(From left to right) Principal Investigator Associate Professor Gautam Sethi and NUS PhD candidate Ms Zhang Jingwen from the Department of Pharmacology at the NUS Yong Loo Lin School of Medicine led a research which found that a bioactive compound from the neem plant could significantly suppress development of prostate cancer.
Credit: National University of Singapore
Image may be NSFW. Clik here to view.
Image may be NSFW. Clik here to view.
Image may be NSFW. Clik here to view.Image may be NSFW. Clik here to view.
Date:September 29, 2016Source:National University of SingaporeSummary:Oral administration of nimbolide, over 12 weeks shows reduction of prostate tumor size by up to 70 per cent and decrease in tumor metastasis by up to 50 per cent, report investigators.
Oral administration of nimbolide, over 12 weeks shows reduction of prostate tumor size by up to 70 per cent and decrease in tumor metastasis by up to 50 per cent
A team of international researchers led by Associate Professor Gautam Sethi from the Department of Pharmacology at the Yong Loo Lin School of Medicine at the National University of Singapore (NUS) has found that nimbolide, a bioactive terpenoid compound derived from Azadirachta indica or more commonly known as the neem plant, could reduce the size of prostate tumor by up to 70 per cent and suppress its spread or metastasis by half.
Prostate cancer is one of the most commonly diagnosed cancers worldwide. However, currently available therapies for metastatic prostate cancer are only marginally effective. Hence, there is a need for more novel treatment alternatives and options.
“Although the diverse anti-cancer effects of nimbolide have been reported in different cancer types, its potential effects on prostate cancer initiation and progression have not been demonstrated in scientific studies. In this research, we have demonstrated that nimbolide can inhibit tumor cell viability — a cellular process that directly affects the ability of a cell to proliferate, grow, divide, or repair damaged cell components — and induce programmed cell death in prostate cancer cells,” said Assoc Prof Sethi.
Image may be NSFW. Clik here to view.
Nimbolide: promising effects on prostate cancer
Cell invasion and migration are key steps during tumor metastasis. The NUS-led study revealed that nimbolide can significantly suppress cell invasion and migration of prostate cancer cells, suggesting its ability to reduce tumor metastasis.
The researchers observed that upon the 12 weeks of administering nimbolide, the size of prostate cancer tumor was reduced by as much as 70 per cent and its metastasis decreased by about 50 per cent, without exhibiting any significant adverse effects.
“This is possible because a direct target of nimbolide in prostate cancer is glutathione reductase, an enzyme which is responsible for maintaining the antioxidant system that regulates the STAT3 gene in the body. The activation of the STAT3 gene has been reported to contribute to prostate tumor growth and metastasis,” explained Assoc Prof Sethi. “We have found that nimbolide can substantially inhibit STAT3 activation and thereby abrogating the growth and metastasis of prostate tumor,” he added.
The findings of the study were published in the April 2016 issue of the scientific journal Antioxidants & Redox Signaling. This work was carried out in collaboration with Professor Goh Boon Cher of Cancer Science Institute of Singapore at NUS, Professor Hui Kam Man of National Cancer Centre Singapore and Professor Ahn Kwang Seok of Kyung Hee University.
Neem — The medicinal plant
The neem plant belongs to the mahogany tree family that is originally native to India and the Indian sub-continent. It has been part of traditional Asian medicine for centuries and is typically used in Indian Ayurvedic medicine. Today, neem leaves and bark have been incorporated into many personal care products such as soaps, toothpaste, skincare and even dietary supplements.
Image may be NSFW. Clik here to view.
Future Research
The team is looking to embark on a genome-wide screening or to perform a large-scale study of proteins to analyse the side-effects and determine other potential molecular targets of nimbolide. They are also keen to investigate the efficacy of combinatory regimen of nimbolide and approved drugs such as docetaxel and enzalutamide for future prostate cancer therapy.
Journal Reference:
Jingwen Zhang, Kwang Seok Ahn, Chulwon Kim, Muthu K. Shanmugam, Kodappully Sivaraman Siveen, Frank Arfuso, Ramar Perumal Samym, Amudha Deivasigamanim, Lina Hsiu Kim Lim, Lingzhi Wang, Boon Cher Goh, Alan Prem Kumar, Kam Man Hui, Gautam Sethi. Nimbolide-Induced Oxidative Stress Abrogates STAT3 Signaling Cascade and Inhibits Tumor Growth in Transgenic Adenocarcinoma of Mouse Prostate Model. Antioxidants & Redox Signaling, 2016; 24 (11): 575 DOI:10.1089/ars.2015.6418
GENENTECH, INC. [US/US]; 1 DNA Way South San Francisco, California 94080-4990 (US). CONSTELLATION PHARMACEUTICALS, INC. [US/US]; 215 First Street Suite 200 Cambridge, Massachusetts 02142 (US)
Chromatin is a complex combination of DNA and protein that makes up chromosomes. It is found inside the nuclei of eukaryotic cells and is divided between heterochromatin (condensed) and euchromatin (extended) forms. The major components of chromatin are DNA and proteins. Histones are the chief protein components of chromatin, acting as spools around which DNA winds. The functions of chromatin are to package DNA into a smaller volume to fit in the cell, to strengthen the DNA to allow mitosis and meiosis, and to serve as a mechanism to control expression and DNA replication. The chromatin structure is controlled by a series of post-translational modifications to histone proteins, notably histones H3 and H4, and most commonly within the “histone tails” which extend beyond the core nucleosome structure. Histone tails tend to be free for protein-protein interaction and are also the portion of the histone most prone to post-translational modification. These modifications include acetylation, methylation, phosphorylation, ubiquitinylation, and SUMOylation. These epigenetic marks are written and erased by specific enzymes that place the tags on specific residues within the histone tail, thereby forming an epigenetic code, which is then interpreted by the cell to allow gene specific regulation of chromatin structure and thereby transcription.
Of all classes of proteins, histones are amongst the most susceptible to post-translational modification. Histone modifications are dynamic, as they can be added or removed in response to specific stimuli, and these modifications direct both structural changes to chromatin and alterations in gene transcription. Distinct classes of enzymes, namely histone acetyltransferases (HATs) and histone deacetylases (HDACs), acetylate or de-acetylate specific histone lysine residues (Struhl K., Genes Dev., 1989, 12, 5, 599-606).
Bromodomains, which are approximately 1 10 amino acids long, are found in a large number of chromatin-associated proteins and have been identified in approximately 70 human proteins, often adjacent to other protein motifs (Jeanmougin F., et al., Trends Biochem. Sc , 1997, 22, 5, 151-153; and Tamkun J.W., et al., Cell, 1992, 7, 3, 561-572).
Interactions between bromodomains and modified histones may be an important mechanism underlying chromatin structural changes and gene regulation. Bromodomain-containing proteins have been implicated in disease processes including cancer, inflammation and viral replication. See, e.g., Prinjha et al,, Trends Pharm. Sci., 33(3):146-153 (2012) and Muller et al , Expert Rev. , 13 (29): 1 -20 (September 201 1 ).
Cell-type specificity and proper tissue functionality requires the tight control of distinct transcriptional programs that are intimately influenced by their environment.
Alterations to this transcriptional homeostasis are directly associated with numerous disease states, most notably cancer, immuno-inflammation, neurological disorders, and metabolic diseases. Bromodomains reside within key chromatin modifying complexes that serve to control distinctive disease-associated transcriptional pathways. This is highlighted by the observation that mutations in bromodomain-containing proteins are linked to cancer, as well as immune and neurologic dysfunction. Hence, the selective inhibition of bromodomains across a specific family, such as the selective inhibition of a bromodomain of CBP/EP300, creates varied opportunities as novel therapeutic agents in human dysfunction.
There is a need for treatments for cancer, immunological disorders, and other
To a solution of (^)-tetrahydrofuran-3-ol (25 g, 253.7 mmol) in DCM (250 mL) at 0 °C was added triethylamine (86 g, 851.2 mmol) and mesyl chloride (39 g, 340.48 mmol) dropwise. The mixture was stirred at room temperature for 12 h. The reaction was quenched with water (100 mL) and extracted with DCM (100 mL x 2). The combined organic layers were dried over anhydrous Na2S04, filtered and concentrated in vacuo to give the title compound (47 g, 99%) as a brown oil. Ή NMR (400 MHz, CDC13) δ 5.35 – 5.27 (m, 1H), 4.05 – 3.83 (m, 4H), 3.04 (s, 3 H), 2.28 – 2.20 (m, 2 H).
To a solution of tert-butyl 3-bromo-6,7-dihydro-lH-pyrazolo[4,3-c]pyridine-5(4H)-carboxylate (Intermediate A, 24.8 g, 82 mmol) in DMF (200 mL) was added Cs2C03 (79 g, 246 mmol) and (/?)-tetrahydrofuran-3-yl methanesulfonate (17.4 g, 98 mmol). The mixture was heated to 80 °C for 12 h. After cooling the reaction to room temperature, the mixture was concentrated in vacuo. The crude residue was purified by silica gel chromatography
(petroleum ether / EtOAc = from 10 : 1 to 3 : 1) to give the title compound (Intermediate F, 50 g, 71 %) as a yellow oil. Ή NMR (400 MHz, DMSO-i ) δ 4.97 – 4.78 (m, 1H), 4.13 (s, 2H), 3.98 – 3.86 (m, 2H), 3.81 – 3.67 (m, 2H), 3.56 (t, J= 5.6 Hz, 2H), 2.68 (t, J= 5.6 Hz, 2H), 2.33 – 2.08 (m, 2H), 1.38 (s, 9H).
To a solution of (S)-tert-buty\ 3-bromo- 1 -(tetrahydrofuran-3-yl)-6,7-dihydro-lH-pyrazolo [4,3 -c]pyridine-5(4H)-carboxy late (29 g, 78 mmol) in DCM (300 mL) was added trifluroacetic acid (70 mL) dropwise. The mixture was stirred at room temperature for 2 h. The solvent was concentrated in vacuo and the crude residue was re -dissolved in DMF (100 mL). The mixture was cooled to 0 °C before triethylamine (30 g, 156 mmol) and acetic anhydride (8.7 g, 86 mmol) were added dropwise. The mixture was stirred at room temperature for an additional 2 h. The reaction was quenched with water (200 mL) at 0 °C and extracted with EtOAc (150 mL x 3). The combined organic layers were dried over anhydrous Na2S0 , filtered and concentrated in vacuo. The crude residue was purified by silica gel chromatography (DCM / MeOH = 30 : 1) to give the title compound (Intermediate G, 21.3 g, 87%) as a white solid. lH NMR (400 MHz, CDC13) δ 4.78 – 4.67 (m, 1H), 4.45 -4.29 (m, 2H), 4.15 – 4.06 (m, 2H), 3.96 – 3.92 (m, 2H), 3.88 – 3.70 (m, 2H), 2.71 – 2.67 (m, 2H), 2.38 – 2.34 (m, 2H), 2.16 (s, 3H).
1-[3-[2-fluoro-4-(1-methylpyrazol-4- yl)anilino]-1-[(3S)-tetrahydrofuran-3- yl]-6,7-dihydro-4H-pyrazolo[4,3- c]pyridin-5-yl]ethanone Image may be NSFW. Clik here to view.
To a solution of (R)-tetrahydrofuran-3-ol (25 g, 253.7 mmol) in DCM (250 mL) at 0° C. was added triethylamine (86 g, 851.2 mmol) and mesyl chloride (39 g, 340.48 mmol) dropwise. The mixture was stirred at room temperature for 12 h. The reaction was quenched with water (100 mL) and extracted with DCM (100 mL×2). The combined organic layers were dried over anhydrous Na2SO4, filtered and concentrated in vacuo to give the title compound (47 g, 99%) as a brown oil. 1H NMR (400 MHz, CDCl3) δ 5.35-5.27 (m, 1H), 4.05-3.83 (m, 4H), 3.04 (s, 3H), 2.28-2.20 (m, 2H).
To a solution of tert-butyl 3-bromo-6,7-dihydro-1H-pyrazolo[4,3-c]pyridine-5(4H)-carboxylate (Intermediate A, 24.8 g, 82 mmol) in DMF (200 mL) was added Cs2CO3 (79 g, 246 mmol) and (R)-tetrahydrofuran-3-yl methanesulfonate (17.4 g, 98 mmol). The mixture was heated to 80° C. for 12 h. After cooling the reaction to room temperature, the mixture was concentrated in vacuo. The crude residue was purified by silica gel chromatography (petroleum ether/EtOAc=from 10:1 to 3:1) to give the title compound (Intermediate F, 50 g, 71%) as a yellow oil. 1H NMR (400 MHz, DMSO-d6) δ 4.97-4.78 (m, 1H), 4.13 (s, 2H), 3.98-3.86 (m, 2H), 3.81-3.67 (m, 2H), 3.56 (t, J=5.6 Hz, 2H), 2.68 (t, J=5.6 Hz, 2H), 2.33-2.08 (m, 2H), 1.38 (s, 9H).
To a solution of (S)-tert-butyl 3-bromo-1-(tetrahydrofuran-3-yl)-6,7-dihydro-1H-pyrazolo[4,3-c]pyridine-5(4H)-carboxylate (29 g, 78 mmol) in DCM (300 mL) was added trifluroacetic acid (70 mL) dropwise. The mixture was stirred at room temperature for 2 h. The solvent was concentrated in vacuo and the crude residue was re-dissolved in DMF (100 mL). The mixture was cooled to 0° C. before triethylamine (30 g, 156 mmol) and acetic anhydride (8.7 g, 86 mmol) were added dropwise. The mixture was stirred at room temperature for an additional 2 h. The reaction was quenched with water (200 mL) at 0° C. and extracted with EtOAc (150 mL×3). The combined organic layers were dried over anhydrous Na2SO4, filtered and concentrated in vacuo. The crude residue was purified by silica gel chromatography (DCM/MeOH=30:1) to give the title compound (Intermediate G, 21.3 g, 87%) as a white solid. 1H NMR (400 MHz, CDCl3) δ 4.78-4.67 (m, 1H), 4.45-4.29 (m, 2H), 4.15-4.06 (m, 2H), 3.96-3.92 (m, 2H), 3.88-3.70 (m, 2H), 2.71-2.67 (m, 2H), 2.38-2.34 (m, 2H), 2.16 (s, 3H).
OTHER ISOMER
Image may be NSFW. Clik here to view.
Example 299
1-[3-[2-fluoro-4-(1-methylpyrazol-4- yl)anilino]-1-[(3R)-tetrahydrofuran-3- yl]-6,7-dihydro-4H-pyrazolo[4,3- c]pyridin-5-yl]ethanone Image may be NSFW. Clik here to view.
The single bromodomain of the closely related transcriptional regulators CBP/EP300 is a target of much recent interest in cancer and immune system regulation. A co-crystal structure of a ligand-efficient screening hit and the CBP bromodomain guided initial design targeting the LPF shelf, ZA loop, and acetylated lysine binding regions. Structure–activity relationship studies allowed us to identify a more potent analogue. Optimization of permeability and microsomal stability and subsequent improvement of mouse hepatocyte stability afforded 59 (GNE-272, TR-FRET IC50 = 0.02 μM, BRET IC50 = 0.41 μM, BRD4(1) IC50 = 13 μM) that retained the best balance of cell potency, selectivity, and in vivo PK. Compound 59 showed a marked antiproliferative effect in hematologic cancer cell lines and modulates MYC expression in vivo that corresponds with antitumor activity in an AML tumor model.
Discovery of a Potent and Selective in Vivo Probe (GNE-272) for the Bromodomains of CBP/EP300
In a similar procedure to59, the title compound was prepared from (S)-tetrahydrofuran-3-yl
methanesulfonate and purified by Prep-TLC (DCM / MeOH = 15 : 1) to give the title
compound as a light yellow solid.
The crude residue was purified by silica gel chromatography (DCM / MeOH = 100:1) to give (S)-1-(3-((2-fluoro-4-(1- methyl-1H-pyrazol-4-yl)phenyl)amino)-1-(tetrahydrofuran-3-yl)-6,7-dihydro-1Hpyrazolo[4,3-c]pyridin-5(4H)-yl)ethanone as a light yellow solid.
Clenbuterol, marketed as Dilaterol, Spiropent, Ventipulmin,[1] is a sympathomimeticamine used by sufferers of breathing disorders as a decongestant and bronchodilator. People with chronic breathing disorders such as asthma use this as a bronchodilator to make breathing easier. It is most commonly available as the hydrochloridesalt, clenbuterol hydrochloride.[2]
Clenbuterol is also prescribed for treatment of horses, but equine use is usually the liquid form.
Human use
Clenbuterol is approved for use in some countries, free or via prescription, as a bronchodilator for asthma patients.[3]
Image may be NSFW. Clik here to view.
Legal status
Clenbuterol is not an ingredient of any therapeutic drug approved by the US Food and Drug Administration[3] and is now banned forIOC-tested athletes.[4] In the US, administration of clenbuterol to any animal that could be used as food for human consumption is banned by the FDA.[5][6]
Clenbuterol is a therapeutic drug for asthma and COPD, approved for human use in some countries in Europe (Bulgaria and Russia) and Asia (China).
Image may be NSFW. Clik here to view.
Weight-loss drug
Although often used by bodybuilders during their “cutting” cycles,[citation needed] the drug has been more recently known to the mainstream, particularly through publicized stories of use by celebrities such as Victoria Beckham,[4]Britney Spears, and Lindsay Lohan, [7] for its off-label use as a weight-loss drug similar to usage of other sympathomimetic amines such as ephedrine, despite the lack of sufficient clinical testing either supporting or negating such use.
Image may be NSFW. Clik here to view.
Image may be NSFW. Clik here to view.
Image may be NSFW. Clik here to view.
By bromination of 4-amino-3,5-dichloroacetophenone (I) with Br2 in CHCl3 to give 4-amino-3,5-dichloro-alpha-bromoacetophenone (II), m.p. 140-5 C, which is condensed with tert-butylamine (III) in CHCl3 to 4-amino-3,5-dichloro-alpha-tertbutylaminoacetophenone hydrochloride (IV), m.p. 252-7 C; this product is finally reduced with NaBH4 in methanol.
Synthesen von neuen Amino-Halogen-substituierten Phenyl-aminothanolen. Arzneim-Forsch Drug Res 1972, 22, 5, 861-869
CLIP
Synthesis and Characterization of Bromoclenbuterol
Ravi Kumar Kannasani*, Srinivasa Reddy Battula, Suresh Babu Sannithi, Sreenu Mula and Venkata Babu VV
R&D Division, RA Chem Pharma Limited, API, Hyderabad, Telangana, India
*Corresponding Author:
Ravi Kumar Kannasani
R&D Division, RA Chem Pharma Limited
API, Prasanth Nagar, Hyderabad, Telangana, India Tel: +919000443184 E-mail: kannasani.ravi@rachempharma.com
Citation: Kannasani RK, Battula SR, Sannithi SB, Mula S, Babu VVV (2016) Synthesis and Characterization of Bromoclenbuterol. Med Chem (Los Angeles) 6:546-549. doi:10.4172/2161-0444.1000397
Clenbuterol, it is most commonly available as the hydrochloride salt, clenbuterol hydrochloride. Clenbuterol, marketed as Dilaterol, Spiropent, Ventipulmin, and also generically as clenbuterol, is a sympathomimetic amine used for breathing disorders as a decongestant and bronchodilator. People with chronic breathing disorders such as asthma use this as a bronchodilator to make breathing easier. Clenbuterol is a β2 agonist with some structural and pharmacological similarities to epinephrine and salbutamol, but its effects are more potent and longerlasting as a stimulant and thermogenic drug. It causes an increase in aerobic capacity, central nervous system stimulation, blood pressure, and oxygen transportation. It increases the rate at which body fat is metabolized while increasing the body’s BMR. It is commonly used for smooth muscle-relaxant properties as a bronchodilator and tocolytic. Clenbuterol is also prescribed for treatment of horses, but equine use is usually the liquid form
Clenbuterol Hydrochloride was first synthesized at Thomae; a Boehringer Ingelheim research facility in Biberach, Germany, in 1967. The synthesis of Clenbuterol Hydrochloride was patented in the United States in 1970. After comprehensive clinical trials, Clenbuterol Hydrochloride was approved for the treatment of reversible airway obstruction in Germany in 1976 and later as a veterinary pharmaceutical for the treatment of bronchiolytic disorders in Germany in 1980. Boehringer Ingelheim markets Clenbuterol Hydrochloride as Spirospent for Human Pharmaceuticals and as Ventipulmin for Veterinary Pharmaceuticals. Clenbuterol Hydrochloride is not approved by the Federal Drug Administration for human use in the United States.
Image may be NSFW. Clik here to view.
As per the available literature [4–7], clenbuterol hydrochloride was synthesized from 4-amino acetophenone (Scheme 1). Initially 4-amino acetophenone (1) was reacted with chlorine to afford 4-amino-3,5- dichloro acetopheneone (2) which was further reacted bromine to give 1-(4-amino-3,5-dichlorophenyl)-2-bromoethanone (3). The obtained bromo compound was reacted tertiary butyl amine to afford 2-(tertbutylamino)- 1-(4-amino-3,5-dichlorophenyl)ethanone (4), which was further reduced with sodium borohydride to give clenbuterol base (5) and converted in to hydrochloride salt by using alcoholic HCl to get clenbuterol hydrochloride (6).
In the synthesis of clenbuterol hydrochloride, first step was a double chlorination of 4-aminoacetophenone (1) through an electrophillic aromatic substitution reaction to yield 4-amino-3,5- dichloroacetophenone (2). Due to the ortho/para directing, amino group and the meta directing, electron withdrawing, acetyl group, chlorination of 4-aminoacetophenone occurs primarily at the 3 and 5 positions over the 2 and 6 positions. Therefore, under chlorination would produce only the mono chlorinated impurity, 4-amino-3- chloroacetophenone. Under these conditions, over chlorination does not result in the addition of chlorine to the 2 and 6 positions because the amino and acetyl groups do not direct that addition. Even though chlorides are ortho/para directing and direct to the 2 and 6 position, chlorides are also deactivating. After close observation on this chlorination reaction, it was noted that the formed mono chlorinated impurity (Scheme 2) (4-amino-3-chloro acetophenone) caused the formation of process related impurity (bromoclenbuterol) in clenbuerol synthesis.
Generalised anxiety disorder; Major depressive disorder
Most Recent Events
10 May 2016Discontinued – Phase-I for Generalised anxiety disorder in USA, Japan (PO)
10 May 2016Discontinued – Phase-I for Major depressive disorder in USA, Japan (PO)
30 Jul 2015Tedatioxetine is still in phase I trials for Major depressive disorders and Generalised anxiety disorder in USA and Japan
Tedatioxetine (Lu AA24530) is an antidepressant that was discovered by scientists at Lundbeck; in 2007 Lundbeck and Takedaentered into a partnership that included tedatioxetine but was focused on another, more advanced Lundbeck drug candidate,vortioxetine.[1]
Tedatioxetine chemical name 4- (2- (4-methylphenyl group)) phenylpiperidine by Lundbeck developed for the treatment of severe depression, it is a monoamine reuptake inhibitor, a monoamine reuptake transporter inhibitors, 5-HT3 antagonists and 5-HT2c receptor antagonist. For the treatment of major depressive disorder and generalized anxiety, II clinical study in. Tedatioxetine has the following structure:
Image may be NSFW. Clik here to view.
According to the literature, the current synthesis routes are the following:
WO 2003/029232 discloses Tedatioxetine first preparation method, as shown in the following Scheme,
Image may be NSFW. Clik here to view.
The method of low yield, the product is not easy purification by column chromatography requires; more important is the preparation of the compound N-Boc- piperidin-4-ol of the need to use butyl lithium, and reaction was carried out at lower temperatures, not conducive to industrial production.
WO 2009109541 provides a, as shown in the above-described method for improved routes following synthetic route,
Image may be NSFW. Clik here to view.
Bn- replaced with Boc-, dehydroxylation switch to TFA and Et 3 of SiH, yield improved despite increased. But there are many shortcomings.Deficiencies mainly reflected in the following aspects: the compound used in the expensive starting 2-bromo benzene iodine source and a catalyst of palladium and a bidentate phosphine ligand 3, an increase of production cost; preparation of compound needed 4:00 butyl lithium reagent to the more dangerous, the need at a low temperature reaction. This will bring in the production of a big security risk, is not conducive to the operation; when dehydroxylation
Preparation of 2- (4-methyl-phenyl mercapto) phenylpiperidine hydrobromide, to use a lot of trifluoroacetate (15eq), post-processing is too much trouble and the environment have a greater pollution.
Given 4- [2- (4-methylphenyl) phenyl] piperidine and salts thereof possess excellent pharmacological properties, and deficiencies of the prior processes, is necessary to develop a suitable industrial production, easy to operate and environmentally friendly preparation process.
Image may be NSFW. Clik here to view.
Image may be NSFW. Clik here to view.
Image may be NSFW. Clik here to view.
Image may be NSFW. Clik here to view.
Image may be NSFW. Clik here to view.
2- (4-methyl-phenylthio) benzaldehyde prepared as in Example 1
Image may be NSFW. Clik here to view.
Direction of Na 2 CO. 3 stirred mixture (11g, 105mmol) and 30mlDMF added 4-methyl-thiophenol (12.4g, 100mmol), stirred for 20 minutes. To the mixture was slowly added 2-bromobenzaldehyde (18.4g, 100mmol); a pending completion of the addition, under nitrogen, was heated to 100 deg.] C for 6 hours. After completion of the reaction, the reaction solution was cooled to room temperature, 100ml of water was added and stirred for 30 minutes. Filtered, washed with water (30ml) and dried in vacuo to give the filter cake was washed with 20.5g pale green solid; After n-hexane to give 18.5g pale yellow solid was recrystallized from 2- (4-phenylthio) benzaldehyde (mp: 52- 54 ℃), 81% yield. 2- (4-methyl-phenylthio) benzaldehyde Example 2 Preparation of
Image may be NSFW. Clik here to view.
To the K 2 CO. 3 stirred mixture (15g, 110mmol) and 30mlDMA added 4-methyl-thiophenol (12.6g, 102mmol), stirred for 20 minutes. To the mixture was slowly added 2-chlorobenzaldehyde (14g, 100mmol); a pending completion of the addition, under nitrogen, the reaction was heated to 100 deg.] C for 7 hours. After completion of the reaction, the reaction solution was cooled to room temperature, 100ml of water was added and stirred for 30 minutes. Filtered, washed with water (30ml) and dried in vacuo to give the filter cake was washed with 19.7g pale green solid; After n-hexane to give 17g as a pale yellow solid was recrystallized from 2- (4-phenylthio) benzaldehyde (melting point: 51-53 ℃), a yield of 77.5%
2- (4-methyl-phenylthio) benzaldehyde Example 3 Preparation of
Image may be NSFW. Clik here to view.
Ask NaOH (4.2g, 105mmol) and stirred 50ml 1,4-dioxane was added 4-methyl-thiophenol (12.4g, 100mmol), stirred for 30 minutes. To the mixture was slowly added 2-iodo-benzaldehyde (23.1g, 100mmol); a pending completion of the addition, under nitrogen, was heated under reflux for 5 hours.After completion of the reaction, the reaction solution was cooled to room temperature, 50ml of water was added, extraction separated; the organic phase was washed with 50ml of ethyl acetate, and the combined organic phases were washed with 20% aqueous ammonium chloride solution and saturated brine, dried over anhydrous magnesium sulfate, filtration and concentration gave 21g viscous liquid, and cooled to solidify; after n-hexane to give 18.1g pale yellow solid was recrystallized from 2- (4-phenylthio) benzaldehyde (m.p.: 53-54 ℃), close rate of 79%.
Example 4 Preparation of 3- [2 (4-methyl) phenyl] pentanedioic acid
Image may be NSFW. Clik here to view.
1) Preparation of ethyl-2-cyano-3- (2- (4-methyl) phenyl) acrylate
Image may be NSFW. Clik here to view.
2- (4-methylphenyl thio) benzaldehyde (4g, 17.5mmol), ethyl cyanoacetate (2.4g 21mmol) and toluene (30ml) was added a mixture of glacial acetic acid (5ml) and piperidine (0.3 ml of) stirred for 10 minutes; heated to reflux, and isolating the resulting water trap. Completion of the reaction, cooled to room temperature; the reaction was washed with 30ml water and 30ml saturated sodium bicarbonate solution, dried over anhydrous magnesium sulfate; filtered, and concentrated to give 5.0g yellow liquid (solidifies on cooling), yield 86%. It was used directly in the next reaction without purification.
2) Preparation of Diethyl 2,4-diethyl-3- (2- (4-methyl) phenyl) glutarate
Image may be NSFW. Clik here to view.
Sodium methoxide (1.9g, 35mmol) and dry THF (30ml) was stirred and cooled to mix 0-5 ℃, was added dropwise diethyl malonate (4.6g, 35mmol), stirred for 15 minutes at room temperature dropwise Bi; dropwise obtained above in step 2-cyano-3- (2- (4-methyl) phenyl) acrylate (5g, 15.4mmol) and dry tetrahydrofuran (40ml) solution; BI dropwise, at room temperature stirred for 13 hours. Completion of the reaction, the reaction mixture was added 150ml20% aqueous ammonium chloride solution, followed by extraction separated; the aqueous phase was extracted with ethyl acetate, the combined organic phase was dried over anhydrous magnesium sulfate; filtered, and concentrated to give 5.4 g of a viscous liquid, yield 78%. It was used directly in the next reaction without purification.
Was added 6N hydrochloric acid (70ml), was heated at reflux for 3 days the material obtained in the above step (5.4 g of); completion of the reaction, slowly cooled to room temperature, added 50ml of ethyl acetate, stirred for 30 minutes to precipitate a solid from the solution, filtered and washed with 20ml washed with ethyl acetate, and dried in vacuo at 50 ℃ 10 hours to give 2.7g of white solid 3- [2 (4-methylphenyl) phenyl] glutaric acid (melting point: 191-195 ℃), in 58% yield.
Example 5 Preparation of 3- [2- (4-phenylthio) phenyl] pentanedioic acid
Image may be NSFW. Clik here to view.
