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DR ANTHONY MELVIN CRASTO Ph.D

DR ANTHONY MELVIN CRASTO Ph.D

DR ANTHONY MELVIN CRASTO, Born in Mumbai in 1964 and graduated from Mumbai University, Completed his Ph.D from ICT, 1991,Matunga, Mumbai, India, in Organic Chemistry, The thesis topic was Synthesis of Novel Pyrethroid Analogues, Currently he is working with GLENMARK PHARMACEUTICALS LTD, Research Centre as Principal Scientist, Process Research (bulk actives) at Mahape, Navi Mumbai, India. Total Industry exp 30 plus yrs, Prior to joining Glenmark, he has worked with major multinationals like Hoechst Marion Roussel, now Sanofi, Searle India Ltd, now RPG lifesciences, etc. He has worked with notable scientists like Dr K Nagarajan, Dr Ralph Stapel, Prof S Seshadri, Dr T.V. Radhakrishnan and Dr B. K. Kulkarni, etc, He did custom synthesis for major multinationals in his career like BASF, Novartis, Sanofi, etc., He has worked in Discovery, Natural products, Bulk drugs, Generics, Intermediates, Fine chemicals, Neutraceuticals, GMP, Scaleups, etc, he is now helping millions, has 9 million plus hits on Google on all Organic chemistry websites. His friends call him Open superstar worlddrugtracker. His New Drug Approvals, Green Chemistry International, All about drugs, Eurekamoments, Organic spectroscopy international, etc in organic chemistry are some most read blogs He has hands on experience in initiation and developing novel routes for drug molecules and implementation them on commercial scale over a 30 year tenure till date Dec 2017, Around 35 plus products in his career. He has good knowledge of IPM, GMP, Regulatory aspects, he has several International patents published worldwide . He has good proficiency in Technology transfer, Spectroscopy, Stereochemistry, Synthesis, Polymorphism etc., He suffered a paralytic stroke/ Acute Transverse mylitis in Dec 2007 and is 90 %Paralysed, He is bound to a wheelchair, this seems to have injected feul in him to help chemists all around the world, he is more active than before and is pushing boundaries, He has 9 million plus hits on Google, 2.5 lakh plus connections on all networking sites, 50 Lakh plus views on dozen plus blogs, He makes himself available to all, contact him on +91 9323115463, email amcrasto@gmail.com, Twitter, @amcrasto , He lives and will die for his family, 90% paralysis cannot kill his soul., Notably he has 19 lakh plus views on New Drug Approvals Blog in 216 countries......https://newdrugapprovals.wordpress.com/ , He appreciates the help he gets from one and all, Friends, Family, Glenmark, Readers, Wellwishers, Doctors, Drug authorities, His Contacts, Physiotherapist, etc

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LY 3104607


imgChemSpider 2D Image | LY3104607 | C27H25N3O3
FDIWCHYTKOPHPS-QFIPXVFZSA-N.png
 LY3104607
(3S)-3-[4-[[2-(2,6-Dimethylphenyl)-[l,2,4]triazolo[l,5-a]pyridin-6-yl]methoxy]phenyl]hex-4-ynoic Acid
(3S)-3-[4-[[2-(2,6-dimethylphenyl)-[1,2,4]triazolo[1,5-a]pyridin-6-yl]methoxy]phenyl]hex-4-ynoic acid
CAS: 1795232-22-2
Chemical Formula: C27H25N3O3
Molecular Weight: 439.515
(3S)-3-(4-{[2-(2,6-Dimethylphenyl)[1,2,4]triazolo[1,5-a]pyridin-6-yl]methoxy}phenyl)-4-hexinsäure
Benzenepropanoic acid, 4-[[2-(2,6-dimethylphenyl)[1,2,4]triazolo[1,5-a]pyridin-6-yl]methoxy]-β-1-propyn-1-yl-, (βS)-

[+]Enlarge

Structure of LY3104607.
Credit: Tien Nguyen/C&EN

Presented by: Chafiq Hamdouchi, founder at Hamdouchi Pharmaceutical Consulting

Target: G-protein-coupled receptor 40 (GPR40), a receptor that modulates insulin secretion in cells

Disease: Type 2 diabetes

Reporter’s notes: Developed by Eli Lilly, LY3104607 joins the handful of GPR40 agonists recently offered by the company. The compound is not exactly a first disclosure as its structure was revealed in a January publication that describes its discovery and pharmacokinetic properties (J. Med. Chem. 2018, DOI: 10.1021/acs.jmedchem.7b01411). Hamdouchi, who worked on the molecule while at Eli Lilly, presented what the team learned about GPR40 and suggested that allosteric binding, binding which happens at a location other than the active site, may be a viable mode of action for GPR40 agonists.

Image result for LY3104607

Chafiq Hamdouchi

Image result

STR OF TITLE LY 3104607

str1 str2 str3

Paper

Discovery of LY3104607: A Potent and Selective G Protein-Coupled Receptor 40 (GPR40) Agonist with Optimized Pharmacokinetic Properties to Support Once Daily Oral Treatment in Patients with Type 2 Diabetes Mellitus

 Lilly Research Laboratories, A Division of Eli Lilly and Company, Lilly Corporate Center, DC: 0540, Indianapolis, Indiana 46285, United States
 Jubilant Biosys Research Center, Bangalore, India
J. Med. Chem.201861 (3), pp 934–945
DOI: 10.1021/acs.jmedchem.7b01411
Publication Date (Web): December 13, 2017
Copyright © 2017 American Chemical Society
*E-mail: hamdouchi_chafiq@lilly.comchafiq.hamdouchi@gmail.com. Phone: 317-797-4751.

Abstract

Abstract Image

As a part of our program to identify potent GPR40 agonists capable of being dosed orally once daily in humans, we incorporated fused heterocycles into our recently disclosed spiropiperidine and tetrahydroquinoline acid derivatives 12, and 3 with the intention of lowering clearance and improving the maximum absorbable dose (Dabs). Hypothesis-driven structural modifications focused on moving away from the zwitterion-like structure. and mitigating the N-dealkylation and O-dealkylation issues led to triazolopyridine acid derivatives with unique pharmacology and superior pharmacokinetic properties. Compound 4 (LY3104607) demonstrated functional potency and glucose-dependent insulin secretion (GDIS) in primary islets from rats. Potent, efficacious, and durable dose-dependent reductions in glucose levels were seen during glucose tolerance test (GTT) studies. Low clearance, volume of distribution, and high oral bioavailability were observed in all species. The combination of enhanced pharmacology and pharmacokinetic properties supported further development of this compound as a potential glucose-lowering drug candidate.

(3S)-3-[4-[[2-(2,6-Dimethylphenyl)-[l,2,4]triazolo[l,5-a]pyridin-6-yl]methoxy]phenyl]hex-4-ynoic Acid (4)

Compound 4 (LY3104607)

title compound as white solid (35.76 kg, 91%). LCMS m/z [M + H]+: calcd, 439.5; found, 439.2.
1H NMR (399.80 MHz, DMSO, δ): 12.22 (s, 1H), 9.13 (dd, J = 0.8, 1.5 Hz, 1H), 7.88 (dd, J = 0.8, 9.2 Hz, 1H), 7.75 (dd, J = 1.7, 9.2 Hz, 1H), 7.29–7.24 (m, 3H), 7.14–7.12 (m, 2H), 7.01–6.99 (m, 2H), 5.18 (s, 2H), 3.96–3.91 (m, 1H), 2.58 (d, J = 7.7 Hz, 2H), 2.06 (s, 6H), 1.75 (d, J = 2.4 Hz, 3H).
PATENT
WO 2015088868
Applicants: ELI LILLY AND COMPANY [US/US]; Lilly Corporate Center Indianapolis, Indiana 46285 (US)
Inventors: HAMDOUCHI, Chafiq; (US)

A Novel Triazolo-Pyridine Compound

This invention relates to triazolo-pyridine compounds or pharmaceutically acceptable salts thereof, and for use of compounds in therapy. Triazolo-pyridine compounds of this invention are activators of GPR-40.

