international nonproprietary name (INN).^[6]

The Role of Interleukin (IL)‑17A and IL‑17F in Psoriatic Arthritis

The IL-17 cytokine family comprises six different members (from A to F), of which IL-17A is the most studied. Known to be produced by a wide range of immune cells, IL-17A is involved in the pathophysiology of several inflammatory diseases including spondyloarthritis.^6–8

Most non-hematopoietic cells possess IL-17 receptors, including fibroblasts, epithelial cells and synoviocytes,⁸ but despite this ubiquitous presence, IL-17 seems to have only moderate inflammatory capability per se, rather recruiting and amplifying other pathways, such as IL-6, IL-8, TNF and inflammatory-cell attracting chemokines.^6,7,9,10

Still, evidence supporting the centrality of the IL-17 pathway in both PsO and PsA is available from a wide range of data.¹¹ Th17 cells, IL-17 protein and related genes are elevated in both skin, blood and synovial fluid of PsO and PsA patients.^11,12 In PsA, increased levels of IL-17+ CD4 and CD8^13,14, as well as IL-17A+Tγδ cells, have been found in the synovial fluid compared with peripheral blood. Specifically, the levels of IL-17+CD8+ cells in the synovial fluid distinguish PsA from rheumatoid arthritis (RA) and correlate with increased DAS28 scores, C-reactive protein levels, power-doppler findings of activity and prevalence of erosions.¹³ Inhibition of this pathway is capable of normalizing almost four times more disease-related genes than anti-TNFα treatments.^11,15

Within the entire IL-17 family, IL-17F is the most structurally homologous (~50%) to IL-17A⁸ (Figure 1). They can both be secreted as homodimers (ie IL-17A/A or IL-17F/F) or as heterodimers of IL-17A/IL-17F,⁹ sharing signaling pathways through the same heterodimeric complex of IL-17 receptors A and C (IL-RA/RC) and biologic function.^7–9

The role of both IL-17A and F in psoriasis pathogenesis has been previously addressed.^6,9,16

In enthesitis, a central pathologic process in PsA, Tγδ cells have recently been described that are capable of producing both IL-17A and IL-17F even independently of IL-23 stimulation.¹⁷ IL-17A and F had already been shown to promote osteogenic differentiation in in vitro models of human periosteum activated through the use of Th17 and Tγδ cells or through culture with serum from patients with ankylosing spondylitis,¹⁸ a mechanism potentially implied in the development of enthesitis. Importantly, both cytokines seem to be equipotent in this role, unlike in inflammatory processes where IL-17F seems to be less potent.¹⁸

Both IL-17A and IL-17F, when synergized with TNF, lead to increased production of pro-inflammatory cytokines, such as IL-8 and IL-6 in synoviocytes of PsA patients.⁹ IL-17A seems to be the most pro-inflammatory of the two cytokines.^9,19 However, despite some inconsistencies in the literature regarding IL-17F detection levels which might be attributable to differences in methodology,¹⁹ IL-17F levels have been reported to be 30–50 times higher in some cytokine microenvironments, such as in psoriatic skin lesions of PsA patients²⁰ or the synovium,²¹ which might dilute differences in relative potency. Additionally, IL-17F seems to be significantly increased in the synovium of PsA compared to osteoarthritis (OA) patients, unlike IL-17A.²¹ Dual neutralization of both IL-17A and IL-17F (using bimekizumab) resulted in greater downregulation of pro-inflammatory cytokine production than a single blockade in synovial fibroblasts.^9,19 Critically, in in vitro models, anti-TNF blockade alone did not reduce the production of IL-8 as much as both IL-17A and F neutralization or even just anti-IL17A alone.^9,19 In in vitro models of human periosteum dual blockade of IL-17A and F was also more effective in suppressing osteogenic differentiation than the blockade of either cytokine individually.¹⁸

Interestingly, in Tγδ cells, the predominant IL-17 production seems to be the F subtype.¹⁸ Also of note is the recent description that the IL-17receptorC (IL-17RC) competes with IL-17RA for IL-17F, IL-17A and IL-17A/F heterodimers,²² suggesting the possibility of IL-17RA-independent signaling pathways (and thus not targeted by brodalumab, an anti-IL17RA monoclonal antibody).

Bimekizumab

Bimekizumab is a humanized monoclonal IgG1 antibody that selectively neutralizes both IL-17A and IL-17F. In in vitro models, bimekizumab appears to be as potent as ixekizumab at inhibiting IL-17A (also more potent than secukinumab)⁸ but, unlike those drugs, also possesses the unique ability to inhibit IL-17F as well, functioning as a dual inhibitor. Unlike brodalumab, an IL-17 receptor A blocker – which targets not only IL-17A and F signaling but also IL-17 C, D and E – bimekizumab spares IL-17E (also known as IL-25), for example, which is believed to have anti-inflammatory properties.⁶

Bimekizumab demonstrates dose-proportional linear pharmacokinetics, with a half-life ranging from 17 to 26 days, and its distribution is restricted to the extravascular compartment.²³ Currently, bimekizumab is in advanced clinical development for psoriasis, but also for psoriatic arthritis, and ankylosing spondylitis (both currently in phase III).

Bimekizumab in PsA – Efficacy

Phase I

The first bimekizumab clinical trial in PsA was a phase Ib randomized, double-blind, placebo-controlled clinical trial that included 53 patients (39 treated with bimekizumab, 14 with placebo) with active psoriatic arthritis who had failed conventional disease-modifying antirheumatic drugs (DMARDs) and/or one biologic DMARD. Patients in the active treatment arm were randomized to four different treatment regimens of varying loading doses (ranging from 80 to 560 mg) and maintenance doses (from 40 to 320 mg) at weeks 0, 3 and 6. Patients were followed for up to 20 weeks.⁹

Patients treated with bimekizumab had a faster response, compared to placebo. This was first detected at week two, with maximal or near-maximal responses maintained up to week 20, for both arthritis and skin psoriasis. ACR20, 50 and 70 responses were maximal at week 8 (80%), week 12 (57%) and week 16 (37%), respectively. For patients with skin involvement, PASI75 and PASI100 responses at week 8 were 100% and 87%, respectively (Table 1).

Phase II

BE ACTIVE¹⁰ was a 48-week multicentric, international, phase 2b dose-ranging, randomized, double-blind placebo-controlled trial to assess the efficacy and safety of bimekizumab. Two hundred and six adult patients (out of 308 screened) with active (tender and swollen count >3) PsA (diagnosed according to CASPAR criteria) were enrolled in 5 treatment arms (placebo, 16 mg, 160 mg with single 320 mg loading dose, 160 mg, 320 mg bimekizumab dose, with SC injections every 4 weeks). Concurrent use of TNF inhibitors was not permitted but conventional DMARDs (if on a stable dose and kept throughout the study), corticosteroids (equal or less 10mg/day) and NSAIDs were allowed. Sixteen-milligram bimekizumab (a much lower dose than other treatment arms) was tested with a programmed re-randomization at week 12 to either 160 or 320 mg dosing (meaning no placebo arm after 12 weeks). All patients received treatment up to week 48.

The primary outcome was ACR50 response at 12 weeks, a much more stringent outcome than used for other IL-17 inhibitors. The prespecified analysis was not possible due to the absence of a statistically significant difference versus placebo for the 320 mg group at week 12. All other outcomes were thus considered exploratory, rendering this a failed primary endpoint with no active comparator group.

At 12 weeks, significant ACR50 responses were present for every bimekizumab group, although lower in both the 16 mg and 320 mg dose group (Table 1 reports average values for all bimekizumab treatment groups). The 160 mg dosing had the greatest ACR and PASI response rates. These were confirmed to be increasing response rates up to week 24 and stability thereafter up to week 48, where the results of both 160 and 320 mg were similar. There were also responses in PASI scores, enthesitis, HAQ-DI and SF-36 across all bimekizumab doses. There was no loss of efficacy by week 48.

At the recent American College of Rheumatology (ACR) congress, additional data on BE ACTIVE were reported. BASDAI scoring was improved on the 93 patients in the treatment arm (160–320 mg bimekizumab) who had a baseline score >4 (mean 6.2 ± 1.42). BASDAI50 response rates were 43% and 56% at week 12 and 48, respectively.²⁴

Regarding patient-reported outcomes (PROs), the Health assessment questionnaire Disability Index (HAQ-DI) and the psoriatic arthritis impact of disease-9 (PsAID-9) questionnaire developed specifically to assess health-related quality of life (QoL) in PsA were used on 206 patients from the BE ACTIVE trial. Rapid improvement was registered by week 12 and this response was sustained up to 48 weeks. Better QoL was associated with the better clinical outcomes reported in that study.^25,26

Open-Label Extension Study (OLE)

Results from the 108 weeks of follow-up in the open-label extension study of BE ACTIVE (BE ACTIVE2, NCT03347110) have been recently presented.^27,28 All patients who completed all 48 weeks of the BE ACTIVE trial were enrolled and switched to the 160 mg dosing regardless of previous treatment dose regimen. Over 108 weeks (an additional 60 weeks of OLE study over the 48 of the original BE ACTIVE trial) there was a 66.7% and 75.4% ACR 50 and body surface area (BSA) 0% response, respectively. Dactylitis and enthesitis were also significantly improved completely resolving in 65.9% and 77.9% of patients, respectively.²⁷ Regarding week 12 responders, ACR20/50/70 and BSA 0% responses were maintained until week 108 in 80/78/81% and 72%, respectively.²⁷ MDA/VLDA responses and DAPSA remission were maintained by 81/72/76% of Week 12 responders, respectively, to Week 120 (MDA/VLDA), and Week 108 (DAPSA remission).

Bimekizumab in PsA – Safety

Phase I

Over 90% of reported adverse events, in both arms, were mild or moderate. In the treatment arm, two fungal infections (oropharyngeal and vulvovaginal candidiasis) were reported, both treated with oral medication. There was no increased incidence of other infections. There were no deaths or severe adverse events resulting from treatment, and no patient discontinued bimekizumab.⁹

Phase II

No difference was found in the frequency of adverse events between placebo and treatment arms by week 12 in the BE ACTIVE trial. After reallocation (after week 12) and up to the 48 weeks of the trial 151 (74%) of the total 204 patients who ever received bimekizumab reported some AE (exposure adjusted incidence rate 166.8/100 patient-years). Most AE were mild or moderate (the most frequently reported were nasopharyngitis and upper respiratory tract infections) and there was no direct association with bimekizumab dose.

Nine patients (8 of which received bimekizumab) had serious adverse effects. These included one patient with drug-induced liver injury. Another patient also had severe liver enzyme elevation. Both had been given the 320 mg dosing. From the hepatic point of view, the other 11 patients were noted to have increased liver enzymes (>3x ULN). There was no relation with bimekizumab dose, and most were on DMARDs and one was on TB prophylaxis. At least two serious adverse events were related to infections across the entire study period (28 weeks) – 1 hepatitis E infection, 1 cellulitis (both with the 160 mg dosing). Non-severe Candida infection was reported in 7% of the patients, none led to treatment discontinuation. Other serious AEs reported were melanoma in situ (160 mg), suicidal ideation (160 mg loading dose), and neutropenia (320 mg dosing) (only in one patient each).¹⁰ In summary, this safety profile overlaps with those of other anti-IL17 therapies.²⁹

In the OLE study, at week 108, serious adverse events occurred in 9.3% of patients (no deaths or major adverse cardiac events) and a total of 8.8% of patients withdrew from the study due to side effects. Full publication is still pending but the authors share that the safety profile observed in the OLE study reflected previous observations.²⁷

Discussion

Dual inhibitor antibodies represent a novel therapeutic strategy, and a logical extension of the success monoclonal antibodies has had over the last couple of decades.

Here we review the most recent information on IL-17A and F inhibition in psoriatic arthritis through the first-of-its-class bimekizumab, a dual inhibitor of both cytokines.

The importance of the IL-17 pathway in psoriatic arthritis, already suggested by preclinical data, was reinforced by the excellent results obtained by secukinumab³⁰ or ixekizumab³¹ in the control of the disease in the last few years.

Indeed, IL-17 seems to be involved in all of the clinical domains of psoriatic arthritis. In preclinical trials, it has been shown that both IL-17A and F are capable of inducing pro-inflammatory cytokines, like IL-8 or IL-6, in synoviocytes, periosteum and the skin,²³ and that this activation was greatly suppressed by blocking both these cytokines simultaneously. Research is expanding on the differential role of IL-17F in different environments,^18,21 compared with the more studied IL-17A, as well as possible alternative signaling pathways.²² Taken together these findings could potentially explain different clinical phenotypes in PsA and treatment responses to anti-IL17A (secukinumab, ixekizumab) and IL-17RA (brodalumab) inhibitors furthering support for the use of dual cytokine blockade such as with bimekizumab (Figure 1).

Phase II trials, specifically BE ACTIVE results, have been encouraging. Bimekizumab has shown to be relatively fast-acting, with initial improvements detected by week 8 and well established by week 12. Additionally, at a dose of 160 mg every 4 weeks, bimekizumab has shown to be capable of retaining this level of response in a high percentage of patients for at least 2 years. These results are independent of prior exposure to anti-TNF therapy.¹⁰

As with all new drugs, there are still pending questions regarding its optimal use. In BE ACTIVE,¹⁰ in which patients received four different dosages through the first 12 weeks, the 160 mg seemed most effective. The initial lower response in the 320 mg group might have been produced by a higher proportion of refractory patients in which bimekizumab took longer to work. This impression is reinforced, in the author’s opinion, by the fact that response rates were different as early as week 4 in both 160 mg (loading dose) and 320 mg dose groups although by that time period both groups had received the same dose. Co-medication was balanced between both groups.

Whichever dose proves best, these results were achieved with mostly mild side-effects that did not lead to treatment discontinuation – most commonly nasopharyngitis, upper respiratory infections and candidiasis. Overall the available data have not revealed any unexpected adverse events. Nonetheless, the number of patients included in the trials is still small. Thirteen out of the 204 patients (6,4%) receiving any dose of bimekizumab in the BE ACTIVE trial had some hepatic adverse effect, raising the need for attentive monitoring by treating physicians. Co-medication needs to be well pondered in this setting as well, but if real-world outcomes of bimekizumab prove as beneficial as in the trials there might be a reduced need for concomitant use of other DMARDs. Although IL-17F has been shown to be associated with increased susceptibility in many forms of human cancer, it has shown a protective role in colon tumorigenesis in mice,^32,33 mainly by regulating tumor angiogenesis.⁶ Longer and bigger trials will be needed to fully ascertain the safety of bimekizumab.

Overall the available results for this new therapeutic option in psoriatic arthritis are encouraging, although it is still early to completely understand the added value offered by bimekizumab. As of yet, however, there are no head-to-head trials directly comparing it to other treatment options in PsA. Anti-IL17A monoclonal antibodies have been evaluated against other therapies, such as anti-TNF inhibitors in the treatment of PsA with mixed results (using different endpoints).^34,35

Right now we can only look to early reports from the more advanced Phase 3 trials in psoriasis, where bimekizumab was first studied, which already encompass hundreds of patients and compare bimekizumab with other biologics. A head-to-head comparison with ustekinumab was recently published³⁶ involving 567 patients (321 randomized to bimekizumab, 163 to ustekinumab and 83 to a placebo arm that was switched to bimekizumab at week 16). Using a 320 mg dose of bimekizumab every 4 weeks (and not the 160 mg shown in BE ACTIVE to be the most efficacious in PsA) bimekizumab was superior to ustekinumab (85% vs 49.7% PASI 90 responses at week 16, p<0.001). This response was also sustained throughout the 52-week duration of the study (81.6% vs 55.8%, p<0.001). Similar responses (86.2% vs 47.2% PASI 90 at week 16, p<0.001) in the BE SURE trial comparing bimekizumab (320 mg every 4 weeks or 320 mg until week 16 and then every 8 weeks) and adalimumab (80 mg week 0, 40 mg week 1 and every 2 weeks) were recently presented.³⁷ Switching adalimumab patients to bimekizumab resulted in increased response rates, comparable to rates in bimekizumab-randomized patients at week 56. UCB, the company developing bimekizumab, have also reported the superiority of bimekizumab against secukinumab.³⁸

If nothing else, bimekizumab is a proof-of-concept for a novel avenue in treating inflammatory diseases. Up until now the clinical practice in inflammatory diseases has been to steer clear of the combination of monoclonal antibodies. The results of the trials reported here using bimekizumab to simultaneously inhibit two cytokines, even if related ones, are an important reminder of the redundant and overlapping nature of the immune system and of the multiple pathways through which one arrives at inflammatory disease.

As of yet, however, there are no head-to-head trials directly comparing bimekizumab to conventional DMARDS or other bDMARDs in PsA although the results reported here seem encouraging. Upcoming trials (see Table 2) will hopefully fill this gap in knowledge.

Conclusion

Psoriatic arthritis can be a severe and disabling disease. Although improvements in its treatment have been achieved in the past decade, its pathogenesis is not completely known, and its treatment is still difficult particularly throughout all disease domains.

The IL-17 pathway has been implicated in disease pathogenesis and targeting IL-17A with secukinumab and ixekizumab has shown good results, although there is still a large proportion of patients that respond only partially. The simultaneous blockade of both IL-17A and IL-17F seems to have a synergistic benefit, with IL-17F inhibition contributing with a differentiated role in both osteogenesis and skin inflammation, important domains of PsA.

Bimekizumab uses a novel approach to biologic treatment in psoriatic arthritis through dual cytokine blockade. Mounting evidence from early trials has shown a good safety and efficacy profile, with rapid onset and sustained response, with results now extending to 108 weeks of follow-up. Moreover, clinical trials in skin psoriasis have also shown that bimekizumab is highly effective, confirming the importance of inhibiting these two cytokines in psoriatic disease.

In the near future, phase III trials will help to better understand the potential of bimekizumab in the treatment of psoriatic arthritis.

References

1. ^ Jump up to:^{a b c d e f} “Bimzelx EPAR”. European Medicines Agency (EMA). 23 June 2021. Retrieved 24 August 2021. Text was copied from this source which is © European Medicines Agency. Reproduction is authorized provided the source is acknowledged.
2. ^ Lim SY, Oon HH (2019-05-13). “Systematic review of immunomodulatory therapies for hidradenitis suppurativa”. Biologics. 13: 53–78. doi:10.2147/BTT.S199862. PMC 6526329. PMID 31190730.
3. ^ “UCB Announces European Commission Approval of Bimzelx (bimekizumab) for the Treatment of Adults with Moderate to Severe Plaque Psoriasis”. UCB (Press release). 24 August 2021. Retrieved 24 August 2021.
4. ^ Warren, Richard B.; Blauvelt, Andrew; Bagel, Jerry; Papp, Kim A.; Yamauchi, Paul; Armstrong, April; Langley, Richard G.; Vanvoorden, Veerle; De Cuyper, Dirk; Cioffi, Christopher; Peterson, Luke (2021-07-08). “Bimekizumab versus Adalimumab in Plaque Psoriasis”. New England Journal of Medicine. 385 (2): 130–141. doi:10.1056/NEJMoa2102388. ISSN 0028-4793. PMID 33891379.
5. ^ Reich, Kristian; Warren, Richard B.; Lebwohl, Mark; Gooderham, Melinda; Strober, Bruce; Langley, Richard G.; Paul, Carle; De Cuyper, Dirk; Vanvoorden, Veerle; Madden, Cynthia; Cioffi, Christopher (2021-07-08). “Bimekizumab versus Secukinumab in Plaque Psoriasis”. New England Journal of Medicine. 385 (2): 142–152. doi:10.1056/NEJMoa2102383. ISSN 0028-4793. PMID 33891380.
6. ^ World Health Organization (2014). “International nonproprietary names for pharmaceutical substances (INN): recommended INN: list 72”. WHO Drug Information. 28 (3). hdl:10665/331112.