To ethyl acetoacetate (13g, 100mmol) and piperidine (1.7g, 10mmol) was added a mixture of 2- (4-methyl-phenylthio) benzaldehyde (11.5g, 50mmol), room temperature for 1 day to give a yellow viscous semi-solid, 2.7g of sodium methoxide was added. after stirring for 1 hour cure, stand for 2 days.To the above mixture was added ethanol (180ml) and 40% aqueous sodium hydroxide (140ml) was stirred and heated to reflux for 4-5 hours the reaction. Completion of the reaction the heating was stopped, and after cooling to room temperature, the solvent was distilled off under reduced pressure; the residue after distillation under cooling in an ice water bath, and treated dropwise with concentrated hydrochloric acid (150ml) adjusted to pH 1-2. 300ml ethyl acetate was added, the aqueous phase was extracted with 300ml of ethyl acetate, and the combined organic phases were washed with 300ml water; the organic phase was dried over anhydrous sodium sulfate, filtered, and concentrated to 500ml of the solvent. The residue was cooled to room temperature, stirred for 2 hours. The title compound was isolated by filtration through with ethyl acetate (20ml) and was washed and dried at 50 deg.] C in vacuo overnight to give 21.5g of white solid 3- [2 (4-methylphenyl) phenyl] glutaric acid (melting point: 194-196 ℃) yield 65%.
Example 6 Preparation of 4- [2- (4-methylphenyl) phenyl] piperidine-2,6-dione
Image may be NSFW. Clik here to view.
Mixing the compound 3- [2 (4-methyl) phenyl] glutaric acid (10g, 30mmol) and urea (5.4g, 90mmol) prepared in Step stirred and heated to 146 deg.] C for 4 hours ; after completion of the reaction was monitored by TLC, cooled to 80 deg.] C, was slowly added 70ml of water and 70ml of ethanol was stirred for 30 minutes; cooled to room temperature and stirred for 1 hour. The title compound was filtered absolute ethanol (170ml) and recrystallized from 50 deg.] C overnight and dried in vacuo to give 8.0g white solid 4- [2- (4-methylphenyl) phenyl] piperidine-2,6-di -one (mp: 164-166 ℃), yield 86%
7 Preparation of 4- [2- (4-methylphenyl) phenyl] piperidine-2,6-dione Example
Image may be NSFW. Clik here to view.
In four of 250ml equipped with a condenser reaction flask was added 3- [2 (4-methyl) phenyl] glutaric acid (10g, 30mmol) and urea (14.4g, 240mmol) and the mixture was stirred and heated to 146 deg.] C for 4 hours; TLC monitoring completion of the reaction, cooled to 100 deg.] C, was slowly added 70ml of water and 70ml of ethanol was stirred for 30 minutes; cooled to room temperature and stirred for 1 hour. The title compound was filtered absolute ethanol (170ml) and recrystallized from 50 deg.] C overnight and dried in vacuo to give 7.8g white solid 4- [2- (4-methylphenyl) phenyl] piperidine-2,6-di -one (mp: 165-166 ℃), yield 84%.
8 Preparation of 4- [2- (4-methylphenyl) phenyl] piperidine-2,6-dione Example
Image may be NSFW. Clik here to view.
In four of 250ml equipped with a condenser reaction flask was added 3- [2 (4-methyl) phenyl] pentanedioic acid (5g, 15mmol) and urea (1.8g, 30mmol) and the mixture was stirred and heated to 143 deg.] C for 4 hours; cool to 100 deg.] C, was slowly added 35ml of water and 35ml of ethanol was stirred for 30 minutes; cooled to room temperature and stirred for 1 hour. The title compound was filtered absolute ethanol (70ml) and recrystallized from 50 deg.] C overnight and dried in vacuo to give an off-white solid 2.9g of 4- [2- (4-methylphenyl) phenyl] piperidine-2,6-dione (Melting point: 163-166 ℃), a yield of 63%.
9 Preparation of 4- [2- (4-methylphenyl) phenyl] piperidine-2,6-dione Example
Image may be NSFW. Clik here to view.
The compound prepared in the step of 3- [2 (4-methyl) phenyl] glutaric acid (10g, 30mmol) and urea (3.6g, 60mmol) were mixed and stirred and heated to 146 deg.] C for 4 hours ; after completion of the reaction was monitored by TLC, cooled to 80 deg.] C, was slowly added 70ml of water and 70ml of ethanol was stirred for 30 minutes; cooled to room temperature and stirred for 1 hour. The title compound was filtered, absolute ethanol (45 ml of) and recrystallized from 50 deg.] C overnight and dried in vacuo to give 8.0g white solid 4- [2- (4-methylphenyl) phenyl] piperidine-2,6 dione (melting point: 164-166 ℃), yield 86%.
10 Preparation of 4- [2- (4-methylphenyl) phenyl] piperidine-2,6-dione Example
Image may be NSFW. Clik here to view.
A step of preparing the compound 3- [2 (4-methylphenyl) phenyl] glutaric acid (19.8g, 60mmol) and urea (21.6g, 360mmol) were mixed and stirred and heated to 144 deg.] C for 4 hours; after completion of the reaction was monitored by TLC, cooled to 100 deg.] C, slowly added water 140ml 140ml ethanol and stirred for 30 min; cooled to room temperature and stirred for 1 hour. The title compound was filtered, absolute ethanol (350ml) and recrystallized from 50 deg.] C overnight and dried in vacuo to give a white solid 16.5g of 4- [2- (4-methylphenyl) phenyl] piperidine-2,6 dione (melting point: 164-166 ℃), yield 88%.
Example 11 Preparation of 4- [2- (4-methylphenyl) phenyl] piperidine
Tetrahydro lithium aluminum (5.1g, 39mmol) with 140ml of tetrahydrofuran were mixed and stirred ice bath cooled to 8 ℃, under nitrogen, was added dropwise 4- (2-mercapto-methylphenyl) piperidine-2,6-phenyl one (7g) in tetrahydrofuran (140ml) solution, so that the temperature does not exceed 20 ℃; dropping was completed, the reaction at room temperature for 5 hours. The reaction solution was cooled in an ice-water bath, was slowly added dropwise 30ml of water, stirred for 20 minutes. The reaction mixture was added sodium sulfate (20g), stirred for 30 minutes. Filtered and the filtrate was concentrated to give a colorless liquid (4.5g), cooled to solidify to a white solid of 4- [2- (4-methylphenyl) phenyl] piperidine.
Example 12 Preparation of 4- [2- (4-methylphenyl) phenyl] piperidine
The reaction flask was added 100ml four 1mol / l borane tetrahydrofuran solution (40ml, 40mmol), cooled to ice bath 5 ℃; under nitrogen was added dropwise 4- (2-mercapto-methylphenyl) piperidine-2-phenyl , 6-dione (3.1g) in tetrahydrofuran (40ml) solution, so that the temperature does not exceed 10 ℃; dropping was completed, the reaction at room temperature for 20 hours. The reaction solution was cooled to 0 deg.] C, and slowly added dropwise 1mol / l HCl (30mL), dropwise finished warming at reflux for 5 hours; of THF was removed and concentrated, 30ml of ethyl acetate and washed with an aqueous solution, a saturated aqueous sodium bicarbonate was added to adjust the pH> 10 , followed by addition of 50ml of ethyl acetate, the organic phase was dried, filtered and concentrated to give 1.8g of a colorless liquid, and cooled to solidify to a white solid of 4- [2- (4-methyl) phenyl] piperidine.
Example 13 Preparation of 4- [2- (4-methylphenyl) phenyl] piperidine
The a 2 mol / L the BH 3 .CH 3 the SCH 3 (20 mL) and diethylene glycol dimethyl ether 20ml were mixed and stirred ice bath cooled to 10 ℃, solution of 4- (2-mercapto-methyl-phenyl) phenylpiperidine pyridine 2,6-dione (3.1g) in diethylene glycol dimethyl ether (60ml) solution, so that the temperature does not exceed 20 ℃; dropping was completed, the reaction at room temperature 0.5 hours, then slowly heated to 120 deg.] C for 10 hours. The reaction solution was cooled to 0 deg.] C, and slowly added dropwise 30ml of methanol, a dropping was completed, the mixture was stirred overnight at room temperature; was added 4mol / l HCl / EA (10ml ), was heated to 100 deg.] C for 4 hours; the resulting residue was distilled under reduced pressure was dissolved in 30ml water, saturated aqueous sodium bicarbonate was added to adjust the pH> 10, followed by addition of 50ml of ethyl acetate, the organic phase was dried, filtered and concentrated to give a pale red liquid; after column chromatography (hexane – acetic acid – ethanol 10 : 1.5: 0.5) to give a white solid (0.9g) 4- [2- (4- methylphenylsulfanyl) phenyl] piperidine after purification.
14 Preparation of 4- [2- (4-methylphenyl) phenyl] piperidine hydrochloride Example
The step resulting 4- [2- (4-methylphenyl) phenyl] piperidine (4g, 14mmol) was added to absolute ethanol (30ml) and heated to 50 deg.] C to dissolve; 4mol slowly added dropwise / l hydrogen chloride – ethyl acetate solution (4ml), 40 minutes with the reaction temperature; cooled to 5-10 ℃ stirred for 2 hours, filtered through a cake when the ethanol (5ml) and washed with 44 ℃ overnight and dried in vacuo to give 3.2 g of white solid 4- [2- (4-methylphenyl) phenyl] piperidine hydrochloride (melting point: 222-225 ℃), 75% yield.
15 Preparation of 4- [2- (4-methylphenyl) phenyl] piperidine hydrochloride Example
4- [2- (4-methylphenyl) phenyl] piperidine (4g, 14mmol) was added to acetone (20ml) and heated to 50 deg.] C to dissolve; 37% was gradually added dropwise concentrated hydrochloric acid ( 1.5ml), 40 minutes with the reaction temperature; cooled with stirring to 5-10 ℃ 2 hours, filtered through a cake of acetone (5ml) and washed with 44 ℃ vacuum dried overnight to give 3.6g of white solid 4- [2- ( 4-methylphenyl) phenyl] piperidine hydrochloride (melting point: 224-227 ℃), in 80% yield.
Example 16 Preparation of 4- [2- (4-methylphenyl) phenyl] piperidine hydrochloride embodiment
Tetrahydro Lithium aluminum (19g, 500mmol) and 200ml of tetrahydrofuran were mixed and stirred at room temperature was added dropwise 4- (2-mercapto-methylphenyl) piperidine-2,6-dione phenyl (31.1g, 100mmol) and tetrahydrofuran ( 200ml) solution, the temperature does not exceed 35 ℃; dropping was completed, the reaction heated under reflux for 3 hours. The reaction solution was cooled in an ice-water bath, was slowly added dropwise 100ml of saturated aqueous sodium sulfate solution, stirred for 60 minutes. The reaction mixture was added ethyl acetate (200ml) and anhydrous magnesium sulphate (50g) was stirred for 60 minutes. Filtered and the filtrate was concentrated to give a colorless liquid. Was added to 80ml of acetone and heated to 40 ℃ dissolved, was added quickly 4mol / l hydrogen chloride – ethyl acetate solution (10ml), seeded, stirred for 20 minutes to precipitate a white solid. 40 ℃, slowly dropping the remaining hydrogen chloride – ethyl acetate solution (20ml). Drop Bi, 5-10 ℃ for 3 hours. The filtered cake in acetone (30ml) and washed with 44 ℃ when dried in vacuo overnight to give 20.8g of white solid 4- [2- (4-methylphenyl) phenyl] piperidine hydrochloride (melting point: 225-228 ℃), yield 66%.
4- [2- (4-METHYLPHENYLSULFANYD-PHENYL] PIPERIDINE WITH COMBINED SEROTONIN AND NOREPINEPHRINE REUPTAKE INHIBITION FOR THE TREATMENT OF ADHD, MELANCHOLIA, TREATMENT RESISTENT DEPRESSION OR RESIDUAL SYMPTOMS IN DEPRESSION
CRYSTALLINE FORMS OF 4- [2- (4-METHYLPHENYLSULFANYL) -PHENYL] PIPERIDINE WITH COMBINED SEROTONIN AND NOREPINEPHRINE REUPTAKE INHIBITION FOR THE TREATMENT OF NEUROPATHIC PAIN
CRYSTALLINE FORMS OF 4-[2-(4-METHYLPHENYLSULFANYL)-PHENYL] PIPERIDINE WITH COMBINED SEROTONIN AND NOREPINEPHRINE REUPTAKE INHIBITION FOR THE TREATMENT OF NEUROPATHIC PAIN
Image may be NSFW. Clik here to view.AKEBIA THERAPEUTICS PRESIDENT & CEO JOHN BUTLER PRESENTING AT THE AMERICAN KIDNEY FUND
Vadadustat, also known as AKB-6548 and PG-1016548, is a potent Hypoxia-inducible factor-proline dioxygenase inhibitor. AKB-6548 works by inhibiting hypoxia inducible factor-prolyl hydroxylase (HIP-PH), leading to stabilization and increased levels of HIFα. In turn HIFα improves production of hemoglobin and red blood cells (RBCs), while maintaining normal levels of erythropoietin (EPO) in patients. We believe this differentiated mechanism of action has the potential to be safer than that of injectable recombinant erythropoietin stimulating agents (rESAs), avoiding supra-physiological levels of EPO and saturation of EPO receptors for prolonged periods of time.
Akebia Therapeutics, under license from Procter & Gamble, and sublicensee Mitsubishi Tanabe Pharma are developing vadadustat, an orally active small-molecule hypoxia-inducible factor prolyl hydroxylase (HIF-PH) inhibitor that stabilize HIF2-α, as a once-daily formulation, for treating anemia. Also the company is investigating AKB-6899, an oral HIF-PH inhibitor, for treating cancer and ocular diseases. In March 2016, the IND application was opened. Aerpio Therapeutics, a spinoff of Akebia, is investigating AKB-4924, a HIF2-α stabilizer, which inhibits HIF prolyl hydroxylase-2, for treating inflammatory bowel disease and wound healing
Hypoxia-inducible factor (HIF) is a transcription factor that is a key regulator of responses to hypoxia. In response to hypoxic conditions, i.e., reduced oxygen levels in the cellular environment, HIF upregulates transcription of several target genes, including those encoding erythropoietin. HIF is a heteroduplex comprising an alpha and beta subunit. While the beta subunit is normally present in excess and is not dependent on oxygen tension, the HIF-alpha subunit is only detectable in cells under hypoxic conditions. In this regard, the accumulation of HIF-alpha is regulated primarily by hydroxylation at two proline residues by a family of prolyl hydroxylases known as HIF prolyl hydroxylases, wherein hydroxylation of one or both of the proline residues leads to the rapid degradation of HIF-alpha. Accordingly, inhibition of HIF prolyl hydroxylase results in stabilization and accumulation of HIF-alpha {i.e., the degradation of HIF-alpha is reduced), thereby leading to an increase in the amount of HIF-alpha available for formation of the HIF heterodimer and upregulation of target genes, such as the Erythropoietin gene. Conversely, activation of HIF prolyl hydroxylase results in destabilization of HIF-alpha {i.e., the degradation of HIF-alpha is increased), thereby leading to a decrease in the amount of HIF-alpha available for formation of the HIF heterodimer and downregulation of target genes, such as VEGF.
The family of hypoxia inducible factors includes HIF- 1 -alpha, HIF-2-alpha, and HIF-3 -alpha.
A new class of prolyl hydroxylase inhibitors and their use to treat or prevent diseases ameliorated by modulation of hypoxia-inducible factor (HIF) prolyl hydroxylase are described in U.S. Patent No. 7,811,595, which is incorporated herein by reference in its entirety. The synthesis of such prolyl hydroxylase inhibitors is described in U.S. Patent Publication No.2012/0309977, which is incorporated herein by reference in its entirety. Such compounds inhibit HIF prolyl hydroxylase, thereby stabilizing HIF-alpha. As a consequence of stabilizing HIF-alpha, endogenous erythropoietin (EPO) production is increased. As with all drugs, proper doses and dosing regimens for treating patients having diseases such as anemia are essential for achieving a desired or optimal therapeutic effect without adverse effects or unwanted side-effects. Indeed, many active compounds fail in clinical trials because an effective and safe dosing regimen cannot be found.
Vadadustat (also known as AKB-6548) in anemia secondary to chronic kidney disease (CKD)
We are developing our lead product candidate, vadadustat, to be the potential best-in-class hypoxia inducible factor–prolyl hydroxylase inhibitor for the treatment of anemia secondary to CKD.
Image may be NSFW. Clik here to view.
PATENT
CN 105837502
HIF inhibitor Vadadustat (Code AKB-6548) The chemical name N- [5- (3- chlorophenyl) -3-hydroxypyridine-2-carbonyl] glycine,
Vadadustat is a treatment for anemia associated with chronic kidney disease oral HIF inhibitor, is an American biopharmaceutical company Akebia Therapeutics invention in the research of new drugs, has completed Phase II pivotal clinical trial treatment studies, successfully met the researchers set given the level of hemoglobin in vivo target and good security, a significant effect, and phase III clinical trials.
U.S. Patent Publication US20120309977 synthetic route for preparing a Vadadustat: A 3-chlorophenyl boronic acid and 3,5_-dichloro-2-cyanopyridine as starting materials, by-catalyzed coupling methoxy substituted, cyano hydrolysis and condensation and ester hydrolysis reaction Vadadustat, process route is as follows:
Since the entire synthetic route 12 steps long, complicated operation, high cost.U.S. Patent No. 1 2 ^ ¥ disclosed 20070299086 & (^ (Scheme 3 1118 seven seven to 3,5-dichloro-2-cyanopyridine starting material, first-dichloro substituted with benzyloxy, then cyano hydrolysis, condensation, hydrogenation and deprotection trifluorosulfonyl, to give N- [5- trifluoromethanesulfonyloxy-3-hydroxypyridine-2-carbonyl) glycine methyl ester, 3-chlorophenyl and then boronic acid catalyzed coupling reactions, the final ester hydrolysis reaction Vadadustat, process route is as follows:
The synthesis steps long, intermediate products and final products contain more impurities and byproducts, thus purified requires the use of large amounts of solvents, complicated operation, low yield, and because the hydrogenation reaction is a security risk on the production, not conducive to the promotion of industrial production, it is necessary to explore a short process, simple operation, low cost synthetic method whereby industrial production Vadadus tat fit.
A) Preparation of N- (3,5_-dichloro-2-carbonyl) glycine methyl ester:
3,5-dichloro-2-pyridinecarboxylic acid (19.2g, 0.10mol) and N, N’_ carbonyldiimidazole (24.3g, 0.15mol) was dissolved in N, N- dimethylformamide (100 mL ), was added glycine methyl ester hydrochloride (15.18,0.12111〇1), 11 was added dropwise diisopropylethylamine (51.7g, 0.40mol), the reaction mixture was stirred 35 ° C for 8 hours, TLC determined the completion of reaction gussets The reaction solution was concentrated by rotary evaporation to dryness, dilute hydrochloric acid was adjusted to neutral by adding ethyl acetate, dried over magnesium sulfate, and concentrated by rotary evaporation to dryness, and recrystallized from methanol to give N- (3,5- dichloro-pyridin-2 – carbonyl) glycine methyl ester, an off-white solid (21.6g), a yield of 82.0%, this reaction step is as follows:Image may be NSFW. Clik here to view.
1234567 B) Preparation of N- [5- (3- chlorophenyl) -3-chloropyridine-2-carbonyl] glycine methyl ester: 2
1 (3,5-dichloro-2-carbonyl) glycine methyl ester (20 (^, 〇1 76111111), 3-chlorophenyl boronic acid (13.18, 3 83.7mmol), [l, l’- bis (diphenylphosphino) ferrocene] dichloropalladium (2.8g, 3.8mmol), potassium carbonate (14.2g, 4 0. lmo 1) and N, N- dimethylformamide (75mL) was added The reaction flask, the reaction mixture was heated to 60 ° C for 20 hours the reaction was stirred for 5:00, point TLC plates to determine completion of the reaction, the reaction solution was cooled to room temperature, was concentrated by rotary evaporation to dryness, extracted with ethyl acetate, washed with brine, sulfuric acid 6 magnesium dried and concentrated by rotary evaporation to dryness, a mixed solvent of ethyl acetate and n-hexane was recrystallized to give N- [5- (3- chlorophenyl) -3-7-chloro-2-carbonyl] glycine methyl ester, white solid (19.7g), yield 76.4%, this reaction step is as follows:
C) Preparation of N_ [5- (3- chlorophenyl) -3-methoxy-pyridine-2-carbonyl] glycine:
N- [5- (3- chlorophenyl) -3-chloropyridine-2-carbonyl] glycine methyl ester (19 (^, 56111 111〇1) and sodium methoxide (7.6g, 0.14mol) was dissolved in methanol (150 mL), the reaction mixture was heated to 65 ° C, the reaction was stirred at reflux for 24 hours, TLC determined gussets completion of the reaction the reaction solution was cooled to room temperature, water (300mL) was stirred for 3h, cooled to 0 ° C, stirred for 2h, precipitated solid was filtered, the filter cake was dried to give N- [5- (3- chlorophenyl) -3-methoxy-pyridine-2-carbonyl] glycine, off-white solid (17.4 g of), a yield of 96.5%, of the reaction steps are as follows:
N- [5- (3- chlorophenyl) -3-methoxy-pyridine-2-carbonyl] glycine (16.68,51.7111111〇1) and 48% hydrobromic acid solution (52mL, 0.46mol) added to the reaction bottle, the reaction mixture was heated to 100 ° C, the reaction was stirred at reflux for 24 hours, TLC determined gussets completion of the reaction the reaction solution cooled square ~ 5 ° C, was slowly added 50% sodium hydroxide solution was adjusted to pH 2 at 0 -5 ° C under crystallization 3h, the filter cake washed with ethyl acetate and n-hexane mixed solvent of recrystallization, in finished Vadadustat, off-white solid (15.6g), a yield of 98.0%, this reaction step is as follow
which has an X-ray powder diffraction pattern as shown in FIG. 1. In certain embodiments, Form A of Compound (I) has an X-ray powder diffraction pattern comprising one, two, three, four, or five peaks at approximately 18.1 , 20.3, 22.9, 24.0, and 26.3 °2Θ; and wherein the crystalline Compound (I) is substantially free of any other crystalline form of Compound (I).
EXAMPLE 1Methyl {[5-(3-chlorophenyl)-3-hydroxypyridin-2-yl]amino}acetate (4)
Preparation of 5-(3-chlorophenyl)-3-chloro-2-cyanopyridine (1): To a 100 mL round bottom flask adapted for magnetic stirring and equipped with a nitrogen inlet was charged (3-chlorophenyl)boronic acid (5 g, 32 mmol), 3,5-dichloro-2-cyanopyridine (5.8 g, 34 mmol), K2CO3 (5.5 g, 40 mmol), [1,1′-bis(diphenyphosphino)ferrocene]dichloro-palladium(II) [PdCl2(dppf)] (0.1 g, 0.13 mmol), dimethylformamide (50 mL) and water (5 mL). The reaction solution was agitated and heated to 45° C. and held at that temperature for 18 hours after which the reaction was determined to be complete due to the disappearance of 3,5-dichloro-2-cyanopyridine as measured by TLC analysis using ethyl acetate/methanol (4:1) as the mobile phase and UV 435 nm to visualize the reaction components. The reaction solution was then cooled to room temperature and the contents partitioned between ethyl acetate (250 mL) and saturated aqueous NaCl (100 mL). The organic phase was isolated and washed a second time with saturated aqueous NaCl (100 mL). The organic phase was dried for 4 hours over MgSO4, the MgSO4 removed by filtration and the solvent removed under reduced pressure. The residue that remained was then slurried in methanol (50 mL) at room temperature for 20 hours. The resulting solid was collected by filtration and washed with cold methanol (50 mL) then hexanes (60 mL) and dried to afford 5.8 g (73% yield) of an admixture containing a 96:4 ratio of the desired regioisomer. 1H NMR (DMSO-d6) δ 9.12 (d, 1H), 8.70 (d, 1H), 8.03 (t, 1H) 7.88 (m, 1H), and 7.58 (m, 2H)
Preparation of 5-(3-chlorophenyl)-3-methoxy-2-cyanopyridine (2): To a 500 mL round bottom flask adapted for magnetic stirring and fitted with a reflux condenser and nitrogen inlet was charged with 5-(3-chlorophenyl)-3-chloro-2-cyanopyridine, 1, (10 g, 40 mmol), sodium methoxide (13.8 mL, 60 mmol) and methanol (200 mL). With stirring, the reaction solution was heated to reflux for 20 hours. The reaction was determined to be complete due to the disappearance of 5-(3-chlorophenyl)-3-chloro-2-cyanopyridine as measured by TLC analysis using hexane/ethyl acetate (6:3) as the mobile phase and UV 435 nm to visualize the reaction components. The reaction mixture was cooled to room temperature and combined with water (500 mL). A solid began to form. The mixture was cooled to 0° C. to 5° C. and stirred for 3 hours. The resulting solid was collected by filtration and washed with water, then hexane. The resulting cake was dried in vacuo at 40° C. to afford 9.4 g (96% yield) of the desired product as an off-white solid. 1H NMR (DMSO-d6) δ 8.68 (d, 1H), 8.05 (d, 1H), 8.01 (s, 1H) 7.86 (m, 1H), 7.59 (s, 1H), 7.57 (s, 1H) and 4.09 (s, 3H).
Preparation of 5-(3-chlorophenyl)-3-hydroxypyridine-2-carboxylic acid (3): To a 50 mL round bottom flask adapted for magnetic stirring and fitted with a reflux condenser was charged 5-(3-chlorophenyl)-3-methoxy-2-cyanopyridine, 2, (1 g, 4 mmol) and a 48% aqueous solution of HBr (10 mL). While being stirred, the reaction solution was heated to reflux for 20 hours. The reaction was determined to be complete due to the disappearance of 5-(3-chlorophenyl)-3-methoxy-2-cyanopyridine as measured by TLC analysis using hexane/ethyl acetate (6:3) as the mobile phase and UV 435 nm to visualize the reaction components. The reaction contents was then cooled to 0° C. to 5° C. with stirring and the pH was adjusted to approximately 2 by the slow addition of 50% aqueous NaOH. Stirring was then continued at 0° C. to 5° C. for 3 hours. The resulting solid was collected by filtration and washed with water, then hexane. The resulting cake was dried in vacuo at 40° C. to afford 1.03 g (quantitative yield) of the desired product as an off-white solid. 1H NMR (DMSO-d6) δ 8.52 (d, 1H), 7.99 (d, 1H), 7.95 (s, 1H) 7.81 (t, 1H), 7.57 (s, 1H), and 7.55 (s, 1H).
Preparation of methyl {[5-(3-chlorophenyl)-3-hydroxypyridin-2-yl]amino}acetate (4): To a 50 mL round bottom flask adapted for magnetic stirring and fitted with a nitrogen inlet tube was charged 5-(3-chlorophenyl)-3-hydroxypyridine-2-carboxylic acid, 3, (1 gm, 4 mmol), N,N′-carbonyldiimidazole (CDI) (0.97 g, 6 mmol) and dimethyl sulfoxide (5 mL). The reaction mixture was stirred at 45° C. for about 1 hour then cooled to room temperature. Glycine methyl ester hydrochloride (1.15 g, 12 mmol) is added followed by the dropwise addition of diisopropylethylamine (3.2 mL, 19 mmol). The mixture was then stirred for 2.5 hours at room temperature after which water (70 mL) was added. The contents of the reaction flask was cooled to 0° C. to 5° C. and 1N HCl was added until the solution pH is approximately 2. The solution was extracted with dichloromethane (100 mL) and the organic layer was dried over MgSO4 for 16 hours. Silica gel (3 g) is added and the solution slurried for 2 hours after which the solids are removed by filtration. The filtrate is concentrated to dryness under reduced pressure and the resulting residue was slurried in methanol (10 mL) for two hours. The resulting solid was collected by filtration and washed with cold methanol (20 mL) then hexane and the resulting cake is dried to afford 0.85 g of the desired product as an off-white solid. The filtrate was treated to afford 0.026 g of the desired product as a second crop. The combined crops afford 0.88 g (68% yield) of the desired product. 1H NMR (DMSO-d6) δ 12.3 (s, 1H), 9.52 (t, 1H), 8.56 (d, 1H), 7.93 (s, 1H), 7.80 (q, 2H), 7.55 (t, 2H), 4.12 (d, 2H), and 3.69 (s, 3H).
The formulator can readily scale up the above disclosed synthesis. Disclosed herein below is a synthesis wherein the disclosed process is scaled up for commercial use
EXAMPLE 2Methyl {[5-(3-chlorophenyl)-3-hydroxypyridin-2-yl]amino}acetate (4)
Preparation of 5-(3-chlorophenyl)-3-chloro-2-cyanopyridine (1): A 20 L reactor equipped with a mechanical stirrer, dip tube, thermometer and nitrogen inlet was charged with (3-chlorophenyl)boronic acid (550 g, 3.52 mol), 3,5-dichloro-2-cyanopyridine (639 g, 3.69 mol), K2CO3 (5.5 g, 40 mmol), [1,1′-bis(diphenyphosphino)ferrocene]dichloro-palladium(II) [PdCl2(dppf)] (11.5 g, 140 mmol), and dimethylformamide (3894 g, 4.125 L). The reaction solution was agitated and purged with nitrogen through the dip-tube for 30 minutes. Degassed water (413 g) was then charged to the reaction mixture while maintaining a temperature of less than 50° C. 25 hours. The reaction was determined to be complete due to the disappearance of 3,5-dichloro-2-cyanopyridine as measured by TLC analysis using ethyl acetate/methanol (4:1) as the mobile phase and UV 435 nm to visualize the reaction components. The reaction solution was then cooled to 5° C. and charged with heptane (940 g, 1.375 L) and agitated for 30 minutes. Water (5.5 L) was charged and the mixture was further agitated for 1 hour as the temperature was allowed to rise to 15° C. The solid product was isolated by filtration and washed with water (5.5 L) followed by heptane (18881 g, 2750 ML). The resulting cake was air dried under vacuum for 18 hours and then triturated with a mixture of 2-propanol (6908 g, 8800 mL0 and heptane (1 g, 2200 mL0 at 50° C. for 4 hours, cooled to ambient temperature and then agitated at ambient temperature for 1 hour. The product was then isolated by filtration and washed with cold 2-propanol (3450 g, 4395 mL) followed by heptane (3010 g, 4400 mL). The resulting solid was dried under high vacuum at 40° C. for 64 hours to afford 565.9 g (65% yield) of the desired product as a beige solid. Purity by HPLC was 98.3. 1H NMR (DMSO-d6) δ 9.12 (d, 1H), 8.70 (d, 1H), 8.03 (t, 1H) 7.88 (m, 1H), and 7.58 (m, 2H).