GPR-40, also known as Free Fatty Acid Receptor 1 (FFA1 or FFAR1), is reported as predominately expressed at high levels in rodent pancreatic beta cells, insulinoma cell lines, and human islets. The glucose modulation of insulin secretion is an important feature of activating GPR-40. Compounds that effectuate GPR-40 activation are associated with stimulation of insulin secretion in a patient with type II diabetes (T2D). Compounds that are GPR-40 activators are desired for use in treatment of GPR-40 mediated conditions.

WO2004/041266 discloses GPR-40 receptor function regulators comprising a compound having an aromatic ring and a group capable of releasing a cation.

The present invention rovides compounds of the Formula la below:

la

Example 1

(3S)-3-[4-[[2-(2,6-Dimethylphenyl)-[l,2,4]triazolo[l,5-a]pyridin-6- yl]methoxy]phenyl]hex-4-ynoic acid

To a solution of ethyl (3S)-3-[4-[[2-(2,6-dimethylphenyl)-[l,2,4]triazolo[l,5-a]pyridin-6-yl]methoxy]phenyl]hex-4-ynoate (0.22 g, 0.47 mmol) in EtOH (20 mL) is added 5 N NaOH (0.3 mL) and the reaction mixture is stirred at 80 °C in a microwave instrument for 30 minutes. The reaction mixture is evaporated to dryness, diluted with water, and acidified with 6 N HC1 solution to pH ~ 3. The precipitated solid is filtered, washed with n-pentane, and dried to give the title compound as a white solid (0.155 g, 75%). LCMS m/z 440 (M+H)+.

Alternate Preparation, Example 1

To a solution of ethyl (3S)-3-[4-[[2-(2,6-dimethylphenyl)-[l,2,4]triazolo[l,5-a]pyridin-6-yl]methoxy]phenyl]hex-4-ynoate (16 g, 34.22 mmol) in ethanol (160 mL) is added aqueous 5 N NaOH (2.73 g, 68.44 mmol in 16 mL water) drop wise at room temperature and the reaction mixture is stirred for 16 hours. The reaction mixture is evaporated to dryness, the residue is dissolved in water (300 mL), washed with diethyl ether (2 χ 200 mL), and the organic extract is discarded. The aqueous layer is cooled to 10 °C- 15 °C, acidified with saturated citric acid solution to pH~5, and extracted with DCM (2 x 300 mL). The combined organic extracts are washed with water (2 x 500 mL), brine solution (500 mL), dried over Na2S04, filtered, and evaporated to dryness to give the title compound as an off-white solid (14 g, 93%). LCMS m/z 440 (M+H)+.

The products from other batches, prepared as in Alternate Preparation of Example 1, are mixed with the product from Alternate Preparation Example 1 DCM (5 L) and warmed to 40 °C to get a clear solution. Then the solvent is evaporated to give an off-white solid. The possibility of trapped DCM is a concern, thus EtOAc (7.5 L) is charged and the resulting mixture is warmed to 65 °C to get a clear solution (-30 minutes). The solvent is evaporated and the resulting solid is dried under vacuum at 50 °C to obtain the desired product as an off-white solid. LCMS m/z 440 (M+H)+.

Form II Seed Crystal, Example 1

A saturated ethanol solution of (3S)-3-[4-[[2-(2,6-Dimethylphenyl)-[l,2,4]triazolo[l,5-a]pyridin-6-yl]methoxy]phenyl]hex-4-ynoic acid is filtered through 0.22 μιη nylon syringe filter into a clean vessel. Slow solvent evaporation at 25°C results in Form II seed crystals of Example 1.

Crystalline Form II (3S)-3-[4-[[2-(2,6-Dimethylphenyl)-[l,2,4]triazolo[l,5-a]pyridin- 6-yl] methoxy] phenyl] hex-4-ynoic acid

(3S)-3-[4-[[2-(2,6-Dimethylphenyl)-[l,2,4]triazolo[l,5-a]pyridin-6-yl]methoxy]phenyl]hex-4-ynoic acid can be prepared as a crystalline anhydrous Form II by dissolving (3S)-3-[4-[[2-(2,6-Dimethylphenyl)-[l,2,4]triazolo[l,5-a]pyridin-6-yl]methoxy]phenyl]hex-4-ynoic acid (580 mg, 132 mmol) in EtOH (1.2 mL) while stirring the mixture at 80 °C for 10 minutes. The solution is filtered and cooled to 70 °C at which point seeds of Form II are introduced. The mixture is then slowly cooled to ambient temperature while stirring overnight. The resulting solid plug is loosened with the addition of heptane (600 μΐ.) and the solids are recovered by vacuum filtration and dried under vacuum at 60 °C to give the crystalline title product (438 mg, 75.5%).

Patent ID

Patent Title

Submitted Date

Granted Date

US9120793 Triazolo-pyridine compound
2014-12-04
2015-09-01
US2015166535 NOVELTRIAZOLO-PYRIDINE COMPOUND
2014-12-04
2015-06-18

/////////LY3104607, LY-3104607, LY 3104607, PRECLINICAL

CC#C[C@H](C1=CC=C(OCC2=CN3C(C=C2)=NC(C4=C(C)C=CC=C4C)=N3)C=C1)CC(O)=O

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Pfizer’s Monobactam PF-?


STR1

Pfizer’s monobactam PF-?

1380110-34-8, C20 H24 N8 O12 S2, 632.58

Propanoic acid, 2-​[[(Z)​-​[1-​(2-​amino-​4-​thiazolyl)​-​2-​[[(2R,​3S)​-​2-​[[[[[(1,​4-​dihydro-​1,​5-​dihydroxy-​4-​oxo-​2-​pyridinyl)​methyl]​amino]​carbonyl]​amino]​methyl]​-​4-​oxo-​1-​sulfo-​3-​azetidinyl]​amino]​-​2-​oxoethylidene]​amino]​oxy]​-​2-​methyl-

2-((Z)-1-(2-Aminothiazol-4-yl)-2-((2R,3S)-2-((((1,5-dihydroxy-4-oxo-1,4-dihydropyridin-2-yl)methoxy)carbonylamino)methyl)-4-oxo-1-sulfoazetidin-3-ylamino)-2-oxoethylideneaminooxy)-2-methylpropanoic Acid

2-[[(Z)-[1-(2-Amino-4-thiazolyl)-2-[[(2R,3S)-2-[[[[[(1,4-dihydro-1,5-dihydroxy-4-oxo-2-pyridinyl)methyl]amino]carbonyl]amino]methyl]-4-oxo-1-sulfo-3-azetidinyl]amino]-2-oxoethylidene]amino]oxy]-2-methylpropanoic acid

Monobactams are a class of antibacterial agents which contain a monocyclic beta-lactam ring as opposed to a beta-lactam fused to an additional ring which is found in other beta-lactam classes, such as cephalosporins, carbapenems and penicillins. The drug Aztreonam is an example of a marketed monobactam; Carumonam is another example. The early studies in this area were conducted by workers at the Squibb Institute for Medical Research, Cimarusti, C. M. & R.B. Sykes: Monocyclic β-lactam antibiotics. Med. Res. Rev. 1984, 4, 1 -24. Despite the fact that selected

monobacatams were discovered over 25 years ago, there remains a continuing need for new antibiotics to counter the growing number of resistant organisms.

Although not limiting to the present invention, it is believed that monobactams of the present invention exploit the iron uptake mechanism in bacteria through the use of siderophore-monobactam conjugates. For background information, see: M. J. Miller, et al. BioMetals (2009), 22(1 ), 61-75.

The mechanism of action of beta-lactam antibiotics, including monobactams, is generally known to those skilled in the art and involves inhibition of one or more penicillin binding proteins (PBPs), although the present invention is not bound or limited by any theory. PBPs are involved in the synthesis of peptidoglycan, which is a major component of bacterial cell walls.