Further reading

• Reis J, Vender R, Torres T (August 2019). “Bimekizumab: The First Dual Inhibitor of Interleukin (IL)-17A and IL-17F for the Treatment of Psoriatic Disease and Ankylosing Spondylitis”. BioDrugs. 33 (4): 391–9. doi:10.1007/s40259-019-00361-6. PMID 31172372. S2CID 174812750.

External links

• “Bimekizumab”. Drug Information Portal. U.S. National Library of Medicine.

//////////Bimekizumab, Bimzelx, EU 2021, APPROVALS 2021, Monoclonal antibody
,  plaque psoriasis,ビメキズマブ (遺伝子組換え) , UCB 4940

NEW DRUF APPROVALS

ONE TIME ANTHONY CRASTO +919321316780 amcrasto@gmail.com

$10.00

Click here to purchase.

4-(2-Fluoro-3chloro-(cyclopropylaminocarbonyl)aminophenoxy)-7-methoxy-6-quinolinecarboxamide

6-Quinolinecarboxamide, 4-[3-chloro-4-[[(cyclopropylamino)carbonyl]amino]-2-fluorophenoxy]-7-methoxy-

N-(4-(6-Aminocarbonyl-7-methoxyquinolin-4-yl)oxy-2-chloro-3-fluorophenyl)-N’-cyclopropylurea

cas 2304405-29-4

C₂₁ H₁₈ Cl F N₄ O₄

444.84CN109134365 discloses an active compound or medicinal salt with multi-target effects of VEGFR1~3, fibroblast growth factor receptor 1~3, RET, Kit and PDGFR, and its chemical structure formula is as follows: Formula I:

Chemical name: 4-(2-Fluoro-3chloro-(cyclopropylaminocarbonyl)aminophenoxy)-7-methoxy-6-quinoline carboxamide, the drug name is fluvatinib. The compound has strong activity and provides a potential new treatment option for patients with tumors such as liver and kidney.

PATENT

CN109134365

PATENT

WO 2020187188

https://patents.google.com/patent/WO2020187188A1/enProcess A

Example 1A

At 20-30°C, 4-chloro-7-methoxyquinoline-6-carboxamide (550.0 g) was added to the reaction kettle. At 20-30°C, DMSO (16.5L) was added to the reactor. At 20-30°C, 2-fluoro-3chloro-4-aminophenol was added to the reactor. At 20-35°C, sodium tert-butoxide (229g) was slowly added to the reaction kettle under stirring for 10-15 minutes. The reaction kettle was heated to 96°C (internal temperature) in 1.5 hours. The reaction was stirred at 96-100°C for 6.5 hours, and no 4-amino-3-chloro-2 fluorophenol remained. The reaction was cooled to 20-30°C. Under stirring, 23.1L of water was slowly added to the reaction solution. During the process, a dark brown solid was precipitated. Keep the internal temperature below 40°C. Stir at 30-40°C for 0.5 hour. Cool to 20-30°C and filter. At 20-30°C, the filter cake and 3.5L of water are added to the reactor. Stir for 0.5 hour at 20-30°C. filter. At 20-30°C, the filter cake and 4.0L of water are added to the reactor. Stir for 0.5 hour at 20-30°C. After filtering, the filter cake was dried in a vacuum dryer at 40°C for 18 hours (phosphorus pentoxide used as a desiccant, and the oil pump was vacuumed). The solid was pulverized to obtain 758 g of off-white solid and dried at 40° C. for 18 hours (phosphorus pentoxide was used as the desiccant, and the oil pump was vacuumed) to obtain Example 1A.LCMS(ESI)m/z:362.0[M+1] ⁺¹ H NMR (400MHz, DMSO-d ₆ ) δppm 8.68 (br s, 2H), 7.82-7.96 (m, 1H), 7.67-7.82 (m, 1H), 7.46-7.59 (m, 1H), 7.12-7.26 (m, 1H), 6.67-6.80 (m, 1H), 6.43-6.58 (m, 1H), 5.84 (s, 2H), 4.04 (s, 3H).Example 1B

Example 1A (6.05g) was added to a three-necked flask containing NMP (60mL), pyridine (1.32g) and phenyl chloroformate (5.20g) were added to the reaction system, and the reaction system was at room temperature (25-30°C). ) After stirring for 1 hour, the reaction was complete. Cyclopropylamine (2.84g) was also added to the reaction system. The reaction solution was stirred at room temperature (25-30°C) for 0.5 hours. The reaction was completed. Add 20 mL of ethanol to the reaction system and stir. Tap water (500 mL) was added to the reaction system, a solid was precipitated, filtered, and the filter cake was spin-dried under reduced pressure to obtain a crude product (orange solid, 5.26 g); the crude product was passed through a chromatography column (DCM: MeOH = 20/1～10 /1) Purification to obtain the product (orange solid, 3.12 g), the product was added with 4 mL of absolute ethanol and stirred at room temperature for 18 hours, filtered, the filter cake was washed with 1 mL of ethanol, and dried under reduced pressure to obtain Example 1B. This compound is obtained by adding 1 equivalent of hydrochloric acid, sulfuric acid or methanesulfonic acid in acetone or ethanol solution to obtain the corresponding salt.LCMS(ESI)m/z:445.0[M+1] ⁺¹ H NMR (400MHz, DMSO-d ₆ ) ppm 8.66-8.71 (m, 2H), 8.12-8.20 (m, 2H), 7.72-7.93 (m, 2H), 7.45 (t, J = 9.16 Hz, 1H) ,7.28(d,J=2.76Hz,1H),6.58(d,J=5.02Hz,1H),4.05(s,3H),2.56-2.64(m,1H),0.38-0.77(m,4H)Example 1

Example 1B (1.5g, 3.37mmol) was added to EtOH (45mL), the reaction temperature was raised to 60°C, at this temperature, CH ₃ SO ₃ H (324.07mg, 3.37mmol, 240.05μL) was added dropwise to the reaction In the solution, after the dripping is completed, the reaction solution is dissolved, and the temperature of the reaction solution is naturally cooled to 15-20°C under stirring, and the reaction solution is stirred at this temperature for 2 hours. A large amount of brown solid precipitated, filtered, and the filter cake was rinsed with absolute ethanol (5 mL), and the obtained filter cake was spin-dried under reduced pressure at 50° C. without purification, and Example 1 was obtained.LCMS(ESI)m/z:445.0[M+1] ⁺¹ H NMR(400MHz,DMSO-d ₆ )δppm 9.02(d,J=6.53Hz,1H)8.72(s,1H)8.18-8.27(m,2H)7.87-8.03(m,2H)7.65(s,1H )7.53(t,J＝9.03Hz,1H)7.32(br s,1H)7.11(d,J＝6.27Hz,1H)4.08(s,3H)2.55-2.62(m,1H)2.35(s,3H) 0.34-0.75(m,4H)

PATENT

https://patentscope.wipo.int/search/en/detail.jsf?docId=WO2021143954&tab=FULLTEXT&_cid=P12-KSZPW4-91508-1Example 1 Preparation of fluvatinib crystal form I 
Add the free base of fluvatinib of formula I (50mg, 112.40umol) to EtOH (2mL), stir at 15-20℃ for 12h, filter to obtain a filter cake, add the filter cake to 200mL acetone, stir at 15-20℃ After 12h, filter and spin-dry the filter cake under reduced pressure at 40°C to obtain fluvatinib solid. The result of XRPD detection is shown in Figure 1, named as the crystalline form I of fluvatinib, and the detection results of DSC and TGA are shown in Figure 2. And Figure 3. 
Example 2 Preparation of crystal form I of fluvatinib mesylate (also referred to herein as “fluvatinib mesylate”) 
The 4-[3-chloro-4-(cyclopropylaminocarbonylamino)-2-fluoro-phenoxy]-7-methoxy-quinoline-6-carboxamide i.e. fluvatinib (0.5g, 1.12mmol) was added to EtOH (10mL) solvent, heated to 55～60℃, and methanesulfonic acid (108.02mg, 1.12mmol, 80.02μL, 1eq) was added to the reaction flask under stirring at this temperature, and the reaction solution was dissolved. , The reaction solution was cooled to 20 ~ 30 ℃, stirred at this temperature for 1 h, a brown solid precipitated out under vacuum filtration, the filter cake was rinsed with ethanol (2mL*2), and the filter cake was spin-dried at 40 ~ 50 ℃ under reduced pressure. The solid product, named as the crystalline form I of fluvatinib mesylate, was tested by XRPD, DSC, and TGA. The XRPD test results are shown in Table 1 and Figure 4 below, and the DSC and TGA test results are shown in Figure 5. Melting point is about 232-237°C.

/////////////

NC(=O)c1cc2c(ccnc2cc1OC)Oc1ccc(NC(=O)NC2CC2)c(Cl)c1F

NEW DRUG APPROVALS

ONE TIME

$10.00

Click here to purchase.

Tegoprazan

RN: 942195-55-3
UNII: W017G7IF4S

ROTATION (-)

Molecular Formula, C20-H19-F2-N3-O3, Molecular Weight, 387.3841

(S)-4-(5,7-difluorochroman-4-yloxy)-N,N,2-trimethyl-lH-benzo[d]imidazole-6-carboxamide).

• 1H-Benzimidazole-5-carboxamide, 7-(((4S)-5,7-difluoro-3,4-dihydro-2H-1-benzopyran-4-yl)oxy)-N,N,2-trimethyl-
• 7-(((4S)-5,7-Difluoro-3,4-dihydro-2H-chromen-4-yl)oxy)-N,N,2-trimethyl-1H-benzimidazole-5-carboxamide
• (S)-4-((5,7-difluorochroman-4-yl)oxy)-N,N,2-trimethyl-1H-benzo(d)imidazole-6-carboxamide

HK inno.N/RaQualia Pharma

Alternative Names: CJ-12420; IN-A001; K-CAB; LXI-15028; RQ-00000004; RQ-4Тегопразан [Russian] [INN]تيغوبرازان [Arabic] [INN]替戈拉生 [Chinese] [INN]

• A novel Potassium-competitive acid blocker.

• OriginatorPfizer
• Tegoprazan, a reversible H+/K+-ATPase inhibitor developed by CJ Healthcare (now inno.N), was first approved and launched in South Korea in 2019 for the treatment of gastroesophageal reflux disease (GERD).
• DeveloperCJ Cheiljedang Corp.; HK inno.N; RaQualia Pharma; Shandong Luoxin Pharmaceutical
• ClassAmides; Anti-inflammatories; Antibacterials; Antiulcers; Benzimidazoles; Benzopyrans; Fluorobenzenes; Small molecules
• Mechanism of ActionH(+) K(+)-exchanging ATPase inhibitors; Potassium-competitive acid blockers

• MarketedErosive oesophagitis; Gastro-oesophageal reflux
• Phase IIIGastric ulcer; Helicobacter infections; Peptic ulcer

• 28 Aug 2021No recent reports of development identified for phase-I development in Gastro-oesophageal-reflux in Japan (PO, Tablet)
• 28 Aug 2021No recent reports of development identified for phase-I development in Gastro-oesophageal-reflux in USA (PO, Tablet)
• 18 Aug 2021Shandong Luoxin Pharmaceutical Group plans a phase III trial for Duodenal ulcer in China (PO, Tablet) (NCT05010954)

PATENT

https://patents.google.com/patent/US20070142448A1/en

• [0370]
  

STEP 1: N-{4-Bromo-2-nitro-6-[(phenylmethyl)oxy]phenyl}acetamide

• [0371]
  To a solution of 4-bromo-2-nitro-6-[(phenylmethyl)oxy]aniline (33.0 g, 102 mmol, WO 2004054984) and acetic anhydride (14.5 mL, 153 mmol) in acetic acid (90 mL) was added concentrated sulfuric acid (2 drops) at 70° C. The mixture was stirred at 70° C. for 20 minutes. After cooling to room temperature, water (800 mL) was added, and the formed precipitate was collected by filtration, and washed with diisopropyl ether to give the title compound as a brown solid (30.9 g, 83%).
• [0372]
  ¹H NMR (CDCl₃, 270 MHz) δ: 7.69 (d, J=2.0 Hz, 1H), 7.56 (br. s, 1H), 7.47-7.38 (m, 5H), 7.34 (d, J=2.0 Hz, 1H), 5.14 (s, 2H), 2.16 (s, 3H) ppm.
• [0373]
  MS (ESI) m/z: 365 (M+H)⁺.

STEP 2: N-{4-Cyano-2-nitro-6-[(phenylmethyl)oxy]phenyl}acetamide

• [0374]
  A mixture of N-{4-bromo-2-nitro-6-[(phenylmethyl)oxy]phenyl}acetamide (6.5 g, 17.8 mmol, STEP 1), zinc cyanide (4.18 g, 35.6 mmol), and tetrakis(triphenylphosphine)palladium (2.06 g, 1.78 mmol) in N,N-dimethylformamide (100 mL) was heated to 170° C. for 20 minutes in the microwave synthesizer (Biotage, Emrys Optimizer). After cooling to room temperature, the suspension was filtered, and washed with ethyl acetate. The organic layers were combined, washed with water, dried over magnesium sulfate, and concentrated in vacuum. The residual solid was purified by column chromatography on silica gel eluting with hexane/ethyl acetate (3:1) to afford the title compound as a white solid (5.5 g, 99%).
• [0375]
  ¹H NMR (CDCl₃, 300 MHz) δ: 7.92 (s, 1H), 7.83 (s, 1H), 7.53-7.33 (m, 5H), 7.39 (s, 1H), 5.21 (s, 2H), 2.21 (s, 3H) ppm.
• [0376]
  MS (ESI) m/z: 312 (M+H)⁺, 310 (M−H)⁻.

STEP 3: 2-Methyl-4-[(phenylmethyl)oxy]-1H-benzimidazole-6-carbonitrile

• [0377]
  A mixture of N-{4-cyano-2-nitro-6-[(phenylmethyl)oxy]phenyl}acetamide (5.5 g, 17.7 mmol, STEP 2) and iron powder (2.96 g, 53.0 mmol) in acetic acid (90 mL) was refluxed with stirring for 2 hours. After cooling to room temperature, the mixture was filtered through a pad of Celite, and the filtrate was concentrated in vacuum. The residue was poured into water, and the aqueous layer was extracted with ethyl acetate/methanol (20:1). The organic layers were combined, washed with brine, dried over magnesium sulfate, and concentrated in vacuum to afford the title compound as a brown solid (3.82 g, 82%).
• [0378]
  ¹H NMR (DMSO-d₆, 300 MHz) δ: 7.64 (s, 1H), 7.64-7.27 (m, 6H), 7.19 (s, 1H), 5.34 (s, 2H), 2.50 (s, 3H) ppm.
• [0379]
  MS (ESI) m/z: 264 (M+H)⁺, 262 (M−H)⁻.

STEP 4: 2-Methyl-4-[(phenylmethyl)oxy]-1H-benzimidazole-6-carboxylic Acid

• [0380]
  A solution of 2-methyl-4-[(phenylmethyl)oxy]-1H-benzimidazole-6-carbonitrile (3.82 g, 14.5 mmol, STEP 3) and potassium hydroxide (85%, 10.2 g, 15.4 mmol) in ethylene glycol (50 mL) was heated to 170° C. for 20 minutes in the microwave synthesizer (Biotage, Emrys Optimizer). After cooling to room temperature, the mixture was acidified with 2M hydrochloric acid aqueous solution (pH=3). The precipitated solid was collected by filtration to afford the title compound as a white solid (3.83 g, 93%).
• [0381]
  ¹H NMR (DMSO-d₆, 270 MHz) δ: 12.68 (br. s, 1H), 7.74 (s, 1H), 7.64-7.01 (m, 7H), 5.33 (s, 2H), 2.50 (s, 3H) ppm.
• [0382]
  MS (ESI) m/z: 283 (M+H)⁺, 281 (M−H)⁻.

STEP 5: N,N,2-Trimethyl-4-[(phenylmethyl)oxy]-1H-benzimidazole-6-carboxamide

• [0383]
  A mixture of 2-methyl-4-[(phenylmethyl)oxy-1H-benzimidazole-6-carboxylic acid (5.0 g, 17.7 mmol, STEP 4), dimethylamine hydrochloride (4.33 g, 53.1 mmol), 2-[1H-benzotriazole-1-yl]-1,1,3,3-tetramethyluronium hexafluorophosphate (10.1 g, 26.6 mmol), and triethylamine (10.7 g, 106 mmol) in N,N-dimethylformamide (80 mL) was stirred at room temperature for 1 hour. The mixture was diluted with ethyl acetate/methanol (20:1), and washed with saturated ammonium chloride aqueous solution. The organic layer was dried over magnesium sulfate, and concentrated in vacuum. The residue was purified by column chromatography on silica gel (gradient elution from ethyl acetate only to ethyl acetate methanol 5:1) to afford the title compound as a white solid (4.90 g, 89%).
• [0384]
  ¹H NMR (CDCl₃, 270 MHz) δ: 7.47-7.23 (m, 5H), 7.20 (s, 1H), 6.75 (s, 1H), 5.22 (s, 2H), 2.95 (br. s, 6H), 2.54 (s, 3H) ppm (—NH was not observed).
• [0385]
  MS (ESI) m/z: 310 (M+H)⁺, 308 (M−H)⁻.

STEP 6: N,N,2-Trimethyl-1-[(4-methylphenyl)sulfonyl]-4-[(phenylmethyl)oxy]-1H-benzimidazole-6-carboxamide

• [0386]
  To a suspension of N,N,2-trimethyl-4-[(phenylmethyl)oxy]-1H-benzimidazole-6-carboxamide (928 mg, 3.0 mmol, STEP 5) in N,N-dimethylformamide (20 mL) was added sodium hydride (60% in mineral oil, 180 mg, 4.50 mmol) at 0° C. After stirring at room temperature for 30 minutes, the reaction mixture was cooled to 0° C. To the mixture was added 4-methylbenzenesulfonyl chloride (572 mg, 3.00 mmol) at 0° C., and the reaction mixture was stirred at room temperature for 2 hours. The mixture was poured into water, and the aqueous layer was extracted with ethyl acetate. The organic layers were combined, washed with water, dried over magnesium sulfate and concentrated in vacuum. The residue was purified by column chromatography on silica gel (gradient elution from dichloromethane only to ethyl acetate only) to afford the title compound as a white solid (1.00 g, 72%).
• [0387]
  ¹H NMR (CDCl₃,270 MHz) δ: 7.80 (d, J=8.1 Hz, 2H), 7.70 (s, 1H), 7.45 (d, J=831 Hz, 2H), 7.40-7.22 (m, 5H), 6.86 (s, 1H), 5.32 (s, 2H), 3.11 (br. s, 3H), 2.89 (br s, 3H), 2.81 (s, 3H), 2.40 (s, 3H) ppm.
• [0388]
  MS (ESI) m/z: 464 (M+H)⁺.