Preparation of 5-(3-chlorophenyl)-3-methoxy-2-cyanopyridine (2): A 20 L reactor equipped with a mechanical stirred, condenser, thermometer and nitrogen inlet was charged with 5-(3-chlorophenyl)-3-chloro-2-cyanopyridine, 1, (558 g, 2.24 mol) and sodium methoxide (25% solution in methanol, 726.0 g, 3.36 mol). With agitation, the reaction solution was heated to reflux for 24 hours, resulting in a beige-colored suspension. The reaction was determined to be complete due to the disappearance of 5-(3-chlorophenyl)-3-chloro-2-cyanopyridine as measured by TLC analysis using hexane/ethyl acetate (6:3) as the mobile phase and UV 435 nm to visualize the reaction components. The reaction mixture was cooled to 5° C. and then charged with water (5580 mL). The resulting slurry was agitated for 3 hours at 5° C. The solid product was isolated by filtration and washed with water (5580 mL) until the filtrate had a pH of 7. The filter cake was air dried under vacuum for 16 hours. The filter cake was then charged back to the reactor and triturated in MeOH (2210 g, 2794 mL) for 1 hour at ambient temperature. The solid was collected by filtration and washed with MeOH (882 g, 1116 mL, 5° C.) followed by heptane (205 mL, 300 mL), and dried under high vacuum at 45° C. for 72 hours to afford 448 g (82% yield) of the desired product as an off-white solid. Purity by HPLC was 97.9%. 1H NMR (DMSO-d6) δ 8.68 (d, 1H), 8.05 (d, 1H), 8.01 (s, 1H) 7.86 (m, 1H), 7.59 (s, 1H), 7.57 (s, 1H) and 4.09 (s, 3H).
Preparation of 5-(3-chlorophenyl)-3-hydroxypyridine-2-carboxylic acid (3): A 20 L reactor equipped with a mechanical stirrer, condenser, thermometer, nitrogen inlet and 25% aqueous NaOH trap was charged 5-(3-chlorophenyl)-3-methoxy-2-cyanopyridine, 2, (440.6 g, 1.8 mol) and 37% aqueous solution of HCl (5302 g). While being agitated, the reaction solution was heated to 102° C. for 24 hours. Additional 37% aqueous HCl (2653 g) was added followed by agitation for 18 hours at 104° C. The reaction contents was then cooled to 5° C., charged with water (4410 g) and then agitated at 0° C. for 16 hours. The resulting precipitated product was isolated by filtration and washed with water until the filtrate had a pH of 6 (about 8,000 L of water). The filter cake was pulled dry under reduced pressure for 2 hours. The cake was then transferred back into the reactor and triturated in THF (1958 g, 2201 mL) at ambient temperature for 2 hours. The solid product was then isolated by filtration and washed with THF (778 g, 875 mL) and dried under reduced pressure at 5° C. for 48 hours to afford 385 g (89% yield) of the desired product as an off-white solid. HPLC purity was 96.2%. 1H NMR (DMSO-d6) δ 8.52 (d, 1H), 7.99 (d, 1H), 7.95 (s, 1H) 7.81 (t, 1H), 7.57 (s, 1H), and 7.55 (s, 1H).
Preparation of methyl {[5-(3-chlorophenyl)-3-hydroxypyridin-2-yl]amino}acetate (4): A 20 L reactor equipped with a mechanical stirrer, condenser, thermometer and nitrogen inlet was charged with 5-(3-chlorophenyl)-3-hydroxypyridine-2-carboxylic acid, 3, (380 g, 1.52 mol) and diisopropylethylamine (DIPEA) (295 g, 2.28 mol). With agitation, the solution was cooled to 3° C. and charged with trimethylacetyl chloride (275.7 g, 2.29 mol) while maintaining a temperature of less than 11° C., The mixture was then agitated at ambient temperature for 2 hours. The mixture was then cooled to 10° C. and charged with a slurry of glycine methyl ester HCl (573.3 g, 4. 57 mol) and THF (1689 g, 1900 mL), then charged with DIPEA (590.2 g, 4.57 mol) and agitated at ambient temperature for 16 hours. The mixture was then charged with EtOH (1500 g, 1900 mL) and concentrated under reduced pressure to a reaction volume of about 5.8 L. The EtOH addition and concentration was repeated twice more. Water (3800 g) was then added and the mixture was agitated for 16 hours at ambient temperature. The resulting solid product was isolated by filtration and washed with a mixture of EtOH (300 g, 380 mL) and water (380 g), followed by water (3800 g), dried under reduced pressure for 18 hours at 50° C. to afforded 443 g (91% yield) of the desired product as an off-white solid. Purity by HPLC was 98.9%. 1H NMR (DMSO-d6) δ 12.3 (s, 1H), 9.52 (t, 1H), 8.56 (d, 1H), 7.93 (s, 1H), 7.80 (q, 2H), 7.55 (t, 2H), 4.12 (d, 2H), and 3.69 (s, 3H).
Scheme II herein below outlines and Example 2 describes a non-limiting example of the disclosed process for preparing a prolyl hydroxylase inhibitor from an ester prodrug.
EXAMPLE 3{[5-(3-Chlorophenyl)-3-hydroxypyridin-2-yl]amino}acetic acid (5)
Preparation of {[5-(3 -chlorophenyl)-3-hydroxypyridin-2-yl]amino}acetic acid (5): To a 50 mL flask is charged methyl {[5-(3-chlorophenyl)-3-hydroxypyridin-2-yl]amino}-acetate, 4, (0.45 g, 1.4 mmol), tetrahydrofuran (4.5 mL) and 1 M NaOH (4.5 mL, 4.5 mmol). The mixture was stirred for 2 hours at room temperature after which it was determined by TLC analysis using hexane/ethyl acetate (6:3) as the mobile phase and UV 435 nm to visualize the reaction components that the reaction was complete. The reaction solution was adjusted to pH 1 with concentrated HCl and the solution was heated at 35° C. under vacuum until all of the tetrahydrofuran had been removed. A slurry forms as the solution is concentrated. With efficient stirring the pH is adjusted to ˜2 with the slow addition of 1 M NaOH. The solid which forms was collected by filtration, washed with water, followed by hexane, then dried under vacuum to afford 0.38 g (88% yield) of the desired product as a white solid. 1H NMR (DMSO-d6) δ 12.84 (s, 1H), 12.39 (s, 1H), 9.39 (t, 1H), 8.56 (d, 1H), 7.94 (s, 1H), 7.81 (m, 2H), 7.55 (q, 2H), and 4.02 (d, 2H).
The formulator can readily scale up the above disclosed synthesis. Disclosed herein below is a synthesis wherein the disclosed process is scaled up for commercial use.
EXAMPLE 4{[5-(3-Chlorophenyl)-3-hydroxypyridin-2-yl]amino}acetic acid (5)
Preparation of {[5-(3-chlorophenyl)-3-hydroxypyridin-2-yl]amino}acetic acid (5): To a 20 L reactor equipped with a mechanical stirrer, condenser, thermometer and nitrogen inlet was charged methyl {[5-(3-chlorophenyl)-3-hydroxypyridin-2-yl]amino}-acetate, 4, (440 g, 1.42 mol), tetrahydrofuran (3912 g, 4400 mL) and 1 M NaOH (4400 mL). The mixture was stirred for 2 hours at room temperature after which it was determined by TLC analysis using hexane/ethyl acetate (6:3) as the mobile phase and UV 435 nm to visualize the reaction components that the reaction was complete. The reaction solution was acidified to a pH of 2 with slow addition of 2M HCl (2359 g). The resulting mixture was concentrated under reduced pressure to a volume of about 7.5 L. Ware (2210 g) was added and the solution cooled to ambient temperature and agitated for 18 hours. The solid product was isolated by filtration and washed with water (6 L). the crude product was transferred back into the reactor and triturated with 2215 g o deionized water at 70° C. for 16 hours. The mixture was cooled to ambient temperature, The solid product was isolated by filtration and washed with water (500 mL) and dried under reduced pressure at 70° C. for 20 hours to afford 368 g (87% yield) of the desired product as an off-white solid. Purity by HPLC was 99.3%. 1H NMR (DMSO-d6) δ 12.84 (s, 1H), 12.39 (s, 1H), 9.39 (t, 1H), 8.56 (d, 1H), 7.94 (s, 1H), 7.81 (m, 2H), 7.55 (q, 2H), and 4.02 (d, 2H).
Scheme III herein below outlines and Example 3 describes a non-limiting example of the disclosed process for preparing a prolyl hydroxylase amide prodrug.
EXAMPLE 55-(3-Chlorophenyl)-N-(2-amino-2-oxoethyl)-3-hydroxylpyridin-2-yl amide
Preparation of 5-(3-chlorophenyl)-N-(2-amino-2-oxoethyl)-3-hydroxylpyridin-2-yl amide (6): To a solution of 5-(3-chlorophenyl)-3-hydroxypyridine-2-carboxylic acid, 3, (749 mg, 3 mmol) in DMF (20 mL) at room temperature under N2 is added 1-(3-dimethyl-aminopropyl)-3-ethylcarbodiimide (EDCI) (0.925 g, 5.97 mmol) and 1-hydroxybenzo-triazole (HOBt) (0.806 g, 5.97 mmol). The resulting solution is stirred for 15 minutes then 2-aminoacetamide hydrochloride (0.66 g, 5.97 mmol) and diisopropylethylamine (1.56 ml, 8.96 mmol) are added. The reaction is monitored by TLC and when the reaction is complete the reaction mixture is concentrated under reduced pressure and H2O added. The product can be isolated by normal work-up: The following data have been reported for compound (6). 1H NMR (250 MHz, DMSO-d6) δ ppm 12.46 (1H, s), 9.17 (1H, t, J=5.9 Hz), 8.55 (1H, d, J=2.0 Hz), 7.93 (1H, d, J=0.9 Hz), 7.75-7.84 (2H, m), 7.49-7.60 (3H, m), 7.18 (1H, s), 3.91 (2H, d, J=5.9 Hz). HPLC-MS: m/z 306 [M+H]+.
Scheme IV herein below depicts a non-limiting example the hydrolysis of an amide pro-drug to a prolyl hydroxylase inhibitor after removal of a R10 protecting group
Beuck S, Schänzer W, Thevis M. Hypoxia-inducible factor stabilizers and other
small-molecule erythropoiesis-stimulating agents in current and preventive doping
analysis. Drug Test Anal. 2012 Nov;4(11):830-45. doi: 10.1002/dta.390. Epub 2012
Feb 24. Review. PubMed PMID: 22362605.
Akebia Reaches Agreement with FDA and EMA on Vadadustat Global Phase 3 Program
Plans to Initiate Phase 3 PRO2TECT™ Clinical Program by Year-End
CAMBRIDGE, Mass.–(BUSINESS WIRE)– Akebia Therapeutics, Inc. (NASDAQ: AKBA), a biopharmaceutical company focused on delivering innovative therapies to patients with kidney disease through the biology of hypoxia inducible factor (HIF), today announced the successful completion of the End-of-Phase 2 Meeting process with the United States Food and Drug Administration (FDA) and the Scientific Advice Process with the European Medicines Agency (EMA) for its lead product, vadadustat (formerly AKB-6548), for patients with anemia related to non-dialysis dependent chronic kidney disease (NDD-CKD). The company has reached agreement with both the FDA and EMA regarding key elements of the Phase 3 program, known as the PRO2TECT™ program, and expects to launch the program later this year.
The PRO2TECT™program includes two separate studies and will collectively enroll approximately 3,100 NDD-CKD patients across 500 sites globally. The correction study will address anemia patients not currently being treated with recombinant erythropoiesis stimulating agents (rESAs). The conversion study includes patients currently receiving rESA who will be converted to either vadadustat or the active control with the goal of maintaining their baseline hemoglobin levels. Both studies will include a 1:1 randomization and an open label, active-control, non-inferiority design. Primary endpoints include an efficacy assessment of the hemoglobin response and an assessment of cardiovascular safety measured by major adverse cardiovascular events.
“Akebia’s Phase 3 program is designed to provide the medical community and regulators with a clear understanding of vadadustat’s potential benefit and safety advantages over rESAs, the current standard of care worldwide and, with a positive outcome, to establish vadadustat as the best-in-class treatment option for patients with renal anemia,” stated John P. Butler, President and Chief Executive Officer of Akebia. “We are pleased that the regulators are in agreement regarding the importance of an active-control trial as this design is the most clinically relevant and commercially valuable, and will allow us the quickest path to full enrollment. We are now moving rapidly to launch these studies and advance our goal of bringing forward new treatment options for patients suffering from renal anemia.”
“This Phase 3 program builds on the positive data from our Phase 2 program in NDD-CKD patients which demonstrated that once-daily vadadustat can control and maintain hemoglobin levels in a clinically relevant range while minimizing fluctuations in hemoglobin levels that are associated with increased cardiovascular safety risks,” stated Brad Maroni, M.D., Chief Medical Officer at Akebia. “These two Phase 3 event-driven studies are designed to establish the safety and efficacy of vadadustat in the setting of contemporary clinical practice patterns, and support regulatory approvals globally.”
In addition, Akebia discussed with the FDA and EMA a parallel Phase 3 program, known as the INNO2VATE™ program, for vadadustat in patients with anemia related to chronic kidney disease who are undergoing dialysis (DD-CKD). Akebia expects to formalize its Phase 3 program in DD-CKD patients after presenting the results from its recently completed Phase 2 study to both regulatory agencies.
About Vadadustat (Formerly AKB-6548)
Vadadustat is an oral therapy currently in development for the treatment of anemia related to chronic kidney disease (CKD). Vadadustat is designed to stabilize HIF, a transcription factor that regulates the expression of genes involved with red blood cell (RBC) production in response to changes in oxygen levels, by inhibiting the hypoxia-inducible factor prolyl hydroxylase (HIF-PH) enzyme. Vadadustat exploits the same mechanism of action used by the body to naturally adapt to lower oxygen availability associated with a moderate increase in altitude. At higher altitudes, the body responds to lower oxygen availability with increased production of HIF, which coordinates the interdependent processes of iron mobilization and erythropoietin (EPO) production to increase RBC production and, ultimately, improve oxygen delivery.
As a HIF stabilizer with best-in-class potential, vadadustat raises hemoglobin levels predictably and sustainably, with a dosing regimen that allows for a gradual and controlled titration. Vadadustat has been shown to improve iron mobilization, potentially eliminating the need for intravenous iron administration and reducing the overall need for iron supplementation.
About Anemia Related to CKD
Approximately 30 million people in the United States have CKD, with an estimated 1.8 million of these patients suffering from anemia. Anemia results from the body’s inability to coordinate RBC production in response to lower oxygen levels due to the progressive loss of kidney function, which occurs in patients with CKD. Left untreated, anemia significantly accelerates patients’ overall deterioration of health with increased morbidity and mortality. Renal anemia is currently treated with injectable rESAs, which are associated with inconsistent hemoglobin responses and well-documented safety risks.
About Akebia Therapeutics
Akebia Therapeutics, Inc. is a biopharmaceutical company headquartered in Cambridge, Massachusetts, focused on delivering innovative therapies to patients with kidney disease through HIF biology. The company has completed Phase 2 development of its lead product candidate, vadadustat, an oral therapy for the treatment of anemia related to CKD in both non-dialysis and dialysis patients.
clip
Akebia Announces Positive Top-Line Results from its Phase 2 Study of Vadadustat in Dialysis Patients with Anemia Related to Chronic Kidney Disease
-Treatment with Vadadustat Successfully Maintained Mean Hemoglobin Levels Following Conversion from rESA Therapy-
-Vadadustat Demonstrated a Favorable Safety Profile with Once Daily and Three Times per Week Dosing-
CAMBRIDGE, Mass.–(BUSINESS WIRE)–Akebia Therapeutics, Inc. (NASDAQ:AKBA), a biopharmaceutical company focused on delivering innovative therapies to patients with kidney disease through the biology of hypoxia inducible factor (HIF), today announced positive top-line results from its Phase 2 study of vadadustat (formerly AKB-6548) in dialysis patients with anemia related to chronic kidney disease (CKD). The study achieved its primary objective, indicating that vadadustat maintained stable hemoglobin (HGB) levels throughout the 16-week treatment period following conversion from recombinant erythropoiesis-stimulating agent (rESA) therapy. Vadadustat demonstrated a favorable safety profile with no drug-related serious adverse events and no deaths. The results highlight the potential of vadadustat, dosed either once daily or three times per week, to safely and predictably manage and sustain HGB levels in CKD patients undergoing dialysis.
“This study was a clear success, demonstrating the potential of vadadustat to effectively and safely treat anemia in dialysis patients switching from injectable rESA therapy”
The open-label, multi-center, 94 patient study was designed to evaluate the ability of vadadustat to maintain hemoglobin levels in patients undergoing hemodialysis who were previously being treated with rESAs. Patients were assigned to one of three dose cohorts: once daily vadadustat at a starting dose of 300mg, once daily vadadustat at a starting dose of 450mg, or vadadustat three times per week in conjunction with the patient’s hemodialysis schedule at a starting dose of 450mg. The study achieved its primary endpoints of maintaining stable hemoglobin levels over 16 weeks of treatment in all three cohorts of patients converting from rESAs to vadadustat.
Vadadustat was well tolerated among patients in all three dose cohorts. Treatment-emergent adverse events (TEAEs) with vadadustat were balanced across the cohorts. Serious adverse events (SAEs) were reported in 13 subjects (13.8%), well within the expected range for this patient population. There were no drug-related SAEs and no deaths reported in the study.
“This study was a clear success, demonstrating the potential of vadadustat to effectively and safely treat anemia in dialysis patients switching from injectable rESA therapy,” said Brad Maroni, M.D., Chief Medical Officer at Akebia. “We are impressed with the consistency in hemoglobin levels across the duration of the study, which highlights the ability of vadadustat to control and maintain hemoglobin levels in this patient population. Furthermore, the results indicate that daily and three times per week dosing regimens are both viable options for patients on dialysis.”
John P. Butler, President and Chief Executive Officer of Akebia, stated, “These results further confirm vadadustat as a potential best-in-class anemia treatment for CKD patients, and reinforce our confidence in this product candidate as we advance toward our Phase 3 program. Adding these results to the 12 other clinical studies we have completed, we are confident in the potential for vadadustat to treat anemia in a broad array of patients with CKD. We are pleased to have successfully completed this stage of our drug development and look forward to initiating Phase 3 studies.”
Complete efficacy and safety data from this Phase 2 study will be presented at an upcoming medical meeting.
About the Phase 2 Study Design of Vadadustat in Dialysis Patients with Anemia Related to CKD
The Phase 2 multi-center, open-label study evaluated 94 patients over 16 weeks of treatment, at 20 dialysis centers in the United States, including an assessment of HGB response to the starting dose of vadadustat during the first 8 weeks, followed by an assessment of HGB response to algorithm-guided dose adjustments of vadadustat during the subsequent 8 weeks of treatment. The study enrolled three cohorts, each consisting of approximately 30 CKD patients with anemia undergoing dialysis who were switched from injectable rESA therapy to vadadustat. Patients in the first two cohorts received once daily doses of vadadustat, while patients in the third cohort received vadadustat three times per week in conjunction with their hemodialysis schedule.
Class Antivirals; Aza compounds; Cyclic ethers; Cyclopentanes; Cyclopropanes; Quinoxalines; Small molecules
Mechanism of Action Hepatitis C virus NS3 protein inhibitors
Phase II Hepatitis C
Most Recent Events
18 Apr 2016 Pooled efficacy and adverse event data from the phase II SURVEYOR-I and SURVEYOR-2 trials for Hepatitis C presented at The International Liver Congress™ 2016 (ILC-2016)
15 Apr 2016 Updated efficacy data from a phase II MAGELLAN 1 study were reported by Enanta Pharmaceuticals
15 Apr 2016 Updated safety and efficacy data from a phase II MAGELLAN 1 study were presented at the International Liver Congress™ (ILC-2016)
AbbVie’s investigational, pan-genotypic regimen of glecaprevir (ABT-493) / pibrentasvir (ABT-530) (G/P) has received breakthrough therapy designation from the US Food and Drug Administration (FDA) to treat chronic hepatitis C virus (HCV).
HCV is the principal cause of non-A, non-B hepatitis and is an increasingly severe public health problem both in the developed and developing world. It is estimated that the virus infects over 200 million people worldwide, surpassing the number of individuals infected with the human immunodeficiency virus (HIV) by nearly five fold. HCV infected patients, due to the high percentage of individuals inflicted with chronic infections, are at an elevated risk of developing cirrhosis of the liver, subsequent hepatocellular carcinoma and terminal liver disease. HCV is the most prevalent cause of hepatocellular cancer and cause of patients requiring liver transplantations in the western world.
There are considerable barriers to the development of anti-HCV therapeutics, which include, but are not limited to, the persistence of the virus, the genetic diversity of the virus during replication in the host, the high incident rate of the virus developing drug-resistant mutants, and the lack of reproducible infectious culture systems and small-animal models for HCV replication and pathogenesis. In a majority of cases, given the mild course of the infection and the complex biology of the liver, careful consideration must be given to antiviral drugs, which are likely to have significant side effects.
Only two approved therapies for HCV infection are currently available. The original treatment regimen generally involves a 3-12 month course of intravenous interferon-α (IFN-α), while a new approved second-generation treatment involves co-treatment with IFN-α and the general antiviral nucleoside mimics like ribavirin. Both of these treatments suffer from interferon related side effects as well as low efficacy against HCV infections. There exists a need for the development of effective antiviral agents for treatment of HCV infection due to the poor tolerability and disappointing efficacy of existing therapies.
In a patient population where the majority of individuals are chronically infected and asymptomatic and the prognoses are unknown, an effective drug would desirably possess significantly fewer side effects than the currently available treatments. The hepatitis C non-structural protein-3 (NS3) is a proteolytic enzyme required for processing of the viral polyprotein and consequently viral replication. Despite the huge number of viral variants associated with HCV infection, the active site of the NS3 protease remains highly conserved thus making its inhibition an attractive mode of intervention. Recent success in the treatment of HIV with protease inhibitors supports the concept that the inhibition of NS3 is a key target in the battle against HCV.
HCV is a flaviridae type RNA virus. The HCV genome is enveloped and contains a single strand RNA molecule composed of circa 9600 base pairs. It encodes a polypeptide comprised of approximately 3010 amino acids.
The HCV polyprotein is processed by viral and host peptidase into 10 discreet peptides which serve a variety of functions. There are three structural proteins, C, E1 and E2. The P7 protein is of unknown function and is comprised of a highly variable sequence. There are six non-structural proteins. NS2 is a zinc-dependent metalloproteinase that functions in conjunction with a portion of the NS3 protein. NS3 incorporates two catalytic functions (separate from its association with NS2): a serine protease at the N-terminal end, which requires NS4A as a cofactor, and an ATP-ase-dependent helicase function at the carboxyl terminus. NS4A is a tightly associated but non-covalent cofactor of the serine protease.
The NS3/4A protease is responsible for cleaving four sites on the viral polyprotein. The NS3-NS4A cleavage is autocatalytic, occurring in cis. The remaining three hydrolyses, NS4A-NS4B, NS4B-NS5A and NS5A-NS5B all occur in trans. NS3 is a serine protease which is structurally classified as a chymotrypsin-like protease. While the NS serine protease possesses proteolytic activity by itself, the HCV protease enzyme is not an efficient enzyme in terms of catalyzing polyprotein cleavage. It has been shown that a central hydrophobic region of the NS4A protein is required for this enhancement. The complex formation of the NS3 protein with NS4A seems necessary to the processing events, enhancing the proteolytic efficacy at all of the sites.
A general strategy for the development of antiviral agents is to inactivate virally encoded enzymes, including NS3, that are essential for the replication of the virus. Current efforts directed toward the discovery of NS3 protease inhibitors were reviewed by S. Tan, A. Pause, Y. Shi, N. Sonenberg, Hepatitis C Therapeutics: Current Status and Emerging Strategies, Nature Rev. Drug Discov. 1, 867-881 (2002).
The acid 1-6a (21 mg, 0.0356 mmol) was dissolved in DCM (1.5 ml), and to this solution was added sulfonamide 1-7e (13.0 mg, 0.0463 mmol), HATU (17.6 mg, 0.0462 mmol) and DIPEA (12.4 ul, 0.0712 mmol). The mixture was stirred for 3 h, and then diluted with DCM. The organic layer was washed with 1 N HCl, water, brine, dried and concentrated in vacuo. The residue was purified by HPLC to afford the title compound. MS-ESI m/z 839.41 (M+H)+.
US FDA grants breakthrough therapy designation to AbbVie’s G/P to treat HCV
Image may be NSFW. Clik here to view.
4 October 2016
AbbVie’s investigational, pan-genotypic regimen of glecaprevir (ABT-493) / pibrentasvir (ABT-530) (G/P) has received breakthrough therapy designation from the US Food and Drug Administration (FDA) to treat chronic hepatitis C virus (HCV).
The HCV is a bloodborne virus commonly transmitted through injecting drug use due to the sharing of injection equipment, reuse or inadequate sterilisation of medical equipment, and the transfusion of unscreened blood and blood products.
The designation facilitates the use of AbbVie’s G/P to treat chronic HCV patients who failed previous therapy with direct-acting antivirals (DAAs) in genotype 1 (GT1), including therapy with an NS5A inhibitor and / or protease inhibitor.
AbbVie research and development executive vice-president Michael Severino said: “AbbVie is committed to advancing HCV care and addressing areas of continued unmet need for people living with chronic HCV.
“AbbVie is committed to advancing HCV care and addressing areas of continued unmet need for people living with chronic HCV.”
“The FDA’s breakthrough therapy designation is an important step in our effort to bring our pan-genotypic regimen to market, which we are also investigating as an eight-week path to virologic cure for the majority of patients.”
AbbVie said that G/P is currently in Phase III trials evaluating the safety and efficacy of the regimen across all major HCV genotypes (genotypes 1-6).
Figures released by the World Health Organisation revealed that an estimated 700,000 people die each year from hepatitis C-related liver diseases.
There is currently no vaccine for hepatitis C, although research in this area is underway at present.
The GDUFA (Generic Drug User Fee Amendments) is a legislative package which came into force in 2012 and entitles the US-American FDA to collect fees from generic drug manufacturers, who strive for a marketing authorisation for the American market. An annual fee has to be paid after the successful registration.
The core of the document is the obligation to “Self-Identify” for those companies that have to submit essential site-related information to the FDA. The details of this self-identification are set in a Guidance for Industry entitled “Self-Identification of Generic Drug Facilities, Sites, and Organizations” published on 22 September 2016 by the FDA in the finalised form.
The Guidance describes the following elements:
1. Which types of generic facilities, sites, and organizations are required to self-identify?
2. What information is requested?
3. What technical standards are to be used for electronically submitting the requested information?
4. What is the penalty for failing to self-identify?
Hereinafter, you will find a short summary of these four topics:
1. Companies that manufacture finished generic medicinal products for human use or the APIs for them, or both are required to self-identify as well as companies that package the finished generic drug into the primary container and label it. Besides, sites that – pursuant to a contract with the applicant (generic drug manufacturer) – repack/redistribute the finished drug from a primary container into a different primary container are also required to submit a self-identification as well as sites that perform bioequivalence/bioavailability studies. Last but not least, the obligation to self-identify also concerns sites that are listed in the application dossier as contract laboratories for the sampling and performing of analytical testing.
2. Essential data are: the D-U-N-S number (a unique nine-digit sequence specific for each site / each distinct physical location of an entity), the “Facility Establishment Identifier, FEI” (an identifier used by the FDA for the planning and tracking of inspections) and general information with regard to the facility (company owner, type of business operation, contact data, information about the manufacture of non generic drugs).
4. Companies that fail to self-identify do not have to expect an explicit penalty. However, such a failure leads to two drawbacks: first, the likelihood of a site inspection by the FDA prior to approval is higher. The second drawback which is much more serious is that all the APIs or finished drugs from a manufacturer who hasn’t self-identified are deemed misbranded. For the FDA, such products are not allowed for importation in the USA.
To the satisfaction of the FDA, the regulations set in the GDUFA and the provisions laid down in the new Guidance represent a major contribution to an enhanced transparency in particular of complex supply chains.
//////////GDUFA, FDA, new Guidance, Self-Identification, Generic Drug Manufacturers
LCMS (method A) tR, 2.01 min, MS Anal. Calcd. for [M+H]+ C23H15ClF3N4: 439.09; found: 439.15.
LC/MS HPLC methods: method A: Column: Phenomenex Luna 30 x 2.0 mm 3um; Mobile Phase A: 10:90 acetonitrile:water with 0.1% TFA; Mobile Phase B: 90:10 acetonitrile:water with 0.1% TFA; Temperature: 40 °C; Gradient: 0-100% B over 2 min; Flow: 1 mL/min.
DETAILS WILL BE UPDATED…………
Image may be NSFW. Clik here to view.Image may be NSFW. Clik here to view.
BMT-145027 is a potent mGluR5 PAM with no inherent mGluR5 agonist activity. BMT-145027 is a non-MPEP site PAM to demonstrate in vivo efficacy. BMT-145027 has mGluR5 PAM EC50 = 47 nM, with fold shit = 3.5, and is effective in mouse NOR. The metabotropic glutamate receptor 5 (mGluR5) is an attractive target for the treatment of schizophrenia due to its role in regulating glutamatergic signaling in association with the N-methyl-D-aspartate receptor (NMDAR).
Image may be NSFW. Clik here to view.
The metabotropic glutamate receptor 5 (mGluR5) is an attractive target for the treatment of schizophrenia due to its role in regulating glutamatergic signaling in association with the N-methyl-d-aspartate receptor (NMDAR). We describe the synthesis of 1H-pyrazolo[3,4-b]pyridines and their utility as mGluR5 positive allosteric modulators (PAMs) without inherent agonist activity. A facile and convergent synthetic route provided access to a structurally diverse set of analogues that contain neither the aryl-acetylene-aryl nor aryl-methyleneoxy-aryl elements, the predominant structural motifs described in the literature. Binding studies suggest that members of our new chemotype do not engage the receptor at the MPEP and CPPHA mGluR5 allosteric sites. SAR studies culminated in the first non-MPEP site PAM, 1H-pyrazolo[3,4-b]pyridine 31 (BMT-145027), to improve cognition in a preclinical rodent model of learning and memory.
Development of 1H-Pyrazolo[3,4-b]pyridines as Metabotropic Glutamate Receptor 5 Positive Allosteric Modulators
Matthew D. Hill*, Haiquan Fang, Jeffrey M. Brown, Thaddeus Molski, Amy Easton, Xiaojun Han, Regina Miller, Melissa Hill-Drzewi, Lizbeth Gallagher, Michele Matchett, Michael Gulianello, Anand Balakrishnan, Robert L. Bertekap, Kenneth S. Santone, Valerie J. Whiterock, Xiaoliang Zhuo, Joanne J. Bronson, John E. Macor, and Andrew P. Degnan
Research and Development, Bristol-Myers Squibb, 5 Research Parkway, Wallingford, Connecticut 06492-7660, United States
ACS Med. Chem. Lett., Article ASAP
DOI: 10.1021/acsmedchemlett.6b00292, http://pubs.acs.org/doi/abs/10.1021/acsmedchemlett.6b00292
CP-456773, also known as MCC950 and CRID3, is a potent and selective cytokine release inhibitor and NLRP3 inflammasome inhibitor for the treatment of inflammatory diseases. CP-456773 inhibits interleukin 1β (IL-1β) secretion and caspase 1 processing. MCC950 blocked canonical and noncanonical NLRP3 activation at nanomolar concentrations. MCC950 specifically inhibited activation of NLRP3 but not the AIM2, NLRC4 or NLRP1 inflammasomes. MCC950 reduced interleukin-1β (IL-1β) production in vivo and attenuated the severity of experimental autoimmune encephalomyelitis (EAE), a disease model of multiple sclerosis. MCC950 is a potential therapeutic for NLRP3-associated syndromes, including autoinflammatory and autoimmune diseases, and a tool for further study of the NLRP3 inflammasome in human health and disease.