WO 2012073138

https://www.google.com/patents/WO2012073138A1?cl=en

Inventors Matthew Frank BrownSeungil HanManjinder LallMark. J. Mitton-FryMark Stephen PlummerHud Lawrence RisleyVeerabahu ShanmugasundaramJeremy T. Starr
Applicant Pfizer Inc.

Example 4, Route 1

2-({[(1Z)-1 -(2-amino-1 ,3-thiazol-4-yl)-2-({(2f?,3S)-2-[({[(1 ,5-dihydroxy-4-oxo-1 ,4- dihydropyridin-2-yl)methyl]carbamoyl}amino)methyl]-4-oxo-1 -sulfoazetidin-3- yl}amino)-2-oxoethylidene]amino}oxy)-2-methylpropanoic acid, bis sodium salt

(C92-Bis Na Salt).

Figure imgf000080_0001

C92-bis Na salt

Step 1 : Preparation of C90. A solution of C26 (16.2 g, 43.0 mmol) in tetrahydrofuran (900 mL) was treated with 1 , 1 ‘-carbonyldiimidazole (8.0 g, 47.7 mmol). After 5 minutes, the reaction mixture was treated with a solution of C9 (15 g, 25.0 mmol) in anhydrous tetrahydrofuran (600 mL) at room temperature. After 15 hours, the solvent was removed and the residue was treated with ethyl acetate (500 mL) and water (500 mL). The layers were separated and the aqueous layer was back extracted with additional ethyl acetate (300 mL). The organic layers were combined, washed with brine solution (500 mL), dried over sodium sulfate, filtered and concentrated in vacuo. The crude product was purified via chromatography on silica gel (ethyl acetate / 2-propanol) to yield C90 as a yellow foam. Yield: 17.44 g, 19.62 mmol, 78%. LCMS m/z 889.5 (M+1 ). 1H NMR (400 MHz, DMSO-d6) 1 1 .90 (br s, 1 H), 9.25 (d, J=8.7 Hz, 1 H), 8.40 (br s, 1 H), 7.98 (s, 1 H), 7.50-7.54 (m, 2H), 7.32-7.47 (m, 8H), 7.28 (s, 1 H), 6.65 (br s, 1 H), 6.28 (br s, 1 H), 5.97 (s, 1 H), 5.25 (s, 2H), 5.18 (dd, J=8.8, 5 Hz, 1 H), 4.99 (s, 2H), 4.16-4.28 (m, 2H), 3.74-3.80 (m, 1 H), 3.29-3.41 (m, 1 H), 3.13-3.23 (m, 1 H), 1.42 (s, 9H), 1.41 (s, 3H), 1.39 (br s, 12H).

Step 2: Preparation of C91. A solution of C90 (8.5 g, 9.6 mmol) in anhydrous N,N- dimethylformamide (100 mL) was treated sulfur trioxide /V,/V-dimethylformamide complex (15.0 g, 98.0 mmol). The reaction was allowed to stir at room temperature for 20 minutes then quenched with water (300 mL). The resulting solid was collected by filtration and dried to yield C91 as a white solid. Yield: 8.1 g, 8.3 mmol, 87%. LCMS m/z 967.6 (M-1 ). 1H NMR (400 MHz, DMSO-d6) δ 1 1.62 (br s, 1 H), 9.29 (d, J=8.8 Hz, 1 H), 9.02 (s, 1 H), 7.58-7.61 (m, 2H), 7.38-7.53 (m, 9H), 7.27 (s, 1 H), 7.07 (s, 1 H), 6.40 (br d, J=8 Hz, 1 H), 5.55 (s, 2H), 5.25 (s, 2H), 5.20 (dd, J=8.8, 5.6 Hz, 1 H), 4.46 (br dd, half of ABX pattern, J=17, 5 Hz, 1 H), 4.38 (br dd, half of ABX pattern, J=17, 6 Hz, 1 H), 3.92-3.98 (m, 1 H), 3.79-3.87 (m, 1 H), 3.07-3.17 (m, 1 H), 1.40 (s, 9H), 1 .39 (s, 3H), 1 .38 (s, 12H).

Step 3: Preparation of C92. A solution of C91 (8.1 g, 8.3 mmol) in anhydrous dichloromethane (200 mL) was treated with 1 M boron trichloride in p-xylenes (58.4 mL, 58.4 mmol) and allowed to stir at room temperature for 15 minutes. The reaction mixture was cooled in an ice bath, quenched with 2,2,2-trifluoroethanol (61 mL), and the solvent was removed in vacuo. A portion of the crude product (1 g) was purified via reverse phase chromatography (C-18 column; acetonitrile / water gradient with 0.1 % formic acid modifier) to yield C92 as a white solid. Yield: 486 mg, 0.77 mmol. LCMS m/z 633.3 (M+1 ). 1H NMR (400 MHz, DMSO-d6) δ 9.22 (d, J=8.7 Hz, 1 H), 8.15 (s, 1 H), 7.26-7.42 (br s, 2H), 7.18-7.25 (m, 1 H), 6.99 (s, 1 H), 6.74 (s, 1 H), 6.32-6.37 (m, 1 H), 5.18 (dd, J=8.7, 5.7 Hz, 1 H), 4.33 (br d, J=4.6 Hz, 2H), 3.94-4.00 (m, 1 H), 3.60-3.68 (m, 1 H), 3.19-3.27 (m, 1 H), 1.40 (s, 3H), 1.39 (s, 3H).

Step 4: Preparation of C92-Bis Na Salt. A flask was charged with C92 (388 mg, 0.61 mmol) and water (5.0 mL). The mixture was cooled in an ice bath and treated dropwise with a solution of sodium bicarbonate (103 mg, 1.52 mmol) in water (5.0 mL). The sample was lyophilized to yield C92-Bis Na Salt as a white solid. Yield: 415 mg, 0.61 mmol, quantitative. LCMS m/z 633.5 (M+1 ). 1H NMR (400 MHz, D20) δ 7.80 (s, 1 H), 6.93 (s, 1 H), 6.76 (s, 1 H), 5.33 (d, J=5.7 Hz, 1 H), 4.44 (ddd, J=6.0, 6.0, 5.7 Hz, 1 H), 4.34 (AB quartet, JAB=17.7 Hz, ΔνΑΒ=10.9 Hz, 2H), 3.69 (dd, half of ABX pattern, J=14.7, 5.8 Hz, 1 H), 3.58 (dd, half of ABX pattern, J=14.7, 6.2 Hz, 1 H), 1.44 (s, 3H), 1.43 (s, 3H).

Alternate preparation of C92

Figure imgf000082_0001

Step 1 : Preparation of C93. An Atlantis pressure reactor was charged with 10% palladium hydroxide on carbon (0.375 g, John Matthey catalyst type A402028-10), C91 (0.75 g, 0.77 mmol) and treated with ethanol (35 mL). The reactor was flushed with nitrogen and pressurized with hydrogen (20 psi) for 20 hours at 20 °C. The reaction mixture was filtered under vacuum and the filtrate was concentrated using the rotary evaporator to yield C93 as a tan solid. Yield: 0.49 g, 0.62 mmol, 80%. LCMS m/z 787.6 (M-1 ). 1H NMR (400 MHz, DMSO-d6) δ 1 1.57 (br s, 1 H), 9.27 (d, J=8.5 Hz, 1 H), 8.16 (s, 1 H), 7.36 (br s, 1 H), 7.26 (s, 1 H), 7.00 (s, 1 H), 6.40 (br s, 1 H), 5.18 (m, 1 H), 4.35 (m, 2H), 3.83 (m, 1 H), 3.41 (m, 1 H), 3.10 (m, 1 H), 1.41 (s, 6H), 1.36 (s, 18H).