STEP 7: 4-Hydroxy-N N,2-trimethyl-1-[(4-methylphenyl)sulfonyl]-1H-benzimidazole-6-carboxamide

• [0389]
  A mixture of N,N,2-trimethyl-1-(4-methylphenyl)sulfonyl]-4-[(phenylmethyl)oxy]-1H-benzimidazole-6-carboxamide (350 mg, 0.756 mmol, STEP 6) and 20% palladium hydroxide (1.20 g) in acetic acid (20 mL) was stirred under hydrogen gas (4 atmospheres) for 4 hours. The resulted mixture was filtered through a pad of Celite, and the filtrate was concentrated in vacuum. The residue was purified by column chromatography on silica gel (gradient elution from ethyl acetate only to ethyl acetate:methanol 5:1) to afford the title compound as a white solid (131 mg, 36%).
• [0390]
  ¹H NMR (CDCl₃, 270 MHz) δ: 7.82 (d, J=8.1 Hz, 2H), 7.63 (s, 1H), 7.31 (d, J=8.1 Hz, 2H), 6.92 (s, 1H), 3.14 (br. s, 3H), 3.01 (br. s, 3H), 2.79 (s, 3H), 2.40 (s, 3H) ppm (—OH was not observed).
• [0391]
  MS (ESI) m/z: 374 (M+H)⁺, 372 (M−H)⁻.

STEP 8: 4-[(5,7-Difluoro-3,4-dihydro-2H-chromen-4-yl)oxy]-N,N,2-trimethyl-1-[(4-methylphenyl)sulfonyl]-1H-benzimidazole-6-carboxamideSTEP 8-1: 5,7-Difluoro-3,4-dihydro-2H-chromen-4-ol

• [0392]
  To a solution of 5,7-difluoro-2,3-dihydro-4H-chromen-4-one (14.2 g, 77.0 mmol, US 20050038032) in methanol (200 mL) was added sodium borohydride (3.50 g, 92.5 mmol) at 0° C. The reaction mixture was stirred at the same temperature for 1 hour, and evaporated to remove methanol. The residue was quenched with water, and extracted with ethyl acetate. The extract was washed with brine, dried over magnesium sulfate, and concentrated in vacuum. The residue was purified by column chromatography on silica gel (hexane:ethyl acetate=3:1 as an eluent) to afford the title compound as a pale gray solid (9.64 g, 67%).
• [0393]
  ¹H NMR (CDCl₃, 270 MHz) δ: 6.47-6.36 (m, 2H), 5.05-4.97 (m, 1H), 4.36-4.20 (m, 2H), 2.16-1.92 (m, 3H) ppm.

STEP 8-2: 4-[(5,7-Difluoro-3,4-dihydro-2H-chromen-4-yl)oxy]-N,N,2-trimethyl-1-[(4-methylphenyl)sulfo nyl]-1H-benzimidazole-6-carboxamide

• [0394]
  To a stirred mixture of 4-hydroxy-N,N,2-trimethyl-1-[(4-methylphenyl)sulfonyl]-1H-benzimidazole-6-carboxamide (110 mg, 0.294 mmol, STEP 7), 5,7-difluoro-3,4-dihydro-2H-chromen-4-ol (164 mg, 0.884 mmol, STEP 8-1) and triphenylphosphine (232 mg, 0.884 mmol) in toluene (5 mL) was added diisopropyl azodicarboxylate (DIAD) (179 mg, 0.884 mmol) at room temperature. The reaction mixture was stirred at room temperature for 6 hours and concentrated in vacuum. The residue was purified by column chromatography on silica gel (ethyl acetate:hexane gradient elution from 1:20 to 10:1) to afford a mixture of the title compound and triphenylphosphine oxide (280 mg, crude) as white solids, which was used in the next step without further purification.
• [0395]
  ¹H NMR (CDCl₃, 270 MHz) δ: 7.81 (d, J=8.1 Hz, 2H), 7.51 (s, 1H), 7.31 (d, J=8.1 Hz, 2H), 7.07 (s, 1H), 6.54-6.22 (m, 2H), 5.93 (br. s, 1H), 4.40 (t, J=10.8 Hz, 1H), 4.27 (t, J=10.8 Hz, 1H), 3.15 (br. s, 3H), 3.03 (br. s, 3H), 2.79 (s, 3H), 2.39 (s, 3H), 2.40-2.21 (m, 1H), 2.19-1.73 (m, 1H) ppm.
• [0396]
  MS (ESI) m/z: 542 (M+H)⁺, 540 (M−H)⁻.

STEP 9: 4-[(5,7-Difluoro-3,4-dihydro-2H-chromen-4-yl)oxy]-N,N,2-trimethyl-1H-benzimidazole-6-carboxamide

• [0397]
  To a solution of 4-[(5,7-difluoro-3,4-dihydro-2H-chromen-4-yl)oxy-N,N,2-trimethyl-1-[(4-methylphenyl)-sulfonyl]-1H-benzi midazole-6-carboxamide (280 mg, crude, STEP 8) in tetrahydrofuran (8 mL) and methanol (4 mL) was added sodium hydroxide (165 mg, 4.1 mmol) at room temperature. After stirring at room temperature for 1 hour, the mixture was quenched with saturated sodium dihydrogenphosphate aqueous solution, and extracted with ethyl acetate. The organic layers were combined, dried over magnesium sulfate and concentrated in vacuum. The residue was purified by column chromatography on silica gel (gradient elution from dichloromethane only to ethyl acetate:methanol 10:1) to afford the title compound as a white solid (74 mg, 65% for 2 steps).
• [0398]
  ¹H NMR (CDCl₃, 270 MHz) δ: 7.27 (s, 1H), 6.95 (s, 1H), 6.51-6.33 (m, 2H), 5.87-5.69 (m, 1H), 4.41-4.25 (m, 2H), 3.10 (br. s, 6H), 2.56 (s, 3H), 2.44-2.34 (m, 1H), 2.14-1.98 (m, 1H) ppm (—NH was not observed).
• [0399]
  MS (ESI) m/z: 388 (M+H)⁺, 386 (M−H)⁻.

Example 2(−)-4-[((4S)-5,7-Difluoro-3,4-dihydro-2H-chromen-4-yl)oxy]-N,N,2-trimethyl-1 fl-benzimidazole-6-carb oxamide andExample 3(−)-4-]((4R)-5,7-Difluoro-3,4-dihydro-2H-chromen-4-yl)oxy]-N,N,2-trimethyl-1H-benximidazole-6-carboxamide

• [0400]
  
• [0401]
  The fraction-1 (582 mg) and fraction-2 (562 mg) were prepared from racemic 4-[(5,7-difluoro-3,4-dihydro-2H-chromen-4-yl)oxy]-N, 1,2-trimethyl-1H-benzimidazole-6-carboxamide (1.63 g, STEP 9 in Example 1) by HPLC as follows.
• Isolation Condition
• [0402]
  Column: CHIRALCEL OJ-H (20 mm×250 mm, DAICEL)
• [0403]
  Mobile phase: n-Hexane/Ethanol/Diethylamine (95/5/0.1)
• [0404]
  Flow fate: 18.9 mL/min

(−)-4-[((4S)-5,7-Difluoro-3,4-dihydro-2H-chromen-4-yl)oxy]-N,N,2-trimethyl-1H-benzimidazole-6-carboxamide (fraction-1)

• [0405]
  ¹H NMR: spectrum data were identical with those of the racemate
• [0406]
  optical rotation: [α]_D ²³=−101.1° (c=1.00, Methanol)
• [0407]
  retention time: 14 min

(+)-4-[((4R)-5,7-Difluoro-3,4-dihydro-2H-chromen-4-yl)oxy]-N,N,2-trimethyl-1H-benzimidazole-6-carboxamide (fraction-2)

• [0408]
  ¹H NMR: spectrum data were identical with those of the racemate
• [0409]
  optical rotation: [α]_D ²³=+104.2° (c=1.00, Methanol)
• [0410]
  retention time: 18 min
• The following is the alternative method for synthesizing (−)-4-[((4S)-5,7-difluoro-3,4-dihydro-2H-chromen-4-yl)oxy]-N,N,2-trimethyl-1H-benzimidazole-6-carboxamide.

STEP 1: 6-Bromo-2-methyl-4-[(phenylmethyl)oxy]-1H-benzimidazole

• [0411]
  A mixture of N-{4-bromo-2-nitro-6-[(phenylmethyl)oxy]phenyl}acetamide (120 g, 329 mmol, STEP 1 in Example 1) and iron powder (55.1 g, 986 mmol) in acetic acid (500 mL) was refluxed with stirring for 6 hours. After cooling to room temperature, the mixture was filtered through a pad of Celite, and the filtrate was concentrated in vacuum. The residue was diluted with ethyl acetate (1.5 L). The resulted precipitates were filtered through a pad of Celite, and washed with ethyl acetate (500 mL). The filterate was concentrated in vacuum, and the residue was diluted with ethyl acetate (200 mL). The brine (800 mL) was added to the organic mixture, the resulted white precipitates were collected by filtration, and washed with water (200 mL) and diethyl ether (200 mL). The white solid was dissolved with dichloromethane/methanol (10:1, 1.0 L), dried over magnesium sulfate, and concentrated. The solid was triturated with diethyl ether (300 mL), collected by filtration, and dried in vacuum to afford the title compound as a white solid (54.7 g, 53%).
• [0412]
  ¹H NMR (DMSO-d₆, 270 MHz) δ: 7.63-7.28 (m, 7H), 5.38 (s, 2H), 2.69 (s, 3H) ppm. (NH was not observed.)
• [0413]
  MS (ESI) m/z: 317 (M+H)⁺, 315 (M−H)⁻.

STEP 2: 6-Bromo-2-methyl-1-[(4-methylphenyl)sulfonyl]-4-[(Phenylmethyl)oxy]-1H-benzimidazole

• [0414]
  To a suspension of 6-bromo-2-methyl-4-[(phenylmethyl)oxy]-1H-benzimidazole (79.2 g, 250 mmol, STEP 1) in N,N-dimethylformamide (500 mL) was added sodium hydride (60% in mineral oil, 12.0 g, 300 mmol) at 0° C. After stirring at room temperature for 20 minutes, the reaction mixture was cooled to 0° C. To the mixture was added 4-methylbenzenesulfonyl chloride (47.6 g, 250 mmol) at 0° C., and the reaction mixture was stirred at room temperature for 30 minutes. The mixture was quenched with water (800 mL), and the white precipitates were collected by filtration, washed with diisopropyl ether (500 mL), and dried in vacuum at 70° C. for 7 hours to afford the title compound as a white solid (116 g, 98%).
• [0415]
  ¹H NMR (DMSO-d₆, 270 MHz) δ: 7.98 (d, J=8.1 Hz, 2H), 7.64 (d, J=1.9 Hz, 1H), 7.53-7.34 (m, 7H), 7.22 (d, J=1.9 Hz, 1H), 5.28 (s, 2H), 2.74 (s, 3H), 2.38 (s, 3H) ppm.
• [0416]
  MS (ESI) m/z: 471 (M+H)⁺, 469 (M−H)⁻.

STEP 3: N,N,2-Trimethyl-1-[(4-methylphenyl)sulfonyl]-4-[(phenylmethyl)oxy]-1H-benzimidazole-6-carboxamide

• [0417]
  A mixture of 6-bromo-2-methyl-1-[(4-methylphenyl)sulfonyl]-4-[(phenylmethyl)oxy]-1H-benzimidazole (53.0 g, 112 mmol, STEP 2) and tetrakis(triphenylphosphine)palladium(0) (25.9 g, 22.4 mmol) in 2M dimethylamine tetrahydrofuran solution (580 mL) was stirred at 65° C. under carbon mono-oxide gas (1 atmosphere) for 32 hours. The mixture was cooled to room temperature, and diluted with ethyl acetate (600 mL). The organic mixture was washed with saturated ammonium chloride aqueous solution (800 mL) and brine (500 mL), dried over magnesium sulfate and concentrated in vacuum. The residue was purified by column chromatography on silica gel (hexane:ethyl acetate gradient elution from 1:2 to 1:3) to afford the title compound as a white solid (21.8 g, 42%).
• [0418]
  ¹H NMR: spectrum data were identical with STEP 6 in Example 1.

STEP 4: 4-Hydroxy-N,N,2-trimethyl-1-[(4-methylphenyl)sulfonyl]-1H-benzimidazole-6-carboxamide

• [0419]
  A mixture of N,N,2-trimethyl-1-[(4-methylphenyl)sulfonyl]-4-[(phenylmethyl)oxy]-1H-benzimidazole-6-carboxamide (29.0 g, 62.6 mmol, STEP 3) and 10% palladium on carbon (6.0 g) in tetrahydrofuran (200 mL) was stirred under hydrogen gas (1 atmosphere) at room temperature for 24 hours. Another 4.0 g of 10% palladium on carbon was added, and the mixture was stirred under hydrogen gas (1 atmosphere) at room temperature for additional 6 hours. The resulted mixture was filtered through a pad of Celite, and the filtrate was concentrated in vacuum to afford the title compound as a white solid (23.0 g, 98%).
• [0420]
  ¹H NMR: spectrum data were identical with STEP 7 in Example 1.

STEP 5: Methyl 3-(3,5-difluorophenoxy)acrylate

• [0421]
  A solution of 3,5-difluorophenol (35.5 g, 273 mmol) and methyl propiolate (25.0 mL, 300 mmol) in acetonitrile (109 mL) was added to a solution of tetrabutylammonium fluoride in tetrahydrofuran (1.0 M commercial solution, 109 mL, 109 mmol) at room temperature over a period of 2 hours. After complete addition of the solution, the mixture was stirred for 1 hour. The reaction mixture was diluted with toluene (350 mL) and the organic mixture was washed twice with water (250 mL×2), dried over magnesium sulfate, and concentrated in vacuum. The residue was purified by column chromatography on amino gel (hexane:ethyl acetate=3:2 as an eluent) to afford the title compound as a yellow solid (60.0 g, quant, 1:1 mixture of cis- and trans-isomers).
• [0422]
  ¹H NMR (CDCl₃, 270 MHz,) δ: 7.72 (d, J=10.8 Hz, 0.5H), 6.83 (d, J=5.4 Hz, 0.5H), 6.74-6.49 (m, 3H), 5.68 (d, J=10.8 Hz, 0.5H), 5.28 (d, J=5.4 Hz, 0.5H), 3.76 (s, 3H) ppm.

STEP 6: Methyl 3-(3,5-difluorophenoxy)propanoate

• [0423]
  A mixture of methyl 3-(3,5-difluorophenoxy)acrylate (60.0 g, 280 mmol, STEP 5), and 10% palladium on carbon (1.0 g) in methanol (300 mL) was stirred under hydrogen gas (1 atmosphere) at room temperature for 18 hours. The reaction mixture was filtered through a pad of Celite, and washed with toluene (100 mL). The filtrate was concentrated in vacuum to afford the title compound (61.0 g, quant) as a colorless oil, which was used in the next step without further purification.
• [0424]
  ¹H NMR (CDCl₃, 270 MHz) δ: 6.56-6.21 (m, 3H), 4.21 (t, J=5.4 Hz, 2H), 3.74 (s, 3H), 2.80 (t, J=5.4 Hz, 2H) ppm.

STEP 7: 5,7-Difluoro-2,3-dihydro-4H-chromen-4-one

• [0425]
  A mixture of methyl 3-(3,5-difluorophenoxy)propanoate (11.6 g, 53.7 mmol, STEP 6) and trifluoromethanesulfonic acid (23.2 mL, 2.0 mL/g of substrate) was stirred at 80° C. for 2 hours. After cooling to room temperature, the reaction mixture was diluted with water (120 mL), and extracted with toluene (120 mL). The organic layer was washed successively with aqueous solution of potassium carbonate (50 mL), water (50 mL), and dried over magnesium sulfate. The organic mixture was concentrated in vacuum to afford the title compound (8.75 g, 88%) as a white solid, which was used in the next step without further purification.
• [0426]
  ¹H NMR (CDCl₃, 270 MHz) δ: 6.51-6.40 (m, 2H), 4.55-4.50 (m, 2H), 2.86-2.75 (m, 2H) ppm.

STEP 8: (+)-5,7-Difluoro-3,4-dihydro-2H-chromen-4-ol

• [0427]
  To a mixture of 1 M (S)-tetrahydro-1-methyl-3,3-diphenyl-1H,3H-pyrrolo[1,2-c][1,3,2]oxazaborole toluene solution (5.43 mL, 5.43 mmol) and tetrahydrofuran (40 mL) was added 2M borane-methyl sulfide complex tetrahydrofuran solution (29.8 mL, 59.7 mmol) at 0° C. and the mixture was stirred for 20 minutes. To the mixture was added a solution of 5,7-difluoro-2,3-dihydro-4H-chromen-4-one (10.0 g, 54.3 mmol, STEP 7) in tetrahydrofuran (70 mL) at 0° C. over a period of 1 hour, and the mixture was stirred at the same temperature for 1 hour. The reaction mixture was quenched with methanol (50 mL) and stirred for 30 minutes at room temperature. The mixture was concentrated in vacuum and the residue was purified by column chromatography on silica gel (hexane:ethyl acetate=4:1 as an eluent) to afford crude white solids (8.85 g, 86% ee). The solids were recrystallized from hexane (300 mL) to give the title compound as a colorless needle crystal (5.90 g, 58%, >99% ee).
• [0428]
  ¹H NMR: spectrum data were identical with those of the racemate (STEP 8-1 in Example 1).
• [0429]
  optical rotation: [α]_D ²⁴=+143.6° (c=1.00, Methanol).

STEP 9: (−)-4-[((4S)-5,7-Difluoro-3,4-dihydro-2H-chromen-4-yl)oxy]-N,N,2-trimethyl-1-[(4-methylphenyl) sulfonyl]-1H-benzimidazole-6-carboxamide

• [0430]
  To a stirred mixture of 4-hydroxy-N,N,2-trimethyl-1-[(4-methylphenyl)sulfonyl]-1H-benzimidazole-6-carboxamide (21.2 g, 56.8 mmol, STEP 4), (+)-5,7-difluoro-3,4-dihydro-2H-chromen-4-ol (15.86 g, 85.1 mmol, STEP 8) and tributylphosphine (22.9 g, 113 mmol) in toluene (840 mL) was added 1,1′-(azodicarbonyl)dipiperidine (ADDP) (19.3 g, 76.5 mmol) at room temperature. After stirring at room temperature for 2 hours, the reaction mixture was filtered through a pad of Celite and washed with ethyl acetate (300 mL). The filtrate was concentrated in vacuum. The residue was purified by column chromatography on silica get (ethyl acetate:hexane gradient elution from 1:20 to 20:1) to afford crude solids (27.0 g). The solids were recrystallized from 2-propanol (130 mL) to give the title compound as a colorless crystal (23.2 g, 75%, >99% ee)
• [0431]
  ¹H NMR: spectrum data were identical with those of the racemate (STEP 8-2 in Example 1).
• [0432]
  optical rotation: [α]_D ²⁴=80.4° (c=0.50, Methanol).