MCC950 is an orally bioavailable small molecule inhibitor of the NOD-like receptor pyrin domain-containing protein 3 (NLRP3) inflammasome that exhibits remarkable activity in multiple models of inflammatory disease. Incubation of MCC950 with human liver microsomes, and subsequent analysis by HPLC–MS/MS, revealed a major metabolite, where hydroxylation of MCC950 had occurred on the 1,2,3,5,6,7-hexahydro-s-indacene moiety. Three possible regioisomers were synthesized, and coelution using HPLC–MS/MS confirmed the structure of the metabolite. Further synthesis of individual enantiomers and coelution studies using a chiral column in HPLC–MS/MS showed the metabolite was R-(+)- N-((1-hydroxy-1,2,3,5,6,7-hexahydro-s-indacen-4-yl)carbamoyl)-4-(2-hydroxypropan-2-yl)furan-2-sulfonamide (2a). Incubation of MCC950 with a panel of cytochrome P450 enzymes showed P450s 2A6, 2C9, 2C18, 2C19, 2J2, and 3A4 catalyze the formation of the major metabolite 2a, with a lower level of activity shown by P450s 1A2 and 2B6. All of the synthesized compounds were tested for inhibition of NLRP3-induced production of the pro-inflammatory cytokine IL-1β from human monocyte derived macrophages. The identified metabolite 2a was 170-fold less potent than MCC950, while one regioisomer had nanomolar inhibitory activity. These findings also give first insight into the SAR of the hexahydroindacene moiety.
A novel synthesis of the anti-inflammatory agent 1-(1,2,3,5,6,7- hexahydro-s-indacen-4-yl)-3-[4-(1-hydroxy-1-methyl-ethyl)-furan-2-sulfonyl] urea 1 is described. Sulfonamide 5 was prepared starting from ethyl 3-furoate 2. Key steps were a one-pot sulfonylation with chlorosulfonic acid in methylene chloride followed by pyridinium salt formation and reaction with phosphorus pentachloride to provide ethyl 2-(chlorosulfonyl)-4-furoate 7. This sulfonyl chloride was treated with ammonium bicarbonate to form sulfonamide 8, followed by treatment with excess methyl magnesium chloride to provide 4-(1-hydroxy-1-methyl-ethyl)-furan-2-sulfonamide 5. 4-Isocyanato-1,2,3,5,6,7-hexahydro-s-indacene 16 was prepared from indan in five steps. The formation of the desired sulfonyl urea was carried out both with the isolated isocyanate 16 and via an in situ method.
1-(1,2,3,5,6,7-Hexahydro-s-indacen-4-yl)-3-[4-(1-hydroxy-1-methylethyl)-furan-2-sulfonylurea 1 has been in development for treatment of inflammation. [1] The synthetic route to furan sulfonamide 5 used by its discoverer Mark Dombroski in Medicinal Chemistry is shown in Sch. 1. The starting material was ethyl 3-furoate 2. This was treated with excess methyl magnesium chloride to provide the 3-furanyl-tertiary alcohol 3. Furan alcohol 3 was deprotonated with methyl lithium followed by s-butyl lithium at low temperature and reacted with liquid sulfur dioxide to generate sulfinic acid 4. Without isolation, sulfinic acid 4 was oxidized to sulfonamide 5 with hydroxylamine O-sulfinic acid via a procedure described by workers at Merck.[2] We were interested in finding a synthesis of furan sulfonamide 5 and its conversion to sulfonylurea 1 that would be suitable for scale up. In this article, we describe the discovery of a better bulk process to sulfonamide 5 from the same starting material and a procedure to form the desired sulfonylurea without isolating the isocyanate of 4-amino-1,2,3,5,6,7-hexahydro-s-indacene.
Dr Rebecca Coll, PostDoctoral Researcher, Inflammasome Lab, UQ Fellow
Rebecca completed her PhD research under Prof. Luke O’Neill in Trinity College Dublin at one of the leading laboratories in the innate immunity field. For her work on the regulation of TLR signalling she received the International Endotoxin and Innate Immunity Society Young Investigator Award in 2012. However, her main research focus has been inflammasomes and their therapeutic targeting by small molecule drugs. Her recent first author publication on MCC950 in Nature Medicine has been widely acclaimed (the subject of seven commentaries in leading journals and attention from 24 international news outlets) and is already a highly cited paper. She joined the Schroder group in May 2014 with the goal of defining the molecular target of MCC950 as part of a broader collaboration between the Schroder, Cooper and O’Neill labs.
Pathogenesis of several diseases, including Alzheimer’s, have a strong inflammatory component. Inflammatory processes can be triggered by molecules of the NOD-like receptor (NLR) family such as NLRP3. Once activated, this molecule leads to a cascade of events known as the NLRP3 inflammasome that ultimately causes the production of inflammatory factors. Aberrant activation of NLRP3 is responsible for increased inflammatory responses in complex diseases such as multiple sclerosis, Muckle-Wells syndrome, type 2 diabetes, Alzheimer’s disease and atherosclerosis.
Targeting this molecule can overcome the side effects of other anti-inflammatory drugs commonly used: “Drugs like aspirin or steroids can work in several diseases, but can have side effects or be ineffective. What we have found is a potentially transformative medicine, which targets what appears to be the common disease-causing process in a myriad of inflammatory diseases,” said Luke A J O’Neill, one of the team leaders.
Previous studies identified NLRP3 inhibitors, though neither very potent nor specific. This research team now identified a specific inhibitor of NLRP3 inflammasome, the molecule MCC950. They observed that it inhibits NLRP3 in mouse models of multiple sclerosis with consequent attenuation of disease progression. MCC950 also blocks production of inflammatory factors in blood samples from patients with a severe inflammatory disorder, Muckle-Wells syndrome. These results demonstrated the pharmaceutical potential of this specific NLRP3 inhibitor.
“MCC950 is blocking what was suspected to be a key process in inflammation. There is huge interest in NLRP3 both among medical researchers and pharmaceutical companies and we feel our work makes a significant contribution to the efforts to find new medicines to limit it,” said Rebecca Coll, the paper’s first author.
The researchers were able to demonstrate the potential of MCC950 in multiple sclerosis, an inflammatory disease of the central nervous system (CNS). However, the target for MCC950 is strongly implicated in other diseases of the CNS such as Alzheimer’s and Parkinson’s diseases indicating that it has the potential to treat all of these conditions. The fact that MCC950 can be orally administered further enhances the potential of this molecule as a therapeutic drug.
“MCC950 is able to be given orally and will be cheaper to produce than current protein-based treatments, which are given daily, weekly, or monthly by injection. Importantly, it will also have a shorter duration in the body, allowing clinicians to stop the anti-inflammatory action of the drug if the patient ever needed to switch their immune response back to 100% in order to clear an infection.” said Matt Cooper, chemist and also co-senior author in this study.
REFERENCES
1: Shao BZ, Xu ZQ, Han BZ, Su DF, Liu C. NLRP3 inflammasome and its inhibitors: a review. Front Pharmacol. 2015 Nov 5;6:262. doi: 10.3389/fphar.2015.00262. eCollection 2015. Review. PubMed PMID: 26594174; PubMed Central PMCID: PMC4633676.
2: Baker PJ, Boucher D, Bierschenk D, Tebartz C, Whitney PG, D’Silva DB, Tanzer MC, Monteleone M, Robertson AA, Cooper MA, Alvarez-Diaz S, Herold MJ, Bedoui S, Schroder K, Masters SL. NLRP3 inflammasome activation downstream of cytoplasmic LPS recognition by both caspase-4 and caspase-5. Eur J Immunol. 2015 Oct;45(10):2918-26. doi: 10.1002/eji.201545655. Epub 2015 Aug 24. PubMed PMID: 26173988.
3: Krishnan SM, Dowling JK, Ling YH, Diep H, Chan CT, Ferens D, Kett MM, Pinar A, Samuel CS, Vinh A, Arumugam TV, Hewitson TD, Kemp-Harper BK, Robertson AA, Cooper MA, Latz E, Mansell A, Sobey CG, Drummond GR. Inflammasome activity is essential for one kidney/deoxycorticosterone acetate/salt-induced hypertension in mice. Br J Pharmacol. 2016 Feb;173(4):752-65. doi: 10.1111/bph.13230. Epub 2015 Jul 31. PubMed PMID: 26103560; PubMed Central PMCID: PMC4742291.
4: Groß CJ, Groß O. The Nlrp3 inflammasome admits defeat. Trends Immunol. 2015 Jun;36(6):323-4. doi: 10.1016/j.it.2015.05.001. Epub 2015 May 16. PubMed PMID: 25991463.
5: Coll RC, Robertson AA, Chae JJ, Higgins SC, Muñoz-Planillo R, Inserra MC, Vetter I, Dungan LS, Monks BG, Stutz A, Croker DE, Butler MS, Haneklaus M, Sutton CE, Núñez G, Latz E, Kastner DL, Mills KH, Masters SL, Schroder K, Cooper MA, O’Neill LA. A small-molecule inhibitor of the NLRP3 inflammasome for the treatment of inflammatory diseases. Nat Med. 2015 Mar;21(3):248-55. doi: 10.1038/nm.3806. Epub 2015 Feb 16. PubMed PMID: 25686105; PubMed Central PMCID: PMC4392179.
BMS-906024 is a novel, potent Notch receptor inhibitor . Cancers have a tendency to relapse or to become resistant to treatments that once worked. A family of proteins called Notch is implicated in that resistance and in cancer progression more generally. BMS-906024 is in Phase I clinical trials, both alone and in combination with other agents. Patients with colon, lung, breast, and other cancers are receiving intravenous doses of the compound to determine its safety and optimum dose ranges.
New Phase I drug structure by Bristol-Myers Squibb disclosed at the spring 2013 American Chemical Society meeting in New Orleans to treat breast, lung, and colon cancers and leukemia.[1] The drug works as an pan-Notch inhibitor. The structure is one of a set patented in 2012,[2] and it currently being studied in clinical trials.[3][4]
useful for the treatment of conditions related to the Notch pathway, such as cancer and other proliferative diseases.
Notch signaling has been implicated in a variety of cellular processes, such as cell fate specification, differentiation, proliferation, apoptosis, and angiogenesis. (Bray, Nature Reviews Molecular Cell Biology, 7:678-689 (2006); Fortini, Developmental Cell 16:633-647 (2009)). The Notch proteins are single-pass heterodimeric transmembrane molecules. The Notch family includes 4 receptors, NOTCH 1-4, which become activated upon binding to ligands from the DSL family (Delta-like 1, 3, 4 and Jagged 1 and 2).
The activation and maturation of NOTCH requires a series of processing steps, including a proteolytic cleavage step mediated by gamma secretase, a multiprotein complex containing Presenilin 1 or Presenilin 2, nicastrin, APH1, and PEN2. Once NOTCH is cleaved, NOTCH intracellular domain (NICD) is released from the membrane. The released NICD translocates to the nucleus, where it functions as a transcriptional activator in concert with CSL family members (RBPSUH, “suppressor of hairless”, and LAG1). NOTCH target genes include HES family members, such as HES- 1. HES- 1 functions as transcriptional repressors of genes such as HERP 1 (also known as HEY2), HERP2 (also known as HEY1), and HATH1 (also known as ATOH1).
The aberrant activation of the Notch pathway contributes to tumorigenesis. Activation of Notch signaling has been implicated in the pathogenesis of various solid tumors including ovarian, pancreatic, as well as breast cancer and hematologic tumors such as leukemias, lymphomas, and multiple myeloma. The role of Notch inhibition and its utility in the treatment of various solid and hematological tumors are described in Miele, L. et al, Current Cancer Drug Targets, 6:313-323 (2006); Bolos, V. et al, Endocrine Reviews, 28:339-363 (2007); Shih, I.-M. et al, Cancer Research, 67: 1879- 1882 (2007); Yamaguchi, N. et al., Cancer Research, 68: 1881-1888 (2008); Miele, L., Expert Review Anti-cancer Therapy, 8: 1 197-1201 (2008); Purow, B., Current Pharmaceutical Biotechnology, 10: 154-160 (2009); Nefedova, Y. et al, Drug Resistance Updates, 1 1 :210-218 (2008); Dufraine, J. et al, Oncogene, 27:5132-5137 (2008); and Jun, H.T. et al, Drug Development Research, 69:319-328 (2008).
There remains a need for compounds that are useful as Notch inhibitors and that have sufficient metabolic stability to provide efficacious levels of drug exposure. Further, there remains a need for compounds useful as Notch inhibitors that can be orally or intravenously administered to a patient.
U.S. Patent No. 7,053,084 Bl discloses succinoylamino benzodiazepine compounds useful for treating neurological disorders such as Alzheimer’s Disease. The reference discloses that these succinoylamino benzodiazepine compounds inhibit gamma secretase activity and the processing of amyloid precursor protein linked to the formation of neurological deposits of amyloid protein. The reference does not disclose the use of these compounds in the treatment of proliferative diseases such as cancer.
Applicants have found potent compounds that have activity as Notch inhibitors and have sufficient metabolic stability to provide efficacious levels of drug exposure upon intravenous or oral administration. These compounds are provided to be useful as pharmaceuticals with desirable stability, bioavailability, therapeutic index, and toxicity values that are important to their drugability.
Image may be NSFW. Clik here to view.Image may be NSFW. Clik here to view.
Image may be NSFW. Clik here to view.
PAPER
Image may be NSFW. Clik here to view.
Structure–activity relationships in a series of (2-oxo-1,4-benzodiazepin-3-yl)-succinamides identified highly potent inhibitors of γ-secretase mediated signaling of Notch1/2/3/4 receptors. On the basis of its robust in vivo efficacy at tolerated doses in Notch driven leukemia and solid tumor xenograft models, 12 (BMS-906024) was selected as a candidate for clinical evaluation.
Discovery of Clinical Candidate BMS-906024: A Potent Pan-Notch Inhibitor for the Treatment of Leukemia and Solid Tumors
[00219] To a stirred solution of 5,5,5-trifluoropentanoic acid (5 g, 32.0 mmol) in THF (30 mL) and hexane (30 mL) at 0 °C, was added tert-butyl 2,2,2-trichloroacetimidate (11.46 mL, 64.1 mmol). The mixture was stirred for 15 min at 0 °C. Boron trifluoride etherate (0.406 mL, 3.20 mmol) was added and the reaction mixture was allowed to warm to room temperature overnight. To the clear reaction mixture was added solid aHC03 (5 g) and stirred for 30 min. The mixture was filtered through MgS04 and washed with hexanes (200 mL). The solution was allowed to rest for 45 min, and the resulting solid material was removed by filtering on the same MgS04 filter again, washed with hexanes (100 mL) and concentrated under reduced pressure without heat. The volume was reduced to about 30 mL, filtered through a clean fritted funnel, washed with hexane (5 mL), and then concentrated under reduced pressure without heat. The resulting neat oil was filtered through a 0.45μηι nylon membrane filter disk to provide tert-butyl 5,5,5- trifluoropentanoate (6.6 g, 31.4 mmol 98% yield) as a colorless oil: XH NMR (400 MHz, CDC13) δ ppm 1.38 (s, 9 H) 1.74-1.83 (m, 2 H) 2.00-2.13 (m, 2 H) 2.24 (t, J=7.28 Hz, 2 H).
[00220] To a stirred solution of 5,5,5-trifluoropentanoic acid (5.04 g, 32.3 mmol) in DCM (50 mL) and DMF (3 drops) was added oxalyl chloride (3.4 mL, 38.8 mmol) dropwise over 5 min and the solution was stirred until all bubbling subsided. The reaction mixture was concentrated under reduced pressure to give pale yellow oil. To a separate flask charged with a solution of (45)-4-(propan-2-yl)-l,3-oxazolidin-2-one (4.18 g, 32.4 mmol) in THF (100 mL) at -78 °C was added n-BuLi (2.5M in hexane) (13.0 mL, 32.5 mmol) dropwise via syringe over 5 min. After stirring for 10 min, the above acid chloride dissolved in THF (20 mL) was added via cannula over 15 min. The reaction mixture was warmed to 0 °C, and was allowed to warm to room temperature as the bath warmed and stirred overnight. To the reaction mixture was added saturated NH4CI, and then extracted with EtOAc (2x). The combined organics were washed with brine, dried (Na2S04), filtered and concentrated under reduced pressure. The crude material was purified by flash chromatography (Teledyne ISCO CombiFlash Rf, 5% to 60% solvent A/B=hexanes/EtOAc, REDISEP® S1O2 120g). Concentration of appropriate fractions provided Preparation IB (7.39 g, 86%) as a colorless oil: XH NMR (400 MHz, CDC13) δ ppm 4.44 (1 H, dt, J=8.31, 3.53 Hz), 4.30 (1 H, t, J=8.69 Hz), 4.23 (1 H, dd, J=9.06, 3.02 Hz), 2.98-3.08 (2 H, m), 2.32-2.44 (1 H, m, J=13.91, 7.02, 7.02, 4.03 Hz), 2.13-2.25 (2 H, m), 1.88-2.00 (2 H, m), 0.93 (3 H, d, J=7.05 Hz), 0.88 (3 H, d, J=6.80 Hz). Preparation 1C: (25′,3R)-tert-Butyl 6,6,6-trifluoro-3-((5)-4-isopropyl-2-oxooxazolidine- 3 -carbonyl)-2-(3 ,3,3 -trifluoropropyl)hexanoate, and
[00221] To a cold (-78 °C), stirred solution of diisopropylamine (5.3 mL, 37.2 mmol) in THF (59 mL) under nitrogen atmosphere was added n-BuLi (2.5M in hexane) (14.7 mL, 36.8 mmol), then warmed to 0 °C to give a 0.5M solution of LDA. A separate vessel was charged with Preparation IB (2.45 g, 9.17 mmol), the material was azeotroped twice with benzene (the RotoVap air inlet was fitted with nitrogen inlet to completely exclude humidity) then toluene (15.3 mL) was added. This solution was added to a flask containing dry lithium chloride (1.96 g, 46.2 mmol). To the resultant mixture, cooled to -78 °C, was added LDA solution (21.0 mL, 10.5 mmol) and stirred at -78 °C for 10 min, warmed to 0 °C for 10 min then recooled to -78 °C. To a separate reaction vessel containing Preparation 1A (3.41 g, 16.07 mmol), also azeotroped twice with benzene, was added toluene (15.3 mL), cooled to -78 °C and LDA (37.0 mL, 18.5 mmol) was added, the resulting solution was stirred at -78° for 25 min. At this time the enolate derived from the ester was transferred via cannula into the solution of the oxazolidinone enolate, stirred at -78 °C for an additional 5 min at which time the septum was removed and solid powdered bis(2-ethylhexanoyloxy)copper (9.02 g, 25.8 mmol) was rapidly added to the reaction vessel and the septum replaced. The vessel was immediately removed from the cold bath and immersed into a warm water bath (40 °C) with rapid swirling with a concomitant color change from the initial turquoise to brown. The reaction mixture was stirred for 20 min, was poured into 5% aqueous NH4OH (360 mL) and extracted with EtOAc (2x). The combined organics were washed with brine, dried (Na2S04), filtered and concentrated under reduced pressure. The residue was purified by flash chromatography (Teledyne ISCO CombiFlash Rf, 0% to 60% solvent A/B=hexanes/EtOAc, REDISEP® S1O2 120g). Concentration of appropriate fractions provided Preparation 1C (2.87 g, 66%) as pale yellow viscous oil. XH NMR showed the product was a 1.6: 1 mixture of diastereoisomers 1C: 1D as determined by the integration of the multiplets at 2.74 & 2.84 ppm: XH NMR (400 MHz, CDC13) δ ppm 4.43-4.54 (2 H, m), 4.23-4.35 (5 H, m), 4.01 (1 H, ddd, J=9.54, 6.27, 3.51 Hz), 2.84 (1 H, ddd, J=9.41, 7.28, 3.64 Hz), 2.74 (1 H, ddd, J=10.29, 6.27, 4.02 Hz), 2.37-2.48 (2 H, m, J=10.38, 6.98, 6.98, 3.51, 3.51 Hz), 2.20-2.37 (3 H, m), 1.92-2.20 (8 H, m), 1.64-1.91 (5 H, m), 1.47 (18 H, s), 0.88-0.98 (12 H, m). Preparation IE: (2R,35)-3-(tert-Butoxycarbonyl)-6,6,6-trifluoro-2-(3,3,3- trifluoropropyl)hexanoic acid, and
[00222] To a cool (0 °C), stirred solution of Preparation 1C and ID (4.54 g, 9.51 mmol) in THF (140 mL) and water (42 mL) was sequentially added hydrogen peroxide (30% in water) (10.3 g, 91 mmol) and LiOH (685.3 mg, 28.6 mmol) and the mixture was stirred for 1 hr. At this time the reaction vessel was removed from the cold bath and then stirred for 1.5 hr. The reaction was judged complete by HPLC. To the reaction mixture was added saturated NaHC03 (45 mL) and saturated a2S03(15 mL), and then partially concentrated under reduced pressure. The resulting crude solution was extracted with DCM (3x). The aqueous phase was acidified to pH~l-2 with IN HC1, extracted with DCM (3x) and EtOAc (lx). The combined organics were washed with brine, dried (Na2S04), filtered and concentrated under reduced pressure to provide a mixture of Preparation IE and IF (3.00 g, 86%) as colorless oil: XH NMR (400 MHz, CDC13) δ ppm 2.76-2.84 (1 H, m, diastereoisomer 2), 2.64-2.76 (3 H, m), 2.04-2.35 (8 H, m), 1.88-2.00 (4 H, m), 1.71-1.83 (4 H, m), 1.48 (9 H, s, diastereoisomer 1), 1.46 (9 H, s, diastereoisomer 2); XH NMR showed a 1.7: 1 mixture of 1E: 1F by integration of the peaks for the ?-butyl groups.
Preparation IE: (2R,35)-3-(tert-Butoxycarbonyl)-6,6,6-trifluoro-2-(3,3,3- trifluoropropyl)hexanoic acid, and
[00223] To a cold (-78 °C), stirred solution of diisopropylamine (1.7 mL, 11.93 mmol) in THF (19 mL) under nitrogen atmosphere was added n-BuLi (2.5M in hexanes) (4.8 mL, 12.00 mmol). The mixture was stirred for 5 min and then warmed to 0 °C. In a separate vessel, to a cold (-78 °C) stirred solution of the mixture of Preparation IE and IF (1.99 g, 5.43 mmol) in THF (18 mL) was added the LDA solution prepared above via cannula slowly over 25 min. The mixture was stirred for 15 min, then warmed to room temperature (placed in a 24 °C water bath) for 15 min, and then again cooled to -78 °C for 15 min. To the reaction mixture was added Et2AlCl (1M in hexane) (11.4 mL, 1 1.40 mmol) via syringe, stirred for 10 min, warmed to room temperature for 15 min and then cooled back to -78 °C for 15 min. Methanol (25 mL) was rapidly added, swirled vigorously while warming to room temperature, then concentrated to ~l/4 original volume. The mixture was dissolved in EtOAc and washed with IN HCl (50 mL) and ice (75 g). The aqueous phase was separated, extracted with EtOAc (2x). The combined organics were washed with a mixture of KF (2.85g in 75 mL water) and IN HCl (13 mL) [resulting solution pH 3-4], then with brine, dried (Na2S04), filtered and concentrated under reduced pressure to give a 9: 1 (IE: IF) enriched diastereoisomeric mixture (as determined by XH NMR) of Preparation IE and Preparation IF (2.13 g, >99%) as a pale yellow viscous oil: XH NMR (400 MHz, CDC13) δ ppm 2.64-2.76 (2 H, m), 2.04-2.35 (4 H, m), 1.88-2.00 (2 H, m), 1.71-1.83 (2 H, m), 1.48 (9 H, s). Preparation 1 G: (35)-3 -Amino- 1 -methyl-5-phenyl- 1 ,3 -dihydro-2H- 1 ,4-benzodiazepin-2- one, and
[00224] Racemic 3-amino-l-methyl-5-phenyl-l,3-dihydro-2H-l,4-benzodiazepin-2- one (Rittle, K.E. et al, Tetrahedron Letters, 28(5):521-522 (1987)) was prepared according to the literature procedure. The enantiomers were separated under chiral-SFC conditions using the following method: CHIRALPAK® AS-H 5×25; Mobile phase: 30% MeOH+ 0.1% DEA in C02; Flow rate: 280 mL/min; Pressure: 100 bar; Temperature: 35 °C.
[00225] Obtained the S-enantiomer (Preparation 1G): HPLC: RT=1.75 min (30% MeOH + 0.1% DEA in C02 on CHIRALPAK® AS-H 4.6×250 mm, 3 mL/min, 35 °C, 100 bar, 230 nm, ΙΟμΙ injection); ¾ NMR (400 MHz, CDC13) δ ppm 7.58-7.63 (2 H, m), 7.55 (1 H, ddd, J=8.50, 7.1 1, 1.76 Hz), 7.40-7.47 (1 H, m), 7.34-7.40 (3 H, m), 7.31 (1 H, dd, J=7.81, 1.51 Hz), 7.14-7.22 (1 H, m), 4.46 (1 H, s), 3.44 (3 H, s), 3.42 (2 H, s); [a]D= -155° (c=1.9, MeOH) (Lit. Rittle, K.E. et al, Tetrahedron Letters, 28(5):521-522 (1987): [a]D=-236°).
[00226] Also obtained the R-enantiomer (Preparation 1H): HPLC: RT=1.71 min; [a]D=+165° (c=2.1, MeOH) (Lit [a]D= +227°).
Alternate procedure to make Preparation 1 G:
Preparation 1G»CSA salt: (35)-3-Amino-l-methyl-5-phenyl-l,3-dihydro-2H-l,4- benzodiazepin-2-one, (15)-(+)-10-camphorsulfonic acid salt
[00227] Preparation lG’CSA was prepared from racemic 3-amino-l-methyl-5-phenyl- l,3-dihydro-2H-l,4-benzodiazepin-2-one (9.98g, 37.6 mmol) (prepared according to the literature as shown above) according to the literature procedure (Reider, P.J. et al, J. Org. Chem., 52:955-957 (1987)). Preparation lG’CSA (16.91g, 99%) was obtained as a colorless solid: Optical Rotation: [a]D = -26.99° (c=l, H20) (Lit. [a]D = -27.8° (c=l,
[00228] To a stirred solution of Preparation 1G (1.45 g, 5.47 mmol) and a 9: 1 mixture of Preparation IE and IF (1.989 g, 5.43 mmol) in DMF (19 mL) was added O- benzotriazol-l-yl-N,N,N’,N’-tetra-methyluronium tetrafluoroborate (1.79 g, 5.57 mmol) and triethylamine (3.0 mL, 21.52 mmol) and stirred overnight. The reaction was judged complete by LCMS. The reaction mixture was poured into water (125 mL) and the precipitated solid was collected by filtration, washed with water and air dried to provide an 8: 1 mixture of Preparation II and Preparation 1J (2.95 g, 89%) as a cream solid: MS (ES): m/z= 614 [M+H]+;XH NMR (400 MHz, CDC13) δ ppm 7.55-7.65 (3 H, m), 7.44- 7.52 (2 H, m), 7.35-7.45 (4 H, m), 5.52 (1 H, d, J=8.03 Hz), 3.48 (3 H, s), 2.63 (2 H, ddd, J=9.35, 3.95, 3.76 Hz), 2.14-2.25 (4 H, m), 1.90-2.03 (3 H, m), 1.69-1.82 (1 H, m), 1.51 (9 H, s).
Preparation IK: (25,,3R)-6,6,6-Trifluoro-3-(((35)-l-methyl-2-oxo-5-phenyl-2,3-dihydro- lH-l,4-benzodiazepin-3-yl)carbamoyl)-2-(3,3,3-trifluoropropyl)hexanoic acid, and
[00229] To a cool (0 °C), stirred solution of the above mixture of Preparation II and Preparation 1 J (2.95 g, 4.81 mmol) in DCM (20 mL) was added TFA (20 mL, 260 mmol). The reaction mixture was stirred for lhr, then allowed to warm to room temperature and stirred for 2.5 hr. The reaction was judged complete by LCMS. The reaction mixture was diluted with toluene (50 mL) and concentrated under reduced pressure. The residue mixture was redissolved in toluene (50 mL) and concentrated under reduced pressure then dried under high vacuum. The crude product was dissolved in DCM, S1O2 (15g) was added, concentrated, then was purified by flash chromatography (Teledyne ISCO CombiFlash Rf, 0% to 45% solvent A/B=DCM/EtOAc, REDISEP® S1O2 80g). Concentration of appropriate fractions provided a mixture of Preparation IK and Preparation 1L (2.00 g, 75%) as a cream solid: HPLC: RT=2.770 min
(CHROMOLITH® SpeedROD 4.6 x 50 mm (4 min grad) eluting with 10-90% aqueous MeOH over 4 minutes containing 0.1% TFA, 4 mL/min, monitoring at 254 nm); MS (ES): m/z= 558 [M+H]+; XH NMR (400 MHz, CDC13) δ ppm 8.32 (1 H, d, J=8.03 Hz), 7.65-7.71 (1 H, m), 7.50-7.60 (3 H, m), 7.41-7.49 (2 H, m), 7.39 (1 H, dd, J=7.91, 1.63 Hz), 7.23-7.35 (2 H, m), 5.59 (1 H, d, J=8.03 Hz), 3.51 (3 H, s), 2.81 (1 H, ddd, J=10.54, 6.90, 3.64 Hz), 2.67-2.76 (1 H, m), 2.22-2.33 (3 H, m), 1.99-2.12 (3 H, m), 1.85-1.94 (1 H, m), 1.79 (1 H, ddd, J=13.87, 7.84, 3.64 Hz). Example 1 :
[00230] To a stirred solution of an 8: 1 mixture of Preparation IK and Preparation 1L (3.46 g, 6.21 mmol) in DMF (25 mL) under nitrogen atmosphere was added ammonium chloride (3.32 g, 62.1 mmol), EDC (3.55 g, 18.52 mmol), HOBT (2.85 g, 18.61 mmol), and triethyl amine (16 mL, 1 15 mmol) and stirred overnight. The reaction was judged complete by LCMS. The reaction mixture was poured into water (200 mL) with vigorous swirling and then allowed to sit. The solid was collected by filtration, washed with water, allowed to dry to afford 3.6 g colorless solid. The solid was purified by preparative SFC chromatography (Lux-Cellulose-2 (3x25cm), 8% methanol in CO2, 140ml/min @220nm and 35 °C; Sample: 3.6g in 50cc methanol, conc.=70mg/ml, Stack injection:
0.5cc/9.2min). Fractions containing product were concentrated, dried overnight under vacuum. Obtained Example 1 (2.74 g, 79%) as a colorless solid (Crystal Form -1): HPLC: RT=9.601 min (H20/CH3CN with TFA, Sunfire CI 8 3.5um, 4.6x150mm, 4.6x150mm, gradient = 15 min, wavelength = 220 and 254 nm). MS (ES): m/z= 557 [M+H]+; XH NMR (400 MHz, DMSO-d6) δ ppm 9.54 (1 H, d, J=7.28 Hz), 7.71-7.80 (1 H, m), 7.68 (2 H, d, J=8.78 Hz), 7.50-7.62 (3 H, m), 7.45 (2 H, t, J=7.28 Hz), 7.29-7.40 (2 H, m), 7.15 (1 H, br. s.), 5.30 (1 H, d, J=7.28 Hz), 3.39 (3 H, s), 2.74-2.86 (1 H, m), 2.02-2.32 (3 H, m), 1.45-1.79 (4 H, m); [a]D = -107.0° (5.73 mg/mL, DMSO).