Step 2: Preparation of C92. A solution of C93 (6.0 g, 7.6 mmol) in anhydrous dichloromethane (45 mL) at 0 °C was treated with trifluoroacetic acid (35.0 mL, 456 mmol). The mixture was warmed to room temperature and stirred for 2 hours. The reaction mixture was cannulated into a solution of methyl ferf-butyl ether (100 mL) and heptane (200 mL). The solid was collected by filtration and washed with a mixture of methyl ferf-butyl ether (100 mL) and heptane (200 mL) then dried under vacuum. The crude product (~5 g) was purified via reverse phase chromatography (C-18 column; acetonitrile / water gradient with 0.1 % formic acid modifier) and lyophilized to yield C92 as a pink solid. Yield: 1.45 g, 2.29 mmol. LCMS m/z 631.0 (M-1). 1H NMR (400 MHz, DMSO-de) δ 9.20 (d, J=8.7 Hz, 1H), 8.13 (s, 1H), 7.24-7.40 (br s, 2H), 7.16-7.23 (m, 1H), 6.97 (s, 1H), 6.71 (s, 1H), 6.31-6.35 (m, 1H), 5.15 (dd, J=8.7, 5.7 Hz, 1H), 4.31 (br d, J=4.6 Hz, 2H), 3.92-3.98 (m, 1H), 3.58-3.67 (m, 1H), 3.17-3.25 (m, 1H), 1.37 (s, 3H), 1.36 (s, 3H).

Example 4, route 2

2-({[(1Z)-1-(2-amino-1,3-thiazol-4-yl)-2-({(2 ?,3S)-2-[({[(1,5-dihydroxy-4-oxo-^ dihydropyridin-2-yl)methyl]carbamoyl}amino)methyl]-4-oxo-1-sulfoazetidin-3- yl}amino)-2-oxoethylidene]amino}oxy)-2-methylpropanoic acid (C92).

lt

Figure imgf000083_0001

single

enantiomer

Figure imgf000083_0002

Step 1. Preparation of C95. A solution of C94 (50.0 g, 189.9 mmol) in

dichloromethane (100 mL) was treated with trifluoroacetic acid (50.0 mL, 661.3 mmol). The reaction mixture was stirred at room temperature for 24 hours. The dichloromethane and trifluoroacetic acid was displaced with toluene (4 x 150 mL) using vacuum, to a final volume of 120 mL. The solution was added to heptane (250 mL) and the solid was collected by filtration. The solid was washed with a mixture of toluene and heptane (1 : 3, 60 mL), followed by heptane (2 x 80 mL) and dried under vacuum at 50 °C for 19 hours to afford C95 as a solid. Yield: 30.0 g, 158 mmol, 84%. 1H NMR (400 MHz, CDCI3) δ 9.66 (s, 1 H), 7.86 – 7.93 (m, 2H), 7.73 – 7.80 (m, 2H), 4.57 (s, 2H). HPLC retention time 5.1 minutes; column: Agilent Extended C-18 column (75 mm x 3 mm, 3.5 μηη); column temperature 45 °C; flow rate 1.0 mL / minute; detection UV 230 nm; mobile phase: solvent A = acetonitrile (100%), solvent B = acetonitrile (5%) in 10 mM ammonium acetate; gradient elusion: 0-1.5 minutes solvent B (100%), 1.5-10.0 minutes solvent B (5%), 10.0-13.0 minutes solvent B (100%); total run time 13.0 minutes.

Step 2: Preparation of C96-racemic. A solution of C95 (32.75 g; 173.1 mmol) in dichloromethane (550 mL) under nitrogen was cooled to 2 °C. The solution was treated with 2,4-dimethoxybenzylamine (28.94 g, 173.1 mmol) added dropwise over 25 minutes, maintaining the temperature below 10 °C. The solution was stirred for 10 minutes at 2 °C and then treated with molecular sieves (58.36 g, UOP Type 3A). The cold bath was removed and the reaction slurry was stirred for 3 hours at room temperature. The slurry was filtered through a pad of Celite (34.5 g) and the filter cake was rinsed with dichloromethane (135 mL). The dichloromethane filtrate (imine solution) was used directly in the following procedure.

A solution of A/-(ferf-butoxycarbonyl)glycine (60.6 g, 346.1 mmol) in

tetrahydrofuran (622 mL) under nitrogen was cooled to -45 °C and treated with triethylamine (38.5 g, 380.8 mmol). The mixture was stirred for 15 minutes at -45 °C and then treated with ethyl chloroformate (48.8 g, 450 mmol) over 15 minutes. The reaction mixture was stirred at -50 °C for 7 hours. The previously prepared imine solution was added via an addition funnel over 25 minutes while maintaining the reaction mixture temperature below -40 °C. The slurry was treated with triethylamine (17.5 g, 173 mmol) and the reaction mixture was slowly warmed to room temperature over 5 hours and stirred for an additional 12 hours. The reaction slurry was charged with water (150 mL) and the volatiles removed using a rotary evaporator. The reaction mixture was charged with additional water (393 mL) and the volatiles removed using a rotary evaporator. The mixture was treated with methyl ferf-butyl ether (393 mL) and vigorously stirred for 1 hour. The solid was collected by vacuum filtration and the filter cake was rinsed with a mixture of methyl ferf-butyl ether and water (1 : 1 , 400 mL). The solid was collected and dried in a vacuum oven at 50 °C for 16 hours to afford C96- racemic. Yield: 55.8 g, 1 13 mmol, 65%. 1H-NMR (400 MHz, DMSO-d6) δ 7.85 (s, NH), 7.80 (s, 4H), 6.78 (d, J=7.8 Hz, 1 H), 6.25 (m, 1 H), 6.10 (m, 1 H), 4.83 (m, 1 H), 4.38 (d, J=9.5 Hz, 1 H), 3.77-3.95 (m, 3H), 3.62 (s, 3H), 3.45 (m, 1 H), 3.40 (s, 3H), 1.38 (s, 9H). HPLC retention time 6.05 minutes; XBridge C8 column (4.6 x 75 mm, 3.5 μηη); column temperature 45 °C; flow rate 2.0 mL/minute; detection UV 210 nm, 230 nm, and 254 nm; mobile phase: solvent A = methanesulfonic acid (5%) in 10 mmol sodium octylsulfonate, solvent B = acetonitrile (100%); gradient elusion: 0-1.5 minutes solvent A (95%) and solvent B (5%), 1.5-8.5 minutes solvent A (5%) and solvent B (95%), 8.5- 10.0 minutes solvent A (5%) and solvent B (95%), 10.01 -12.0 minutes solvent A (95%) and solvent B (5%); total run time 12.0 minutes.

Step 3: Preparation of C97-racemic. A solution of C96-racemic (15.0 g, 30.3 mmol) in ethyl acetate (150 mL) under nitrogen was treated with ethanolamine (27.3 mL, 454.1 mmol). The reaction mixture was heated at 90 °C for 3 hours and then cooled to room temperature. The mixture was charged with water (150 mL) and the layers separated. The aqueous layer was extracted with ethyl acetate (75 mL) and the combined organic layers washed with water (2 x 150 mL) followed by saturated aqueous sodium chloride (75 mL). The organic layer was dried over magnesium sulfate, filtered and the filtrate concentrated to a volume of 38 mL. The filtrate was treated with heptane (152 mL) and the solid was collected by filtration. The solid was washed with heptane and dried at 50 °C in a vacuum oven overnight to yield C97-racemic as a solid. Yield: 9.68 g, 26.5 mmol, 88%. LCMS m/z 967.6 (M-1 ). 1H NMR (400 MHz, DMSO-d6) δ 7.64 (d, J=9.4 Hz, 1 H), 7.14 (d, J=8.2 Hz, 1 H), 6.56 (s, 1 H), 6.49 (dd, J=8.20, 2.3 Hz, 1 H), 4.78 (dd, J=9.37, 5.1 Hz, 1 H), 4.30 (d, J=14.8 Hz, 1 H), 4.14 (d, J=14.8 Hz, 1 H), 3.77 (s, 3H), 3.75 (s, 3H), 3.45 – 3.53 (m, 1 H), 2.65 – 2.75 (m, 1 H), 2.56 – 2.64 (m, 1 H), 1.38 (s, 9H), 1.30 – 1.35 (m, 2H). HPLC retention time 5.1 minutes; column: Agilent Extended C-18 column (75 mm x 3 mm, 3.5 μΐη); column temperature 45 °C; flow rate 1.0 mL / minute;