STEP 10: (−)-4-[((4S)-5,7-Difluoro-3,4-dihydro-2H-chromen-4-yl)oxy]-N,N,2-trimethyl-1H-benzimidazole-6-carboxamide

• [0433]
  To a solution of (−)-4-[((4S)-5,7-difluoro-3,4-dihydro-2H-chromen-4-yl)oxy]-N,N,2-trimethyl-1-[(4-methylphenyl)-sulfonyl]-1H-benzimidazole-6-carboxamide (24.2 g, 44.7 mmol, STEP 9) in tetrahydrofuran (65 mL) and 2-propanol (220 mL) was added 2M sodium hydroxide aqueous solution (220 mL, 440 mmol) at room temperature. After stirring at room temperature for 4 hours, the mixture was diluted with ethyl acetate (1.20 L) and washed with saturated ammonium chloride aqueous solution (500 mL). The organic solution was dried over magnesium sulfate and concentrated in vacuum. The residue was purified by column chromatography on amino gel (ethyl acetate:methanol gradient elution from 50:1 to 20:1) to afford the title compound as a white solid (15.2 g, 87%, >99% ee).
• [0434]
  ¹H NMR: spectrum data were identical with those of the racemate (STEP 9 in Example 1).
• [0435]
  Optical rotation and retention time were identical with the above.

PATENT

WO2021171239

Tegoprazan is the world’s first potassium-competitive acid blocker (P-CAB), has a mechanism similar to that of an acid pump antagonist (APA), and blocks gastric acid secretion by competing with potassium ions for binding to the enzyme H⁺/K⁺– ATPase (proton pump) that secretes H⁺ ions, which are a component of gastric acid, from the gastric parietal cells into the gastric lumen. Since tegoprazan is not a prodrug such as a proton pump inhibitor (PPI), it does not require an activation process, and thus acts not only on an active proton pump but also on an inactive proton pump. Thus, tegoprazan has the advantages of exhibiting its effect rapidly and reaching the maximum effect within one hour.

Meanwhile, in general, in order for a drug to exhibit an expected effect, the blood concentration of the drug needs to be maintained at a certain level or higher. To maintain the blood concentration of the drug, a patient is required to take the prescribed drug repeatedly according to a certain schedule.

In this case, taking the drug frequently decreases the patient’s medication compliance, and as a result, there are many cases where the expected therapeutic effect is not obtained. Thus, in a disease for which a drug needs to be taken for a long period of time or the blood concentration of the drug at a time when the patient cannot take the drug needs to be maintained at a certain level or higher, the frequency and method of taking the drug is also an important factor to be considered for increasing the therapeutic effect of the drug.

Accordingly, there is a need to develop a formulation capable of maintaining a therapeutically effective blood concentration of a drug because there is no problem in the absorption rate of the drug while modifying the release of the drug.

DISCLOSURE

PATENT

CN326416556

Gastric acid-related gastrointestinal diseases, such as gastroesophageal reflux disease, non-erosive reflux disease, gastric ulcers, and ulcers caused by non-steroidal anti-inflammatory drugs are the most common diseases of the gastrointestinal tract. Histamine 2 receptor blockers and proton pump inhibitors (PPIs) are used in the treatment of the above symptoms, showing good curative effects and greatly improving the quality of life of patients. However, the degree of satisfaction with existing drugs for the treatment of gastrointestinal diseases related to gastric acid is still not high. For example, during the process of taking proton pump inhibitors, the symptoms of heartburn and esophageal reflux at night are still difficult to overcome, and the related symptoms cannot be effectively relieved 3 days before taking the medicine.

Potassium ion competitive acid blocker (P-CAB) is a new mechanism of H ⁺ -K ⁺ -ATPase inhibitor, which is a reversible proton pump inhibitor. Currently on the market are Revaprazan, Vonoprazan and Tegoprazan.

Tegoprazan’s chemical name is (S)-4-((5,7-difluorochroman-4-yl)oxy)-N,N,2-trimethyl-1H-benzo [d] Imidazole-6-carboxamide, the structure is shown in formula (1):

WO2007072146 and CN101341149B quote the synthesis method of WO2004054984 to prepare A-3 compound, then acetylate under concentrated sulfuric acid/acetic anhydride, introduce cyano group through microwave reaction to obtain A-5 compound, and then undergo reduction, ring closure, hydrolysis, condensation, and Toluenesulfonyl protection, ether hydrogenolysis, Mitsunobu reaction (Mitsunobu reaction) to obtain A-11 compound, after hydrolysis to remove the p-toluenesulfonyl protecting group to obtain A-12 compound, namely Tegorazan racemate, and finally through a chiral column Split to obtain Tegorazan with optical activity.

This synthetic route requires 12 steps of reactions (not including the preparation of 5,7-difluorochroman-4-ol), and the synthesis yield is only 2.0%; zinc cyanide is used in the reaction, which requires special treatment of wastewater; In the reaction, the protecting group (benzyl protection, p-toluenesulfonyl protection) and the removal of the protecting group need to be carried out twice. Suitable for industrial production.

Method two (ten-gram preparation method):

The obtained A-4 compound is reduced and fused under the condition of iron powder/acetic acid to obtain A-13 compound, which is protected by p-toluenesulfonyl, amidation, and debenzyl protection to obtain A-10 compound, and finally combined with a chiral alcohol The Tegorazan precursor is obtained by the Mitsunobu reaction, and then the Tegorazan is obtained by hydrolysis to protect it.

Although method 2 has been shortened compared with method 1, the synthetic route still requires 9-step reaction (excluding the preparation of chiral alcohol), the route is still longer, and the total yield is 6.8%; carbon monoxide gas is used in the reaction to pass through the coupling Co-preparation of amides requires special equipment to carry out the reaction, which poses a safety hazard; two protective groups (benzyl protection, p-toluenesulfonyl protection) and two removal of protective groups are still required in the reaction, and the reaction steps are also added. This results in low synthesis efficiency, which is not conducive to industrial production.

The comparative document CN101341149B discloses the preparation method of compound 5, that is, the tetrahydrofuran solution of 5,7-difluorochroman-4-one is added to the chiral reagent (S)-1-methyl-3,3- Diphenyl-1H,3H-pyrrolo[1,2-c][1,3,2]oxazolborane, borane-dimethyl sulfide complex and tetrahydrofuran in a mixed solution, wait until the reaction is complete After purification by column chromatography, the chiral purity was 86% ee, and then recrystallized with hexane to obtain compound 5, the optical purity of which was >99% ee, and the yield was 58%.

The comparative document CN107849003A discloses the preparation methods of compounds 3 and 5, that is, 5,7-difluorochroman-4-one is used as a raw material for reduction with a chiral ruthenium catalyst, and the yield of compound 3 is 85%. The purity is 100% ee, the yield of the obtained compound 5 is 91%, and the chiral purity is 100% ee. This method involves ruthenium reagents that are difficult to purchase commercially and are expensive.

Patent EP2390254A1 discloses the preparation method of compound 2, which uses 3-fluoro-4nitrobenzoic acid in dichloromethane with oxalyl chloride and N,N-dimethylformamide to obtain acid chloride after concentration, and then the obtained The acid chloride is dissolved in dichloromethane, and then added dropwise to a mixed solution containing dimethylamine hydrochloride and triethylamine for preparation, and the purification method adopts column chromatography for purification.

PATENT

CN297688244

Patents

CN 112851646

CN 111303131,

US 20070142448

Tegoprazan was approved by the Ministry of Food and Drug Safety (MFDS) for marketing in July 2018 for the treatment of gastroesophageal reflux disease and erosive esophagitis. Tegoprazan was originally developed by Pfizer. In 2008, it was licensed to RaQualiaPharma (separated from Pfizer) for joint development. In 2014, Tegoprazan was licensed to CJHealthCare by RaQualiaPharma. Finally, CJHealthCare was successfully developed and marketed in Korea. Tegoprazan is a competitive potassium ion acid blocker (P-CAB) and hydrogen ion/potassium ion exchange ATPase (H+/K+ATPase) inhibitor. The drug was first marketed in South Korea. Medicines for treating gastroesophageal reflux disease and erosive esophagitis. Proton pump hydrogen ion/potassium ion exchange ATPase is the main pharmacological target for the treatment of gastric acid-related diseases. Potassium-competitive acid blocker (P-CAB) can inhibit gastric acid secretion by competitively binding to K+ with H+/K+-ATPase. Research finds that Tegoprazan is such a potassium-competitive acid blocker and is considered to be the most advanced drug for treating gastroesophageal reflux disease, because proton pump inhibitors are the most commonly used drugs for treating gastroesophageal reflux disease. Tegoprazan The shortcomings of proton pump inhibitors can be just overcome. Tegoprazan’s effectiveness and safety are mainly based on two phase III clinical trials. One of them is a double-blind, active-controlled phase III study. This study was conducted in South Korea. The study used 280 patients with erosive esophagitis as the primary endpoint[1].

Fig 1. Chemical structure formula and three-dimensional structure of Tegoprazan

Tegoprazan, a potassium-competitive acid blocker, is a potent, oral active and highly selective inhibitor of gastric H+/K+-ATPase that could control gastric acid secretion and motility, with IC50 values ranging from 0.29-0.52 μM for porcine, canine, and human H+/K+-ATPases in vitro.

Tegoprazan inhibits porcine, canine, and human H+/K+-ATPase activity. Tegoprazan inhibits gastric H+/K+-ATPase in a potassium-competitive and reversible manner. Tegoprazan (3 μM) inhibits 86% of H+/K+-ATPase activity, whereas the inhibition is decreased to 34% after the dilution of Tegoprazan concentration to 0.15 μM[2].

Tegoprazan (1.0 mg/kg, p.o.) potently and completely inhibits histamine-induced gastric acid secretion in dogs. Tegoprazan (1.0-3.0 mg/kg, p.o.) reverses the pentagastrin-induced acidified gastric pH to the neutral range. Tegoprazan (3 mg/kg, p.o.) immediately evokes a gastric phase III contraction of the migrating motor complex in pentagastrin-treated dogs[3].

The invention relates to a method for preparing Tegoprazan chiral alcohols, in particular to the preparation method of (S) 5,7 difluoro 3,4 dihydro 2H chromogenic ene 4 alcohol. Using 5,7-difluoro-4H-benzopyran-4-ketone as starting material, the method realizes the preparation of (S)5,7-difluoro-3,4-dihydro-2H-chromogenic enone-4-alcohol by asymmetric reduction of ketone carbonyl with chiral reagent and subsequent conventional hydrogenation reaction[4].

Tegoprazan, a reversible H+/K+-ATPase inhibitor developed by CJ Healthcare (now inno.N), was first approved and launched in South Korea in 2019 for the treatment of gastroesophageal reflux disease (GERD). In 2020, the product attained supplemental approval for the treatment of gastric ulcers and Helicobacter pylori infection. Additional phase III clinical trials are being conducted by Shandong Luoxin Pharmacy Group, CJ Healthcare’s Chinese licensee. Tegoprazan was originally developed by RaQualia and licensed to CJ CheilJedang (the parent company of CJ Healthcare) in 2010 in Southeastern Asian markets; this agreement was later extended to Europe and North America in 2019. In 2015, a Chinese sublicense was granted to Shandong Luoxin Pharmacy Group. CJ Healthcare was acquired by Kolmar Korea in 2018, and renamed as inno.N in 2020.

NEW DRUG APPROVALS

one time

$10.00

Click here to purchase.

References

[1]  Takahashi N, et al. Tegoprazan, a Novel Potassium-Competitive Acid Blocker to Control Gastric Acid Secretion and Motility. J Pharmacol Exp Ther. 2018 Feb;364(2):275-286.

[2] Nobuyuki Takahashi and Yukinori Take.Journal of Pharmacology and Experimental Therapeutics February 2018, 364 (2) 275-286.

[3] Kim HK, Park SH, Cheung DY, Cho YS, Kim JI, Kim SS, Chae HS, Kim JK, and Chung IS (2010) Clinical trial: inhibitory effect of revaprazan on gastric acid secretion in healthy male subjects. J Gastroenterol Hepatol 25:1618–1625.

Mikami T, Ochi Y, Suzuki K, Saito T, Sugie Y, and Sakakibara M (2008) 5-Amino-6-chloro-N-[(1-isobutylpiperidin-4-yl)methyl]-2-methylimidazo[1,2-α]pyridine-8-carboxamide (CJ-033,466), a novel and selective 5-hydroxytryptamine4 receptor partial agonist: pharmacological profile in vitro and gastroprokinetic effect in conscious dogs. J Pharmacol Exp Ther 325:190–199.

/////// tegoprazan, Тегопразан , تيغوبرازان , 替戈拉生 ,  CJ-12420, IN-A001, K-CAB, LXI-15028, RQ-00000004,  RQ-4, CJ 12420, IN A001, K CAB, LXI 15028, RQ 00000004,  RQ 4, korea 2019

CN(C)C(=O)c1cc(O[C@H]2CCOc3cc(F)cc(F)c23)c4[nH]c(C)nc4c1

THIAMINE

• Molecular FormulaC₁₂H₁₇N₄OS
• Average mass265.354 Da

• 59-43-8    has cl atom
• 70-16-6 [RN]  no cl atom

• Thiazolium, 3-[(4-amino-2-methyl-5-pyrimidinyl)methyl]-5-(2-hydroxyethyl)-4-methyl-, chloride, hydrochloride (1:1:1), Thiamine CL  hcl, 67-03-8, (Component: 70-16-6) 1;1;1,
• C₁₂ H₁₇ N₄ O S . Cl H . Cl

3595616 [Beilstein]

3-[(4-Amino-2-methyl-5-pyrimidinyl)methyl]-5-(2-hydroxyethyl)-4-methylthiazolium

thiamin hydrochloride
Vitamin B1 hydrochloride
thiamine hydrochloride
aneurin hydrochloride
3-(4-amino-2-methyl-5-pyrimidinyl)methyl-5-(2-hydroxyethyl)-4-methylthiazolium chloride hydrochlorideThiamineCAS Registry Number: 59-43-8CAS Name: 3-[(4-Amino-2-methyl-5-pyrimidinyl)methyl]-5-(2-hydroxyethyl)-4-methylthiazolium chlorideAdditional Names: vitamin B1; aneurin; thiamine monochloride; thiaminium chlorideMolecular Formula: C12H17ClN4OSMolecular Weight: 300.81Percent Composition: C 47.91%, H 5.70%, Cl 11.79%, N 18.63%, O 5.32%, S 10.66%Literature References: Essential nutrient required for carbohydrate metabolism; also involved in nerve function. Biosynthesized by microorganisms and plants. Dietary sources include whole grains, meat products, vegetables, milk, legumes and fruit. Also present in rice husks and yeast. Converted in vivo to thiamine diphosphate, a coenzyme in the decarboxylation of a-keto acids. Chronic deficiency may lead to neurological impairment, beriberi, Wernicke-Korsakoff syndrome. Isoln from rice bran: B. C. P. Jansen, W. F. Donath, Chem. Weekbl.23, 201 (1926). 
Structure: R. R. Williams, J. Am. Chem. Soc.58, 1063 (1936); R. R. Williams, J. K. Cline, ibid. 1504; R. R. Williams et al.,ibid.59, 526 (1937). Review of syntheses: Knobloch in H. Vogel, Chemie und Technik der Vitaminevol. II (Stuttgart, 1953) pp 1-128. Toxicity data: D. Winter et al.,Int. Z. Vitaminforsch.37, 82 (1967). HPLC determn in foods, pharmaceuticals, body tissues: T. Kawaski, Methods Enzymol.122, 15 (1986); in plasma and pharmacokinetics: H. Mascher, C. Kikuta, J. Pharm. Sci.82, 56 (1993). 
Review of bioavailability, absorption, and role in nutrition: F. L. Iber et al.,Am. J. Clin. Nutr.36, 1067-1082 (1982). Reviews: “Thiamin: Twenty Years of Progress”, Ann. N.Y. Acad. Sci.378, H. Z. Sable, C. J. Grubier, Eds. (1982) 470 pp; “Thiamin, Vitamin B1, Aneurin” in Vitamins, W. Friedrich, Ed. (de Gruyter, Berlin, 1988) pp 339-401. 
Derivative Type: HydrochlorideCAS Registry Number: 67-03-8Additional Names: Thiamine chloride hydrochloride; thiamine dichlorideTrademarks: Benerva (Roche); Betabion (Merck KGaA); Betalin S (Lilly); Betaxin (Sterling Winthrop); Bewon (Wyeth); Metabolin (Takeda); Vitaneurin (Mepha)Molecular Formula: C12H17ClN4OS.HClMolecular Weight: 337.27Percent Composition: C 42.73%, H 5.38%, Cl 21.02%, N 16.61%, O 4.74%, S 9.51%Literature References: Comprehensive description: K. A. M. Al-Rashood et al.,Anal. Profiles Drug Subs.18, 413-458 (1989).Properties: Monoclinic plates in rosette-like clusters. Slight thiazole odor. Bitter taste. dec 248°. One gram dissolves in ~1 ml water, 18 ml glycerol, 100 ml 95% alcohol, 315 ml abs alcohol; more sol in methanol. Sol in propylene glycol. Practically insol in ether, benzene, hexane, chloroform. pH of a 1% w/v soln in water 3.13; pH of a 0.1% w/v soln in water 3.58. 
On exposure to air of average humidity, the vitamin absorbs an amount of water corresponding to nearly one mol, forming a hydrate. LD50 in mice (mg/kg): 89.2 i.v.; 8224 orally (Winter).Toxicity data: LD50 in mice (mg/kg): 89.2 i.v.; 8224 orally (Winter) 
Derivative Type: MononitrateCAS Registry Number: 532-43-4Molecular Formula: C12H17N5O4SMolecular Weight: 327.36Percent Composition: C 44.03%, H 5.23%, N 21.39%, O 19.55%, S 9.80%Literature References: Prepn: R. J. Turner, G. J. Schmitt, US2844579 (1958 to Am. Cyanamid).Properties: Crystals, mp 196-200° (dec). Practically nonhygroscopic. pKa 4.8. Soly in water (g/100 ml): 2.7 (25°); ~30 (100°). pH of 2% aq soln 6.5 to 7.1. More stable than the hydrochloride; suitable for enrichment of flours and feeds, multivitamin prepns.Melting point: mp 196-200° (dec)pKa: pKa 4.8 
Therap-Cat: Vitamin (enzyme cofactor).Therap-Cat-Vet: Vitamin (enzyme cofactor).Keywords: Enzyme Cofactor; Vitamin/Vitamin Source; Vitamin B1.