[00231] Crystal Form A-2 was prepared by adding approximately 1 mg of Example 1 to approximately 0.7 mL of acetone/acetonitrile/water solution (2:2: 1). A mixture of colorless needles and thin blades crystals were obtained after one day of slow evaporation of the solution at room temperature. The thin blade crystals were separated to provide crystal Form A-2.
[00232] Crystal Form EA-3 was prepared by adding approximately 1 mg of Example 1 to approximately 0.7 mL of ethyl acetate/heptane solution (1 : 1). Colorless blade crystals were obtained after three days of slow evaporation of the solution at room temperature.
[00233] Crystal Form THF-2 was obtained by adding approximately 5 mg of Example 1 to approximately 0.7 mL of THF/water solution (4: 1). Colorless blade-like crystals were obtained after one day of solvent evaporation at room temperature.
Alternate Procedure to Make Example 1 : Preparation 1M: 3,3,3-Trifluoropropyl trifluoromethanesulfonate
[00234] To a cold (-25 °C), stirred solution of 2,6-lutidine (18.38 mL, 158 mmol) in CH2CI2 (120 mL) was added Tf20 (24.88 mL, 147 mmol) over 3 min, and stirred for 5 min. To the reaction mixture was added 3,3,3-trifluoropropan-l-ol (12 g, 105 mmol) over an interval of 3 min. After 2 hr, the reaction mixture was warmed to room temperature and stirred for 1 hr. The reaction mixture was concentrated to half volume, then purified by loading directly on silica gel column (330g ISCO) and eluted with CH2C12. Obtained Preparation 1M (13.74 g, 53%) as a colorless oil. XH NMR (400 MHz, CDCI3) δ ppm 4.71 (2 H, t, J=6.15 Hz), 2.49-2.86 (2 H, m).
[00235] Preparation IN was prepared from 5,5,5-trifluoropentanoic acid (3.35 g, 21.46 mmol) and (45)-4-benzyl-l,3-oxazolidin-2-one (3.80 g, 21.46 mmol) by the general methods shown for Preparation IB. Preparation IN (5.67 g, 84%) was obtained as a colorless viscous oil: XH NMR (400 MHz, CDC13) δ ppm 7.32-7.39 (2 H, m), 7.30 (1 H, d, J=7.05 Hz), 7.18-7.25 (2 H, m), 4.64-4.74 (1 H, m), 4.17-4.27 (2 H, m), 3.31 (1 H, dd, J=13.35, 3.27 Hz), 3.00-3.1 1 (2 H, m), 2.79 (1 H, dd, J=13.35, 9.57 Hz), 2.16-2.28 (2 H, m), 1.93-2.04 (2 H, m).
[00236] To a cold (-78 °C), stirred solution of Preparation IN (3.03 g, 9.61 mmol) in THF (20 mL) was added NaHMDS (1.0M in THF) (10.6 mL, 10.60 mmol) under nitrogen atmosphere. After 2 hours, tert-butyl 2-bromoacetate (5.62 g, 28.8 mmol) was added neat via syringe at -78 °C and stirring was maintained at the same temperature. After 6 hours, the reaction mixture was warmed to room temperature. The reaction mixture was partitioned between saturated NH4C1 and EtOAc. The organic phase was separated, and the aqueous was extracted with EtOAc (3x). The combined organics were washed with brine, dried (Na2S04), filtered and concentrated under reduced pressure. The residue was purified by flash chromatography (Teledyne ISCO CombiFlash Rf, 5% to 100% solvent A/B=hexanes/EtO Ac, REDISEP® Si02 120g). Concentration of appropriate fractions provided Preparation 10 (2.79 g, 67.6%) as a colorless viscous oil: XH NMR (400 MHz, CDC13) δ ppm 7.34 (2 H, d, J=7.30 Hz), 7.24-7.32 (3 H, m), 4.62- 4.75 (1 H, m, J=10.17, 6.89, 3.43, 3.43 Hz), 4.15-4.25 (3 H, m), 3.35 (1 H, dd, J=13.60, 3.27 Hz), 2.84 (1 H, dd, J=16.62, 9.57 Hz), 2.75 (1 H, dd, J=13.35, 10.07 Hz), 2.47 (1 H, dd, J=16.62, 4.78 Hz), 2.1 1-2.23 (2 H, m), 1.90-2.02 (1 H, m), 1.72-1.84 (1 H, m), 1.44 (9 H, s). -2-(2-tert-Butoxy-2-oxoethyl)-5,5,5-trifluoropentanoic acid
[00237] Preparation IP was prepared from Preparation 10 (2.79 g, 6.50 mmol) by the general methods shown for Preparation IE. Preparation IP (1.45 g, 83%) was obtained as a colorless oil: XH NMR (400 MHz, CDC13) δ ppm 2.83-2.95 (1 H, m), 2.62-2.74 (1 H, m), 2.45 (1 H, dd, J=16.62, 5.79 Hz), 2.15-2.27 (2 H, m), 1.88-2.00 (1 H, m), 1.75-1.88 (1 H, m), 1.45 (9 H, s). Preparation IE: (2R,35′)-3-(tert-Butoxycarbonyl)-6,6,6-trifluoro-2-(3,3,3- trifluoropropyl)hexanoic acid, and
[00238] To a cold (-78 °C), stirred solution of Preparation IP (5.44 g, 20.13 mmol) in THF (60 mL) was slowly added LDA (24.60 mL, 44.3 mmol) over 7 min. After stirring for 2 hr, Preparation 1M (6.44 g, 26.2 mmol) was added to the reaction mixture over 3 min. After 45 min, the reaction mixture was warmed to -25 °C bath (ice/MeOH/dry ice) for 1 hr, and then warmed to 0 °C. After 45 min, Preparation 1M (lg) was added and the reaction mixture was stirred for 20 min. The reaction was quenched with water and IN NaOH and was extracted with (¾(¾. The organic layer was again extracted with IN NaOH (2x) and the aqueous layers were combined. The aqueous layer was cooled in ice/water bath and then acidified with concentrated HCl to pH 2. Next, the aqueous layer was extracted with EtOAc. The combined organics were washed with brine, dried over anhydrous sodium sulphate, and concentrated under reduced pressure. The residue was dried under high vacuum to provide a 1 :5 (IE: IF) mixture (as determined by XH NMR) of Preparation IE and Preparation IF (5.925 g, 80%) as a pale yellow solid. XH NMR (500 MHz, CDC13) 8 ppm 2.81 (1 H, ddd, J=10.17, 6.32, 3.85 Hz), 2.63-2.76 (1 H, m), 2.02- 2.33 (4 H, m), 1.86-1.99 (2 H, m), 1.68-1.85 (2 H, m), 1.47 (9 H, s).
Preparation IE: (2R,35)-3-(tert-Butoxycarbonyl)-6,6,6-trifluoro-2-(3,3,3- trifluoropropyl)hexanoic acid, and
[00239] A mixture of Preparation IE and Preparation IF (64 mg, 1.758 mmol) was taken in THF (6 mL) to give a colorless solution which was cooled to -78 °C. Then, LDA (2.149 mL, 3.87 mmol) (1.8M in heptane/THF/ethylbenzene) was slowly added to the reaction mixture over 10 min. After stirring for 15 min the reaction mixture was placed in a room temperature water bath. After 15 min the reaction mixture was placed back in -78 °C bath and then diethylaluminum chloride (3.87 mL, 3.87 mmol) (1M in hexane) was added slowly over 5 min. The reaction mixture was stirred at -78 °C. After 15 min the reaction mixture was placed in a room temperature water bath for 10 min and then cooled back to -78 °C bath. After 15 min the reaction was quenched with MeOH (8 mL, 198 mmol), removed from the -78 °C bath and concentrated. To the reaction mixture was added ice and HC1 (16 mL, 16.00 mmol), followed by extraction with EtOAc (2x). The organic layer was washed with potassium fluoride (920 mg, 15.84 mmol) (in 25 mL FLO) and HC1 (4.5 mL, 4.50 mmol). The organics were dried over anhydrous magnesium sulphate and concentrated under reduced pressure to provide a 9: 1 (IE: IF) enriched mixture of Preparation IE and Preparation IF (540 mg, 1.583 mmol, 90% yield) as light yellow/orange solid. ¾ NMR (400 MHz, CDC13) δ ppm 2.64-2.76 (2 H, m), 2.04-2.35 (4 H, m), 1.88-2.00 (2 H, m), 1.71-1.83 (2 H, m), 1.48 (9 H, s). It was converted to Example 1 by the sequence of reactions as outlined above.
(1Q) [00240] A clean and dry 5 L four neck round bottom flask equipped with mechanical stirring, thermometer socket and nitrogen bubbler at room temperature was charged with Ν,Ν-dimethyl formamide (2.07 L), a 1.2: 1 mixture of Preparation IE and Preparation IF (207 g, 0.5651 moles), potassium carbonate (1 17.1 g, 0.8476 moles) followed by benzyl bromide (116 g, 0.6781 moles) over 15-20 min. The reaction mixture was stirred for 2-3 hr. After completion of the reaction, the reaction mixture was concentrated to dryness at 50-55 °C under vacuum. Ethyl acetate (3.1 L, 30 Vol.) was charged into the concentrated reaction mass and then washed with water (2.07 L), brine (0.6 L) then dried over anhydrous sodium sulfate (207 g), filtered and concentrated to dryness at 40-45 °C under vacuum. The residue was dissolved in dichloromethane (1.035 L, 5 vol.) and then absorbed onto silica gel (60-120) (607 g, 3.0 w/w), then was purified with column chromatography using petroleum ether and ethyl acetate as solvents. After pooling several batches, Preparation 1Q (235 g) was obtained. HPLC purity: 99.77%, Preparation IE: (2R,35)-3-(tert-Butoxycarbonyl)-6,6,6-trifluoro-2-(3,3,3- trifluoropropyl)hexanoic acid
[00241] A clean and dry 2 L autoclave was charged with methanol (540 mL) and was purged with nitrogen for 5-10 minutes. To the autoclave was added 10% palladium on carbon (12 g, 20%), purged with nitrogen once again for 5-10 min then was charged with Preparation 1Q (60g, 0.1315 moles), the autoclave was flushed with methanol (60mL) and stirred for 4-6 hr at 20-25 °C under 5Kg hydrogen pressure. After completion of the reaction, the reaction mass was filtered through CELITE®, washed with methanol (180 mL), dried with anhydrous sodium sulfate (60 g), filtered and concentrated to dryness at 45-50 °C under vacuum. Obtained Preparation IE (45.8 g, 95%) as a colorless solid: HPLC purity: 98.9%.
Alternate procedure to make Preparation IE: Preparation IE: (2R,35)-3-(te^Butoxycarbonyl)-6,6,6-trifluoro-2-(3,3,3- trifluoropropyl)hexanoic acid
[00242] Preparation IE was prepared in a procedure identical as above from a mixture of Preparations IE and IF (200g, 0.5460 moles) using LDA (1.8 M solution in THF, ethyl benzene and heptane) (698mL, 2.3equiv.) and diethyl aluminum chloride (1.0 M solution in hexane) (1256mL, 2.3equiv) in THF (2.0L). After workup as explained above, the resulting residue was treated as follows: The crude material was added to a 2L four neck round bottom flask, followed by the addition of MTBE (1.0L) charged below 30 °C. The resulting mixture was stirred for 5-10 minutes to obtain a clear solution.
Hexanes (600mL) was charged to the reaction mixture at a temperature below 30 °C. The reaction mixture was stirred for 10 min. Next, tert-butylamine (43.8g, l. leq) was charged slowly over a period of 15 minutes below 30 °C. This addition was observed to be exothermic. The reaction mixture was stirred for 2 hrs below 30 °C and filtered. The solid material was washed with 5:3 MTBE: hexane (200mL), the filtrate was
concentrated and transferred to an amber color bottle. The filtered solid was dissolved in dichloromethane (2.0L), washed with IN HC1 (2.0), the organic layer was washed with brine (1.0L x 2), then was concentrated under reduced pressure below 45 °C. This material was found to be 91.12% pure. The material was repurified by the above t- butylamine crystallization purification procedure. Obtained Preparation IE (78 g, 39%): HPLC purity: 99.54%.
[00243] A clean and dry 2 L four neck round bottom flask equipped with mechanical stirring, thermometer socket and nitrogen bubbler was charged with N,N- dimethylformamide (457 mL), Preparation IE (45.7g, 0.1248moles) and Preparation lG’CSA (62.08g, 0.1248moles) under nitrogen atmosphere at 20-25 °C. The reaction mixture was stirred for 15-20 minutes to make clear solution at 20-25 °C. To the reaction mixture was added TBTU (48.16g, 0.1498 moles) at 20-25 °C followed by triethylamine (50.51g, 0.4992 moles) over 15-20 minutes at 20-25 °C. The reaction mixture was stirred for 60-120 minutes at 20-25 °C under nitrogen atmosphere. After completion of the reaction, the reaction was quenched into water (1.37L, 30 Vol.) at 20-25 °C under stirring. The reaction mixture was stirred for 30 minutes at 20-25 °C. The reaction mixture was filtered and washed with water (228 mL). The resulting solid material was dissolved in ethyl acetate (457 mL), washed with water (2×137 mL), brine (137 mL), and then dried with anhydrous sodium sulfate (45.7g). Activated charcoal (9.14 g, 20%) was charged into the reaction mixture and stirred for 30 minutes. The mixture was filtered through CELITE® bed and 1 micron filter cloth, washed charcoal bed with ethyl acetate (137 mL), concentrated to 1.0 Vol. stage and then petroleum ether (457 mL, 10 Vol.) was charged and stirred for 30 minutes at 20-25 °C. The solid was collected by filtration, washed with petroleum ether (137 mL) and then dried under vacuum at 40-45 °C for 8 hr until loss on drying was less than 3.0%. Obtained Preparation II (65.2 g, 85%): HPLC purity: 98.26%.
[00244] A clean and dry 3 L four neck round bottom flask equipped with mechanical stirring, thermometer socket and nitrogen bubbler was charged with dichloromethane (980 mL) under nitrogen atmosphere followed by Preparation II (140 g, 0.2282 moles) at 20-25 °C. The reaction mixture was cooled to 0-5 °C and trifluoroacetic acid (980 mL) was charged slowly for 30-40 minutes. The resulting mixture was stirred for 2 hr at 0-5 °C under nitrogen atmosphere. The reaction temperature was raised to 20 to 25 °C, and the reaction mixture was stirred for 1-2 hr at 20 to 25 °C. After completion of the reaction, the reaction mixture was concentrated to dryness at 50 to 55 °C under vacuum. Toluene (3×700 mL,) was charged into the concentrated reaction mass, and then distilled off at 50 to 55 °C under vacuum. After complete concentration from toluene, ethyl acetate (280 mL) was charged into the reaction mass at 20 to 25 °C, stirred for 60 minutes, then the solid was collected by filtration, washed with ethyl acetate (140 mL), dried under vacuum at 50 to 55 °C for 12 hr until loss on drying was less than 2.0%. Obtained Preparation IK (106 g, 84%): HPLC purity: 98.43%.
Example 1 :
[00245] A reaction vessel was charged with Preparation IK (30 g, 53.81 mmol), HOBt (8.7g, 64.38 mmol), and THF (150 mL) at room temperature. To the homogeneous solution was added EDCI (12.4g, 64.68 mmol), stirred for 15 min, then cooled to 8 °C. To the reaction mixture was added ammonia (2M in IP A) (81 mL, 162 mmol) over 5 min so as to maintain a temperature below 10 °C. The resulting heavy slurry was stirred for 10 min, warmed to room temperature over 30 min, then stirred for 4 hr. At the completion of the reaction, water (230 mL) was slowly added over 15 min to maintain a temperature below 20 °C, and then stirred for 2 hr. The solid was collected by filtration, washed with water (3X60 mL), then dried under vacuum 48 hr at 55 °C. The above crude product was charged into a 1 L 3 -necked round flask. IP A (200 mL) was added, then heated to 80 °C resulting in a homogeneous solution. Water (170 mL) was slowly added (15 min) to maintain an internal temperature >75 °C. The resulting slurry was stirred and cooled to room temperature for 2 hr. The solid was collected by filtration, washed with water (2 X 50 mL), then dried under vacuum (55 °C for 24 h, and 30 °C for 48 h).
Obtained Example 1 (23.4 g, 78% yield): HPLC purity: 99.43%.
[00246] Racemic 3-amino-5-phenyl-l,3-dihydro-2H-l,4-benzodiazepin-2-one (J. Med. Chem., 49:231 1-2319 (2006), compound# 5) was prepared according to the literature procedure. The enantiomers were separated on Berger SFC MGIII Column: Lux 25X3 cm, 5cm; Mobile phase: 30% MeOH+ 0.1% DEA in C02; Flow rate: 150 mL/min;
Temperature: 40 °C; Detector wavelength: 250 nM. Obtained the S-enantiomer
Preparation 2A as a white solid: XH NMR (400 MHz, DMSO-d6) δ ppm 10.67 (1 H, br. s.), 7.58 (1 H, td, J=7.65, 1.76 Hz), 7.37-7.53 (5 H, m), 7.23-7.30 (2 H, m), 7.14-7.22 (1 H, m), 4.23 (1 H, s), 2.60 (2 H, br. s.); HPLC: RT=3.0625 min (30% MeOH + 0.1% DEA in C02 on OD-H Column, 3 mL/min, 35 °C, 96 bar, 230 nm, ΙΟμΙ inj); [a]D = -208.3° (5.05 mg/niL, MeOH). Also obtained the R-enantiomer Preparation 2B as an off white solid: HPLC: RT=3.970 min; [a]D = 182.1° (2.01 mg/mL, MeOH).
[00247] Preparation 2C was prepared from Preparation 2A (564 mg, 2.244 mmol) and a mixture of Preparation IE and Preparation IF (822 mg, 2.244 mmol) according to the general procedure shown for Preparation II. Obtained Preparation 2C and Preparation 2D (1.31 g, 97%): HPLC: RT=3.443 min (CHROMOLITH® ODS 4.6 x 50 mm (4 min grad) eluting with 10-90% aqueous MeOH over 4 minutes containing 0.% TFA, 4 mL/min, monitoring at 220 nm); MS (ES): m/z= 600.3 [M+H]+.
Preparation 2E: (25′,3R)-6,6,6-Trifluoro-3-(((35)-2-oxo-5-phenyl-2,3-dihydro-lH-l,4- benzodiazepin-3-yl)carbamoyl)-2-(3,3,3-trifluoropropyl)hexanoic acid, and
(2E) (2F) [00248] A mixture of Preparation 2E and Preparation 2F was prepared from a mixture of Preparation 2C and Preparation 2D (1.3 lg, 2.185 mmol) by the general methods shown for Preparation IK. Obtained a mixture of Preparation 2E and Preparation 2F (1.18 g, 99%): HPLC: RT=2.885 min (CHROMOLITH® ODS 4.6 x 50 mm (4 min grad) eluting with 10-90% aqueous MeOH over 4 minutes containing 0.% TFA, 4 mL/min, monitoring at 220 nm). MS (ES): m/z= 544.2 [M+H]+.
Example 2:
[00249] Example 2 was prepared from a mixture of Preparation 2E and Preparation 2F (354 mg, 0.651 mmol) by the general methods shown for Example 1. After separation of the diastereoisomers, Example 2 was obtained (188 mg, 52%) as a white solid: HPLC: RT=9.063 min (H20/CH3CN with TFA, Sunfire C18 3.5um, 4.6x150mm, 4.6x150mm, gradient = 15 min, wavelength = 220 and 254 nm); MS (ES): m/z= 543 [M+H]+; XH NMR (400 MHz, DMSO-d6) δ ppm 10.87 (1 H, br. s.), 9.50-9.55 (1 H, m), 7.62-7.69 (2 H, m), 7.40-7.57 (5 H, m), 7.29-7.36 (2 H, m), 7.22-7.28 (1 H, m), 7.16 (1 H, br. s.), 5.25 (1 H, d), 3.30-3.32 (1 H, m), 2.75-2.86 (1 H, m), 2.44-2.48 (1 H, m), 2.06-2.34 (3 H, m), 1.51- 1.77 (4 H, m); [a]D = -114.4° (8.04 mg/mL, DMSO).
[00250] Crystal Form M2- 1 was prepared by adding approximately 1 mg of Example 2 to approximately 0.7 mL of MeOH/fluorobenzene solution (3 : 1). Colorless plate-like crystals were obtained after 2 days of solvent evaporation at room temperature.
Clip
For some disease targets, an indirect approach may be best. Or so Ashvinikumar V. Gavai and his colleagues atBristol-Myers Squibbfound in their quest toward a potential cancer drug. Gavai unveiled BMS-906024, which is an experimental—and slightly roundabout—treatment for a number of cancers, including breast, lung, and colon cancers, and leukemia.
Cancers have a tendency to relapse or to become resistant to treatments that once worked. Research at BMS and elsewhere had suggested that a family of proteins called Notch is implicated in that resistance and in cancer progression more generally. Gavai, director of oncology chemistry at BMS in Princeton, N.J., and his team set out to block Notch family signaling.
Notch family members lack enzymatic activity, so blocking them directly is difficult. Instead, BMS developed inhibitors of an enzyme that is essential for activating Notch signaling—γ-secretase.
Image may be NSFW. Clik here to view.
Company: Bristol-Myers Squibb
Target: pan-Notch
Disease: breast, lung, colon cancer; leukemia
Interfering with Notch, even in this indirect way, can have detrimental effects on the gastrointestinal tract. Only two of the four Notch family members are linked to that side effect, Gavai says. But he and his team think their drug will be most effective if it acts on all four family members roughly equally—a so-called pan-Notch inhibitor. By selecting a molecule that’s well tolerated in animals and carefully scheduling doses of the drug in humans, it could be possible to minimize side effects, he says.
The BMS team relied on Notch signaling assays in leukemia and breast cancer cell lines to find leads. They soon learned that for their molecules to work, three chiral centers had to be in the S,R,Sconfiguration. After that, they strove to make the molecules last in the bloodstream. They removed an isobutyl group and tweaked some other parts of their candidate’s succinamide side chain. It was tough to retain both a long half-life and activity against Notch, Gavai told C&EN. “You’d optimize one and lose the other.”
His team threaded the needle with BMS-906024. Their studies with mice suggest that a dose of 4–6 mg once a week could be effective in people. That’s lower than doses being tested for other Notch-targeted agents, according to the website clinicaltrials.gov. The mouse studies also back the idea that Notch is involved in cancer drug resistance and suggest that Notch could be a target for taking on cancer stem cells, which are notoriously resistant to chemotherapy.
BMS-906024 is in Phase I clinical trials, both alone and in combination with other agents. Patients with colon, lung, breast, and other cancers are receiving intravenous doses of the compound to determine its safety and optimum dose ranges.
Image may be NSFW. Clik here to view.
(From left, front row) Gavai, Weifeng Shan, (second row) Aaron Balog, Patrice Gill, Gregory Vite, (third row) Francis Lee, Claude Quesnelle, (rear row) Wen-Ching Han, Richard Westhouse.
BMS-906024
Company: Bristol-Myers Squibb
Meant to treat: cancers including breast, lung, colon, and leukemia
Mode of action: pan-Notch inhibitor
Medicinal chemistry tidbit: The BMS team used an oxidative enolate heterocoupling en route to the candidate– a procedure from Phil Baran’s lab at Scripps Research Institute. JACS 130, 11546
Status in the pipeline: Phase I
Relevant documents: WO 2012/129353
PAPER
Image may be NSFW. Clik here to view.
An enantioselective synthesis of (S)-7-amino-5H,7H-dibenzo[b,d]azepin-6-one (S–1) is described. The key step in the sequence involved crystallization-induced dynamic resolution (CIDR) of compound 7 using Boc-d-phenylalanine as a chiral resolving agent and 3,5-dichlorosalicylaldehyde as a racemization catalyst to afford S–1 in 81% overall yield with 98.5% enantiomeric excess.
Crystallization-Induced Dynamic Resolution toward the Synthesis of (S)-7-Amino-5H,7H-dibenzo[b,d]-azepin-6-one: An Important Scaffold for γ-Secretase Inhibitors
† Department of Discovery Synthesis, Biocon Bristol-Myers Squibb Research Centre, Biocon Park, Bommasandra IV Phase, Jigani Link Road, Bengaluru 560099, India
‡Bristol-Myers Squibb Company, P.O Box 4000, Princeton, New Jersey 08543-4000, United States
1. Quesnelle, Claude; Kim, Soong-Hoon; Lee, Francis; Gavai, Ashvinikumar. Bis(fluoroalkyl)-1,4-benzodiazepinone compounds as Notch receptor inhibitors and their preparation and use in the treatment of cancer. PCT Int. Appl. (2012), WO 2012129353 A1 20120927.
Mechanism of Action Amyloid precursor protein secretase inhibitors; Notch signalling pathway inhibitors
Phase I Solid tumours
Most Recent Events
30 Aug 2016Bristol-Myers Squibb terminates a phase I trial for Solid tumours (late-stage disease, second-line therapy or greater) in USA, Australia and Canada (NCT01986218)
25 Jan 2016Bristol-Myers Squibb completes enrolment in its phase I trial for Solid tumours in USA, Australia and Canada (NCT01986218)
31 Dec 2013Phase-I clinical trials in Solid tumours (late-stage disease) in Canada & Australia (Oral)
DETAILS WILL BE UPDATED SOON………….
BMS-986115 is an orally bioavailable, gamma secretase (GS) and pan-Notch inhibitor, with potential antineoplastic activity. Upon administration, GS/pan-Notch inhibitor BMS 986115 binds to GS and blocks the proteolytic cleavage and release of the Notch intracellular domain (NICD), which would normally follow ligand binding to the extracellular domain of the Notch receptor. This prevents both the subsequent translocation of NICD to the nucleus to form a transcription factor complex and the expression of Notch-regulated genes. This results in the induction of apoptosis and the inhibition of growth of tumor cells that overexpress Notch. Overexpression of the Notch signaling pathway plays an important role in tumor cell proliferation and survival
Dan O’Malley (Rice University) Currently: Bristol-Myers Squibb
PICTURES WILL BE UPDATED………….
useful for the treatment of conditions related to the Notch pathway, such as cancer and other proliferative diseases.
Notch signaling has been implicated in a variety of cellular processes, such as cell fate specification, differentiation, proliferation, apoptosis, and angiogenesis. (Bray, Nature Reviews Molecular Cell Biology, 7:678-689 (2006); Fortini, Developmental Cell 16:633-647 (2009)). The Notch proteins are single-pass heterodimeric transmembrane molecules. The Notch family includes 4 receptors, NOTCH 1-4, which become activated upon binding to ligands from the DSL family (Delta-like 1, 3, 4 and Jagged 1 and 2).
The activation and maturation of NOTCH requires a series of processing steps, including a proteolytic cleavage step mediated by gamma secretase, a multiprotein complex containing Presenilin 1 or Presenilin 2, nicastrin, APH1, and PEN2. Once NOTCH is cleaved, NOTCH intracellular domain (NICD) is released from the membrane. The released NICD translocates to the nucleus, where it functions as a transcriptional activator in concert with CSL family members (RBPSUH, “suppressor of hairless”, and LAG1). NOTCH target genes include HES family members, such as HES- 1. HES- 1 functions as transcriptional repressors of genes such as HERP 1 (also known as HEY2), HERP2 (also known as HEY1), and HATH1 (also known as ATOH1).
The aberrant activation of the Notch pathway contributes to tumorigenesis. Activation of Notch signaling has been implicated in the pathogenesis of various solid tumors including ovarian, pancreatic, as well as breast cancer and hematologic tumors such as leukemias, lymphomas, and multiple myeloma. The role of Notch inhibition and its utility in the treatment of various solid and hematological tumors are described in Miele, L. et al, Current Cancer Drug Targets, 6:313-323 (2006); Bolos, V. et al, Endocrine Reviews, 28:339-363 (2007); Shih, I.-M. et al, Cancer Research, 67: 1879- 1882 (2007); Yamaguchi, N. et al., Cancer Research, 68: 1881-1888 (2008); Miele, L., Expert Review Anti-cancer Therapy, 8: 1 197-1201 (2008); Purow, B., Current Pharmaceutical Biotechnology, 10: 154-160 (2009); Nefedova, Y. et al, Drug Resistance Updates, 1 1 :210-218 (2008); Dufraine, J. et al, Oncogene, 27:5132-5137 (2008); and Jun, H.T. et al, Drug Development Research, 69:319-328 (2008).
There remains a need for compounds that are useful as Notch inhibitors and that have sufficient metabolic stability to provide efficacious levels of drug exposure. Further, there remains a need for compounds useful as Notch inhibitors that can be orally or intravenously administered to a patient.
U.S. Patent No. 7,053,084 Bl discloses succinoylamino benzodiazepine compounds useful for treating neurological disorders such as Alzheimer’s Disease. The reference discloses that these succinoylamino benzodiazepine compounds inhibit gamma secretase activity and the processing of amyloid precursor protein linked to the formation of neurological deposits of amyloid protein. The reference does not disclose the use of these compounds in the treatment of proliferative diseases such as cancer.