detection UV 230 nm; mobile phase: solvent A = acetonitrile (100%), solvent B = acetonitrile (5%) in 10 mM ammonium acetate; gradient elusion: 0-1 .5 minutes solvent B (100%), 1 .5-10.0 minutes solvent B (5%), 10.0-13.0 minutes solvent B (100%); total run time 13.0 minutes. Step 4: Preparation of C97-(2R,3S) enantiomer. A solution of C97-racemic (20.0 g, 54.7 mmol) in ethyl acetate (450 mL) was treated with diatomaceous earth (5.0 g) and filtered through a funnel charged with diatomaceous earth. The filter cake was washed with ethyl acetate (150 mL). The filtrate was charged with diatomaceous earth (20.0 g) and treated with (-)-L-dibenzoyltartaric acid (19.6 g, 54.7 mmol). The slurry was heated at 60 °C for 1.5 hours and then cooled to room temperature. The slurry was filtered and the solid washed with ethyl acetate (90 mL). The solid was collected and dried at 50 °C in a vacuum oven for 17 hours to yield C97-(2R,3S) enantiomer as a solid (mixed with diatomaceous earth). Yield: 17.3 g, 23.9 mmol, 43.6%, 97.6% ee. 1H NMR (400 MHz, DMSO-de) δ 7.89 – 7.91 (m, 4H), 7.59 – 7.65 (m, 3H), 7.44 – 7.49 (m, 4H), 7.09 (d, J=8.3 Hz, 1 H), 6.53 (d, J=2.3 Hz, 1 H), 6.49 (dd, J=8.3, 2.3 Hz, 1 H), 5.65 (s, 2H), 4.85 (dd, J=9.3, 4.9 Hz, 1 H), 4.30 (d, J=15.3 Hz, 1 H), 4.10 (d, J=15.3 Hz, 1 H), 3.74 (s, 3H), 3.72 (s, 3H), 3.68 – 3.70 (m, 1 H), 2.92 – 2.96 (dd, J=13.6, 5.4 Hz, 1 H), 2.85 – 2.90 (dd, J=13.6, 6.3 Hz, 1 H), 1.36 (s, 9H). HPLC retention time 5.1 minutes; column: Agilent Extended C-18 column (75 mm x 3 mm, 3.5 μηη); column temperature 45 °C; flow rate 1.0 mL / minute; detection UV 230 nm; mobile phase: solvent A = acetonitrile (100%), solvent B = acetonitrile (5%) in 10 mM ammonium acetate; gradient elusion: 0-1 .5 minutes solvent B (100%), 1.5-10.0 minutes solvent B (5%), 10.0-13.0 minutes solvent B (100%); total run time 13.0 minutes. Chiral HPLC retention time 9.1 minutes; column: Chiralcel OD-H column (250 mm x 4.6 mm); column temperature 40 °C; flow rate 1 .0 mL / minute; detection UV 208 nm; mobile phase: solvent A = ethanol (18%), solvent B = heptane (85%); isocratic elusion; total run time 20.0 minutes.

Step 5: Preparation of C98-(2R,3S) enantiomer. A solution of C97-(2R,3S) enantiomer. (16.7 g, 23.1 mmol) in ethyl acetate (301 mL) was treated with diatomaceous earth (18.3 g) and 5% aqueous potassium phosphate tribasic (182 mL). The slurry was stirred for 30 minutes at room temperature, then filtered under vacuum and the filter cake washed with ethyl acetate (2 x 67 mL). The filtrate was washed with 5% aqueous potassium phosphate tribasic (18 mL) and the organic layer dried over magnesium sulfate. The solid was filtered and the filter cake washed with ethyl acetate (33 mL). The filtrate was concentrated to a volume of 42 mL and slowly added to heptane (251 mL) and the resulting solid was collected by filtration. The solid was washed with heptane and dried at 50 °C in a vacuum oven for 19 hours to yield C98- (2R,3S) enantiomer as a solid. Yield: 6.4 g, 17.5 mmol, 76%, 98.8% ee. 1H NMR (400 MHz, DMSO-de) δ 7.64 (d, J=9.4 Hz, 1 H), 7.14 (d, J=8.2 Hz, 1 H), 6.56 (s, 1 H), 6.49 (dd, J=8.20, 2.3 Hz, 1 H), 4.78 (dd, J=9.37, 5.1 Hz, 1 H), 4.30 (d, J=14.8 Hz, 1 H), 4.14 (d, J=14.8 Hz, 1 H), 3.77 (s, 3H), 3.75 (s, 3H), 3.45 – 3.53 (m, 1 H), 2.65 – 2.75 (m, 1 H), 2.56 – 2.64 (m, 1 H), 1.38 (s, 9H), 1.30 – 1.35 (m, 2H). HPLC retention time 5.2 minutes; column: Agilent Extended C-18 column (75 mm x 3 mm, 3.5 μηη); column temperature 45 °C; flow rate 1.0 mL / minute; detection UV 230 nm; mobile phase: solvent A = acetonitrile (100%), solvent B = acetonitrile (5%) in 10 mM ammonium acetate; gradient elusion: 0-1 .5 minutes solvent B (100%), 1.5-10.0 minutes solvent B (5%), 10.0-13.0 minutes solvent B (100%); total run time 13.0 minutes. Chiral HPLC retention time 8.7 minutes; column: Chiralcel OD-H column (250 mm x 4.6 mm); column temperature 40 °C; flow rate 1.0 mL / minute; detection UV 208 nm; mobile phase: solvent A = ethanol (18%), solvent B = heptane (85%); isocratic elusion; total run time 20.0 minutes.

Step 6: Preparation of C99. A solution of potassium phosphate tribasic N-hydrate (8.71 g, 41 .05 mmol) in water (32.0 mL) at 22 °C was treated with a slurry of C26- mesylate salt (12.1 g, 27.4 mmol, q-NMR potency 98%) in dichloromethane (100.00 mL). The slurry was stirred for 1 hour at 22 °C. The reaction mixture was transferred to a separatory funnel and the layers separated. The aqueous layer was back extracted with dichloromethane (50.0 mL). The organic layers were combined, dried over magnesium sulfate, filtered under vacuum and the filter cake washed with

dichloromethane (2 x 16 mL). The filtrate (-190 mL, amine solution) was used directly in the next step.