Vitamin B₁ (Thiamine)

Deficiency of this causes beriberi

Some symptoms of ‘dry’ beriberi. There is also a ‘wet’ version of beriberi which mainly affects the heart and circulatory system,
with shortness of breath, swelling of the lower legs, and increased heart rate. 
According to the global “Vitamin B1 (Thiamine Mononitrate) Market 2020” research report, the global vitamin B1 market revenue was USD 648.8 million in 2020 and will be projected to reach USD 854.7 million by 2026.Global Vitamin B1 (Thiamine Mononitrate) Sales Market Report 2020, 2020. Fully Continuous Flow Synthesis of 3-Chloro-4-oxopentyl Acetate: An Important Intermediate for Vitamin B1
M Jiang, M Liu, C Yu, D Cheng… – … Process Research & …, 2021 – ACS Publications
… Journal Logo. Fully Continuous Flow Synthesis of 3-Chloro-4-oxopentyl Acetate:
An Important Intermediate for Vitamin B1. Meifen Jiang* Meifen Jiang. Shanghai
Engineering Center of Industrial Asymmetric Catalysis for Chiral … 
SPECTROSCOPY 

Compound Name:
Thiamin hydrochlorideMolecular Formula: C₁₂H₁₇ClN₄OSMolecular Weight: 300.8CAS Registry No.:
67-03-8 MASS

13C NMR D2O 

¹H NMR : 400 MHz in DMSO-d₆

IR 

SynCN108239084 – PRODUCTION DEVICE OF MEDICINE THIAMINE HYDROCHLORIDE FOR TREATING NEURITIS 

https://patentscope.wipo.int/search/en/detail.jsf?docId=CN223080274&_cid=P12-KT00YC-33991-1
 The production device for the treatment drug thiamine hydrochloride for neuritis. The production process is as follows: add acetamidine hydrochloride and α-dimethoxymethyl-β-methoxymethylpropionitrile into the reactor D101, and condense in an alkaline medium Is 3,6-dimethyl 1,2-dihydro-2,4,5,7-tetrazine (Ⅱ), which is then hydrolyzed to obtain the intermediate product (Ⅲ), which is then closed to form 2-methyl in alkaline 4-amino-5-aminomethyl pyrimidine (IV), introduced into D102, continue to react with carbon disulfide and ammonia to obtain (Ⅴ), then condense with acetic acid-γ-chloro-γ propyl acetate, and then in hydrochloric acid After hydrolysis and cyclization, thiothiamine hydrochloride is obtained, which is pumped into D103, neutralized with ammonia water, oxidized by hydrogen peroxide, and then converted into ammonium nitrate thiamine with nitric acid, and finally hydrochloric acid is added to obtain the product. The invention has the advantages of reducing the intermediate links of the reaction, reducing the reaction temperature and the reaction time, and improving the reaction yield.

SYNhttps://pubs.acs.org/doi/abs/10.1021/jo00277a036 Journal of Organic Chemistry, 54(16), 3941-5; 1989 

NEW DRUG APPROVALS

ONE TIME

$10.00

Click here to purchase.

SYNhttps://pubs.acs.org/doi/abs/10.1021/acs.oprd.1c00065A fully continuous flow synthesis of 3-chloro-4-oxopentyl acetate (2), an important intermediate for vitamin B₁ (1), was developed. This continuous flow manufacturing included two chemical transformations and an inline extraction step without intermediate purification and solvent exchange. In this work, the traditional synthetic route for batch operation was efficiently simplified via a series of separated screening tests in flows under various conditions. We found that the chlorination reaction can be carried out in only 30 s at room temperature by flow. We also simplified the decarboxylation/acylation step by using a cross-mixer, so that acetic anhydride was no longer required in the acylation reaction. A computational fluid dynamics simulation was carried out to study the improved micromixing of liquid–liquid two-phase streams. Finally, 3-chloro-4-oxopentyl acetate (2) was obtained in a 90% isolated yield with a product purity of 96% and a total residence time of approximately 32 min. This fully continuous process was operated smoothly for 12 h, and approximately 19.1 g of the desired product was generated with a production rate of 1.79 g h^–1.

Batch operation for the decarboxylation/acylation reaction Procedure: 1) Mix acetic acid (3.2 eq.), water (1.1 eq.), and 35 % hydrochloric acid (0.1eq.); 2) Add 1 eq. of 3-acetyl-3-chlorodihydrofuran-2(3H)-one (3) into the mixture at room temperature; 3) Increase the reaction temperature to 120 ℃ to reflux for about 2 hours; 4) Add 2 eq. of acetic anhydride to the mixture; 5) Keep reluxing for another 3 hours; 6) After reaction (analysed by GC-MS), add saturated sodium bicarbonate solution for neutralization to make the pH to be around 7; 7) Add DCM solvent to extract the product for 3 times; 8) Concentrate the DCM solution and distill under vacuum distillation to collect the highly pure product of 3-chloro-4-oxopentyl acetate (2). Distillation condition: 90 ℃, 3-7 mmHg. After 6 hours reaction,the yield of crude product is obtained as 63 % and the purity is around 92 %. After distillation, the purity increases to 95% with an isolation yield of 60%.The production rate for batch is about 1.47 g/h, which is less than the continuous process(1.79g/h). 
syn

CN108239084 – PRODUCTION DEVICE OF MEDICINE THIAMINE HYDROCHLORIDE FOR TREATING NEURITIS

SYN

 Bulletin of the Chemical Society of Japan, 45(7), 2010-15; 1972

https://www.journal.csj.jp/doi/10.1246/bcsj.45.2010

The reaction of 2-dimethoxymethyl-3-methoxypropionitrile (1) with acetamidine produces pyrimidopyrimidine (8) via the consecutive process of 1→an intermediate→8. The intermediate was not isolated, but two structures have been proposed for it. We have now succeeded in the isolation of the intermediate and determined it to be 2-methyl-4-amino-5-dimethoxymethyl-5,6-dihydropyrimidine (4). Several key intermediates were also successfully isolated. The novel reaction pathway for the title reaction was concluded to be as follows: the elimination of methanol from 1, followed by the addition of acetamidine affords 3-acetamidinopropionitrile (3), the subsequent quick cyclization of which produces the intermediate, 4; the further elimination of methanol from 4, followed by a replacement reaction with acetamidine, gives an acetamidinomethylene compound (6), which is converted into the final product, 8, via an intermediate (7). Some minor pathways will also be presented.

syn

CN109467553-PURIFICATION METHOD OF FORMYL PYRIMIDINE AND SYNTHETIC METHOD OF VITAMIN B1

90 Williams, R.R. and Cline, J.K. (1936) Synthesis of vitamin B1. J. Am. Chem. Soc. 58, 1504–1505, https://doi.org/10.1021/ja01299a505SYN

Thiaminpyrophosphate (11) (Figure 1) is an essential cofactor in all forms of life and it plays a key role in carbohydrate and amino acid metabolism by stabilizing acyl carbanion biosynthons. The mechanistic enzymology of thiamin pyrophosphate-dependent enzymes is described in detail in the chapter by Frank Jordan.¹ Here, we will review recent progress on the biosynthesis of thiamin pyrophosphate in bacteria and Saccharomyces cerevisiae with an emphasis on some of the novel organic chemistry that has emerged from these studies. Recent reviews describing the regulation of the pathway,^2,3 the identification of biosynthetic precursors,⁴ and the structural biology of the pathway^5–7 have been published.

SYN

Vitamin B1 338 Commercial production involves a six-step synthetic procedure (Williams & Cline, 1936). Beginning with 339 ethyl 3-ethoxypropionate as the feedstock for vitamin B1 production, the synthetic reactions include (1) 340 formylation using ethyl formate, (2) reaction with acetamidine hydrochloride leading to aminopyrimidine 341 ring formation, (3) replacement of aminopyrimidine hydroxyl group with a chlorine atom (chlorination) 342 using phosphorus(V) oxychloride, (4) replacement of the labile chlorine atom with an amino group using 343 alcoholic ammonia, (5) ammonium salt formation using hydrobromic acid, (6) introduction of the thiazole 344 ring using 4-methyl 5-hydroxyethyl thiazole.

A search of the patent literature revealed two methods for vitamin B1 (thiamine) production by 349 fermentative methods. The first patent describes the development of mutants of the genus Saccharomyces 350 Meyen emend Reess (yeast) for synthesizing vitamin B1 from sugars and inorganic salts (Silhankova, 1980). A 351 more recent invention provides a method for producing thiamine products using a microorganism of the 352 genus Bacillus containing a mutation (i.e., gene deletions or other mutations) that causes it to overproduce 353 and release thiamine products into the medium (Goese, 2012).

PATENT

CN109467553 – PURIFICATION METHOD OF FORMYL PYRIMIDINE AND SYNTHETIC METHOD OF VITAMIN B1

The invention relates to the field of vitamin B1 synthesis, and particularly relates to a purification method of formyl pyrimidine and a synthetic method of vitamin B1. The purification method of formyl pyrimidine comprises the following steps: washing formyl pyrimidine with alcohol; washing formyl pyrimidine with water; dissolving formyl pyrimidine with alcohol, and decoloring formyl pyrimidine with activated carbon to obtain a formyl pyrimidine solution; and separating out formyl pyrimidine in the formyl pyrimidine solution and separating the formyl pyrimidine from the solution to obtain purified formyl pyrimidine. According to the purification method of formyl pyrimidine, by washing the formyl pyrimidine with alcohol and water, decoloring the formyl pyrimidine with activated carbon in an alcohol solution and separation the purification method of formyl pyrimidine by water, impurities in the formyl pyrimidine are removed, the content of the formyl pyrimidine reaches 99.5% over, and agood basis is provided for further synthesizing vitamin B1.

PAPER

HELVETICA CHIMICA ACTA ~ Vol. 73 (1990)

1. 3-Mercapto-4-oxopentyl Acetate (5a). Anh. KSH (7.22 g, 0.1 mol) was suspended in 50 ml of abs. MeOH. The mixture was cooled to 0″ in an ice-bath and 3-chloro-4-oxopentyl acetate (3; 17.9 g, 0.1 mol), previously dissolved in 50 ml of abs. MeOH, was added dropwise in order to maintain the temp. in the mixture between 0 and 5″. After complete addition, stirring was continued at r.t. for 1 h, while a slow stream of N, was passed through the mixture to remove residual H2S. The precipitated KC1 was filtered off and the solvent evaporated under reduced pressure. The residue was taken up in 50 ml of CH,C12 and the insoluble material removed by filtration. Evaporation of the solvent in uamo at 30″ gave 14.9 g of slightly yellow liquid. Bulb-to-bulb distillation of the crude mixture at 120″/0.3 mm yielded 12.95 g (0.07 mol, 73.5%) of 5a as a colourless liquid7). IR (film): 2960w, 2550~. 1740s, 17153, 1370m, 1245s, 1050m. ‘H-NMR (CDCI,): 1.74 (d, J= 12, SH); 1.95-2.25 (m, CH,); 2.05 (s, AcO); 2.35(s,Me);3.42(td,J= 12,5.7,SCH);4.2(t,J=5.7,CH20).EI-MS: 134(2), 116(36),74(21),73(58),43(100). Anal. calc. for C7HI2O,S (176.23): C 47.71, H 6.86, S 18.19; found: C 47.94, H 6.95, S 17.24.

2. 3,4-Dihydro-7-methylpyrimido[4,5-d]pyrimidine (4). From 4-amino-2-methyl-5-(aminomethyl)pyrimidine (Za) and DMF-DMA. In a flask equipped with a Vigreux column and a Liebig condenser, Zag) (69 g, 0.5 mol) was suspended in dimethylformamide dimethyl acetal(59.6 g, 0.5 mol). The stirred suspension was slowly heated to ca. 8&85″, until the temp. at the head of the Vigreux column reached 60°9). The MeOH/Me,NH mixture was then distilled off, until the mixture in the flask became a thick mass. The temp. was increased to 90″ for 30 min, 250 ml of toluene were added, and the obtained suspension was further stirred for 1 h at 90°. It was then allowed to cool to r.t., filtered, and washed twice with 100 ml of hexane. The crude material was dried at SOo under reduced pressure: 69.6 g of a tan solid was obtained, which was then sublimated at 1 SOo (oil-bath temp.) under high vacuum (0.2 mm) togive65.5g(0.44mol,88.5%)of4asawhitesolid. M.p. 173″(dec.).UV:202(4),298(3,7).1R(KBr): 3430m(br.), 2860m, 2840s, 16703, 1620s, 15803, 15303, 1450s. 1210s. ‘H-NMR ((D,)DMSO): 2.4 (s, Me); 4.5 (s, CH,); 7.2 (br. s, vinyl. CH); 8.03 (s, arom. H); 9.9 (br. s, NH). EI-MS: 148 (50, M’), 147 (loo), 106 (12), 53 (17), 42 (20). Anal. calc. for C7H,N, (148.169): C 56.74, H 5.44, N 37.81; found: C 56.79, H 5.44, N 37.75.

From 2a and Triethyl Orthoformate. In a flask equipped with a 20-cm Vigreux column and a Liebig condenser, Zag) (69 g, 0.5 mol), triethyl orthoformate (148.2 g, 1 rnol), and TsOH (2.5 g)”) were introduced. The stirred suspension was slowly heated to ca. 110″ so that the temp. at the head of the Vigreux column reached 80-85″. The EtOH was then distilled off, until the mixture in the flask became a thick mass. The temp. was maintained at 100-1 10″ for 30 min, then 250 ml of toluene were added, and theobtained suspension was further stirred for 1 h at90°. It was cooled to r.t. and placed overnight in the refrigerator. The light-brown precipitate was filtered and washed twice with 50 ml of toluene. The crude material was dried at 50″ under reduced pressure to give 59.3 g of a beige solid which was sublimated at 150″ (oil-bath temp.) under high vacuuni (0.2 mm) to yield 52.5 g (0.35 mol, 71 %) of 4 as a white solid. M.p. 182O (dec.).

3. 3-1 (4-Amino-2-methylpyrimidin-5-yl)methyl]-5-(2-hydroxyethyl)-4-methylthiazolium Chloride Hydrochloride (Thiamine Hydrochloride, la). Compound 4 (7.4 g, 0.05 mol) was dissolved in 100 ml of HCOOH. To this slightly yellow soh, 5a (9.25 g, 0.052 mol) was immediately added at such a rate so that the temp. did not exceed 3540″. The mixture was further stirred for 30 min at r.t. and then 25 ml of a freshly prepared sat. soh. of HCI in abs. EtOH was added dropwise. The temp. rose to 35-36O, and the mixture was further stirred for 30 min at r.t.”), The crude mixture was then poured into a 500-ml flask and evaporated at 50″ under reduced pressure to give 26.07 g of a green-yellow solid residue, which was taken up in 100 ml of ahs. EtOH. Aq. HCI soh. (25%, 30 ml) was then added and the crude mixture heated on a steam-bath, until a clear soln. was obtained. The soln. was cooled to r.t. and placed overnight in the refrigerator. The resulting white crystals were collected and dried in vucuo to yield 14.56 g (86.3%) of la. M.p. 245-246′ (dec.). The mother-liquor was then evaporated at 50O under reduced pressure and the residue taken up in 50 ml of H,O. The aq. phase was then washed twice with 25 ml of CH2C1, and evaporated under reduced pressure to give 3.29 g of a still slightly greenish residue, which was again taken up in 20 ml of abs. EtOH. Aq. HCI soln. (25%, 5 ml) was added and the mixture heated on a steam-bath, until a clear soln. was obtained. It was then cooled to r.t. and kept overnight in the refrigerator. The white crystals were filtered to give 1.42 g (8.4%) of la. M.p. 244-24So(dec.) (combined yieldI2) of la: 94.7% based on 4).

Recrystallization. The two crops of la were combined and dissolved in 100 ml of warm abs. EtOH. Aq. HCI soh (25 %, 40 ml) was added. The soln. was then allowed to cool slowly to r.t. and kept at Oo overnight. The white crystals were filtered and dried in vucuo at 50″ to give 13.6 g (0.04 mol, 80.6 %) of la.

M.p. 243-244″ (dec.). UV: 234 (4.1), 266 (3.9).

IR (KBr): 3500m, 3430m. 3340m. 3240m. 3065s. 2615m. 1660s, 1607m, 1380m.

‘H-NMR (D,O): 2.54(s,Me);2.62(s,Me);3.19(t,J= 5.8,CH2);3.88(t,J= 5.8,CH20);5.56(s,1H,CH2N);8.02(s,1arom.H); proton of thiazole ring is exchanged with deuterium of D,O.

FAB-MS: 265 (100, M+), 181 (18), 144 (30), 123 (65), 122 (65), 91 (78).

Anal. calc. for C,2H18C1,N40S (337.27): C 42.74, H 5.38, N 16.61, S 9.51, CI 21.02; found: C 42.93, H 5.28, N 16.70, S 9.61, C121.17.