Applicants have found potent compounds that have activity as Notch inhibitors and have sufficient metabolic stability to provide efficacious levels of drug exposure upon intravenous or oral administration. These compounds are provided to be useful as pharmaceuticals with desirable stability, bioavailability, therapeutic index, and toxicity values that are important to their drugability.
[00180] To a cold (-25 °C) stirred solution of 2,6-lutidine (18.38 mL, 158 mmol) in DCM (120 mL) was added Tf20 (24.88 mL, 147 mmol) over 3 min, and the mixture was stirred for 5 min. To the reaction mixture was added 3,3,3-trifluoropropan-l-ol (12 g, 105 mmol) over an interval of 3 min. After 2 hr, the reaction mixture was warmed to room temperature and stirred for 1 hr. The reaction mixture was concentrated to half its volume, then purified by loading directly on a silica gel column (330g ISCO) and the product was eluted with DCM to afford Intermediate S-IA (13.74 g, 53%) as a colorless oil. 1H NMR (400 MHz, CDC13) δ ppm 4.71 (2 H, t, J= 6.15 Hz), 2.49-2.86 (2 H, m).
[00181] To a stirring solution of 5,5,5-trifluoropentanoic acid (14.76 g, 95 mmol) and DMF (0.146 rriL) in DCM (50 mL) was slowly added oxalyl chloride (8.27 mL, 95 mmol). After 2h, the mixture was concentrated to dryness. A separate flask was changed with (S)-4-benzyloxazolidin-2-one (16.75 g, 95 mmol) in THF (100 mL) and then cooled to -78 °C. To the solution was slowly added n-BuLi (2.5M, 37.8 mL, 95 mmol) over 10 min, stirred for 10 min, and then a solution of the above acid chloride in THF (50 mL) was slowly added over 5 min. The mixture was stirred for 30 min, and then warmed to room temperature. The reaction was quenched with sat aq NH4C1. Next, 10% aq LiCl was then added to the mixture, and the mixture was extracted with Et20. The organic layer was washed with sat aq NaHC03 then with brine, dried (MgSC^), filtered and concentrated to dryness. The residue was purified by Si02 chromatography (ISCO, 330 g column, eluting with a gradient from 100% hexane to 100% EtOAc) to afford the product Intermediate S-IB; (25.25 g, 85%): 1H NMR (400 MHz, CDC13) δ ppm 7.32-7.39 (2 H, m), 7.30 (1 H, d, J= 7.05 Hz), 7.18-7.25 (2 H, m), 4.64-4.74 (1 H, m), 4.17-4.27 (2 H, m), 3.31 (1 H, dd, J= 13.35, 3.27 Hz), 3.00-3.11 (2 H, m), 2.79 (1 H, dd, J= 13.35, 9.57 Hz), 2.16-2.28 (2 H, m), 1.93-2.04 (2 H, m).
[00182] To a cold (-78 °C), stirred solution of Intermediate S-IB (3.03 g, 9.61 mmol) in THF (20 mL) was added NaHMDS (1.0M in THF) (10.6 mL, 10.60 mmol) under a nitrogen atmosphere. After 2 hours, tert-butyl 2-bromoacetate (5.62 g, 28.8 mmol) was added neat via syringe at -78 °C and stirring was maintained at the same temperature. After 6 hours, the reaction mixture was warmed to room temperature. The reaction mixture was partitioned between saturated NH4C1 and EtOAc. The organic phase was separated, and the aqueous phase was extracted with EtOAc (3x). The combined organics were washed with brine, dried (Na2s04), filtered and concentrated under reduced pressure. The residue was purified by flash chromatography (Teledyne ISCO
CombiFlash Rf, 5% to 100% solvent A/B = hexanes/EtOAc, REDISEP® Si02 120g). Concentration of the appropriate fractions provided Intermediate S-1C (2.79 g, 67.6%) as a colorless viscous oil: 1H NMR (400 MHz, CDC13) δ ppm 7.34 (2 H, d, J= 7.30 Hz), 7.24-7.32 (3 H, m), 4.62-4.75 (1 H, m, J= 10.17, 6.89, 3.43, 3.43 Hz), 4.15-4.25 (3 H, m), 3.35 (1 H, dd, J= 13.60, 3.27 Hz), 2.84 (1 H, dd, J= 16.62, 9.57 Hz), 2.75 (1 H, dd, J = 13.35, 10.07 Hz), 2.47 (1 H, dd, J= 16.62, 4.78 Hz), 2.11-2.23 (2 H, m), 1.90-2.02 (1 H, m), 1.72-1.84 (1 H, m), 1.44 (9 H, s).
[00183] To a cool (0 °C), stirred solution of Intermediate S-1C (2.17 g, 5.05 mmol) in THF (50 mL) and water (15 mL) was added a solution of LiOH (0.242 g, 10.11 mmol) and H202 (2.065 mL, 20.21 mmol) in H20 (2 mL). After 10 min, the reaction mixture was removed from the ice bath, stirred for lh, and then cooled to 0 °C. Saturated aqueous NaHCC”3 (25 mL) and saturated aqueous Na2s03 (25 mL) were added to the reaction mixture, and the mixture was stirred for 10 min, and then partially concentrated. The resulting mixture was extracted with DCM (2x), cooled with ice and made acidic with cone. HC1 to pH 3. The mixture was saturated with solid NaCl, extracted with EtOAc (3x), and then dried over MgS04, filtered and concentrated to a colorless oil to afford Intermediate S-ID, 1.2514g, 92%): 1H NMR (400 MHz, CDCI3) δ ppm 2.83-2.95 (1 H, m), 2.62-2.74 (1 H, m), 2.45 (1 H, dd, J= 16.62, 5.79 Hz), 2.15-2.27 (2 H, m), 1.88-2.00 (1 H, m), 1.75-1.88 (1 H, m), 1.45 (9 H, s). Intermediate S-l : (2R,3S)-3-(fert-Butoxycarbonyl)-6,6,6-trifluoro-2-(3,3,3- trifluoropropyl)hexanoic acid, and Intermediate S-1E: (2R,3R)-3-(tert-butoxycarbonyl)- 6,6,6-trifluoro-2-(3,3,3-trifluoropropyl)hexanoic acid
[00184] To a cold (-78 °C) stirred solution of Intermediate S-1D (5 g, 18.50 mmol) in THF (60 mL) was slowly added LDA (22.2 mL, 44.4 mmol, 2.0M) over 7 min. After stirring for 2 hr, Intermediate S- 1 A (6.38 g, 25.9 mmol) was added to the reaction mixture over 3 min. After 60 min, the reaction mixture was warmed to -25 °C
(ice/MeOH/dry ice) and stirred for an additional 60 min at which time sat aq NH4C1 was added. The separated aqueous phase was acidified with IN HC1 to pH 3, and then extracted with Et20. The combined organic layers were washed with brine (2x), dried over MgS04, filtered and concentrated to provide a 1 :4 (II :I1E) mixture (as determined by 1H NMR) of Intermediate S-l and Intermediate S-1E (6.00 g, 89%) as a pale yellow solid. 1H NMR (500 MHz, CDC13) δ ppm 2.81 (1 H, ddd, J = 10.17, 6.32, 3.85 Hz), 2.63- 2.76 (1 H, m), 2.02-2.33 (4 H, m), 1.86-1.99 (2 H, m), 1.68-1.85 (2 H, m), 1.47 (9 H, s).
[00185] To a cold (-78 °C), stirred solution of a mixture of Intermediate S-l and Intermediate S-1E (5.97 g, 16.30 mmol) in THF (91 mL) was added LDA (19 mL, 38.0 mmol, 2.0M in THF/hexane/ethyl benzene) dropwise via syringe over 10 min (internal temperature never exceeded -65 °C, J-KEM® probe in reaction solution). The mixture was stirred for 15 min, and then warmed to room temperature (24 °C water bath), stirred for 15 min, and then cooled to -78 °C for 15 min. To the reaction mixture was added Et2AlCl (41 mL, 41.0 mmol, 1M in hexane) via syringe (internal temperature never exceeded -55 °C), and the mixture was stirred for 10 min, and then warmed to room temperature (24 °C bath) for 15 min and then back to -78 °C for 15 min. Meanwhile, a 1000 mL round bottom flask was charged with MeOH (145 mL) and precooled to -78 °C. With vigorous stirring the reaction mixture was transferred via cannula over 5 min to the MeOH. The flask was removed from the bath, ice was added followed by the slow addition of IN HC1 (147 mL, 147 mmol). Gas evolution was observed as the HC1 was added. The reaction mixture was allowed to warm to room temperature during which the gas evolution subsided. The reaction mixture was diluted with EtOAc (750 mL), saturated with NaCl, and the organic phase was separated, washed with a solution of potassium fluoride (8.52 g, 147 mmol) and IN HC1 (41 mL, 41.0 mmol) in water (291 mL), brine (100 mL), and then dried (Na2s04), filtered and concentrated under vacuum. 1H NMR showed the product was a 9: 1 mixture of Intermediate S-l and Intermediate S- 1E. The enriched mixture of Intermediate S-l and Intermediate S-1E (6.12 g, >99% yield) was obtained as a dark amber solid: 1H NMR (400 MHz, CDC13) δ ppm 2.64-2.76 (2 H, m), 2.04-2.35 (4 H, m), 1.88-2.00 (2 H, m), 1.71-1.83 (2 H, m), 1.48 (9 H, s).
[00186] To a stirred solution of a 9: 1 enriched mixture of Intermediate S-l and Intermediate S-1E (5.98 g, 16.33 mmol) in DMF (63 mL) were added potassium carbonate (4.06 g, 29.4 mmol) and benzyl bromide (2.9 mL, 24.38 mmol), the mixture was then stirred overnight at room temperature. The reaction mixture was diluted with EtOAc (1000 mL), washed with 10% LiCl (3×200 mL), brine (200 mL), dried (Na2S04), filtered, concentrated, and then dried under vacuum. The residue was purified by Si02 chromatography using a toluene:hexane gradient. Diastereomerically purified
[00187] To a solution of Intermediate S-1F (4.81 g, 10.54 mmol) in MeOH (100 mL) was added 10% palladium on carbon (wet, Degussa type, 568.0 mg, 0.534 mmol) in a H2– pressure flask. The vessel was purged with N2 (4x), then purged with H2 (2x), and finally, pressurized to 50 psi and shaken overnight. The reaction vessel was
depressurized and purged with nitrogen. The mixture was filtered through CELITE®, washed with MeOH and then concentrated and dried under vacuum. Intermediate S-1 (3.81 g, 99% yield)) was obtained as a colorless solid: 1H NMR (400 MHz, chloroform-d) δ 2.62-2.79 (m, 2H), 2.02-2.40 (m, 4H), 1.87-2.00 (m, 2H), 1.67-1.84 (m, 2H), 1.48 (s, 9H).
[00188] Intermediate S-1 as a mixture with Intermediate S-IE was prepared in a similar procedure as above from Intermediate S-1D to afford a 1 :2.2 mixture of
Intermediate S-1 and Intermediate S-IE (8.60 g, 23.48 mmol), which was enriched using LDA (2.0 M solution in THF, ethyl benzene and heptane, 28.2 mL, 56.4 mmol) and diethyl aluminum chloride (1.0 M solution in hexane, 59 mL, 59.0 mmol) in THF (91 mL). After workup as described above, the resulting residue was found to be a 13.2: 1 (by 1H NMR) mixture of Intermediate S-1 and Intermediate S-IE, which was treated as follows: The crude material was dissolved in MTBE (43 mL). Hexanes (26 mL) were slowly charged to the reaction mixture while maintaining a temperature below 30 °C. The reaction mixture was stirred for 10 min. Next, tert-butylamine (2.7 mL, 1.1 eq) was charged slowly over a period of 20 minutes while maintaining a temperature below 30 °C. This addition was observed to be exothermic. The reaction mixture was stirred for 2 hrs below 30 °C and then filtered. The solid material was washed with 5:3 MTBE: hexane (80 mL), and the filtrate was concentrated and set aside. The filtered solid was dissolved in dichloromethane (300 mL), washed with IN HC1 (lOOmL), and the organic layer was washed with brine (100 mL x 2), and then concentrated under reduced pressure below 45 °C to afford Intermediate S-l (5.46 g, 64%).
A second alternate procedure for preparing Intermediate S-l :
[00189] To a stirred solution of 5,5,5-trifluoropentanoic acid (5 g, 32.0 mmol) in THF (30 mL) and hexane (30 mL) at 0 °C, was added tert-butyl 2,2,2-trichloroacetimidate (11.46 mL, 64.1 mmol). The mixture was stirred for 15 min at 0 °C. Boron trifluoride etherate (0.406 mL, 3.20 mmol) was added and the reaction mixture was allowed to warm to room temperature overnight. To the clear reaction mixture was added solid NaHC03 (5 g) and stirred for 30 min. The mixture was filtered through MgSC^ and washed with hexanes (200 mL). The solution was allowed to rest for 45 min, and the resulting solid material was removed by filtering on the same MgSC^ filter again, washed with hexanes (100 mL) and concentrated under reduced pressure without heat. The volume was reduced to about 30 mL, filtered through a clean fritted funnel, washed with hexane (5 mL), and then concentrated under reduced pressure without heat. The resulting neat oil was filtered through a 0.45μιη nylon membrane filter disk to provide Intermediate S-1G (6.6 g, 31.4 mmol 98% yield) as a colorless oil: 1H NMR (400 MHz, CDC13) δ ppm 1.38 (s, 9 H) 1.74-1.83 (m, 2 H) 2.00-2.13 (m, 2 H) 2.24 (t, J= 7.28 Hz, 2 H). Intermediate S-1H: (4S)-4-(Propan-2-yl)-3-(5,5,5-trifluoropentanoyl)-l,3-oxazolidin-2- one
[00190] To a stirred solution of 5,5,5-trifluoropentanoic acid (5.04 g, 32.3 mmol) in DCM (50 mL) and DMF (3 drops) was added oxalyl chloride (3.4 mL, 38.8 mmol) dropwise over 5 min. The solution was stirred until all bubbling subsided. The reaction mixture was concentrated under reduced pressure to give pale yellow oil. To a separate flask charged with a solution of (4S)-4-(propan-2-yl)-l,3-oxazolidin-2-one (4.18 g, 32.4 mmol) in THF (100 mL) at -78 °C was added n-BuLi (2.5M in hexane) (13.0 mL, 32.5 mmol) dropwise via syringe over 5 min. After stirring for 10 min, the above acid chloride, dissolved in THF (20 mL), was added via cannula over 15 min. The reaction mixture was warmed to 0 °C, and was allowed to warm to room temperature as the bath warmed and stirred overnight. To the reaction mixture was added saturated NH4C1, and the mixture was extracted with EtOAc (2x). The combined organics were washed with brine, dried (Na2s04), filtered and concentrated under reduced pressure. The crude material was purified by flash chromatography (Teledyne ISCO CombiFlash Rf, 5% to 60% solvent A/B = hexanes/EtOAc, REDISEP® Si02 120g). Concentration of the appropriate fractions provided Intermediate S-1H (7.39 g, 86%) as a colorless oil: 1H NMR (400 MHz, CDC13) δ ppm 4.44 (1 H, dt, J= 8.31, 3.53 Hz), 4.30 (1 H, t, J= 8.69 Hz), 4.23 (1 H, dd, J= 9.06, 3.02 Hz), 2.98-3.08 (2 H, m), 2.32-2.44 (1 H, m, J= 13.91, 7.02, 7.02, 4.03 Hz), 2.13-2.25 (2 H, m), 1.88-2.00 (2 H, m), 0.93 (3 H, d, J= 7.05 Hz), 0.88 (3 H, d, J= 6.80 Hz).
[00191] To a cold (-78 °C), stirred solution of diisopropylamine (5.3 mL, 37.2 mmol) in THF (59 mL) under a nitrogen atmosphere was added n-BuLi (2.5M in hexane) (14.7 mL, 36.8 mmol). The mixture was then warmed to 0 °C to give a 0.5M solution of LDA. A separate vessel was charged with Intermediate S-1H (2.45 g, 9.17 mmol). The material was azeotroped twice with benzene (the RotoVap air inlet was fitted with a nitrogen inlet to completely exclude humidity), and then toluene (15.3 mL) was added. This solution was added to a flask containing dry lithium chloride (1.96 g, 46.2 mmol). To the resultant mixture, cooled to -78 °C, was added the LDA solution (21.0 mL, 10.5 mmol) and the mixture was stirred at -78 °C for 10 min, then warmed to 0 °C for 10 min., and then cooled to -78 °C. To a separate reaction vessel containing Intermediate S-1G (3.41 g, 16.07 mmol), also azeotroped twice with benzene, was added toluene (15.3 mL), cooled to -78 °C and LDA (37.0 mL, 18.5 mmol) was added. The resulting solution was stirred at -78 °C for 25 min. At this time the enolate derived from the ester was transferred via cannula into the solution of the oxazolidinone enolate and stirred at -78 °C for an additional 5 min, at which time the septum was removed and solid powdered bis(2- ethylhexanoyloxy)copper (9.02 g, 25.8 mmol) was rapidly added to the reaction vessel and the septum was replaced. The vessel was immediately removed from the cold bath and immersed into a warm water bath (40 °C) with rapid swirling and with a concomitant color change from the initial turquoise to brown. The reaction mixture was stirred for 20 min, was then poured into 5% aqueous NH4OH (360 mL) and extracted with EtOAc (2x). The combined organics were washed with brine, dried (Na2s04), filtered and concentrated under reduced pressure. The residue was purified by flash chromatography (Teledyne ISCO CombiFlash Rf, 0% to 60% solvent A/B = hexanes/EtOAc, REDISEP® Si02 120g). Concentration of the appropriate fractions provided a mixture of Intermediate S- II and Intermediate S-1J (2.87 g, 66%) as a pale yellow viscous oil. 1H NMR showed the product was a 1.6: 1 mixture of diastereomers S-1LS-1J as determined by the integration of the multiplets at 2.74 and 2.84 ppm: 1H NMR (400 MHz, CDC13) δ ppm 4.43-4.54 (2 H, m), 4.23-4.35 (5 H, m), 4.01 (1 H, ddd, J= 9.54, 6.27, 3.51 Hz), 2.84 (1 H, ddd, J = 9.41, 7.28, 3.64 Hz), 2.74 (1 H, ddd, J= 10.29, 6.27, 4.02 Hz), 2.37-2.48 (2 H, m, J = 10.38, 6.98, 6.98, 3.51, 3.51 Hz), 2.20-2.37 (3 H, m), 1.92-2.20 (8 H, m), 1.64-1.91 (5 H, m), 1.47 (18 H, s), 0.88-0.98 (12 H, m). Intermediate S-1 : (2R,3S)-3-(fert-Butoxycarbonyl)-6,6,6-trifluoro-2-(3,3,3- trifluoropropyl)hexanoic acid, and Intermediate S-IE: (2R,3R)-3-(tert-Butoxycarbonyl)- 6,6,6-trifluoro-2-(3,3,3-trifluoropropyl)hexanoic acid
[00192] To a cool (0 °C), stirred solution of Intermediate S-1I and Intermediate S-1 J (4.54 g, 9.51 mmol) in THF (140 mL) and water (42 mL) were sequentially added hydrogen peroxide (30% in water) (10.3 g, 91 mmol) and LiOH (685.3 mg, 28.6 mmol). The mixture was stirred for 1 hr. At this time the reaction vessel was removed from the cold bath and then stirred for 1.5 hr. To the reaction mixture were added saturated NaHC03 (45 mL) and saturated Na2s03 (15 mL), and then the mixture was partially concentrated under reduced pressure. The resulting crude solution was extracted with DCM (3x). The aqueous phase was acidified to pH~l-2 with IN HC1, extracted with DCM (3x) and then EtOAc (lx). The combined organics were washed with brine, dried (Na2s04), filtered and concentrated under reduced pressure to provide a mixture of Intermediates S-1 and S-IE (3.00 g, 86%) as a colorless oil: 1H NMR (400 MHz, CDC13) δ ppm 2.76-2.84 (1 H, m, diastereomer 2), 2.64-2.76 (3 H, m), 2.04-2.35 (8 H, m), 1.88- 2.00 (4 H, m), 1.71-1.83 (4 H, m), 1.48 (9 H, s, diastereomer 1), 1.46 (9 H, s,
diastereomer 2); 1H NMR showed a 1.7: 1 mixture of S-1E:S-1F by integration of the peaks for the t-butyl groups. Intermediate S-1 : (2R,3S)-3-(fert-Butoxycarbonyl)-6,6,6-trifluoro-2-(3,3,3- trifluoropropyl)hexanoic acid, and Intermediate S-IF: (2R,3R)-3-(fert-Butoxycarbonyl)- 6,6,6-trifluoro-2-(3,3,3-trifluoropropyl)hexanoic acid
[00193] To a cold (-78 °C) stirred solution of diisopropylamine (1.7 mL, 11.93 mmol) in THF (19 mL) under a nitrogen atmosphere was added n-BuLi (2.5M in hexanes) (4.8 mL, 12.00 mmol). The mixture was stirred for 5 min and then warmed to 0 °C. In a separate vessel, to a cold (-78 °C) stirred solution of the mixture of Intermediates S-1 and S-1E (1.99 g, 5.43 mmol) in THF (18 mL) was added the LDA solution prepared above via cannula slowly over 25 min. The mixture was stirred for 15 min, then warmed to room temperature (placed in a 24 °C water bath) for 15 min, and then again cooled to -78 °C for 15 min. To the reaction mixture was added Et2AlCl (1M in hexane) (11.4 mL, 11.40 mmol) via syringe. The mixture was stirred for 10 min, warmed to room
temperature for 15 min and then cooled back to -78 °C for 15 min. Methanol (25 mL) was rapidly added, swirled vigorously while warming to room temperature, and then concentrated to ~l/4 the original volume. The mixture was dissolved in EtOAc and washed with IN HC1 (50 mL) and ice (75 g). The aqueous phase was separated and extracted with EtOAc (2x). The combined organics were washed with a mixture of KF (2.85g in 75 mL water) and IN HC1 (13 mL) [resulting solution pH 3-4], then with brine, dried (Na2s04), filtered and concentrated under reduced pressure to give a 9: 1 (S-LS-1E) enriched diastereomeric mixture (as determined by 1H NMR) of Intermediate S-1 and Intermediate S-1E (2.13 g, >99%) as a pale yellow viscous oil: 1H NMR (400 MHz, CDC13) δ ppm 2.64-2.76 (2 H, m), 2.04-2.35 (4 H, m), 1.88-2.00 (2 H, m), 1.71-1.83 (2 H, m), 1.48 (9 H, s).
[00194] To a cold (-78 °C), stirred solution of Intermediate S-1D (1.72 g, 6.36 mmol) in THF (30 mL) was slowly added LDA (7.32 mL, 14.6 mmol) over 7 min. After stirring for 1 h, 4,4,4-trifluorobutyltrifluoromethanesulfonate (2.11 g, 8.11 mmol) was added to the reaction mixture over 2 min. After 15 min, the reaction mixture was warmed to -25 °C (ice/MeOH/dry ice) for lh, and then cooled to -78 °C. After 80 min, the reaction was quenched with a saturated aqueous NH4C1 solution (10 mL). The reaction mixture was further diluted with brine and the solution was adjusted to pH 3 with IN HC1. The aqueous layer was extracted with ether. The combined organics were washed with brine, dried over anhydrous magnesium sulfate, and concentrated under reduced pressure to provide a mixture of Intermediates S-2 and S-2A (2.29 g, 95%) as a colorless oil. 1H NMR (400MHz, chloroform-d) δ 2.83-2.75 (m, 1H), 2.64 (ddd, J = 9.9, 6.7, 3.6 Hz, 1H), 2.32-2.03 (m, 5H), 1.98-1.70 (m, 3H), 1.69-1.52 (m, 3H), 1.50-1.42 (m, 9H). 1H NMR showed a 1 :4.5 mixture (S-2:S-2A) of diastereomers by integration of the peaks for the t- Bu groups.
[00195] A mixture of Intermediate S-2 and Intermediate S-2A (2.29 g, 6.02 mmol) was dissolved in THF (38 mL) to give a colorless solution which was cooled to -78 °C. Then, LDA (7.23 mL, 14.5 mmol) (2.0M in heptane/THF/ethylbenzene) was slowly added to the reaction mixture over 3 min. After stirring for 15 min, the reaction mixture was placed in a room temperature water bath. After 15 min the reaction mixture was placed back in a -78 °C bath and then diethylaluminum chloride (14.5 mL, 14.5 mmol) (1M in hexane) was added slowly over 5 min. The reaction mixture was stirred at -78 °C. After 15 min, the reaction mixture was placed in a room temperature water bath for 10 min, and then cooled back to -78 °C. After 15 min, the reaction was quenched with MeOH (30.0 mL, 741 mmol), removed from the -78 °C bath and concentrated. To the reaction mixture was added ice and HC1 (60.8 mL, 60.8 mmol) and the resulting mixture was extracted with EtOAc (2x 200 mL). The organic layer was washed with potassium fluoride (3.50g, 60.3 mmol) in 55 mL H20 and 17.0 mL of IN HC1. The organics were dried over anhydrous magnesium sulfate and concentrated under reduced pressure to provide an enriched mixture of Intermediate S-2 and Intermediate S-2A (2.25g, 98% yield) as a light yellow oil. 1H NMR (400MHz, chloroform-d) δ 2.83-2.75 (m, 1H), 2.64 (ddd, J= 9.9, 6.7, 3.6 Hz, 1H), 2.32-2.03 (m, 5H), 1.98-1.70 (m, 3H), 1.69-1.52 (m, 3H), 1.50-1.42 (m, 9H). 1H NMR showed a 9: 1 ratio in favor of the desired diastereomer Intermediate S-2.
[00196] To a stirred 9: 1 mixture of Intermediate S-2 and Intermediate S-2A (2.24 g, 5.89 mmoL) and potassium carbonate (1.60 g, 11.58 mmoL) in DMF (30 mL) was added benzyl bromide (1.20 mL, 10.1 mmoL)). The reaction mixture was stirred at room temperature for 19 h. The reaction mixture was diluted with ethyl acetate (400 mL) and washed with 10% LiCl solution (3 x 100 mL), brine (50 mL), and then dried over anhydrous magnesium sulfate, filtered and concentrated to dryness under vacuum. The residue was purified by flash chromatography (Teledyne ISCO CombiFlash 0%> to 100% solvent A/B = hexane/EtOAc, REDISEP® Si02 220 g, detecting at 254 nm, and monitoring at 220 nm). Concentration of the appropriate fractions provided Intermediate S-2B (1.59 g, 57.5%). HPLC: RT = 3.863 min (CHROMOLITH® SpeedROD column 4.6 x 50 mm, 10-90% aqueous methanol over 4 minutes containing 0.1% TFA, 4 mL/min, monitoring at 220 nm), 1H NMR (400MHz, chloroform-d) δ 7.40-7.34 (m, 5H), 5.17 (d, J= 1.8 Hz, 2H), 2.73-2.64 (m, 1H), 2.55 (td, J= 10.0, 3.9 Hz, 1H), 2.16-1.82 (m, 5H), 1.79-1.57 (m, 3H), 1.53-1.49 (m, 1H), 1.45 (s, 9H), 1.37-1.24 (m, 1H).
[00197] To a stirred solution of Intermediate S-2B (1.59 g, 3.37 mmoL) in MeOH (10 mL) and EtOAc (10 mL) under nitrogen was added 10%> Pd/C (510 mg). The atmosphere was replaced with hydrogen and the reaction mixture was stirred at room temperature for 2.5 h. The palladium catalyst was filtered off through a 4 μΜ polycarbonate film and rinsed with MeOH. The filtrate was concentrated under reduced pressure to give intermediate S-2 (1.28 g, 99%). 1H NMR (400MHz, chloroform-d) δ 2.76-2.67 (m, 1H), 2.65-2.56 (m, 1H), 2.33-2.21 (m, 1H), 2.17-2.08 (m, 3H), 1.93 (dtd, J= 14.5, 9.9, 5.2 Hz, 1H), 1.84-1.74 (m, 2H), 1.70-1.52 (m, 3H), 1.48 (s, 9H).
[00198] In a 1 L round-bottomed flask was added 2-amino-3-methylbenzoic acid (11.2 g, 74.1 mmol) and Ν,Ο-dimethylhydroxylamine hydrochloride (14.45 g, 148 mmol) in DCM (500 mL) to give a pale brown suspension. The reaction mixture was treated with Et3N (35 mL), HOBT (11.35 g, 74.1 mmol) and EDC (14.20 g, 74.1 mmol) and then stirred at room temperature for 24 hours. The mixture was then washed with 10% LiCl, and then acidified with IN HCl. The organic layer was washed successively with 10%> LiCl and aq NaHC03. The organic layer was decolorized with charcoal, filtered, and the filtrate was dried over MgSC^. The mixture was filtered and concentrated to give 13.22 g (92% yield) of Intermediate A-1A. MS(ES): m/z = 195.1 [M+H+]; HPLC: RT = 1.118 min. (H20/MeOH with TFA, CHROMOLITH® ODS S5 4.6 x 50 mm, gradient = 4 min, wavelength = 220 nm); 1H NMR (500MHz, chloroform-d) δ 7.22 (dd, J= 7.8, 0.8 Hz, 1H), 7.12-7.06 (m, 1H), 6.63 (t, J= 7.5 Hz, 1H), 4.63 (br. s., 2H), 3.61 (s, 3H), 3.34 (s, 3H), 2.17 (s, 3H).
[00199] In a 500 mL round-bottomed flask, a solution of l-fluoro-3-iodobenzene (13.61 mL, 116 mmol) in THF (120 mL) was cooled in a -78 °C bath. A solution of n- BuLi, (2.5M in hexane, 46.3 mL, 116 mmol) was added dropwise over 10 minutes. The solution was stirred at -78 °C for 30 minutes and then treated with a solution of
Intermediate A-1 A (6.43 g, 33.1 mmol) in THF (30 mL). After 1.5 hours, the reaction mixture was added to a mixture of ice and IN HCl (149 mL, 149 mmol) and the reaction flask was rinsed with THF (5 ml) and combined with the aqueous mixture. The resulting mixture was diluted with 10% aq LiCl and the pH was adjusted to 4 with IN NaOH. The mixture was then extracted with Et20, washed with brine, dried over MgS04, filtered and concentrated. The resulting residue was purified by silica gel chromatography (220g ISCO) eluting with a gradient from 10% EtOAc/hexane to 30% EtOAc/hexane to afford Intermediate A-l (7.11 g, 94% yield) as an oil. MS(ES): m/z = 230.1 [M+H+]; HPLC: RT = 2.820 min Purity = 99%. (H20/MeOH with TFA, CHROMOLITH® ODS S5 4.6 x 50 mm, gradient = 4 min, wavelength = 220 nm).