A solution of 1 ,1 ‘-carbonyldiimidazole (6.66 g, 41 .0 mmol) in dichloromethane (100 mL) at 22 °C under nitrogen was treated with the previously prepared amine solution (-190 mL) added dropwise using an addition funnel over 3 hour at 22 °C with stirring. After the addition, the mixture was stirred for 1 hour at 22 °C, then treated with C98-(2R,3S) enantiomer. (10.0 g, 27.4 mmol) followed by /V,/V-dimethylformamide (23.00 mL). The reaction mixture was stirred at 22 °C for 3 hours and then heated at 40 °C for 12 hours. The solution was cooled to room temperature and the dichloromethane was removed using the rotary evaporator. The reaction mixture was diluted with ethyl acetate (216.0 mL) and washed with 10% aqueous citric acid (216.0 mL), 5% aqueous sodium chloride (2 x 216.0 mL), dried over magnesium sulfate and filtered under vacuum. The filter cake was washed with ethyl acetate (3 x 13 mL) and the ethyl acetate solution was concentrated on the rotary evaporator to a volume of (-1 10.00 mL) providing a suspension. The suspension (~1 10.00 mL) was warmed to 40 °C and transferred into a stirred solution of heptane (22 °C) over 1 hour, to give a slurry. The slurry was stirred for 1 hour and filtered under vacuum. The filter cake was washed with heptane (3 x 30 mL) and dried under vacuum at 50 °C for 12 hours to afford C99 as a solid. Yield: 18.1 g, 24.9 mmol, 92%. LCMS m/z 728.4 (M+1 ). 1H NMR (400 MHz, DMSO-d6) δ 8.09 (s, 1 H), 7.62 (d, J=9.4 Hz, 1 H), 7.33-7.52 (m, 10H), 7.07 (d, J=8.3 Hz, 1 H), 6.51 (d, J=2.3 Hz, 1 H), 6.50 (m, 1 H), 6.44 (dd, J=8.3, 2.3 Hz, 1 H), 6.12 (m, 1 H), 6.07 (s, 1 H), 5.27 (s, 2H), 5.00 (s, 2H), 4.73 (dd, J=9.4, 5.2 Hz, 1 H), 4.38 (d, J=15.0 Hz, 1 H), 4.19 (m, 2H), 3.99 (d, J=15.0 Hz, 1 H), 3.72 (s, 3H), 3.71 (s, 3H), 3.48 (m, 1 H), 3.28 (m, 1 H), 3.12 (m, 1 H), 1 .37 (s, 9H).

Step 7: Preparation of C100. A solution of C99 (46.5 g, 63.9 mmol) in acetonitrile (697 mL and water (372 mL) was treated with potassium persulfate (69.1 g, 255.6 mmol) and potassium phosphate dibasic (50.1 g, 287.5 mmol). The biphasic mixture was heated to 75 °C and vigorously stirred for 1.5 hours. The pH was maintained between 6.0-6.5 by potassium phosphate dibasic addition (-12 g). The mixture was cooled to 20 °C, the suspension was filtered and washed with acetonitrile (50 mL). The filtrate was concentrated using the rotary evaporator and treated with water (50 mL) followed by ethyl acetate (200 mL). The slurry was stirred for 2 hours at room temperature, filtered and the solid dried under vacuum at 40 °C overnight. The solid was slurried in a mixture of ethyl acetate and water (6 : 1 , 390.7 mL) at 20 °C for 1 hour then collected by filtration. The solid was dried in a vacuum oven to yield C100. Yield: 22.1 g, 38.3 mmol, 60%. 1H NMR (400 MHz, DMSO-d6) δ 8.17 (br s, 1 H), 7.96 (s, 1 H), 7.58 (d, J=9.6 Hz, 1 H), 7.29-7.50 (m, 10H), 6.49 (dd, J=8.0, 6.0 Hz, 1 H), 6.08 (dd, J=5.6, 5.2 Hz, 1 H), 5.93 (s, 1 H), 5.22 (s, 2H), 4.96 (s, 2H), 4.77 (dd, J=9.6, 5.0 Hz, 1 H), 4.16 (m, 2H), 3.61 (m, 1 H), 3.1 1 (m, 2H), 1.36 (s, 9H). HPLC retention time 6.17 minutes; XBridge C8 column (4.6 x 75 mm, 3.5 μηη); column temperature 45 °C; flow rate 2.0 mL/minute; detection UV 210 nm, 230 nm, and 254 nm; mobile phase: solvent A = methanesulfonic acid (5%) in 10 mmol sodium octylsulfonate, solvent B = acetonitrile (100%); gradient elusion: 0-1 .5 minutes solvent A (95%) and solvent B (5%), 1.5-8.5 minutes solvent A (5%) and solvent B (95%), 8.5-10.0 minutes solvent A (5%) and solvent B (95%), 10.01- 12.0 minutes solvent A (95%) and solvent B (5%); total run time 12.0 minutes.

Step 8: Preparation of C101. A solution of trifluoroacetic acid (120 mL, 1550 mmol) under nitrogen was treated with methoxybenzene (30 mL, 269 mmol) and cooled to -5 °C. Solid C100 (17.9 g, 31.0 mmol) was charged in one portion at -5 °C and the resulting mixture stirred for 3 hours. The reaction mixture was cannulated with nitrogen pressure over 15 minutes to a stirred mixture of Celite (40.98 g) and methyl ferf-butyl ether (550 mL) at 10 °C. The slurry was stirred at 16 °C for 30 minutes, then filtered under vacuum. The filter cake was rinsed with methyl ferf-butyl ether (2 x 100 mL). The solid was collected and slurried in methyl ferf-butyl ether (550 mL) with vigorous stirring for 25 minutes. The slurry was filtered by vacuum filtration and washed with methyl ferf-butyl ether (2 x 250 mL). The solid was collected and dried in a vacuum oven at 60 °C for 18 hours to afford C101 on Celite. Yield: 57.6 g total = C101 + Celite; 16.61 g C101 , 28.1 mmol, 91%. 1H NMR (400 MHz, DMSO-d6) δ 8.75-8.95 (br s, 2H), 8.65 (s, 1 H), 8.21 (s, 1 H), 7.30-7.58 (m, 10H), 6.83 (br s, 1 H), 6.65 (br s, 1 H), 6.17 (s, 1 H), 5.30 (s, 2H), 5.03 (s, 2H), 4.45 (br s, 1 H), 4.22 (br s, 2H), 3.77 (m, 1 H), 3.36 (m, 1 H), 3.22 (m, 1 H). 19F NMR (376 MHz, DMSO-d6) δ -76.0 (s, 3F). HPLC retention time 5.81 minutes; XBridge C8 column (4.6 x 75 mm, 3.5 μηη); column temperature 45 °C; flow rate 2.0 mL/minute; detection UV 210 nm, 230 nm, and 254 nm; mobile phase: solvent A = methanesulfonic acid (5%) in 10 mmol sodium octylsulfonate, solvent B = acetonitrile (100%); gradient elusion: 0-1.5 minutes solvent A (95%) and solvent B (5%), 1.5-8.5 minutes solvent A (5%) and solvent B (95%), 8.5-10.0 minutes solvent A (5%) and solvent B (95%), 10.01-12.0 minutes solvent A (95%) and solvent B (5%); total run time 12.0 minutes.

Step 9: Preparation of C90. A suspension of C101 (67.0 g, 30% activity on Celite = 33.9 mmol) in acetonitrile (281 .4 mL) was treated with molecular sieves 4AE (40.2 g), C5 (17.9 g, 33.9 mmol), 4-dimethylaminopyridine (10.4 g, 84.9 mmol) and the mixture was stirred at 40°C for 16 hours. The reaction mixture was cooled to 20 °C, filtered under vacuum and the filter cake washed with acetonitrile (2 x 100 mL). The filtrate was concentrated under vacuum to a volume of -50 mL. The solution was diluted with ethyl acetate (268.0 mL) and washed with 10% aqueous citric acid (3 x 134 mL) followed by 5% aqueous sodium chloride (67.0 mL). The organic layer was dried over magnesium sulfate and filtered under vacuum. The filter cake was washed with ethyl acetate (2 x 50 mL) and the filtrate was concentrated to a volume of -60 mL. The filtrate was added slowly to heptane (268 mL) with stirring and the slurry was stirred at 20 °C for 1 hour. The slurry was filtered under vacuum and the filter cake washed with a mixture of heptane and ethyl acetate (4: 1 , 2 x 27 mL). The solid was collected and dried under vacuum for 12 hours at 50 °C to afford a solid. The crude product was purified via chromatography on silica gel (ethyl acetate / 2-propanol), product bearing fractions were combined and the volume was reduced to -60 mL. The solution was added dropwise to heptane (268 mL) with stirring. The slurry was stirred at room temperature for 3 hours, filtered and washed with heptane and ethyl acetate (4: 1 , 2 x 27 mL). The solid was collected and dried under vacuum for 12 hours at 50 °C to afford C90 as a solid. Yield: 16.8 g, 18.9 mmol, 58%. LCMS m/z 889.4 (M+1 ). 1H NMR (400 MHz, DMSO-cfe) 1 1.90 (br s, 1 H), 9.25 (d, J=8.7 Hz, 1 H), 8.40 (br s, 1 H), 7.98 (s, 1 H), 7.50-7.54 (m, 2H), 7.32- 7.47 (m, 8H), 7.28 (s, 1 H), 6.65 (br s, 1 H), 6.28 (br s, 1 H), 5.97 (s, 1 H), 5.25 (s, 2H), 5.18 (dd, J=8.8, 5 Hz, 1 H), 4.99 (s, 2H), 4.16-4.28 (m, 2H), 3.74-3.80 (m, 1 H), 3.29-3.41 (m, 1 H), 3.13-3.23 (m, 1 H), 1 .42 (s, 9H), 1 .41 (s, 3H), 1.39 (br s, 12H).