////////////////////////////////////////////////////////////////////////////////////////////////////

Thiamine, also known as thiamin or vitamin B₁, is a vitamin found in food and manufactured as a dietary supplement and medication.^[1][4] Food sources of thiamine include whole grains, legumes, and some meats and fish.^[1] Grain processing removes much of the thiamine content, so in many countries cereals and flours are enriched with thiamine.^[1][5] Supplements and medications are available to treat and prevent thiamine deficiency and disorders that result from it, including beriberi and Wernicke encephalopathy.^[3] Other uses include the treatment of maple syrup urine disease and Leigh syndrome.^[3] They are typically taken by mouth, but may also be given by intravenous or intramuscular injection.^[3][6]

Thiamine supplements are generally well tolerated.^[3][7] Allergic reactions, including anaphylaxis, may occur when repeated doses are given by injection.^[3][7] Thiamine is in the B complex family.^[3] It is an essential micronutrient, which cannot be made in the body.^[8] Thiamine is required for metabolism including that of glucose, amino acids, and lipids.^[1]

Thiamine was discovered in 1897, was the first B vitamin to be isolated in 1926, and was first made in 1936.^[9] It is on the World Health Organization’s List of Essential Medicines.^[10] Thiamine is available as a generic medication, and as an over-the-counter drug.^[3]

Medical uses

Thiamine deficiency

Main article: Thiamine deficiency

Thiamine is used to treat thiamine deficiency which when severe can prove fatal.^[11] In less severe cases, non-specific signs include malaise, weight loss, irritability and confusion.^[12] Well-known disorders caused by thiamine deficiency include beriberi, Wernicke–Korsakoff syndrome, optic neuropathy, Leigh’s disease, African seasonal ataxia (or Nigerian seasonal ataxia), and central pontine myelinolysis.^[13]

In Western countries, thiamine deficiency is seen mainly in chronic alcoholism.^[14] Thiamine deficiency is often present in alcohol misuse disorder. Also at risk are older adults, persons with HIV/AIDS or diabetes, and persons who have had bariatric surgery.^[1] Varying degrees of thiamine deficiency have been associated with the long-term use of high doses of diuretics, particularly furosemide in the treatment of heart failure.^[15]

Prenatal supplementation

See also: Prenatal vitamins

Women who are pregnant or lactating require more thiamine. For pregnant and lactating women, the consequences of thiamine deficiency are the same as those of the general population but the risk is greater due to their temporarily increased need for this nutrient. In pregnancy, this is likely due to thiamine being preferentially sent to the fetus and placenta, especially during the third trimester. For lactating women, thiamine is delivered in breast milk even if it results in thiamine deficiency in the mother.^[16] Pregnant women with hyperemesis gravidarum are also at an increased risk for thiamine deficiency due to losses when vomiting.^[17]

Thiamine is important for not only mitochondrial membrane development, but also synaptosomal membrane function.^[18] It has also been suggested that thiamine deficiency plays a role in the poor development of the infant brain that can lead to sudden infant death syndrome (SIDS).^[19]

Other uses

Thiamine is a treatment for some types of maple syrup urine disease and Leigh disease.^[3]

Adverse effects

Thiamine is generally well tolerated and non-toxic when administered orally.^[3] Rarely, adverse side effects have been reported when thiamine is given intravenously including allergic reactions, nausea, lethargy, and impaired coordination.^[20][21]

Chemistry

Thiamine is a colorless organosulfur compound with an unpleasant sulfur odor and the chemical formula C₁₂H₁₇N₄O S. Its structure consists of an aminopyrimidine and a thiazolium ring linked by a methylene bridge. The thiazole is substituted with methyl and hydroxyethyl side chains. Thiamine is soluble in water, methanol, and glycerol and practically insoluble in less polar organic solvents. As a base it can form salts with acids, such as hydrochloride. It is stable at acidic pH, but is unstable in alkaline solutions.^[11][22] Thiamine, which is a persistent carbene, is used by enzymes to catalyze benzoin condensations in vivo.^[23] Thiamine is unstable to heat, but stable during frozen storage.^[24] It is unstable when exposed to ultraviolet light^[22] and gamma irradiation.^[25][26] Thiamine reacts strongly in Maillard-type reactions.^[11]

Biosynthesis

A 3D representation of the TPP riboswitch with thiamine bound

Complex thiamine biosynthesis occurs in bacteria, some protozoans, plants, and fungi.^[27][28] The thiazole and pyrimidine moieties are biosynthesized separately and then combined to form thiamine monophosphate (ThMP) by the action of thiamine-phosphate synthase (EC 2.5.1.3). The biosynthetic pathways may differ among organisms. In E. coli and other enterobacteriaceae, ThMP may be phosphorylated to the cofactor thiamine diphospate (ThDP) by a thiamine-phosphate kinase (ThMP + ATP → ThDP + ADP, EC 2.7.4.16). In most bacteria and in eukaryotes, ThMP is hydrolyzed to thiamine, which may then be pyrophosphorylated to ThDP by thiamine diphosphokinase (thiamine + ATP → ThDP + AMP, EC 2.7.6.2).

The biosynthetic pathways are regulated by riboswitches.^[21] If there is sufficient thiamine present in the cell then the thiamine binds to the mRNAs for the enzymes that are required in the pathway and prevents their translation. If there is no thiamine present then there is no inhibition, and the enzymes required for the biosynthesis are produced. The specific riboswitch, the TPP riboswitch (or ThDP), is the only riboswitch identified in both eukaryotic and prokaryotic organisms.^[29]

Nutrition

Occurrence in foods

Thiamine is found in a wide variety of processed and whole foods. Whole grains, legumes, pork, fruits, and yeast are rich sources.^[30][31]

The salt thiamine mononitrate, rather than thiamine hydrochloride, is used for food fortification, as the mononitrate is more stable, and does not absorb water from natural humidity (is non-hygroscopic), whereas thiamine hydrochloride is hygroscopic.^{[citation needed]} When thiamine mononitrate dissolves in water, it releases nitrate (about 19% of its weight) and is thereafter absorbed as the thiamine cation.

Dietary recommendations

In the U.S. the Estimated Average Requirements (EARs) and Recommended Dietary Allowances (RDAs) for thiamine were updated in 1998, by the Institute of Medicine now known as the National Academy of Medicine (NAM).^[32]

The European Food Safety Authority (EFSA) refers to the collective set of information as Dietary Reference Values, with Population Reference Intake (PRI) instead of RDA, and Average Requirement instead of EAR. AI and UL defined the same as in United States. For women (including those pregnant or lactating), men and children the PRI is 0.1 mg thiamine per megajoule (MJ) of energy consumed. As the conversion is 1 MJ = 239 kcal, an adult consuming 2390 kilocalories should be consuming 1.0 mg thiamine. This is slightly lower than the U.S. RDA.^[33] The EFSA reviewed the same safety question and also reached the conclusion that there was not sufficient evidence to set a UL for thiamine.^[20]

To aid with adequate micronutrient intake, pregnant women are often advised to take a daily prenatal multivitamin. While micronutrient compositions vary among different vitamins, a typical prenatal vitamin contains around 1.5 mg of thiamine.^[34]

For U.S. food and dietary supplement labeling purposes the amount in a serving is expressed as a percentage of Daily Value (%DV). For thiamine labeling purposes 100% of the Daily Value was 1.5 mg, but as of 27 May 2016 it was revised to 1.2 mg to bring it into agreement with the RDA.^[35][36] Compliance with the updated labeling regulations was required by 1 January 2020 for manufacturers with US$10 million or more in annual food sales, and by 1 January 2021 for manufacturers with lower volume food sales.^[37][38] A table of the old and new adult daily values is provided at Reference Daily Intake.

Antagonists

Thiamine in foods can be degraded in a variety of ways. Sulfites, which are added to foods usually as a preservative,^[39] will attack thiamine at the methylene bridge in the structure, cleaving the pyrimidine ring from the thiazole ring.^[12] The rate of this reaction is increased under acidic conditions. Thiamine is degraded by thermolabile thiaminases (present in raw fish and shellfish).^[11] Some thiaminases are produced by bacteria. Bacterial thiaminases are cell surface enzymes that must dissociate from the membrane before being activated; the dissociation can occur in ruminants under acidotic conditions. Rumen bacteria also reduce sulfate to sulfite, therefore high dietary intakes of sulfate can have thiamine-antagonistic activities.

Plant thiamine antagonists are heat-stable and occur as both the ortho- and para-hydroxyphenols. Some examples of these antagonists are caffeic acid, chlorogenic acid, and tannic acid. These compounds interact with the thiamine to oxidize the thiazole ring, thus rendering it unable to be absorbed. Two flavonoids, quercetin and rutin, have also been implicated as thiamine antagonists.^[12]

Food fortification

Main article: Food fortification

Refining grain removes its bran and germ, and thus subtracts its naturally occurring vitamins and minerals. In the United States, B-vitamin deficiencies became common in the first half of the 20th century due to white flour consumption. The American Medical Association successfully lobbied for restoring these vitamins by enrichment of grain, which began in the US in 1939. The UK followed in 1940 and Denmark in 1953. As of 2016, about 85 countries had passed legislation mandating fortification of wheat flour with at least some nutrients, and 28% of industrially milled flour was fortified, often with thiamine and other B vitamins.^[40]

Absorption and transport

Absorption

Thiamine is released by the action of phosphatase and pyrophosphatase in the upper small intestine. At low concentrations, the process is carrier-mediated. At higher concentrations, absorption also occurs via passive diffusion. Active transport is greatest in the jejunum and ileum, but it can be inhibited by alcohol consumption or by folate deficiency.^[11] Decline in thiamine absorption occurs at intakes above 5 mg/day.^[41] On the serosal side of the intestine, discharge of the vitamin by those cells is dependent on Na⁺-dependent ATPase.^[12]

Bound to serum proteins

The majority of thiamine in serum is bound to proteins, mainly albumin. Approximately 90% of total thiamine in blood is in erythrocytes. A specific binding protein called thiamine-binding protein (TBP) has been identified in rat serum and is believed to be a hormone-regulated carrier protein important for tissue distribution of thiamine.^[12]

Cellular uptake

Uptake of thiamine by cells of the blood and other tissues occurs via active transport and passive diffusion.^[11] About 80% of intracellular thiamine is phosphorylated and most is bound to proteins. Two members of the SLC gene family of transporter proteins, SLC19A2 and SLC19A3, are capable of the thiamine transport.^[19] In some tissues, thiamine uptake and secretion appears to be mediated by a soluble thiamine transporter that is dependent on Na⁺ and a transcellular proton gradient.^[12]

Tissue distribution

Human storage of thiamine is about 25 to 30 mg, with the greatest concentrations in skeletal muscle, heart, brain, liver, and kidneys. ThMP and free (unphosphorylated) thiamine is present in plasma, milk, cerebrospinal fluid, and, it is presumed, all extracellular fluid. Unlike the highly phosphorylated forms of thiamine, ThMP and free thiamine are capable of crossing cell membranes. Calcium and magnesium have been shown to affect the distribution of thiamine in the body and magnesium deficiency has been shown to aggravate thiamine deficiency.^[19] Thiamine contents in human tissues are less than those of other species.^[12][42]

Excretion

Thiamine and its acid metabolites (2-methyl-4-amino-5-pyrimidine carboxylic acid, 4-methyl-thiazole-5-acetic acid, and thiamine acetic acid) are excreted principally in the urine.^[22]

Function

Its phosphate derivatives are involved in many cellular processes. The best-characterized form is thiamine pyrophosphate (TPP), a coenzyme in the catabolism of sugars and amino acids. In yeast, TPP is also required in the first step of alcoholic fermentation. All organisms use thiamine, but it is made only in bacteria, fungi, and plants. Animals must obtain it from their diet, and thus, for humans, it is an essential nutrient. Insufficient intake in birds produces a characteristic polyneuritis.

Thiamine is usually considered as the transport form of the vitamin. Five natural thiamine phosphate derivatives are known: thiamine monophosphate (ThMP), thiamine diphosphate (ThDP), also sometimes called thiamine pyrophosphate (TPP), thiamine triphosphate (ThTP), the most recently discovered adenosine thiamine triphosphate (AThTP), and adenosine thiamine diphosphate (AThDP). While the coenzyme role of thiamine diphosphate is well-known and extensively characterized, the non-coenzyme action of thiamine and derivatives may be realized through binding to a number of recently identified proteins which do not use the catalytic action of thiamine diphosphate.^[43]

Thiamine diphosphate

No physiological role is known for thiamine monophosphate (ThMP); however, the diphosphate is physiologically relevant. The synthesis of thiamine diphosphate (ThDP), also known as thiamine pyrophosphate (TPP) or cocarboxylase, is catalyzed by an enzyme called thiamine diphosphokinase according to the reaction thiamine + ATP → ThDP + AMP (EC 2.7.6.2). ThDP is a coenzyme for several enzymes that catalyze the transfer of two-carbon units and in particular the dehydrogenation (decarboxylation and subsequent conjugation with coenzyme A) of 2-oxoacids (alpha-keto acids). Examples include:

• Present in most species
  • pyruvate dehydrogenase and 2-oxoglutarate dehydrogenase (also called α-ketoglutarate dehydrogenase)
  • branched-chain α-keto acid dehydrogenase
  • 2-hydroxyphytanoyl-CoA lyase
  • transketolase
• Present in some species:
  • pyruvate decarboxylase (in yeast)
  • several additional bacterial enzymes

The enzymes transketolase, pyruvate dehydrogenase (PDH), and 2-oxoglutarate dehydrogenase (OGDH) are all important in carbohydrate metabolism. The cytosolic enzyme transketolase is a key player in the pentose phosphate pathway, a major route for the biosynthesis of the pentose sugars deoxyribose and ribose. The mitochondrial PDH and OGDH are part of biochemical pathways that result in the generation of adenosine triphosphate (ATP), which is a major form of energy for the cell. PDH links glycolysis to the citric acid cycle, while the reaction catalyzed by OGDH is a rate-limiting step in the citric acid cycle. In the nervous system, PDH is also involved in the production of acetylcholine, a neurotransmitter, and for myelin synthesis.^[44]

Thiamine triphosphate

Thiamine triphosphate (ThTP) was long considered a specific neuroactive form of thiamine, playing a role in chloride channels in the neurons of mammals and other animals, although this is not completely understood.^[19] However, recently it was shown that ThTP exists in bacteria, fungi, plants and animals suggesting a much more general cellular role.^[45] In particular in E. coli, it seems to play a role in response to amino acid starvation.^[46]

Adenosine thiamine triphosphate

Adenosine thiamine triphosphate (AThTP) or thiaminylated adenosine triphosphate has recently been discovered in Escherichia coli, where it accumulates as a result of carbon starvation.^[47] In E. coli, AThTP may account for up to 20% of total thiamine. It also exists in lesser amounts in yeast, roots of higher plants and animal tissue.^[48]

Adenosine thiamine diphosphate

Adenosine thiamine diphosphate (AThDP) or thiaminylated adenosine diphosphate exists in small amounts in vertebrate liver, but its role remains unknown.^[48]

History

Further information: Vitamin § History

Thiamine was the first of the water-soluble vitamins to be described,^[11] leading to the discovery of more essential nutrients and to the notion of vitamin.

In 1884, Takaki Kanehiro (1849–1920), a surgeon general in the Japanese navy, rejected the previous germ theory for beriberi and hypothesized that the disease was due to insufficiencies in the diet instead.^[49] Switching diets on a navy ship, he discovered that replacing a diet of white rice only with one also containing barley, meat, milk, bread, and vegetables, nearly eliminated beriberi on a nine-month sea voyage. However, Takaki had added many foods to the successful diet and he incorrectly attributed the benefit to increased protein intake, as vitamins were unknown substances at the time. The Navy was not convinced of the need for so expensive a program of dietary improvement, and many men continued to die of beriberi, even during the Russo-Japanese war of 1904–5. Not until 1905, after the anti-beriberi factor had been discovered in rice bran (removed by polishing into white rice) and in barley bran, was Takaki’s experiment rewarded by making him a baron in the Japanese peerage system, after which he was affectionately called “Barley Baron”.

The specific connection to grain was made in 1897 by Christiaan Eijkman (1858–1930), a military doctor in the Dutch Indies, who discovered that fowl fed on a diet of cooked, polished rice developed paralysis, which could be reversed by discontinuing rice polishing.^[50] He attributed beriberi to the high levels of starch in rice being toxic. He believed that the toxicity was countered in a compound present in the rice polishings.^[51] An associate, Gerrit Grijns (1865–1944), correctly interpreted the connection between excessive consumption of polished rice and beriberi in 1901: He concluded that rice contains an essential nutrient in the outer layers of the grain that is removed by polishing.^[52] Eijkman was eventually awarded the Nobel Prize in Physiology and Medicine in 1929, because his observations led to the discovery of vitamins.

In 1910, a Japanese agricultural chemist of Tokyo Imperial University, Umetaro Suzuki (1874-1943), first isolated a water-soluble thiamine compound from rice bran and named it as aberic acid (He renamed it as Orizanin later). He described the compound is not only anti beri-beri factor but also essential nutrition to human in the paper, however, this finding failed to gain publicity outside of Japan, because a claim that the compound is a new finding was omitted in translation from Japanese to German.^[53] In 1911 a Polish biochemist Casimir Funk isolated the antineuritic substance from rice bran (the modern thiamine) that he called a “vitamine” (on account of its containing an amino group).^[54][55] However, Funk did not completely characterize its chemical structure. Dutch chemists, Barend Coenraad Petrus Jansen (1884–1962) and his closest collaborator Willem Frederik Donath (1889–1957), went on to isolate and crystallize the active agent in 1926,^[56] whose structure was determined by Robert Runnels Williams (1886–1965), a US chemist, in 1934. Thiamine was named by the Williams team as “thio” or “sulfur-containing vitamin”, with the term “vitamin” coming indirectly, by way of Funk, from the amine group of thiamine itself (by this time in 1936, vitamins were known to not always be amines, for example, vitamin C). Thiamine was synthesized in 1936 by the Williams group.^[57]

Thiamine was first named “aneurin” (for anti-neuritic vitamin).^[58] Sir Rudolph Peters, in Oxford, introduced thiamine-deprived pigeons as a model for understanding how thiamine deficiency can lead to the pathological-physiological symptoms of beriberi. Indeed, feeding the pigeons upon polished rice leads to an easily recognizable behavior of head retraction, a condition called opisthotonos. If not treated, the animals died after a few days. Administration of thiamine at the stage of opisthotonos led to a complete cure within 30 minutes. As no morphological modifications were observed in the brain of the pigeons before and after treatment with thiamine, Peters introduced the concept of a biochemical lesion.^[59]

When Lohman and Schuster (1937) showed that the diphosphorylated thiamine derivative (thiamine diphosphate, ThDP) was a cofactor required for the oxydative decarboxylation of pyruvate,^[60] a reaction now known to be catalyzed by pyruvate dehydrogenase, the mechanism of action of thiamine in the cellular metabolism seemed to be elucidated. At present, this view seems to be oversimplified: pyruvate dehydrogenase is only one of several enzymes requiring thiamine diphosphate as a cofactor; moreover, other thiamine phosphate derivatives have been discovered since then, and they may also contribute to the symptoms observed during thiamine deficiency. Lastly, the mechanism by which the thiamine moiety of ThDP exerts its coenzyme function by proton substitution on position 2 of the thiazole ring was elucidated by Ronald Breslow in 1958.^[61]

• Some contributors to the discovery of thiamine
• Takaki Kanehiro
• Christiaan Eijkman
• Gerrit Grijns
• Umetaro Suzuki
• Casimir Funk
• Rudolph Peters
• Ronald Breslow