[00225] In a 1 L round-bottomed flask, a solution of 2-(lH-benzo[d][l,2,3]triazol-l- yl)-2-((phenoxycarbonyl)amino)acetic acid (J. Org. Chem., 55:2206-2214 (1990)) (19.37 g, 62.0 mmol) in THF (135 mL) was cooled in an ice/water bath and treated with oxalyl chloride (5.43 mL, 62.0 mmol) and 4 drops of DMF. The reaction mixture was stirred for 4 hours. Next, a solution of Intermediate A- 1 (7.11 g, 31.0 mmol) in THF (35 mL) was added and the resulting solution was removed from the ice/water bath and stirred at room temperature for 1.5 hours. The mixture was then treated with a solution of ammonia, (7M in MeOH) (19.94 mL, 140 mmol). After 15 mins, another portion of ammonia, (7M in MeOH) (19.94 mL, 140 mmol) was added and the resulting mixture was sealed under N2 and stirred overnight at room temperature. The reaction mixture was then concentrated to ~l/2 volume and then diluted with AcOH (63 mL) and stir at room temperature for 4 hours. The reaction mixture was then concentrated, and the residue was diluted with 500 mL water to give a precipitate. Hexane and Et20 were added and the mixture was stirred at room temperature for 1 hour to form an orange solid. Et20 was removed under a stream of nitrogen and the aqueous layer was decanted. The residue was triturated with 40 mL of iPrOH and stirred at room temperature to give a white precipitate. The solid was filtered and washed with iPrOH, then dried on a filter under a stream of nitrogen to give racemic Intermediate B-1A (5.4 g, 41.7%yield).
[00226] Racemic Intermediate B-1A (5.9 g, 14.3 mmol) was resolved using the Chiral SFC conditions described below. The desired stereoisomer was collected as the second peak in the elution order: Instrument: Berger SFC MGIII, Column: CHIRALPAK® IC 25 x 3 cm, 5 cm; column temp: 45 °C; Mobile Phase: C02/MeOH (45/55); Flow rate: 160 mL/min; Detection at 220 nm.
[00227] After evaporation of the solvent, Intermediate B-1A (2.73 g, 46% yield) was obtained as a white solid. HPLC: RT = 3.075 min. (H20/MeOH with TFA,
CHROMOLITH® ODS S5 4.6 x 50 mm, gradient = 4 min, wavelength = 220 nm).
[00228] In a 100 mL round-bottomed flask, a solution of Intermediate B-1A (2.73 g, 6.54 mmol) in acetic acid (12 mL) was treated with HBr, 33% in HOAc (10.76 mL, 65.4 mmol) and the mixture was stirred at room temperature for 1 hour. The solution was diluted with Et20 to give a yellow precipitate. The yellow solid was filtered and rinsed with Et20 under nitrogen. The solid was transferred to 100 mL round bottom flask and water was added (white precipitate formed). The slurry was slowly made basic with saturated NaHC03. The resulting tacky precipitate was extracted with EtOAc. The organic layer was washed with water, dried over MgS04, and then filtered and
concentrated to dryness to give Intermediate B-l (1.68 g, 91% yield) as a white foam solid. MS(ES): m/z = 284.2 [M+H+]; HPLC: RT = 1.72 min (H20/MeOH with TFA, CHROMOLITH® ODS S5 4.6 x 50 mm, gradient = 4 min, wavelength = 220 nm). 1H NMR (400MHz, DMSO-d6) δ 10.01 (br. s., 1H), 7.56-7.44 (m, 2H), 7.41-7.26 (m, 3H), 7.22-7.11 (m, 2H), 4.24 (s, 1H), 2.55 (br. s., 2H), 2.41 (s, 3H). [00229] The compounds listed below in Table 6 (Intermediates B-2 to B-3) were prepared according to the general synthetic procedure described for Intermediate B-l , using the starting materials Intermediate A- 10 and Intermediate A-4, respectively.
[00240] In a 100 mL round-bottomed flask, a solution of Intermediate B-l (1683 mg, 5.94 mmol), Et3N (1.656 mL, 11.88 mmol), and Intermediate S-l in DMF (20 mL) was treated with o-benzotriazol-l-yl-A .A .N’.N’-tetramethyluronium tetrafluoroborate (3815 mg, 11.88 mmol) and stirred at room temperature for 1 hour. The reaction mixture was diluted with water and saturated aqueous NaHC03. An off white precipitate formed and was filtered and washed with water. The resulting solid was dried on the filter under a stream of nitrogen to give Intermediate 1A (3.7 g, 99% yield). MS(ES): m/z =
632.4[M+H+]; HPLC: RT = 3.635 min Purity = 98%. (H20/MeOH with TFA,
[00241] In a 250 mL round-bottomed flask, a solution of Intermediate 1A (3.7 g, 5.86 mmol) in DCM (25 mL) was treated with TFA (25 mL) and the resulting pale orange solution was stirred at room temperature for 1.5 hours. The reaction mixture was then concentrated to give Intermediate IB. HPLC: RT = 3.12 min (H20/MeOH with TFA, CHROMOLITH® ODS S5 4.6 x 50 mm, gradient = 4 min, wavelength = 220 nm).
[00242] In a 250 mL round-bottomed flask, a solution of Intermediate IB (4.04 g, 5.86 mmol) in THF (50 mL) was treated with ammonia (2M in iPrOH) (26.4 mL, 52.7 mmol), followed by HOBT (1.795 g, 11.72 mmol) and EDC (2.246 g, 11.72 mmol). The resulting white suspension was stirred at room temperature overnight. The reaction mixture was diluted with water and saturated aqueous NaHC03. The resulting solid was filtered, rinsed with water and then dried on the filter under a stream of nitrogen. The crude product was suspended in 20 mL of iPrOH and stirred at room temperature for 20 min and then filtered and washed with iPrOH and dried under vacuum to give 2.83 g of solid. The solid was dissolved in re fluxing EtOH(100 mL) and slowly treated with 200 mg activated charcoal added in small portions. The hot mixture was filtered through CELITE® and rinsed with hot EtOH. The filtrate was reduced to half volume, allowed to cool and the white precipitate formed was filtered and rinsed with EtOH to give 2.57 g of white solid. A second recrystallization from EtOH (70 mL) afforded Example 1 (2.39 g, 70% yield) as a white solid. HPLC: RT = 10.859 min (H20/CH3CN with TFA, Sunfire C18 3.5μπι, 3.0x150mm, gradient = 15 min, wavelength = 220 and 254 nm); MS(ES): m/z = 575.3 [M+H+]; 1H NMR (400MHz, methanol-d4) δ 7.57-7.50 (m, 1H), 7.47-7.30 (m, 3H), 7.29-7.15 (m, 3H), 5.38 (s, 1H), 2.85-2.75 (m, 1H), 2.59 (td, J= 10.5, 4.0 Hz, 1H), 2.53-2.41 (m, 4H), 2.31-2.10 (m, 3H), 1.96-1.70 (m, 4H).
PAPER RELATED
Structure–activity relationships in a series of (2-oxo-1,4-benzodiazepin-3-yl)-succinamides identified highly potent inhibitors of γ-secretase mediated signaling of Notch1/2/3/4 receptors. On the basis of its robust in vivo efficacy at tolerated doses in Notch driven leukemia and solid tumor xenograft models, 12 (BMS-906024) was selected as a candidate for clinical evaluation.
Discovery of Clinical Candidate BMS-906024: A Potent Pan-Notch Inhibitor for the Treatment of Leukemia and Solid Tumors
For some disease targets, an indirect approach may be best. Or so Ashvinikumar V. Gavai and his colleagues atBristol-Myers Squibbfound in their quest toward a potential cancer drug. Gavai unveiled BMS-906024, which is an experimental—and slightly roundabout—treatment for a number of cancers, including breast, lung, and colon cancers, and leukemia.
Cancers have a tendency to relapse or to become resistant to treatments that once worked. Research at BMS and elsewhere had suggested that a family of proteins called Notch is implicated in that resistance and in cancer progression more generally. Gavai, director of oncology chemistry at BMS in Princeton, N.J., and his team set out to block Notch family signaling.
Notch family members lack enzymatic activity, so blocking them directly is difficult. Instead, BMS developed inhibitors of an enzyme that is essential for activating Notch signaling—γ-secretase.
Image may be NSFW. Clik here to view.
Company: Bristol-Myers Squibb
Target: pan-Notch
Disease: breast, lung, colon cancer; leukemia
Interfering with Notch, even in this indirect way, can have detrimental effects on the gastrointestinal tract. Only two of the four Notch family members are linked to that side effect, Gavai says. But he and his team think their drug will be most effective if it acts on all four family members roughly equally—a so-called pan-Notch inhibitor. By selecting a molecule that’s well tolerated in animals and carefully scheduling doses of the drug in humans, it could be possible to minimize side effects, he says.
The BMS team relied on Notch signaling assays in leukemia and breast cancer cell lines to find leads. They soon learned that for their molecules to work, three chiral centers had to be in the S,R,Sconfiguration. After that, they strove to make the molecules last in the bloodstream. They removed an isobutyl group and tweaked some other parts of their candidate’s succinamide side chain. It was tough to retain both a long half-life and activity against Notch, Gavai told C&EN. “You’d optimize one and lose the other.”
His team threaded the needle with BMS-906024. Their studies with mice suggest that a dose of 4–6 mg once a week could be effective in people. That’s lower than doses being tested for other Notch-targeted agents, according to the website clinicaltrials.gov. The mouse studies also back the idea that Notch is involved in cancer drug resistance and suggest that Notch could be a target for taking on cancer stem cells, which are notoriously resistant to chemotherapy.
BMS-906024 is in Phase I clinical trials, both alone and in combination with other agents. Patients with colon, lung, breast, and other cancers are receiving intravenous doses of the compound to determine its safety and optimum dose ranges.
Image may be NSFW. Clik here to view.
(From left, front row) Gavai, Weifeng Shan, (second row) Aaron Balog, Patrice Gill, Gregory Vite, (third row) Francis Lee, Claude Quesnelle, (rear row) Wen-Ching Han, Richard Westhouse.
An enantioselective synthesis of (S)-7-amino-5H,7H-dibenzo[b,d]azepin-6-one (S–1) is described. The key step in the sequence involved crystallization-induced dynamic resolution (CIDR) of compound 7 using Boc-d-phenylalanine as a chiral resolving agent and 3,5-dichlorosalicylaldehyde as a racemization catalyst to afford S–1 in 81% overall yield with 98.5% enantiomeric excess.
Crystallization-Induced Dynamic Resolution toward the Synthesis of (S)-7-Amino-5H,7H-dibenzo[b,d]-azepin-6-one: An Important Scaffold for γ-Secretase Inhibitors
† Department of Discovery Synthesis, Biocon Bristol-Myers Squibb Research Centre, Biocon Park, Bommasandra IV Phase, Jigani Link Road, Bengaluru 560099, India
‡Bristol-Myers Squibb Company, P.O Box 4000, Princeton, New Jersey 08543-4000, United States
Organizational Initiatives Towards Developing Greener Processes for Generic Active Pharmaceutical Ingredients
– Dr. Vilas H. Dahanukar, Chief Scientist-Process R&D, Integerated Product Development Organization, Dr. Reddy’s Laboratories Ltd., India
Image may be NSFW. Clik here to view.
Image may be NSFW. Clik here to view.
presented at
4th Industrial Green Chemistry World Convention & Ecosystem (IGCW-2015) on 4th – 5th December 2015
A PRESENTATION
Image may be NSFW. Clik here to view.The presentation will load below
Please scroll with mouse to see
A PRESENTATION
Innovative Techniques, To Synthesize Breakthrough Molecules, See DOE On pae 4 onwards
Image may be NSFW. Clik here to view.The presentation will load below
Introduction to drug product development – setting the scene
Drug product development at a glance – from first in man to marketing authorization
Pharmaceutical QbD: Quo vadis?
Application of QbD principles to drug product development
Expectations from regulatory agencies
Regulatory initiatives and approaches for supporting emerging technologies
Concepts of Real Time Release Testing (Draft Annex 17 EU GMP Guideline)
Harmonization of regulatory requirements (QbD parallel-assessment FDA-EMA, ICH Q8 -> Draft Q12?)
Regulatory expectations: Lessons learned from applications so far
Knowledge Management
Knowledge Management (KM) System – Definition and Reason
Knowledge Management Cycle
Explicit and Tacit Knowledge – The Knowledge Spiral
Correlation between KM and other Processes
Enabling Knowledge Management
Knowledge Review – integral part of the Management Review (ICH Q10)
Quality Risk Assessment and Control Strategy
Objectives of Quality Risk Assessment (QRA) as part of development
Overview to risk assessment tools
Introduction of Process Risk Map
Introduction of risk based control strategy development
QbD Toolbox: Case studies DoE, PAT, and Basic Statistics
Value-added use of QbD tools – generic approaches and tailored solutions
Case studies and examples for different unit operations and variable problems
Reports and Documentation
Development Reports
Transfer protocols and reports
Control Strategy and link to the submission dossier
Wrap-up & Final Discussion
The concepts and tools used over the two days will be summarized and future implications and opportunities of applying QbD principles to process development will be discussed. Delegates will be given time to ask questions on how they can apply what they have learned to their own drug product development and manufacturing.
Workshop Process Risk Map & link to Control Strategy
Based on a risk assessment tool tailored to cover development needs, delegates will work on case studies of process development for a solid oral dosage form.
From QTPP and CQA to relationship analysis of process parameters and material attributes
Process mapping for integrated documentation of the development work
Process Risk Map as a tool for development-focussed risk assessment
Quality by Design in API Manufacturing
General framework and key elements of QbD for APIs – background and potential strategies
What is it all about?
What are the benefits?
When and how should you use it?
Practical examples with typical points of discussion
How to identify and control Critical Quality Attributes (CQAs) in API synthesis – a risk-based approach to developing a control strategy
Severity assessment of quality attributes
Impact levels for critical process parameters (CPPs) and critical material attributes (CMAs)
Considerations for the API Starting material
Design of an effective risk-based control strategy
Examples
How to provide information on the development of the API manufacturing process – dossier requirements
What should be done at which stage?
Which information is relevant for the dossier?
What are the key-points to be considered for APIs (NCE/Biotech) and their formulations
Typical questions from Authorities
Process Evaluation and Design Space
Changing Validation Approach
Validation Life Cycle
Design Space Concept
Application of PAT in the API industry
PAT at development stages of a QbD-based development
PAT as part of the Control Strategy in a GMP environment
Practical examples of PAT implementations at a commercial scale in a GMP environmen
Festinavir is a nucleoside reverse transcriptase inhibitor
(NRTI) which is being developed for the treatment of HIV infection. The drug has shown considerable efficacy in early development, and with perhaps less toxicity than some other NRTIs, such as the drug stavudine (marketed under the trade name ZERIT®).
Festinavir has the chemical form and the structural formula:
Image may be NSFW. Clik here to view.
Festinavir was developed by Yale University in conjunction with two Japanese research scientists, and is protected by U.S. Patent No. 7,589,078, the contents of which are incorporated herein by reference. The ‘078 patent sets forth the synthesis of the primary compound, and other structural analogs. In addition, Oncolys BioPharma, Inc. of Japan has now published US 2010/0280235 for the production of 4′ ethynyl D4T. As starting raw material, the Oncolys method utilizes a substituted furan compound, furfuryl alcohol. In another publication by Nissan Chemical Industries of Japan, and set forth in WO 201 1/099443, there is disclosed a method for producing a beta-dihydrofuran deriving compound or a beta-tetrahydrofuran deriving compound. In this process, a diol compound is used as the starting material. Nissan has also published WO 2011/09442
directed to a process for the preparation of a β-glycoside compound. Two further publications, each to Hamari Chemicals of Japan, WO 2009/1 19785 and
WO 2009/125841, set forth methods for producing and purifying ethynyl thymide compounds. Pharmaset, Inc. of the U.S. has also published US 2009/0318380,
WO 2009/005674 and WO 2007/038507 for the production of 4’ -nucleoside analogs for treating HIV infection. Reference is also made to the BMS application entitled
“Sulfilimine and Sulphoxide Methods for Producing Festinavir” filed as a PCT application, PCT/US2013/042150 on May 22, 2013 (now WO2013/177243).
Compounds having methyl, vinyl, and ethynyl groups at the 4′-position of stavudine (d4T: 2′,3′-didehydro-3′-deoxythymidine) were synthesized. The compounds were assayed for their ability to inhibit the replication of HIV in cell culture. The 4′-ethynyl analogue (15) was found to be more potent and less toxic than the parent compound stavudine.
FAB-MS m/z 249 (M++H). Anal. calcd for C12H12N2O4·1/6H2O: C, 57.37; H, 4.95; N, 11.15. Found: C, 57.36; H, 4.69; N, 10.98.
PAPER
Scalable Synthesis of the Potent HIV Inhibitor BMS-986001 by Non-Enzymatic Dynamic Kinetic Asymmetric Transformation (DYKAT)
Angewandte Chemie, International Edition (2015), 54, (24), 7185-7188.
Chemical Development, Bristol-Myers Squibb, 1 Squibb Drive, New Brunswick, NJ 08903 (USA)
Chemical Development, Bristol-Myers Squibb, 1 Squibb Drive, New Brunswick, NJ 08903 (USA)
Described herein is the synthesis of BMS-986001 by employing two novel organocatalytic transformations: 1) a highly selective pyranose to furanose ring tautomerization to access an advanced intermediate, and 2) an unprecedented small-molecule-mediated dynamic kinetic resolution to access a variety of enantiopure pyranones, one of which served as a versatile building block for the multigram, stereoselective, and chromatography-free synthesis of BMS-986001. The synthesis required five chemical transformations and resulted in a 44 % overall yield.
The starting material is 5-methylurdine, which is commercially available. The first step of the process is an acetal formation. 5-methyluridine is utilized and is treated with H2SO4 and acetaldehyde. Other acids available to the scientist, such as perchloric acid, will also work for this transformation. The solvent utilized for this step is acetonitrile (ACN), and other solvents may also be utilized as well. Once the starting material is consumed, a slurry is obtained and the product can be simply filtered off and dried to provide Compound 1 as a solid.
Image may be NSFW. Clik here to view.
Acetal formation
Preparation of l-((3aR,4R,6R,6aR)-6-(hydroxymethyl)-2-methyltetrahydrofuro [3,4-d] [1,3] dioxol-4-yl)-5-methylpyrimidine-2,4(lH,3H)-dione
The following were added to a flask: 5-methyluridine (10 g, 38.70 mmol), acetonitrile (20 mL) and 70% perchloric acid (4.01 mL, 47.63 mmol). A solution of acetaldehyde (3.26 mL, 58.10 mmol) in acetonitrile (20 mL) was added dropwise over 1 h. The resulting solution was allowed to stir at 20 °C for 18 h. The resulting slurry was filtered and dried (50 °C, 25 mmHg) to afford Acetal (9.30 g, 84% yield) as white solid
The next step of the sequence is installation of a 4-biphenylacetate. Without being bound by any particular theory, this protecting step may be chosen for two reasons:
1) To provide a solid intermediate that can be easily isolated, and
2) Act as a directing group in the next step (set forth later on).
This reaction consists of reacting Compound 1 with 4-biphenyl acid chloride and pyridine in acetonitrile. In this reaction, pyridine is preferred as it allows the reaction to occur only at the -OH moiety of the molecule. It should also be noted that other polar solvents could be used, but acetonitrile allowed the desired product Compound 2 to be isolated as s solid.
Image may be NSFW. Clik here to view.
Ac lation
Preparation of ((3aR,4R,6R,6aR)-2-methyl-6-(5-methyl-2,4-dioxo-3,4-dihydropyrimidin-l(2H)-yl)tetrahydrofuro[3,4-d] [l,3]dioxol-4-yl)methyl [1,1′-biphenyl]-4-carboxylate.
Acetal (9.30 g, 32 mmol) was dissolved into acetonitrile (100 mL). Pyridine (1.3 eq) was added followed by the addition of 4-biphenylcarbonyl chloride (1.05 eq). The solution was heated to 50 °C and held for 2 h. The slurry was cooled to 20 °C and held for 2 h. The slurry was filtered and washed with acetonitrile (100 mL). The solids were dried (50 °C, 25 mmHg) to Compound 2 (85% yield).
With the acetal and 4-biphenylacetate groups in place, the next reaction is a regioselective acetal opening utilizing TMSOTf (Trimethylsilyl trifluoromethane sulfonate, or other available Lewis acids)/Et3N to afford the corresponding silyl ether, which is cleaved in situ, to afford the 2-vinyloxy compound as Compound 3. Compound 3 may be prepared in a step-wise fashion (shown below), but in order to reduce the number of steps, it is possible to take Compound 3 and selectively form the desired 2-vinyl oxy regioisomer Compound 3. Those skilled in the art may recognize that the 4-biphenylacetate can be important to obtain high selectivity for this transformation.
Image may be NSFW. Clik here to view.
Image may be NSFW. Clik here to view.
Image may be NSFW. Clik here to view.
Although a variety of Lewis acids may be utilized, TMSOTf is generally found to be more effective. Et3 is also a preferred reactant, as other amine bases are generally less effective. The ratio of TMSOTf to Ets is preferably within the range of about 1 : 1.3; if the reaction medium became acidic, Compound 3 would revert back to Compound 2. In terms of solvents, DCM (Dichloromethane) may be particularly effective, but toluene, CF3-PI1, sulfolane, and DCE (Dichloroethene) are also effective. The reaction can be worked up using aqueous acid, preferably K2HP04, or methanolic NH4F to quench the reaction, as well as remove the TMS-ether in situ.
Image may be NSFW. Clik here to view.
TMSOTf-opening
Preparation of ((2R,3R,4R,5R)-3-hydroxy-5-(5-methyl-2,4-dioxo-3,4-dihydropyrimidin-l(2H)-yl)-4-(vinyloxy)tetrahydrofuran-2-yl)methyl [1,1′-biphenyl]-4-carboxylate
Compound 2 (20 g, 43.06 mmol) was dissolved into DCM (160 mL). Triethylamine (78 mL, 560 mmol) was added followed by the addition of TMSOTf (80.30 mL, 431 mmol). This solution was heated to 45 °C and held there until complete by HPLC analysis (6 h). Once complete, this solution was added to ammonium acetate (66.40 g, 861 mmol) in water (200 mL). After stirring for 20 min, the layers were separated. The organics were concentrated and the resulting residue was dissolved into EtOAc (200 mL). The organics were washed with the following solution (potassium phosphate monobasic (118 g, 861 mmol) in water (400 mL). The organics were then dried ( a2S04), filtered and concentrated. The resulting residue was purified by column chromatography [Silica gel; 20% to 90% EtOAc in Hexanes] to afford Compound 3 (15.8 g, 79% yield) as a solid.
Next, Compound 3 is transformed into the iodide compound which is Compound 4. This can be accomplished by treating Compound 3 with (2.0 eq), PPI13 (2.0 eq.) and imidazole (4.0 eq). Other methods to install the iodide may also be utilized, such as mesylation/Nal, etc., but these may be less preferred. In addition, other halogen-bearing compounds such as Br2 and CI2 may be considered by the skilled scientist. Premixing imidazole, , and PPh3, followed by addition of Compound 3 in THF and heating at 60 °C allows smooth conversion to Compound 4. It is highly preferred to add all reagents prior to the addition of Compound 3; if not, the vinyloxy group will be cleaved. Other solvents, such as 2-MeTHF and PhMe may be utilized, but THF often provides the best yield.
Image may be NSFW. Clik here to view.
Iodiiiation
Preparation of ((2R,3S,4S,5R)-3-iodo-5-(5-methyl-2,4-dioxo-3,4-dihydropyrimidin-l(2H)-yl)-4-(vinyloxy)tetrahydrofuran-2-yl)methyl [l,l’-biphenyl]-4-carboxylate
The following were added to a flask: imidazole (8.79 g, 129 mmol),
triphenylphosphine (16.94 g, 65 mmol), iodine 16.39 g, 65 mmol) and THF (525 mL). A solution of Compound 3 (15 g, 32 mmol) in THF (375 mL) was added. The solution was heated to 60 °C and was held at 60 °C for 4 h. Once complete by HPLC analysis (4 h), the solution was concentrated and the residue was purified by column chromatography [Silica gel; 10% to 60% EtOAc in Hexanes] to afford Compound 4 (17.0 g, 92% yield) as a solid.
The next step of the sequence is to install the allyic moiety. Heating a solution of Compound 4 in toluene in the presence of DABCO (l,4-Diazabicyclo[2.2.2]octane) allows for elimination of the iodide. Other solvents, such as THF and DCE may be utilized, but toluene often provides the best conversion and yield. Other amine bases may be used in this transformation, but generally DABCO is preferred.
Image may be NSFW. Clik here to view.
Elimination
Preparation of ((4R,5R)-5-(5-methyl-2,4-dioxo-3,4-dihydropyrimidin-l (2H)-yl)-4-(vinyloxy)-4,5-dihydrofuran-2-yl)methyl [l,l’-biphenyl]-4-carboxylate
Compound 4 (17 g, 30 mmol) was dissolved into toluene (255 niL), and DABCO (10 g, 89 mmol) was added. The solution was heated to 90 °C and held there for 2 h. Once complete, the organics were washed with sat. aq. a2S203 (200 mL). The organics were then dried ( a2S04), filtered, and concentrated. The resulting residue was purified by column chromatography [Silica gel; 5% to 60% EtOAc in Hexanes] to yield
An important reaction in the sequence is the Claisen rearrangement. This reaction is utilized to install the quaternary stereocenter and the olefin geometry in the ring. Heating Compound 5 in benzonitrile at 190 °C for 2-3 hours allows for smooth conversion to Compound 6, and after chromatography, a 90% yield can be achieved.
Toluene (110 °C, 8 h) also works to provide the desired Compound 6 as a solid by simply cooling the reaction to 20 °C (no chromatography). Other solvents with boiling points over about 100°C may also be utilized.
Image may be NSFW. Clik here to view.
Claisen Rearrangement
Preparation of ((2S,5R)-5-(5-methyl-2,4-dioxo-3,4-dihydropyrimidin-l(2H)-yl)-2-(2-oxoethyl)-2,5-dihydrofuran-2-yl)methyl [l,l’-biphenyl]-4-carboxylate
Compound 5 (1 mmol) was dissolved into benzonitrile (10 mL). The solution was heated to 190 °C for 3 h. After cooling to 20 °C, the solution was purified by column chromatography [silica gel, 50:50 Hexanes:EtOAc] to afford Compound 6 (1 mmol).
Alternatively, Compound 5 (1 mmol) was dissolved into toluene (10 mL). The solution was heated to 110 °C and held for 12 h. Upon cooling to 20 °C, a slurry formed. The solids were filtered, washed (PhMe) and dried (50 °C, 25 mmHg) to afford
The alkyne formation is performed by first treating Compound 6 with TMSCl (Trimethylsilyl chloride)/Et3N. NfF (Nonafluoro- 1 -butanesulfonyl fluoride) and P-base () are then added at -20 °C. After warming to 20 °C, the desired alkyne Compound 7 can be isolated in about 80 % yield. Initially, TMSCl is presumed to react at the NH moiety. NfF/P-base then reacts with the aldehyde to form the enol Nonaflate. Upon warming to 20 °C in the presence of P-base, the enol Nonaflate eliminates smoothly to the alkyne Compound 7. Without the TMSCl/Et3N, the yields are only -25%.
Image may be NSFW. Clik here to view.
Alkyne formation
Preparation of ((2R,5R)-2-ethynyl-5-(5-methyl-2,4-dioxo-3,4-dihydropyrimidin-l(2H)-yl)-2,5-dihydrofuran-2-yl)methyl [l,l’-biphenyl]-4-carboxylate
Compound 6 (1 g, 2.24 mmol) was dissolved into DMF (Dimethylformamide) (5 mL). (Other polar solvents could also have been used.) Triethylamine (406 uL, 2.91 mmol) was added and the solution was cooled to 0 °C. TMSCl (314 uL, 2.46 mmol) was added and the solution was allowed to stir at 0 °C for 30 min. The solution was then cooled to -20 °C, and NfF (484 uL, 2.69 mmol) was added and the solution was allowed to stir at -20 °C for 5 min. Phosphazane P l-base (1.54 mL, 4.93 mmol) was added
dropwise over 20 min. The solution was then allowed to warm to 20 °C and held for 20 h. The solution was then poured into water (50 mL) and extracted with DCM (100 mL). The organics were concentrated and the resulting residue was purified by column chromatography [Silica gel; 10% to 60% EtOAc in Hexanes] to afford Compound 7 (816 mg, 85% yield) as a solid.
The final step of the sequence is to remove the aromatic ester protecting group. This consists of hydrolysis by NaOH in aq. THF solution. The API is extracted into THF and then crystallized from THF/PhMe.
Image may be NSFW. Clik here to view.
Deprotection
Preparation of l-((2R,5R)-5-ethynyl-5-(hydroxymethyl)-2,5-dihydrofuran-2-yl)-5-methylpyrimidine-2,4(lH,3H)-dione (Ed4T)
Compound 7 (10 g, 23.40 mmol) was dissolved into THF (100 mL). 3N NaOH (10 mL) was added. The solution was allowed to stir at 20 °C for 12 h. The layers were split and the organics were kept. The organics were concentrated to reach a KF <1 wt%. Toluene (100 mL) was added, and solids crashed out of solution. The solids were filtered and washed with Toluene (100 mL). The solids were then dried (50 °C, 25 mmHg) to afford Festinavir (5.21 g, 90% yield) as a white solid.
Synthesis of (±)-4′-ethynyl-5′,5′-difluoro-2′,3′-dehydro-3′-deoxy- carbocyclic thymidine: a difluoromethylidene analogue of promising anti-HIV agent Ed4T
For the synthesis of 2‘,3‘-didehydro-3‘-deoxy-4‘-ethynylthymidine (8: 4‘-Ed4T), a recently reported promising anti-HIV agent, a new approach was developed. Since treatment of 1-(2,5-dideoxy-β-l–glycero-pent-4-enofuranosyl)thymine with Pb(OBz)4 allowed the introduction of the 4‘-benzoyloxy leaving group, nucleophilic substitution at the 4‘-position became feasible for the first time. Thus, reaction between the 4‘-benzoyloxy derivative (14) and Me3SiC⋮CAl(Et)Cl as a nucleophile led to the isolation of the desired 4‘-“down”-ethynyl derivative (18) stereoselectively in 62% yield. As an application of this approach, other 4‘-substituted nucleosides, such as the 4‘-allyl (24a) and 4‘-cyano (26a) derivatives, were synthesized using organosilicon reagents. In these instances, pretreatment of 14 with MeAlCl2 was necessary.