Step 10: Preparation of C91. A solution of C90 (14.5 g, 16.3 mmol) in anhydrous N,N- dimethylformamide (145.0 mL) was treated with sulfur trioxide /V,/V-dimethylformamide complex (25.0 g, 163.0 mmol). The reaction mixture was stirred at room temperature for 45 minutes, then transferred to a stirred mixture of 5% aqueous sodium chloride (290 mL) and ethyl acetate (435 mL) at 0 °C. The mixture was warmed to 18 °C and the layers separated. The aqueous layer was extracted with ethyl acetate (145 mL) and the combined organic layers washed with 5% aqueous sodium chloride (3 x 290 mL) followed by saturated aqueous sodium chloride (145 mL). The organic layer was dried over magnesium sulfate, filtered through diatomaceous earth and the filter cake washed with ethyl acetate (72 mL). The filtrate was concentrated to a volume of 36 mL and treated with methyl ferf-butyl ether (290 mL), the resulting slurry was stirred at room temperature for 1 hour. The solid was collected by filtration, washed with methyl ferf- butyl ether (58 mL) and dried at 50 °C for 2 hours followed by 20 °C for 65 hours in a vacuum oven to yield C91 as a solid. Yield: 15.0 g, 15.4 mmol, 95%. LCMS m/z 967.6 (M-1 ). 1H NMR (400 MHz, DMSO-d6) δ 1 1.62 (br s, 1 H), 9.29 (d, J=8.8 Hz, 1 H), 9.02 (s, 1 H), 7.58-7.61 (m, 2H), 7.38-7.53 (m, 9H), 7.27 (s, 1 H), 7.07 (s, 1 H), 6.40 (br d, J=8.0 Hz, 1 H), 5.55 (s, 2H), 5.25 (s, 2H), 5.20 (dd, J=8.8, 5.6 Hz, 1 H), 4.46 (br dd, half of ABX pattern, J=17.0, 5.0 Hz, 1 H), 4.38 (br dd, half of ABX pattern, J=17.0, 6.0 Hz, 1 H), 3.92- 3.98 (m, 1 H), 3.79-3.87 (m, 1 H), 3.07-3.17 (m, 1 H), 1.40 (s, 9H), 1.39 (s, 3H), 1.38 (s, 12H).

Step 11 : Preparation of C92. A solution of C91 (20.0 g, 20.6 mmol) in

dichloromethane (400 mL) was concentrated under reduced pressure (420 mmHg) at 45 °C to a volume of 200 mL. The solution was cooled to -5 °C and treated with 1 M boron trichloride in dichloromethane (206.0 mL, 206.0 mmol) added dropwise over 40 minutes. The reaction mixture was warmed to 15 °C over 1 hour with stirring. The slurry was cooled to -15 °C and treated with a mixture of 2,2,2-trifluoroethanol (69.2 mL) and methyl ferf-butyl ether (400 mL), maintaining the temperature at -15 °C. The reaction mixture was warmed to 0 °C over 1 hour. The suspension was filtered using nitrogen pressure and the solid washed with methyl ferf-butyl ether (2 x 200 mL).

Nitrogen was passed over the solid for 2 hours. The solid was collected and suspended in methyl ferf-butyl ether (400 mL) for 1 hour with stirring at 18 °C. The suspension was filtered using nitrogen pressure and the solid washed with methyl ferf-butyl ether (2 x 200 mL). Nitrogen was passed over the resulting solid for 12 hours. A portion of the crude product was neutralized with 1 M aqueous ammonium formate to pH 5.5 with minimal addition of /V,/V-dimethylformamide to prevent foaming. The feed solution was filtered and purified via reverse phase chromatography (C-18 column; acetonitrile / water gradient with 0.2% formic acid modifier). The product bearing fractions were combined and concentrated to remove acetonitrile. The solution was captured on a GC-161 M column, washed with deionized water and blown dry with nitrogen pressure. The product was released using a mixture of methanol / water (10: 1 ) and the product bearing fractions were added to a solution of ethyl acetate (6 volumes). The solid was collected by filtration to afford C92 as a solid. Yield: 5.87 g, 9.28 mmol. LCMS m/z 633.3 (M+1 ). 1H NMR (400 MHz, DMSO-d6) δ 9.22 (d, J=8.7 Hz, 1 H), 8.15 (s, 1 H), 7.26-7.42 (br s, 2H), 7.18-7.25 (m, 1 H), 6.99 (s, 1 H), 6.74 (s, 1 H), 6.32-6.37 (m, 1 H), 5.18 (dd, J=8.7, 5.7 Hz, 1 H), 4.33 (br d, J=4.6 Hz, 2H), 3.94-4.00 (m, 1 H), 3.60-3.68 (m, 1 H), 3.19-3.27 (m, 1 H), 1.40 (s, 3H), 1.39 (s, 3H).

PAPER

Journal of Medicinal Chemistry (2014), 57(9), 3845-3855

Siderophore Receptor-Mediated Uptake of Lactivicin Analogues in Gram-Negative Bacteria

Medicinal Chemistry, Computational Chemistry, §Antibacterials Research Unit, and Structural Biology, Pfizer Global Research and Development, Eastern Point Road, Groton, Connecticut 06340, United States
J. Med. Chem.201457 (9), pp 3845–3855
DOI: 10.1021/jm500219c
Publication Date (Web): April 2, 2014
Copyright © 2014 American Chemical Society
*Phone: (860)-686-1788. E-mail: seungil.han@pfizer.com.

Abstract

Abstract Image

Multidrug-resistant Gram-negative pathogens are an emerging threat to human health, and addressing this challenge will require development of new antibacterial agents. This can be achieved through an improved molecular understanding of drug–target interactions combined with enhanced delivery of these agents to the site of action. Herein we describe the first application of siderophore receptor-mediated drug uptake of lactivicin analogues as a strategy that enables the development of novel antibacterial agents against clinically relevant Gram-negative bacteria. We report the first crystal structures of several sideromimic conjugated compounds bound to penicillin binding proteins PBP3 and PBP1a from Pseudomonas aeruginosa and characterize the reactivity of lactivicin and β-lactam core structures. Results from drug sensitivity studies with β-lactamase enzymes are presented, as well as a structure-based hypothesis to reduce susceptibility to this enzyme class. Finally, mechanistic studies demonstrating that sideromimic modification alters the drug uptake process are discussed.

PAPER

Pyridone-Conjugated Monobactam Antibiotics with Gram-Negative Activity

Worldwide Medicinal Chemistry, Computational Chemistry, §Antibacterials Research Unit, Pharmacokinetics, Dynamics & Metabolism, Structural Biology, Pfizer Global Research and Development, Eastern Point Road, Groton, Connecticut 06340, United States
J. Med. Chem.201356 (13), pp 5541–5552
DOI: 10.1021/jm400560z
Publication Date (Web): June 11, 2013
Copyright © 2013 American Chemical Society
*Phone: 860-441-3522. E-mail: matthew.f.brown@pfizer.com.
Abstract Image

Herein we describe the structure-aided design and synthesis of a series of pyridone-conjugated monobactam analogues with in vitro antibacterial activity against clinically relevant Gram-negative species including Pseudomonas aeruginosaKlebsiella pneumoniae, and Escherichia coli. Rat pharmacokinetic studies with compound 17 demonstrate low clearance and low plasma protein binding. In addition, evidence is provided for a number of analogues suggesting that the siderophore receptors PiuA and PirA play a role in drug uptake in P. aeruginosa strain PAO1.