See also

• Vitamin B₁ analogue

References

1. ^ Jump up to:^{a b c d e f} “Office of Dietary Supplements – Thiamin”. ods.od.nih.gov. 11 February 2016. Archived from the original on 30 December 2016. Retrieved 30 December 2016.
2. ^ Royer-Morrot MJ, Zhiri A, Paille F, Royer RJ (1992). “Plasma thiamine concentrations after intramuscular and oral multiple dosage regimens in healthy men”. European Journal of Clinical Pharmacology. 42 (2): 219–22. doi:10.1007/BF00278489. PMID 1618256. S2CID 19924442.
3. ^ Jump up to:^{a b c d e f g h i j} American Society of Health-System Pharmacists. “Thiamine Hydrochloride”. Drugsite Trust (Drugs.com). Retrieved 17 April 2018.
4. ^ “Thiamine: MedlinePlus Drug Information”. medlineplus.gov. Retrieved 30 April 2018.
5. ^ Guidelines on food fortification with micronutrients (PDF). WHO and FAO. 2006. pp. 13–14. ISBN 92-4-159401-2. Retrieved 5 May2018.
6. ^ “Thiamine”. drugbank.ca. Retrieved 30 April 2018.
7. ^ Jump up to:^a b Kliegman RM, Stanton B (2016). Nelson Textbook of Pediatrics. Elsevier Health Sciences. p. 322. ISBN 9781455775668. There are no cases of adverse effects of excess thiamine… A few isolated cases of puritis…
8. ^ Constable PD, Hinchcliff KW, Done SH, Gruenberg W (2017). Diseases of the Nervous System – Veterinary Medicine (Eleventh Edition) – 14. pp. 1155–1370. ISBN 978-0-7020-5246-0. Thiamine (vitamin B1) is synthesized only in bacteria, fungi, and plants but is an essential nutrient for animals.
9. ^ Squires VR (2011). The Role of Food, Agriculture, Forestry and Fisheries in Human Nutrition – Volume IV. EOLSS Publications. p. 121. ISBN 9781848261952. Archived from the original on 30 December 2016.
10. ^ World Health Organization (2019). World Health Organization model list of essential medicines: 21st list 2019. Geneva: World Health Organization. hdl:10665/325771. WHO/MVP/EMP/IAU/2019.06. License: CC BY-NC-SA 3.0 IGO.
11. ^ Jump up to:^{a b c d e f g} Mahan LK, Escott-Stump S, eds. (2000). Krause’s food, nutrition, & diet therapy (10th ed.). Philadelphia: W.B. Saunders Company. ISBN 978-0-7216-7904-4.
12. ^ Jump up to:^{a b c d e f g} Combs Jr GF (2008). The Vitamins: Fundamental Aspects in Nutrition and Health (3rd ed.). Ithaca, NY: Elsevier Academic Press. ISBN 978-0-12-183493-7.
13. ^ McCandless D (2010). Thiamine Deficiency and Associate Clinical Disorders. New York, NY: Humana Press. pp. 157–159. ISBN 978-1-60761-310-7.
14. ^ The Editors of Encyclopaedia Britannica (19 December 2017). “Beriberi”. Encyclopædia Britannica. Retrieved 13 April 2018.
15. ^ Katta N, Balla S, Alpert MA (July 2016). “Does Long-Term Furosemide Therapy Cause Thiamine Deficiency in Patients with Heart Failure? A Focused Review”. The American Journal of Medicine. 129 (7): 753.e7–753.e11. doi:10.1016/j.amjmed.2016.01.037. PMID 26899752.
16. ^ Butterworth RF (December 2001). “Maternal thiamine deficiency: still a problem in some world communities”. The American Journal of Clinical Nutrition. 74 (6): 712–3. doi:10.1093/ajcn/74.6.712. PMID 11722950.
17. ^ Oudman E, Wijnia JW, Oey M, van Dam M, Painter RC, Postma A (May 2019). “Wernicke’s encephalopathy in hyperemesis gravidarum: A systematic review”. European Journal of Obstetrics, Gynecology, and Reproductive Biology. 236: 84–93. doi:10.1016/j.ejogrb.2019.03.006. PMID 30889425.
18. ^ Kloss O, Eskin NA, Suh M (April 2018). “Thiamin deficiency on fetal brain development with and without prenatal alcohol exposure”. Biochemistry and Cell Biology. 96 (2): 169–177. doi:10.1139/bcb-2017-0082. hdl:1807/87775. PMID 28915355.
19. ^ Jump up to:^a b c d Lonsdale D (March 2006). “A review of the biochemistry, metabolism and clinical benefits of thiamin(e) and its derivatives”. Evidence-Based Complementary and Alternative Medicine. 3 (1): 49–59. doi:10.1093/ecam/nek009. PMC 1375232. PMID 16550223.
20. ^ Jump up to:^a b c d Tolerable Upper Intake Levels For Vitamins And Minerals (PDF), European Food Safety Authority, 2006, archived (PDF) from the original on 16 March 2016
21. ^ Jump up to:^a b Bettendorff L (2020). “Thiamine”. In BP Marriott, DF Birt, VA Stallings, AA Yates (eds.). Present Knowledge in Nutrition, Eleventh Edition. London, United Kingdom: Academic Press (Elsevier). pp. 171–88. ISBN 978-0-323-66162-1.
22. ^ Jump up to:^a b c Tanphaichitr V (1999). “Thiamin”. In Shils ME, Olsen JA, Shike M, et al. (eds.). Modern Nutrition in Health and Disease(9th ed.). Baltimore: Lippincott Williams & Wilkins.
23. ^ “Archived copy” (PDF). Archived (PDF) from the original on 14 February 2012. Retrieved 18 March 2011.
24. ^ “Vitamin B1 (Thiamine)”. Medicine LibreTexts. 12 May 2017.
25. ^ Luczak M (1968). “Changes occurring in milk powder subjected to gamma rays”. Zeszyty Problemowe Postepow Nauk Rolniczych. 80(497–501).Chem Abstr 1969;71,2267g
26. ^ Syunyakova ZM, Karpova IN (1966). “The effect of γ-rays and thermal sterilization on the content of thiamine, riboflavine, nicotinic acid, and tocopherol in beef”. Vop Pitan. 25 (2): 52–5. Chem Abstr1966;65,1297b
27. ^ Webb ME, Marquet A, Mendel RR, Rébeillé F, Smith AG (October 2007). “Elucidating biosynthetic pathways for vitamins and cofactors”. Natural Product Reports. 24 (5): 988–1008. doi:10.1039/b703105j. PMID 17898894.
28. ^ Begley TP, Chatterjee A, Hanes JW, Hazra A, Ealick SE (April 2008). “Cofactor biosynthesis–still yielding fascinating new biological chemistry”. Current Opinion in Chemical Biology. 12(2): 118–25. doi:10.1016/j.cbpa.2008.02.006. PMC 2677635. PMID 18314013.
29. ^ Bocobza SE, Aharoni A (October 2008). “Switching the light on plant riboswitches”. Trends in Plant Science. 13 (10): 526–33. doi:10.1016/j.tplants.2008.07.004. PMID 18778966.
30. ^ “Thiamin content per 100 grams; select food subset, abridged list by food groups”. United States Department of Agriculture, Agricultural Research Service, USDA Branded Food Products Database v.3.6.4.1. 17 January 2017. Archived from the original on 2 February 2017. Retrieved 27 January 2017.
31. ^ “Thiamin, Food sources”. Micronutrient Information Center, Linus Pauling Institute, Oregon State University, Corvallis, OR. 2013. Archived from the original on 2 February 2017. Retrieved 27 January 2017.
32. ^ Jump up to:^a b c Institute of Medicine (1998). “Thiamin”. Dietary Reference Intakes for Thiamin, Riboflavin, Niacin, Vitamin B6, Folate, Vitamin B12, Pantothenic Acid, Biotin, and Choline. Washington, DC: The National Academies Press. pp. 58–86. ISBN 978-0-309-06554-2. Archived from the original on 16 July 2015. Retrieved 29 August2017.
33. ^ “Overview on Dietary Reference Values for the EU population as derived by the EFSA Panel on Dietetic Products, Nutrition and Allergies” (PDF). 2017. Archived (PDF) from the original on 28 August 2017.
34. ^ Kominiarek MA, Rajan P (November 2016). “Nutrition Recommendations in Pregnancy and Lactation”. The Medical Clinics of North America. 100 (6): 1199–1215. doi:10.1016/j.mcna.2016.06.004. PMC 5104202. PMID 27745590.
35. ^ “Federal Register May 27, 2016 Food Labeling: Revision of the Nutrition and Supplement Facts Labels. FR page 33982” (PDF). Archived (PDF) from the original on 8 August 2016.
36. ^ “Daily Value Reference of the Dietary Supplement Label Database (DSLD)”. Dietary Supplement Label Database (DSLD). Retrieved 16 May 2020.
37. ^ “Changes to the Nutrition Facts Label”. U.S. Food and Drug Administration (FDA). 27 May 2016. Retrieved 16 May 2020.  This article incorporates text from this source, which is in the public domain.
38. ^ “Industry Resources on the Changes to the Nutrition Facts Label”. U.S. Food and Drug Administration (FDA). 21 December 2018. Retrieved 16 May 2020.  This article incorporates text from this source, which is in the public domain.
39. ^ McGuire M, Beerman KA (2007). Nutritional Sciences: From Fundamentals to Foods. California: Thomas Wadsworth.
40. ^ Annemarie Hoogendoorn, Corey Luthringer, Ibrahim Parvanta and Greg S. Garrett (2016). “Food Fortification Global Mapping Study” (PDF). European Commission. pp. 121–128.
41. ^ Hayes KC, Hegsted DM. Toxicity of the Vitamins. In: National Research Council (U.S.). Food Protection Committee. Toxicants Occurring Naturally in Foods. 2nd ed. Washington DCL: National Academy Press; 1973.
42. ^ Bettendorff L, Mastrogiacomo F, Kish SJ, Grisar T (January 1996). “Thiamine, thiamine phosphates, and their metabolizing enzymes in human brain”. Journal of Neurochemistry. 66 (1): 250–8. doi:10.1046/j.1471-4159.1996.66010250.x. PMID 8522961. S2CID 7161882.
43. ^ Molecular mechanisms of the non-coenzyme action of thiamin in brain: biochemical, structural and pathway analysis : Scientific Reports Archived 31 July 2015 at the Wayback Machine
44. ^ Butterworth RF (2006). “Thiamin”. In Shils ME, Shike M, Ross AC, Caballero B, Cousins RJ (eds.). Modern Nutrition in Health and Disease (10th ed.). Baltimore: Lippincott Williams & Wilkins.
45. ^ Makarchikov AF, Lakaye B, Gulyai IE, Czerniecki J, Coumans B, Wins P, et al. (July 2003). “Thiamine triphosphate and thiamine triphosphatase activities: from bacteria to mammals”. Cellular and Molecular Life Sciences. 60 (7): 1477–88. doi:10.1007/s00018-003-3098-4. PMID 12943234. S2CID 25400487.
46. ^ Lakaye B, Wirtzfeld B, Wins P, Grisar T, Bettendorff L (April 2004). “Thiamine triphosphate, a new signal required for optimal growth of Escherichia coli during amino acid starvation”. The Journal of Biological Chemistry. 279 (17): 17142–7. doi:10.1074/jbc.M313569200. PMID 14769791.
47. ^ Bettendorff L, Wirtzfeld B, Makarchikov AF, Mazzucchelli G, Frédérich M, Gigliobianco T, et al. (April 2007). “Discovery of a natural thiamine adenine nucleotide”. Nature Chemical Biology. 3(4): 211–2. doi:10.1038/nchembio867. PMID 17334376.
48. ^ Jump up to:^a b Frédérich M, Delvaux D, Gigliobianco T, Gangolf M, Dive G, Mazzucchelli G, et al. (June 2009). “Thiaminylated adenine nucleotides. Chemical synthesis, structural characterization and natural occurrence”. The FEBS Journal. 276 (12): 3256–68. doi:10.1111/j.1742-4658.2009.07040.x. PMID 19438713. S2CID 23313946.
49. ^ McCollum EV. A History of Nutrition. Cambridge, Massachusetts: Riverside Press, Houghton Mifflin; 1957.
50. ^ Eijkman C (1897). “Eine Beriberiähnliche Krankheit der Hühner”[A disease of chickens which is similar to beri-beri]. Archiv für Pathologische Anatomie und Physiologie und für Klinische Medicin. 148 (3): 523–532. doi:10.1007/BF01937576. S2CID 38445999.
51. ^ “The Nobel Prize and the Discovery of Vitamins”. nobelprize.org.
52. ^ Grijns G (1901). “Over polyneuritis gallinarum” [On polyneuritis gallinarum]. Geneeskundig Tijdschrift voor Nederlandsch-Indië (Medical Journal for the Dutch East Indies). 41 (1): 3–110.
53. ^ Suzuki U, Shimamura T (1911). “Active constituent of rice grits preventing bird polyneuritis”. Tokyo Kagaku Kaishi. 32: 4–7, 144–146, 335–358. doi:10.1246/nikkashi1880.32.4.
54. ^ Funk, Casimir (1911). “On the chemical nature of the substance which cures polyneuritis in birds induced by a diet of polished rice”. The Journal of Physiology. 43 (5): 395–400. doi:10.1113/jphysiol.1911.sp001481. PMC 1512869. PMID 16993097.
55. ^ Funk, Casimir (1912). “The etiology of the deficiency diseases. Beri-beri, polyneuritis in birds, epidemic dropsy, scurvy, experimental scurvy in animals, infantile scurvy, ship beri-beri, pellagra”. Journal of State Medicine. 20: 341–368. The word “vitamine” is coined on p. 342: “It is now known that all these diseases, with the exception of pellagra, can be prevented and cured by the addition of certain preventative substances; the deficient substances, which are of the nature of organic bases, we will call “vitamines”; and we will speak of a beri-beri or scurvy vitamine, which means a substance preventing the special disease.”
56. ^ Jansen BC, Donath WF (1926). “On the isolation of antiberiberi vitamin”. Proc. Kon. Ned. Akad. Wet. 29: 1390–1400.
57. ^ Williams RR, Cline JK (1936). “Synthesis of vitamin B₁“. J. Am. Chem. Soc. 58 (8): 1504–1505. doi:10.1021/ja01299a505.
58. ^ Carpenter KJ (2000). “Beriberi, white rice, and vitamin B: a disease, a cause, and a cure”. Berkeley, CA: University of California Press.
59. ^ Peters RA (1936). “The biochemical lesion in vitamin B₁deficiency. Application of modern biochemical analysis in its diagnosis”. Lancet. 230 (5882): 1161–1164. doi:10.1016/S0140-6736(01)28025-8.
60. ^ Lohmann K, Schuster P (1937). “Untersuchungen über die Cocarboxylase”. Biochem. Z. 294: 188–214.
61. ^ Breslow R (1958). “On the mechanism of thiamine action. IV.1 Evidence from studies on model systems”. J Am Chem Soc. 80(14): 3719–3726. doi:10.1021/ja01547a064.

External links

• “Thiamine”. Drug Information Portal. U.S. National Library of Medicine.

Bibliography

• Wikipedia: Beriberi; Christiaan Eijkman; Adolphe_Vorderman; Casimir_Funk; Rice Polishing; White rice; Thiamine; Thiamine_pyrophosphate; Citric Acid Cycle
• A. Bay, “Beriberi in Modern Japan: The Making of a National Disease”, University of Rochester Press (2012).
• K.J. Carpenter. Beriberi, White Rice and Vitamin B. University of California Press, 2000
• http://www.healthline.com/health/beriberi
• M.C. Latham, . “Chapter 16. Beriberi and thiamine deficiency” in Human nutrition in the developing world, 29 [Rome, Food and Agriculture Organization of the United Nations, 1997].
• D.-T. Nguyen-Khoa, Beriberi (Thiamine Deficiency) Treatment & Management
• M. Golden, Mike . “Diagnosing Beriberi in Emergency Situations”. Field Exchange 1 (1997) 18.
• Y. Itokawa, . “Kanehiro Takaki (1849–1920): A Biographical Sketch”. J. Nutrit. 106 (1976) 581–8.
• R. Breslow. “On the mechanism of thiamine action. IV.1 Evidence from studies on model systems”. J. Am. Chem. Soc. 80 (1958) 3719–3726.
• R.R. Williams, J.K. Cline,. “Synthesis of vitamin B₁“. J. Am. Chem. Soc. 58 (1936) 1504–1505.
• T.P. Begley, A.Chatterjee, J.W. Hanes, A. Hazra, S.E. Ealick,. “Cofactor biosynthesis—still yielding fascinating new biological chemistry”. Curr. Opin. in Chem. Biol. 12 (2008) 118–125.
• L. Bettendorff, F. Mastrogiacomo, S.J. Kish, T. Grisar, “Thiamine, thiamine phosphates and their metabolizing enzymes in human brain”. J. Neurochem. 66 (1996) 250–258.
• B.C.P. Jansen, W.F. Donath, “On the isolation of antiberiberi vitamin”. Proc. Kon. Ned. Akad. Wet. 29 (1926) 1390–1400.
• C. Nordqvist, “What is Thiamin, or Vitamin B₁?“, Medical News Today, (2016)
• Thiamin, NIH Fact Sheet for Health Professionals.
• Thiamine, Oregon State University

//////////THIAMINE, aneurin hydrochloride, vitamin b1

Cc2ncc(C[n+]1csc(CCO)c1C)c(N)n2

ABX-464

• Molecular FormulaC₁₆H₁₀ClF₃N₂O
• Averrage mass338.712 Da

SPL-4641258453-75-6[RN]26RU378B9V2-Quinolinamine, 8-chloro-N-[4-(trifluoromethoxy)phenyl]-8-Chloro-N-[4-(trifluoromethoxy)phenyl]-2-quinolinamine

EX-A3322, DB14828, SB18690, BS-14770

Abivax is developing ABX464 a lead from HIV-1 splicing inhibitors, which modulates biogenesis of viral RNA, and acts by targeting the Rev protein, for treating HIV infection, rheumatoid arthritis, ulcerative colitis and COVID-19 infection.

In August 2021, ABX464 was reported to be in phase 3 clinical development.

ABX464 is an oral, first-in-class, small molecule that has demonstrated safety and profound anti-inflammatory activity in preclinical trials and in Phase 2a and Phase 2b induction trials to treat ulcerative colitis (UC). Patients who completed the induction studies had the option to roll over into the respective open-label extension studies.
In May 2021, Abivax communicated the top-line results of its randomized, double-blind and placebo-controlled Phase 2b induction trial conducted in 15 European countries, the US and Canada in 254 patients. The primary endpoint (statistically significant reduction of Modified Mayo Score) was met with once-daily ABX464 (25mg, 50mg, 100mg) at week 8.

Further, all key secondary endpoints, including endoscopic improvement, clinical remission, clinical response and the reduction of fecal calprotectin showed significant difference in patients dosed with ABX464 compared to placebo. Importantly, ABX464 also showed rapid efficacy in patients who were previously exposed to biologics and/or JAK inhibitors treatment.

In addition to the top-line induction results, preliminary data from the first 51 patients treated with 50mg ABX464 in the Phase 2b open-label maintenance study showed increased and durable clinical remission and endoscopic improvement after 48 weeks of treatment.

Based on the positive results from the Phase 2a and Phase 2b studies, Abivax plans to advance ABX464 into a Phase 3 clinical program by the end of 2021.

• Originator Splicos
• Developer Abivax
• Class Anti-inflammatories; Antirheumatics; Antivirals; Small molecules
• Mechanism of Action MicroRNA stimulants; Rev gene product inhibitors; RNA cap-binding protein modulators

• Phase II/III COVID 2019 infections
• Phase II Crohn’s disease; Rheumatoid arthritis; Ulcerative colitis
• DiscontinuedHIV infections

• 24 Jun 2021 Discontinued – Phase-II for HIV infections (Adjunctive treatment, Treatment-experienced) in France (PO) (Abivax pipeline, June 2021)
• 24 Jun 2021 Discontinued – Phase-II for HIV infections (Treatment-experienced, Adjunctive treatment) in Belgium (PO) (Abivax pipeline, June 2021)
• 24 Jun 2021
• Discontinued – Phase-II for HIV infections (Treatment-experienced, Adjunctive treatment) in Spain (PO) (Abivax pipeline, June 2021)

Evotec and Abivax in small-molecule pact

by Michael McCoy

September 18, 2017 | A version of this story appeared in Volume 95, Issue 37

The contract research firm Evotec will work with Abivax, a French biotech company, to develop new treatments for viral diseases. Abivax has developed a library of more than 1,000 small molecules designed to inhibit mRNA biogenesis. At its facility in Toulouse, France, Evotec will optimize Abivax’s drug candidates and help develop new drugs for influenza, Dengue, and other viral infections. Abivax’s lead candidate, ABX464, is in Phase II clinical trials as an HIV/AIDS treatment.