Image may be NSFW. Clik here to view.
PATENTS
US75890782009-09-15Anti-viral nucleoside analogs and methods for treating viral infections, especially HIV infections
Register now for the inaugural ACS Industry Symposium, 11-12 November 2016 in Hyderabad, India. Be sure to secure your seat today as rates will increase on 27 October!
http://acssymposium.org.in/
The theme of the Symposium is Recent Advances in Drug Development. The event will feature lectures by the world’s leading researchers and experts in the pharma industry, including:
Dr. Peter Senter of Seattle Genetics
Dr. Jagath Reddy Junutula of Cellerant Therapeutics, Inc.
Dr. Ming-Wei Wang of the Shanghai Institute of Materia Medica, Chinese Academy of Sciences
This is an exclusive event being organized in partnership with Dr. Reddy’s Laboratories for pharma professionals throughout India. Space is limited so register today!
Please visit our website to learn more about the speakers and the program.
Register today to ensure your access to the ACS Industry Symposium. We look forward to seeing you in Hyderabad in November.
sodium salt of (2S,5R)-sulfuric acid mono-(2-[1,3,4]oxadiazol-2-yl-7-oxo-1,6-diaza-bicyclo[3.2.1]oct-6-yl)ester
CAS 1628207-16-8
C8 H10 N4 O6 S . Na
Sulfuric acid, mono[(1R,2S,5R)-2-(1,3,4-oxadiazol-2-yl)-7-oxo-1,6-diazabicyclo[3.2.1]oct-6-yl] ester, sodium salt (1:1)
PATENTS
WO2013149121
US 20140275001
US 20150094472
WO 2016081452
Infection, multidrug resistant bacteria (MDR) in phase 1 at Merck
CB-618 is in phase I clinical trails by Cubist for the treatment of resistant bacterial infections, including carbapenem-resistant Enterobacteriaceae and Klebsiella pneumonia carbapenemases infection.
CB-618 is a beta-Lactamase inhibitor in phase I clinical trials at Merck & Co. for the treatment of multidrug resistant bacterial infections, including those caused by carbapenem-resistant Enterobacteriaceae and Klebsiella pneumoniae carbapenemases.
The product was originally developed at Cubist. In 2015, Merck & Co. acquired the company
Originator Cubist Pharmaceuticals
Class Antibacterials
Mechanism of Action Beta lactamase inhibitors
Highest Development Phases
Phase I Gram-negative infections
Most Recent Events
01 Apr 2015 Cubist Pharmaceuticals completes a phase-I clinical trial in Gram-negative infections in USA (IV) (NCT02341599)
21 Jan 2015 Cubist Pharmaceuticals has been acquired by Merck & Co
14 Jan 2015 Phase-I clinical trials in Gram-negative infections in USA (IV)
Bacterial resistance to β-lactam antibiotics, especially in Gram-negative bacteria, is most commonly mediated by β-lactamases. β-lactamases are enzymes that catalyze the hydrolysis of the β-lactam ring, which inactivates the antibacterial activity of the β-lactam antibiotic and allows the bacteria to become resistant. Inhibition of the β-lactamase with a BLI slows or prevents degradation of the β-lactam antibiotic and restores β-lactam antibiotic susceptibility to β-lactamase producing bacteria. Many of these β-lactamases are not effectively inhibited by BLIs currently on the market rendering the β-lactam antibiotics ineffective in treating bacteria that produce these β-lactamases. There is an urgent need for novel BLIs that inhibit β-lactamases that are not effectively inhibited by the current clinical BLIs (e.g. KPC, class C and class D β-lactamases) and that could be used in combination with β-lactam antibiotics to treat infections caused by β-lactam resistant bacteria.
Step 1: Ι,Γ-Carbonyldiimidazole (5.8 g, 36.2 mmol) was added to a 0 °C solution of (2S,5R)- 6-(benzyloxy)-7-oxo-l,6-diazabicyclo[3.2.1]octane-2-carboxylic acid (5.0 g, 18.1 mmol) in dry THF (200 mL). The reaction mixture was allowed to warm to rt then was stirred at rt for 3 hrs. Formohydrazide (5.4 g, 90.5 mmol) was added in one portion, and the reaction mixture was stirred for additional 3 hrs. The mixture was then diluted with saturated sodium chloride and exatracted with EtOAc (3x). The combined organic layer was washed with saturated sodium chloride (2x), dried over Na2S04, and concentrated to afford crude (25,5 ?)- 6-(benzyloxy)-N-formyl-7-oxo-l,6-diazabicyclo[3.2.1]octane-2-carbohydrazide (-11 g), which was directly used in the next step. ESI-MS (Ef , m/z): 319.1 [M+H]+.
Step 2: To a -10 °C solution of (25′,5«)-6-(benzyloxy)-N-formyl-7-oxo-l,6- diazabicyclo[3.2.1]octane-2-carbohydrazide (11 g) in dry DCM (200 mL) was added pyridine (28 mL), followed by dropwise addition of (CF3S02)20 (28 mL). The reaction mixture was allowed to warm to rt and was stirred for 3 hrs. The reaction mixture was then cooled to -10 °C and quenched with sat. NaHCC>3. The organic layer was separated and the aqueous layer was extracted with EtOAc (3x). The combined organic layer was dried over Na2S04, concentrated and purified by silica gel column chromatography (gradient elution 1 :3 to 2: 1 EtOAc/hexanes) to give (25,5/?)-6-(benzyloxy)-2-(l,3,4-oxadiazol-2-yl)-l ,6- diazabicyclo[3.2.1]octan-7 -one (4.6 g, 86% for two steps) as a slightly yellow solid. ESI-MS (EI+, m/z): 301.0 [M+H]+.
Step 3: To a solution of (25,5/?)-6-(benzyloxy)-2-(l,3,4-oxadiazol-2-yl)-l ,6- diazabicyclo[3.2.1]octan-7-one (4.6 g, 15.3 mmol) in THF (150 mL) was added 10% Pd/C (1 g). The mixture was stirred under H2 atmosphere at rt for 3 hrs. The reaction mixture was then filtered and concentrated to afford (25,5/?)-6-hydroxy-2-(l,3,4-oxadiazol-2-yl)-l,6- diazabicyclo[3.2.1]octan-7-one (2.9 g, 91 %), which was used directly in the next step. ESI- MS (EI+, m/z): 211.1 [M+H]+. Step 4: To a solution of (25,5fl)-6-hydroxy-2-(l,3,4-oxadiazol-2-yl)-l,6- diazabicyclo[3.2.1]octan-7-one (2.9 g, 13.8 mmol) in dry pyridine (60 mL) was added SC>3- Py (11.0 g, 69.0 mmol). The reaction mixture was stirred at rt for 8 hrs and then concentrated under vacuum. The residue was re-dissolved in aqueous NaH2PC>4 (1.5 M, 100 mL) then tetrabutylammonium hydrogensulphate (5.88 g, 17.3 mmol) was added. The mixture was stirred at rt for 20 minutes, then was extracted with EtOAc (4x). The combined organic layer was dried and concentrated and the residue was purified by silica gel column chromatography (gradient elution 10:1 to 2:1 DCM/acetone) to afford tetrabutylammonium (25,5/?)-2-(l ,3,4-oxadiazol-2-yl)-7-oxo-l,6-diazabicyclo[3.2.1]octan-6-yl sulfate (4.1 g, 97%) as a white solid. ESI-MS (EL, m/z): 289.0 [M-H]\ lH NMR (400 MHz, CDC13): δ 8.48 (s, 1H), 4.75 (d, / = 6.5 Hz, 1H), 4.40 (br s, 1H), 3.34-3.26 (m, 9H), 2.82 (d, / = 12.0 Hz, 1H), 2.37-2.25 (m, 3H), 2.06-1.98 (m, 1H), 1.71-1.65 (m, 8H), 1.49-1.42 (m, 8H), 1.01 (t, / = 7.5 Hz, 12H).
Step 5: Resin Exchange: Tetrabutylammonium (25, 5R)-2-(l, 3, 4-oxadiazol-2-yl)-7-oxo-l, 6-diaza-bicyclo[3.2.1]octan-6-yl sulfate (4.1 g, 7.72 mmol) was dissolved in a minimum amount of HPLC grade water (~ 40 mL) and passed through a column of 80 g of DOWEX 50WX 8 Na+ resin (the resin was prewased with >4 L of HPLC grade water) and eluted with HPLC grade water to afford sodium (25,5fl)-2-(l,3,4-oxadiazol-2-yl)-7-oxo-l,6- diazabicyclo[3.2.1]octan-6-yl sulfate (2.2 g, 91 %) as a white solid after lyophilization. ESI- MS (EI+, m/z): 291.2 [M+H]+. lH NMR (300 MHz, D20) δ 8.92 (s, 1H), 4.84 (d, J = 6.1 Hz, 1H), 4.20 (br s, 1H), 3.25-3.16 (m, 1H), 2.92 (d, / = 12.3 Hz, 1H), 2.41-2.26 (m, 1H), 2.26- 2.11 (m, 2H), 2.04-1.89 (m, 1H).
A compound of Formula (I), chemically known as sodium salt of 2S, 5R) mono-(2-[l,3,4]oxadiazol-2-yl-7-oxo-l,6-diazabicyclo[3.2.1 ]oct-6-yl)ester has antibacterial properties and is disclosed in PCT International Patent Application No. PCT/US2013/034562. The compound of Formula (I) is also generically disclosed in PCT International Patent Application No. PCT/IB2012/054296. The present invention discloses a process for preparation of a compound of Formula (I).
Formula (I)
Image may be NSFW. Clik here to view.
Scheme 1.
Image may be NSFW. Clik here to view.
Image may be NSFW. Clik here to view.
(VI) Compound of Formula (I)
Example 1
Sodium salt of (25, 5R) sulfuric acid mono-(2-[l,3,4]oxadiazol-2-yl-7-oxo-l,6-diaza-bicyclo[3.2.1]oct-6-yl) ester
Step I: Synthesis of (25,5R)-2-(iV’-formyl-hydrazinocarbonyl)-6-benzyloxy-7-oxo-l,6-diaza-bicyclo[3.2.1] octane (III):
To a turbid solution of sodium salt of (2<S’,5i?)-6-benzyloxy-7-oxo-l,6-diaza-bicyclo[3.2.1 ] octane-2-carboxylic acid (II, 20 g, 0.067 mol) (prepared according to process disclosed in PCT/IB2013/059264) in dimethylformamide (200 ml) was added EDC hydrochloride (19.44 g, 0.10 mol) followed by formyl hydrazide (4.02 g, 0.067 mol) and N-hydroxybenzotriazole (9 g, 0.67 mol) at about 25°C under stirring. Diisopropylethylamine (35.62 ml, 0.20 mol) was added to the reaction mixture and stirred at 25°C temperature for 18 hours. The reaction mixture was evaporated under vacuum to provide a residue. The residue was dissolved in ethyl acetate (500 ml) and washed with water (500 ml χ 2), followed by saturated aqueous sodium bicarbonate solution. The organic layer was dried over anhydrous sodium sulphate and evaporated under vacuum to provide a crude intermediate, which was purified by silica gel column chromatography to provide 11 g of the titled compound as solid in 52% yield.
Analysis:
Mass: 319.1 (M+l); for Molecular Formula of C15H18N4O4 and Molecular Weight of 318.34;
Step II: Synthesis of (25,5R)-2-([l,3,4]-oxadiazol-2-yl)-6-benzyloxy-7-oxo-l,6-diaza-bicyclo[3.2.1] octane (IV):
To a clear solution of (2<S’,5i?)-2-(N’-formyl-hydrazinocarbonyl)-6-benzyloxy-7-oxo-l ,6-diaza-bicyclo[3.2.1 ] octane (III, 11 g, 0.0345 mol) in chloroform (120 ml) was added diisopropylethylamine (18.31 ml, 0.1035 mol) and p-tolylsulfonylchloride (9.83 g, 0.0517 mol). The solution was stirred at 60°C for 15 hours. Reaction mixture was cooled to room temperature and water (100 ml) was added. Organic layer was dried over anhydrous sodium sulphate and evaporated under vacuum to provide a crude residue, which was purified by silica gel column chromatography to provide 7 g of the titled compound as a solid in 68% yield.
Analysis:
Mass: 301.3 (M+l); for Molecular Formula of Ci5Hi6N403 and Molecular Weight of 300.32;
Step III: Synthesis of (25,5R)-2-([l,3,4]-oxadiazol-2-yl)-6-hydroxy-7-oxo-l,6-diaza-bicyclo[3.2.1] octane (V):
To a clear solution of (2<S’,5i?)-2-([l,3,4]-oxadiazol-2-yl)-6-benzyloxy-7-oxo-l,6-diaza-bicyclo[3.2.1 ] octane (IV, 7.0 g, 0.0233 mmol) in methanol (70 ml) was added 10% palladium on carbon (2.5 g). The suspension was stirred under atmospheric hydrogen pressure at a temperature 25° C for 2 hrs. The catalyst was filtered over a celite bed and the bed was washed with methanol (30 ml). The filtrate was concentrated under vacuum to provide an oily residue. The residue was triturated with cyclohexane (100 ml) to effect solid formation. The suspension was filtered under suction and the wet cake was washed with additional cyclohexane (50 ml). The soild was dried under vacuum to provide 4.5 g of the titled compound as a whitish solid in 92% yield, which was used for the next reaction immediately.
Analysis:
Mass: 211.2 (M+l); for Molecular Formula of C8Hi0N4O3 and Molecular Weight of 210.19; 1H NMR (DMSO-d6): δ 9.88 (br s, 1H), 9.29 (s, 1H), 4.65 (d, 1H ), 4.64 (br s, 1H), 2.94-2.97 (br d, 1H), 2.63-2.66 (d, 1H), 1.89-2.09 (m,3H), 1.82-1.86 (m, 1H).
Step IV: Synthesis of tetrabutylammonium salt of (25, 5R)-sulfuric acid mono-(2-[l,3,4]oxadiazol-2-yl-7-oxo-l,6-diaza-bicyclo[3.2.1]oct-6-yl) ester (VI):
To a clear solution of (2<S’,5i?)-2-([l,3,4]-oxadiazol-2-yl)-6-hydroxy-7-oxo-l ,6-diaza-bicyclo[3.2.1 ] octane (V, 4.5 g, 0.0214 mol) in dichloromethane (50 ml) was added triethylamine (9 ml, 0.642 mol), followed by the addition of sulfur trioxide pyridine complex (6.83 g, 0.428 mol). The resulting reaction mixture was stirred for 2 hours. Tetrabutylammonium hydrogen sulfate (7.26 g,
0.0214 mol) was added to the reaction mixture and it was stirred for 1.5 hours. A solution of aqueous 0.5 N KH2PO4 (100 ml) was added to the reaction mixture. Layers were separated and the aqueous layer was washed with dichloromethane (125 ml). Combined organic layer was dried over Na2S04, and was evaporated under vacuum to yield crude foam, which was purified on silica gel column chromatography to give 7 g of the titled compound as white foam in 98% yield.
Analysis:
Mass: 289.1 (M-l); for Molecular Formula Image may be NSFW. Clik here to view.
and Molecular Weight of 517.26;
Step V: Sodium salt of (25, 5R)-sulfuric acid mono-(2-[l,3,4]oxadiazol-2-yl-7-oxo-l,6-diaza-bicyclo[3.2.1]oct-6-yl) ester (I):
The compound sodium salt of (2S, 5i?)-sulfuric acid mono-(2-[l,3,4]oxadiazol-2-yl-7-oxo-l,6-diaza-bicyclo[3.2.1]oct-6-yl) ester of Formula (I) was prepared by loading tetrabutylammonium salt of sulfuric acid mono-(2-[l,3,4]oxadiazol-2-yl-7-oxo-l,6-diaza-bicyclo[3.2.1]oct-6-yl) ester (VI, 7 g) on a column packed with Amberlite IR 120 Na form of resin, and by eluting the column with methanol water mixture (9: 1). Fractions containing compound were collected and solvent was evaporated under vacuum below 40°C, to provide formula- 1 compound in 4 gm (62%) quantity as a white solid.
Analysis:
Mass: 289.3 (M-l) as free acid; for Molecular Formula Image may be NSFW. Clik here to view.
and Molecular Weight 290.26;
Ranitidine Synthetic procedure/method of synthesis
The reaction of 5-dimethylaminomethyl-2-furanylmethanol (I) with 2-mercaptoethylamine (II) by means of aqueous HCl gives 2-[[(5-dimethylamino-methyl-2-furanyl)methylthio]ethaneamine (III), which is then condensed with N-methyl-1-methylthio-2-nitrotheneamine (IV) by heating at 120 C. Compound (IV) is obtained by reaction of 1,1-bis(methylthio)-2-nitroethene (V) with methylamine in refluxing ethanol
aDepartment of Chemistry, University of California Davis, 1 Shields Avenue, Davis, US E-mail: mascal@chem.ucdavis.edu Fax: 530-752-8995 Tel: 530-754-5373
The biomass-derived platform chemical 5-(chloromethyl)furfural is converted into the blockbuster antiulcer drug ranitidine (Zantac) in four steps with an overall 68% isolated yield.
Image may be NSFW. Clik here to view.
Image may be NSFW. Clik here to view.
Image may be NSFW. Clik here to view.
PROCESS
Image may be NSFW. Clik here to view.
2. Experimental Procedures
5-[[(2-Acetamidoethyl)thio]methyl]furfural 14
Sodium hydride (95%) (103 mg, 4.08 mmol) was added to a solution of Nacetylcysteamine (0.4051 g, 3.40 mmol) in dry THF (20 mL) under argon. The resulting suspension was stirred at RT for 30 min and a solution of CMF 12 (0.4912 g, 3.40 mmol) in dry THF (10 mL) was added dropwise over a 10 min period. The resulting light yellow solution was allowed to stir overnight at RT. The solvent was evaporated and saturated brine (50 mL) was added. The mixture was extracted with CH2Cl2 (2 × 50 mL) and the organic layers were combined and washed with saturated brine (100 mL). The organic layer was dried over Na2SO4. Charcoal (100 mg) was added and the mixture was stirred for 20 min and filtered. The solvent was evaporated to give 14 as a yellow liquid (0.7042 g, 91 %). 1H NMR (CDCl3, 300 MHz) 9.58 (1H, s), 7.21 (1H, d, J = 3.6 Hz), 6.48 (1H, s, br), 5.95 (1H, d, J = 3.6 Hz), 3.79 (2H, s), 3.45 (2H, q, J = 6.3 Hz), 2.72 (2H, t, J = 6.6 Hz), 2.00 (3H, s); 13C NMR (CDCl3, 75 MHz) 23.1, 27.8, 31.7, 38.4, 110.7, 121.9, 152.2, 158.9, 170.7, 177.4; IR (neat) 3298, 3101, 1663, 1548, 1512, 1287, 1022, 772 cm-1; HRMS (ESI): calculated for C10H14O3NS: [M+H]+ 228.0694: found 228.0690.
Me2NH (1.0 mL) was added to a solution of 14 (0.2105 g, 0.926 mmol) in dry methanol (20 mL) and the mixture was stirred at RT for 1 h. The resulting red solution was cooled to 0 °C and NaBH4 (98 %) (55 mg, 1.42 mmol) was added over a 5 min period. The mixture was allowed to come to RT and stirred for 30 min. The solvent was evaporated while keeping the bath temperature below 45 °C. The residue was dissolved in CH2Cl(50 mL) and filtered to remove inorganic impurities. The solvent was evaporated to give 15 (0.2145 g, 90 %) as a pale yellow oil. 1H NMR (CDCl3, 300 MHz) 6.42 (1H, s, br), 6.09 (1H, s), 3.67 (2H, s), 3.37 (2H, s), 3.26 (2H, q, J = 6.0 Hz), 2.62 (2H, t, J = 6.4 Hz) 2.21 (6H, s), 1.93 (3H, s); 13C NMR (CDCl3, 75 MHz) 23.5, 28.4, 31.9, 38.7, 45.4, 56.2, 108.4, 109.9, 151.4, 152.1, 170.5; IR (neat) 3273, 2944, 1656, 1545, 1291, 1019, 729 cm- 1 ; HRMS (ESI): calculated for C12H21O2N2S: [M+H]+ 257.1322: found 257.1323.
A solution of 15 (0.2473 g, 0.965 mmol) in freshly prepared 2N aq NaOH (10 mL) was heated at reflux for 2 h. The mixture was cooled to RT and extracted with CH2Cl2 (3×30 mL). The organic layers were combined and washed with saturated brine, dried over Na2SO4, and evaporated to give 5 (0.1934 g, 94 %) as a pale yellow oil. 1H NMR (CDCl3, 300 MHz) 6.02 (2H, s), 3.61 (2H, s), 3.33 (2H, s), 2.74 (2H, t, J = 6.3 Hz), 2.52 (2H, t, J = 6.6 Hz), 2.16 (6H, s); 13C NMR (CDCl3, 75 MHz) 28.2, 35.9, 40.9, 45.1, 55.9, 108.1, 109.5, 151.4, 152.1; IR (neat) 3359 cm-1, 2947, 2769, 1559, 1459, 1015, 797 cm-1; HRMS (ESI): calculated for C10H19ON2S: [M+H]+ 215.1212: found 215.1218.
N-[2-[[[5-[(dimethylamino)methyl]-2-furanyl]methyl]thio]ethyl]-N’-methyl-2-nitro- 1-Ethenediamine (Ranitidine) 1 The experimental procedure is modified from existing literature:2 A solution of 5 (0.1501 g, 0.700 mmol ) in distilled water (10 mL) was added dropwise over a period of 10 min to a suspension of 1-methylthio-1-methylamino-2-nitroethylene 7 (0.1041 g, 0.703 mmol) in distilled water (5 mL) with stirring. The resulting light yellow solution was placed in an oil bath at 55 °C and the mixture was stirred at that temperature overnight. Saturated brine (30 mL) was added and the mixture was extracted with CHCl3 (3×20 mL). The combined organic layer was dried over Na2SO4. Evaporation of the solvent gave 1 as a pale yellow oil (0.1935 g, 88 %). 1H NMR (CDCl3, 300 MHz, 56 oC) 10.23-10.15 (1H, br, NH), 6.57 (1H, s), 6.13 (2H, d, 6.0 Hz), 5.04 (1H, br, NH), 3.73 (2H, s), 3.41 (4H, s), 2.92 (2H, s), 2.76 (2H, t, 6.0 Hz), 2.24 (6H, s); 13C NMR (CDCl3, 75 MHz, 56 °C) 28.2, 30.6, 40.7, 44.6, 55.6, 97.9, 108.1, 109.1, 150.4, 152.1, 156.6; IR (neat) 3209, 2944, 2815, 2776, 1620, 1574, 1384, 1230, 1019, 761 cm-1; HRMS (ESI): calculated for C13H23O3N4S: [M+H]+ 315.1491: found 315.1497.
The paper was found in Green Chemistry,“Synthesis of ranitidine (Zantac) from cellulose-derived 5-(chloromethyl)furfural” by Mark Mescal et al, Green Chemistry, 2011,13, 3101-3102, DOI: 10.1039/c1gc15537g. Once again, I am beating the press before they print so I supplied the Digital Object Identifier. I am sure the sales for Ranitidine are quite large; who doesn’t get heartburn at one time or another. I think it is very fortunate the author shows you can use a starting material that can be derived from just about any source of cellulose. I find it interesting how renewable feedstocks can be utilized in industry and become part of important commodities, such as plastics, pharmaceuticals, etc. This paper refers to another discussing where the starting material was derived from. Starting material can be sugars, cellulose or raw cellulosic biomass and the reaction can produce yields of 80-90 %. M.Mascal and E. B. Nikitin, Angew. Chem., Int. Ed., 2008, 47, 7924;Image may be NSFW. Clik here to view.On with the show, though. The original synthetic route was provided in the paper and I will provide it to you.
Image may be NSFW. Clik here to view.
Furfural 1 was reduced to give the furfuryl alcohol 2. The furfuryl alcohol is methylaminated to give 3, which is reacted with cysteamine in concentrated HCl to give 4. This is condensed with 1-methylthio-1-methylamino-2-nitroethylene to give the final product. The patent literature has the yield < 50 % for the aminomethylation and subsequent reaction with cysteamine, but recently, these steps have been reported to have higher conversions.
Image may be NSFW. Clik here to view.
This new synthesis, apart from using a renewable feedstock as a starting material, has synthetic steps with an average yield of 91 %, and requires no chromatography. Note that N-acetylcysteamine was used as opposed to cysteamine in the first step, in high yield. A reductive amination with methylamine gives 8 again in high yield. Treatment with KOH provides the free amine 9 and the final step is the condensation with the nitroethylene used in the previous synthesis
Critical influence of 5-hydroxymethylfurfural aging and decomposition on the utility of biomass conversion in organic synthesis
Angewandte Chemie, International Edition (2016), 55, (29), 8338-8342
Spectral studies revealed the presence of a specific arrangement of 5-hydroxymethylfurfural (5-HMF) molecules in solution as a result of a hydrogen–bonding network, and this arrangement readily facilitates the aging of 5-HMF. Deterioration of the quality of this platform chemical limits its practical applications, especially in synthesis/pharma areas. The model drug Ranitidine (Zantac®) was synthesized with only 15 % yield starting from 5-HMF which was isolated and stored as an oil after a biomass conversion process. In contrast, a much higher yield of 65 % was obtained by using 5-HMF isolated in crystalline state from an optimized biomass conversion process. The molecular mechanisms responsible for 5-HMF decomposition in solution were established by NMR and ESI-MS studies. A highly selective synthesis of a 5-HMF derivative from glucose was achieved using a protecting group at O(6) position.
PAPER
Phytochemical screening and investigation of antiulcer activity of Tridax procumbens
International Journal of Pharmacy and Technology (2015), 6, (4), 7679-7690
Lavanya Asula* , A. Sony John, Deepthi Kotturi, P. Srividyalaxmi, R. Soni and Y. Mamatha Kalyani Department of Pharmacy, Jawaharlal Nehru Technological University, Holy Mary Institute of Technology and Science College of Pharmacy Hyderabad, India. Email: lavanya.asula@gmail.com
Waste gas treatment and methyl mercaptan recovery process in production process of cimetidine and ranitidine
Image may be NSFW. Clik here to view.
cimetidine and ranitidine terms widely used in the treatment of stomach is bound to promote the continuous mass production of APIs, however, the raw material in the manufacturing process of the drug inevitably produce methyl mercaptan, dimethyl sulfide, a methylamine, carbon disulfide and nitromethane workshop emissions.Because of methyl mercaptan, dimethyl sulfide into the atmosphere having foul odor.Resulting in the production shop around smelling, and even affect the normal life of residents of several kilometers around.So some manufacturers use incineration method expects to dispose of the waste gas combustion, which reduces air pollution to some extent.But using incineration method has two drawbacks: one gas methyl mercaptan, dimethyl sulfide gas combustion higher value produce a few meters of flames burning heat generated while it is easy to burn incinerator, security posed by the chemical production big risk; on the other hand by a combustion method can not solve the odor problem, air pollution is still grim, because incomplete combustion, odor difficult to eliminate people’s sense of smell is particularly sensitive to the perception of mercaptans, while burning a large amount of sulfur dioxide in the same air pollution.There’s manufacturers to adopt authoritarian incinerator burning after the first use of chlorine dioxide generator eliminate odor, although this method has a certain smell to eliminate the effect of improving, but requires authoritarian equipment, increasing the cost of gas treatment and discharge sulfur dioxide into the air is still there.
CAS Name:N-[2-[[[-5-[(Dimethylamino)methyl]-2-furanyl]methyl]thio]ethyl]-N¢-methyl-2-nitro-1,1-ethenediamine
Molecular Formula: C13H22N4O3S
Molecular Weight: 314.40
Percent Composition: C 49.66%, H 7.05%, N 17.82%, O 15.27%, S 10.20%
Literature References: Histamine H2-receptor antagonist which inhibits gastric acid secretion. Prepn: B. J. Price et al.,FR2384765; eidem,US4128658 (both 1978 to Allen & Hanburys). HPLC determn in plasma: P. F. Carey, L. E. Martin, J. Liq. Chromatogr.1979, 1291. Pharmacological studies: J. Bradshaw et al.,Br. J. Pharmacol.66, 464 (1979); M. J. Daly et al.,Gut21,408 (1980). Efficacy in treatment of duodenal ulcers: A. Berstad et al.,Scand. J. Gastroenterol.15, 637 (1980); R. P. Walt et al.,Gut22, 49 (1981). Review of pharmacology and therapeutic use: R. N. Brogden et al.,Drugs24, 267-303 (1982). Comprehensive description: M. Hohnjec et al.,Anal. Profiles Drug Subs.15, 533-561 (1986).
Percent Composition: C 44.50%, H 6.61%, N 15.97%, O 13.68%, S 9.14%, Cl 10.10%
Properties: Off-white solid, mp 133-134°. Freely sol in acetic acid and water, sol in methanol, sparingly sol in ethanol. Practically insol in chloroform.
Melting point: mp 133-134°
Derivative Type: Bismuth citrate
CAS Registry Number: 128345-62-0
Additional Names: Ranitidine bismutrex
Manufacturers’ Codes: GR-122311X
Trademarks: Pylorid (GSK); Tritec (GSK)
Molecular Formula: C13H22N4O3S.C6H5BiO7
Molecular Weight: 712.48
Percent Composition: C 32.03%, H 3.82%, N 7.86%, O 22.46%, S 4.50%, Bi 29.33%
Literature References: Pharmacology and activity vs Helicobacter sp: R. Stables et al.,Aliment. Pharmacol. Ther.7, 237 (1993).
Jump up^Fedorowicz, Z; van Zuuren, EJ; Hu, N (14 March 2012). “Histamine H2-receptor antagonists for urticaria.”. The Cochrane database of systematic reviews. 3: CD008596.doi:10.1002/14651858.CD008596.pub2. PMID22419335.
The GMP/GDP Inspectors Working Group of the European Medicines Agency (EMA) has published a concept paper to summarise the GMP responsibilities of the Marketing Authorisation Holders (MAH). It is not intended to introduce any new responsibilities on MAHs but to document existing requirements in a better way.
The current EU GMP-Guidelines define in several chapters and annexes GMP tasks and responsibilities of the MAH. However, there seems to be a lack of clarity and understanding as to what these responsibilities actually are in their totality, and what they mean for MAHs at a practical level. All these tasks and responsibilities have now been summarised in this concept paper:
Chapter 1: responsibility to evaluate the results of the PQR review
Chapter 7: responsibility to put contracts in place
Green Chemistry International
Image may be NSFW.
Image may be NSFW.
Image may be NSFW.
Image may be NSFW.
Image may be NSFW.
Image may be NSFW.
Image may be NSFW.