PATENT

WO 2010143170

WO 2010143168

WO 2010143169

EP 2974729

WO 2016009065

WO 2017158201

PATENT

WO2016009065

https://patentscope.wipo.int/search/en/detail.jsf?docId=WO2016009065

Buchwald-Hartwig coupling of 2,8-dichloroquinoline (I) with 4-(trifluoromethoxy)aniline (II) using Pd(OAc)2, Cs2CO3 and xantphos or Pd2dba3, K2CO3 and xphos in t-BuOH

PATENT

https://patents.google.com/patent/US10253020B2/en

US 20170226095

COMPD 90

• (90) 8-chloro-N-[4-(trifluoromethoxy)phenyl]quinolin-2-amine

Example 5: Compound (90) of the Table IAccording to route (A), a mixture of 2,8-dichloroquinoline (984 mg) and 4-(trifluoromethoxy)aniline (743 μL), Pd(OAc)₂ (22 mg), XantPhos (58 mg) and Cs₂CO₃ (4.6 g) in 20 mL of t-BuOH gave compound (90) (1.1 g).¹H NMR (300 MHz, CDCl₃) δ 7.84 (d, J=9.1, 2H), 7.79 (d, J=8.9, 1H), 7.67 (dd, J=1.2, 7.6, 1H), 7.48 (dd, J=1.1, 8.0, 1H), 7.18 (s, 3H), 6.89 (s, 1H), 6.75 (d, J=8.9, 1H).¹³C NMR (75 MHz, CDCl₃) δ 153.88, 144.30, 143.91, 139.00, 138.25, 131.13, 130.13, 126.55, 125.42, 123.45, 122.50, 122.17, 120.49, 119.10, 113.24.

PAPER

Tetrahedron Letters (2018), 59(23), 2277-2280.

https://www.sciencedirect.com/science/article/abs/pii/S0040403918305641

Abstract

A solvent-free Buchwald-Hartwig amination had been developed under high-speed ball-milling conditions, which afforded the desired products with moderate to high yields. The addition of sodium sulfate was found to be crucial for improving both the performance and the reproducibility. Comparative solvent-free stirring experiments implicated the importance of mechanical interaction for the transformation, and the inert gas was proved to be unnecessary for this amination.

Graphical abstract

PATENT

WO2015001518

COMPD 90

PATENT

WO-2021152131

Novel co-crystalline polymorphic forms and salts of ABX464 , useful for treating inflammatory diseases, cancer, and diseases caused by viruses eg HIV, severe acute respiratory syndrome caused by SARS-CoV or SARS-CoV-2 infection including strains responsible for COVID-19 and their mutants.

W02010/143169 application describes the preparation and use of compounds, and in particular quinoline derivatives including certain pharmaceutically acceptable salts useful in the treatment of HIV infection. Said application in particular discloses 8-Chloro-N-(4-(trifluoromethoxy)phenyl)quinolin-2-amine also named (8-chloro-quinoline-2-yl)-(4-trifluoromethoxy-phenyl) -amine which is currently under clinical development. The inventors have stated that ABX464 is naturally highly crystalliferous and thus is spontaneously present under a specific unique stable and crystalline form named “crystalline form I”.

W02017/158201 application deals with certain mineral acid or sulfonic acid salts of ABX464.

ABX464 has a poor solubility in aqueous solutions. The main drawback of said poor solubility is that the active ingredient cannot entirely reach their targets in the body if the drug remains undissolved in the gastrointestinal system.

PATENT

WO2021152129 ,

amorphous solid dispersion (eg tablet) comprising ABX464.

PATENT

WO2020127839

https://patentscope.wipo.int/search/en/detail.jsf?docId=WO2020127839

use of quinoline derivatives (ie ABX464) for treating cancer and dysplasia.

///////////ABX464, ABX 464, phase 3 ,  SPL 464, EX A3322, DB14828, SB18690, BS 14770

NEWDRUG APPROVALS

ONE TIME

$10.00

Click here to purchase.

Plasminogen

FDA  APPROVED 2021, Ryplazim, 2021/6/4

Plasminogen;
Glu-plasminogen;
Plasminogen, human-tvmh;
Ryplazim (TN)

RYPLAZIM (plasminogen, human-tvmh)

Enzyme replacement (plasminogen), Plasminogen deficiency type 1

CAS: 9001-91-6

STN: 125659
Proper Name: plasminogen, human-tvmh
Tradename: RYPLAZIM
Manufacturer: Prometic Biotherapeutics Inc.
Indication: 

For the treatment of patients with plasminogen deficiency type 1 (hypoplasminogenemia)

READ  https://diapharma.com/plasminogen-plg/

On August 11, 2017 Prometic Biotherapeutics submitted a BLA (STN 125659) for a Drug Product (DP) RYPLAZIM, Plasminogen (Human). This drug product is indicated for replacement therapy in children and adults with plasminogen deficiency.

Plasmin is an important enzyme (EC 3.4.21.7) present in blood that degrades many blood plasma proteins, including fibrin clots. The degradation of fibrin is termed fibrinolysis. In humans, the plasmin protein is encoded by the PLG gene.^[5]

Function

 Fibrinolysis (simplified). Blue arrows denote stimulation, and red arrows inhibition.

Plasmin is a serine protease that acts to dissolve fibrin blood clots. Apart from fibrinolysis, plasmin proteolyses proteins in various other systems: It activates collagenases, some mediators of the complement system, and weakens the wall of the Graafian follicle, leading to ovulation. Plasmin is also integrally involved in inflammation.^[6] It cleaves fibrin, fibronectin, thrombospondin, laminin, and von Willebrand factor. Plasmin, like trypsin, belongs to the family of serine proteases.

Plasmin is released as a zymogen called plasminogen (PLG) from the liver into the systemic circulation. Two major glycoforms of plasminogen are present in humans – type I plasminogen contains two glycosylation moieties (N-linked to N289 and O-linked to T346), whereas type II plasminogen contains only a single O-linked sugar (O-linked to T346). Type II plasminogen is preferentially recruited to the cell surface over the type I glycoform. Conversely, type I plasminogen appears more readily recruited to blood clots.

In circulation, plasminogen adopts a closed, activation-resistant conformation. Upon binding to clots, or to the cell surface, plasminogen adopts an open form that can be converted into active plasmin by a variety of enzymes, including tissue plasminogen activator (tPA), urokinase plasminogen activator (uPA), kallikrein, and factor XII (Hageman factor). Fibrin is a cofactor for plasminogen activation by tissue plasminogen activator. Urokinase plasminogen activator receptor (uPAR) is a cofactor for plasminogen activation by urokinase plasminogen activator. The conversion of plasminogen to plasmin involves the cleavage of the peptide bond between Arg-561 and Val-562.^[5][7][8][9]

Plasmin cleavage produces angiostatin.

Mechanism of plasminogen activation

Full length plasminogen comprises seven domains. In addition to a C-terminal chymotrypsin-like serine protease domain, plasminogen contains an N-terminal Pan Apple domain (PAp) together with five Kringle domains (KR1-5). The Pan-Apple domain contains important determinants for maintaining plasminogen in the closed form, and the kringle domains are responsible for binding to lysine residues present in receptors and substrates.

The X-ray crystal structure of closed plasminogen reveals that the PAp and SP domains maintain the closed conformation through interactions made throughout the kringle array .^[9] Chloride ions further bridge the PAp / KR4 and SP / KR2 interfaces, explaining the physiological role of serum chloride in stabilizing the closed conformer. The structural studies also reveal that differences in glycosylation alter the position of KR3. These data help explain the functional differences between the type I and type II plasminogen glycoforms.^{[citation needed]}

In closed plasminogen, access to the activation bond (R561/V562) targeted for cleavage by tPA and uPA is blocked through the position of the KR3/KR4 linker sequence and the O-linked sugar on T346. The position of KR3 may also hinder access to the activation loop. The Inter-domain interactions also block all kringle ligand-binding sites apart from that of KR-1, suggesting that the latter domain governs pro-enzyme recruitment to targets. Analysis of an intermediate plasminogen structure suggests that plasminogen conformational change to the open form is initiated through KR-5 transiently peeling away from the PAp domain. These movements expose the KR5 lysine-binding site to potential binding partners, and suggest a requirement for spatially distinct lysine residues in eliciting plasminogen recruitment and conformational change respectively.^[9]

Mechanism of plasmin inactivation

Plasmin is inactivated by proteins such as α2-macroglobulin and α2-antiplasmin.^[10] The mechanism of plasmin inactivation involves the cleavage of an α2-macroglobulin at the bait region (a segment of the aM that is particularly susceptible to proteolytic cleavage) by plasmin. This initiates a conformational change such that the α2-macroglobulin collapses about the plasmin. In the resulting α2-macroglobulin-plasmin complex, the active site of plasmin is sterically shielded, thus substantially decreasing the plasmin’s access to protein substrates. Two additional events occur as a consequence of bait region cleavage, namely (i) a h-cysteinyl-g-glutamyl thiol ester of the α2-macroglobulin becomes highly reactive and (ii) a major conformational change exposes a conserved COOH-terminal receptor binding domain. The exposure of this receptor binding domain allows the α2-macroglobulin protease complex to bind to clearance receptors and be removed from circulation.

Pathology

Plasmin deficiency may lead to thrombosis, as the clots are not adequately degraded. Plasminogen deficiency in mice leads to defective liver repair,^[11] defective wound healing, reproductive abnormalities.^{[citation needed]}

In humans, a rare disorder called plasminogen deficiency type I (Online Mendelian Inheritance in Man (OMIM): 217090) is caused by mutations of the PLG gene and is often manifested by ligneous conjunctivitis.

Interactions

Plasmin has been shown to interact with Thrombospondin 1,^[12][13] Alpha 2-antiplasmin^[14][15] and IGFBP3.^[16] Moreover, plasmin induces the generation of bradykinin in mice and humans through high-molecular-weight kininogen cleavage.^[17]

References

1. ^ Jump up to:^a b c GRCh38: Ensembl release 89: ENSG00000122194 – Ensembl, May 2017
2. ^ Jump up to:^a b c GRCm38: Ensembl release 89: ENSMUSG00000059481 – Ensembl, May 2017
3. ^ “Human PubMed Reference:”. National Center for Biotechnology Information, U.S. National Library of Medicine.
4. ^ “Mouse PubMed Reference:”. National Center for Biotechnology Information, U.S. National Library of Medicine.
5. ^ Jump up to:^a b “Entrez Gene: plasminogen”.
6. ^ Atsev S, Tomov N (December 2020). “Using antifibrinolytics to tackle neuroinflammation”. Neural Regeneration Research. 15(12): 2203–2206. doi:10.4103/1673-5374.284979. PMC 7749481. PMID 32594031.
7. ^ Miyata T, Iwanaga S, Sakata Y, Aoki N (October 1982). “Plasminogen Tochigi: inactive plasmin resulting from replacement of alanine-600 by threonine in the active site”. Proc. Natl. Acad. Sci. U.S.A. 79 (20): 6132–6. Bibcode:1982PNAS…79.6132M. doi:10.1073/pnas.79.20.6132. PMC 347073. PMID 6216475.
8. ^ Forsgren M, Råden B, Israelsson M, Larsson K, Hedén LO (March 1987). “Molecular cloning and characterization of a full-length cDNA clone for human plasminogen”. FEBS Lett. 213 (2): 254–60. doi:10.1016/0014-5793(87)81501-6. PMID 3030813. S2CID 9075872.
9. ^ Jump up to:^a b c Law RH, Caradoc-Davies T, Cowieson N, Horvath AJ, Quek AJ, Encarnacao JA, Steer D, Cowan A, Zhang Q, Lu BG, Pike RN, Smith AI, Coughlin PB, Whisstock JC (2012). “The X-ray crystal structure of full-length human plasminogen”. Cell Rep. 1 (3): 185–90. doi:10.1016/j.celrep.2012.02.012. PMID 22832192.
10. ^ Wu, Guojie; Quek, Adam J.; Caradoc-Davies, Tom T.; Ekkel, Sue M.; Mazzitelli, Blake; Whisstock, James C.; Law, Ruby H.P. (2019-03-05). “Structural studies of plasmin inhibition”. Biochemical Society Transactions. 47 (2): 541–557. doi:10.1042/bst20180211. ISSN 0300-5127. PMID 30837322.
11. ^ Bezerra JA, Bugge TH, Melin-Aldana H, Sabla G, Kombrinck KW, Witte DP, Degen JL (December 21, 1999). “Plasminogen deficiency leads to impaired remodeling after a toxic injury to the liver”. Proc. Natl. Acad. Sci. U.S.A. Proceedings of the National Academy of Sciences of the United States of America. 96 (26): 15143–8. Bibcode:1999PNAS…9615143B. doi:10.1073/pnas.96.26.15143. PMC 24787. PMID 10611352.
12. ^ Silverstein RL, Leung LL, Harpel PC, Nachman RL (November 1984). “Complex formation of platelet thrombospondin with plasminogen. Modulation of activation by tissue activator”. J. Clin. Invest. 74 (5): 1625–33. doi:10.1172/JCI111578. PMC 425339. PMID 6438154.
13. ^ DePoli P, Bacon-Baguley T, Kendra-Franczak S, Cederholm MT, Walz DA (March 1989). “Thrombospondin interaction with plasminogen. Evidence for binding to a specific region of the kringle structure of plasminogen”. Blood. 73 (4): 976–82. doi:10.1182/blood.V73.4.976.976. PMID 2522013.
14. ^ Wiman B, Collen D (September 1979). “On the mechanism of the reaction between human alpha 2-antiplasmin and plasmin”. J. Biol. Chem. 254 (18): 9291–7. doi:10.1016/S0021-9258(19)86843-6. PMID 158022.
15. ^ Shieh BH, Travis J (May 1987). “The reactive site of human alpha 2-antiplasmin”. J. Biol. Chem. 262 (13): 6055–9. doi:10.1016/S0021-9258(18)45536-6. PMID 2437112.
16. ^ Campbell PG, Durham SK, Suwanichkul A, Hayes JD, Powell DR (August 1998). “Plasminogen binds the heparin-binding domain of insulin-like growth factor-binding protein-3”. Am. J. Physiol. 275 (2 Pt 1): E321-31. doi:10.1152/ajpendo.1998.275.2.E321. PMID 9688635.
17. ^ Marcos-Contreras OA, Martinez de Lizarrondo S, Bardou I, Orset C, Pruvost M, Anfray A, Frigout Y, Hommet Y, Lebouvier L, Montaner J, Vivien D, Gauberti M (August 2016). “Hyperfibrinolysis increases blood brain barrier permeability by a plasmin and bradykinin-dependent mechanism”. Blood. 128 (20): 2423–2434. doi:10.1182/blood-2016-03-705384. PMID 27531677.

Further reading

• Shanmukhappa K, Mourya R, Sabla GE, Degen JL, Bezerra JA (July 2005). “Hepatic to pancreatic switch defines a role for hemostatic factors in cellular plasticity in mice”. Proc. Natl. Acad. Sci. U.S.A. 102 (29): 10182–7. Bibcode:2005PNAS..10210182S. doi:10.1073/pnas.0501691102. PMC 1177369. PMID 16006527.
• Anglés-Cano E, Rojas G (2002). “Apolipoprotein(a): structure-function relationship at the lysine-binding site and plasminogen activator cleavage site”. Biol. Chem. 383 (1): 93–9. doi:10.1515/BC.2002.009. PMID 11928826. S2CID 29248198.
• Ranson M, Andronicos NM (2003). “Plasminogen binding and cancer: promises and pitfalls”. Front. Biosci. 8 (6): s294-304. doi:10.2741/1044. PMID 12700073.

External links

• The MEROPS online database for peptidases and their inhibitors: S01.233
• Plasmin at the US National Library of Medicine Medical Subject Headings (MeSH)

///////////Plasminogen, FDA 2021, APPROVALS 2021, Ryplazim

NEW DRUG APPROVALS

ONE TIME

$10.00

Click here to purchase.

Benzonatate

• Molecular FormulaC₃₀H₅₃NO₁₁
• Average mass603.742 Da

104-31-4[RN]2,5,8,11,14,17,20,23,26-Nonaoxaoctacosan-28-yl 4-(butylamino)benzoateбензонататبنزوناتات苯佐那酯ベンゾナテート;KM 652,5,8,11,14,17,20,23,26-nonaoxaoctacosan-28-yl 4-(butylamino)benzoate2-[2-[2-[2-[2-[2-[2-[2-(2-methoxyethoxy)ethoxy]ethoxy]ethoxy]ethoxy]ethoxy]ethoxy]ethoxy]ethyl 4-(butylamino)benzoate

Benzonatate bulk and Benzonatate capsules 100mg, cdsco india 2021, 15.07.2021

For the treatment of refractory coughCAS Registry Number: 104-31-4CAS Name: 4-(Butylamino)benzoic acid 3,6,9,12,15,18,21,24,27-nonaoxaoctacos-1-yl esterAdditional Names: nonaethyleneglycol monomethyl ether p-n-butylaminobenzoate; p-butylaminobenzoic acid w-O-methylnonaethyleneglycol ester; benzononatineTrademarks: Exangit; Tessalon (Forest)Molecular Formula: C30H53NO11Molecular Weight: 603.74Percent Composition: C 59.68%, H 8.85%, N 2.32%, O 29.15%Literature References: Prepn: Matter, US2714608 (1955 to Ciba).Properties: Colorless to faintly yellow oil. Soluble in most organic solvents except aliphatic hydrocarbons.Therap-Cat: Antitussive.Keywords: Antitussive.

Synthesis Reference

Matter, M.; U.S. Patent 2,714,608; August 2, 1955; assigned to Ciba Pharmaceutical Products, Inc.

Synthesis Path

Substances Referenced in Synthesis Path

Benzonatate, sold under the brand name Tessalon among others, is a medication used to try to help with the symptoms of cough and hiccups.^[1][2] It is taken by mouth.^[1] Use is not recommended in those under the age of ten.^[3] Effects generally begin within 20 minutes and last up to eight hours.^[1][4]

Side effects include sleepiness, dizziness, headache, upset stomach, skin rash, hallucinations, and allergic reactions.^[1] Excessive doses may cause seizures, irregular heartbeat, and death.^[3] Chewing or sucking on the capsule can lead to laryngospasm, bronchospasm, and circulatory collapse.^[1] It is unclear if use in pregnancy or breastfeeding is safe.^[5] It works by numbing stretch receptors in the lungs and suppressing the cough reflex in the brain.^[1]