STR1

17 as a solid. Yield: 5.87 g, 9.28 mmol. LCMS m/z 633.3 (M+1). 1H NMR (400 MHz, DMSOd6) δ 9.22 (d, J=8.7 Hz, 1H), 8.15 (s, 1H), 7.26-7.42 (br s, 2H), 7.18-7.25 (m, 1H), 6.99 (s, 1H), 6.74 (s, 1H), 6.32-6.37 (m, 1H), 5.18 (dd, J=8.7, 5.7 Hz, 1H), 4.33 (br d, J=4.6 Hz, 2H), 3.94-4.00 (m, 1H), 3.60-3.68 (m, 1H), 3.19-3.27 (m, 1H), 1.40 (s, 3H), 1.39 (s, 3H).

Nc1nc(cs1)\C(=N\OC(C)(C)C(=O)O)C(=O)N[C@@H]3C(=O)N([C@@H]3CNC(=O)NCC2=CC(=O)C(O)=CN2O)S(=O)(=O)O

PAPER

Process Development for the Synthesis of Monocyclic β-Lactam Core 17

Pfizer Worldwide Research and Development, Eastern Point Road, Groton, Connecticut 06340, United States
Org. Process Res. Dev., Article ASAP
DOI: 10.1021/acs.oprd.7b00359
Publication Date (Web): January 4, 2018
Copyright © 2018 American Chemical Society
Abstract Image

Process development and multikilogram synthesis of the monocyclic β-lactam core 17 for a novel pyridone-conjugated monobactam antibiotic is described. Starting with commercially available 2-(2,2-diethoxyethyl)isoindoline-1,3-dione, the five-step synthesis features several telescoped operations and direct isolations to provide significant improvement in throughput and reduced solvent usage over initial scale-up campaigns. A particular highlight in this effort includes the development of an efficient Staudinger ketene–imine [2 + 2] cycloaddition reaction of N-Boc-glycine ketene 12 and imine 9 to form racemic β-lactam 13 in good isolated yield (66%) and purity (97%). Another key feature in the synthesis involves a classical resolution of racemic amine 15 to afford single enantiomer salt 17 in excellent isolated yield (45%) with high enantiomeric excess (98%).

Figure

https://pubs.acs.org/doi/suppl/10.1021/acs.oprd.7b00359/suppl_file/op7b00359_si_001.pdf

Nc1nc(cs1)\C(=N\OC(C)(C)C(=O)O)C(=O)N[C@@H]3C(=O)N([C@@H]3CNC(=O)NCC2=CC(=O)C(O)=CN2O)S(=O)(=O)O

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J. Med. Chem.201356 (13), pp 5541–5552
DOI: 10.1021/jm400560z

OXYGEN ANALOGUE…………..

STR2
 1380110-45-1, C20 H23 N7 O13 S2, 633.57
Propanoic acid, 2-​[[(Z)​-​[1-​(2-​amino-​4-​thiazolyl)​-​2-​[[(2R,​3S)​-​2-​[[[[(1,​4-​dihydro-​1,​5-​dihydroxy-​4-​oxo-​2-​pyridinyl)​methoxy]​carbonyl]​amino]​methyl]​-​4-​oxo-​1-​sulfo-​3-​azetidinyl]​amino]​-​2-​oxoethylidene]​amino]​oxy]​-​2-​methyl-
2-[[(Z)-[1-(2-Amino-4-thiazolyl)-2-[[(2R,3S)-2-[[[[(1,4-dihydro-1,5-dihydroxy-4-oxo-2-pyridinyl)methoxy]carbonyl]amino]methyl]-4-oxo-1-sulfo-3-azetidinyl]amino]-2-oxoethylidene]amino]oxy]-2-methylpropanoic acid

STR2

18 as a light yellow solid. Yield: 43 mg, 0.068 mmol, 51%. LCMS m/z 634.4 (M+1). 1H NMR (400 MHz, DMSO-d6), characteristic peaks: δ 9.29 (d, J=8.5 Hz, 1H), 8.10 (s, 1H), 7.04-7.10 (m, 1H), 7.00 (s, 1H), 6.75 (s, 1H), 5.05-5.30 (m, 3H), 4.00-4.07 (m, 1H), 1.42 (s, 3H), 1.41 (s, 3H).

Nc1nc(cs1)\C(=N\OC(C)(C)C(=O)O)C(=O)N[C@@H]3C(=O)N([C@@H]3CNC(=O)OCC2=CC(=O)C(O)=CN2O)S(=O)(=O)O

Step 4: Preparation of 18-Bis Na salt. A suspension of 5 (212 mg, 0.33 mmol) in water (10 mL) was cooled to 0 oC and treated with a solution of sodium bicarbonate (56.4 mg, 0.67 mmol) in water (2 mL), added dropwise. The reaction mixture was cooled to -70 oC (frozen) and lyophilized to afford 18-Bis Na salt as a white solid. Yield: 210 mg, 0.31 mmol, 93%. LCMS m/z 632.5 (M-1). 1H NMR (400 MHz, D2O) δ 7.87 (s, 1H), 6.94 (s, 1H), 6.92 (s, 1H), 5.35 (d, J=5 Hz, 1H), 5.16 (s, 2H), 4.46-4.52 (m, 1H), 3.71 (dd, half of ABX pattern, J=14.5, 6 Hz, 1H), 3.55 (dd, half of ABX pattern, J=14.5, 6 Hz, 1H), 1.43 (s, 3H), 1.42 (s, 3H).

WO 2012073138

Inventors Matthew Frank BrownSeungil HanManjinder LallMark. J. Mitton-FryMark Stephen PlummerHud Lawrence RisleyVeerabahu ShanmugasundaramJeremy T. Starr
Applicant Pfizer Inc.

Example 5

disodium 2-({[(1Z)-1 -(2-amino-1 ,3-thiazol-4-yl)-2-({(2R,3S)-2-[({[(1 ,5-dihydroxy-4- oxo-1 ,4-dihydropyridin-2-yl)methoxy]carbonyl}amino)methyl]-4-oxo-1 – sulfonatoazetidin-3-yl}amino)-2-oxoethylidene]amino}oxy)-2-methylpropanoate

(C104-Bis Na salt).

Figure imgf000092_0001

Step 1 : Preparation of C102. A solution of C28 (300 mg, 0.755 mmol) in

tetrahydrofuran (10 mL) was treated with 1 , 1 ‘-carbonyldiimidazole (379 mg, 2.26 mmol) at room temperature and stirred for 20 hours. The yellow reaction mixture was treated with a solution of C9 (286 mg, 0.543 mmol) in tetrahydrofuran (25 mL). The mixture was stirred for 6 hours at room temperature, then treated with water (20 mL) and extracted with ethyl acetate (3 x 25 mL). The combined organic layers were dried over sodium sulfate, filtered and concentrated in vacuo. The crude material was purified via chromatography on silica gel (heptane / ethyl acetate / 2-propanol) to afford C102 as a light yellow solid. Yield: 362 mg, 0.381 mmol, 62%. LCMS m/z 950.4 (M+1 ). 1H NMR (400 MHz, DMSO-de), characteristic peaks: δ 9.31 (d, J=8.4 Hz, 1 H), 8.38 (s, 1 H), 8.00 (s, 1 H), 7.41 (br d, J=8.2 Hz, 2H), 7.36 (br d, J=8.8 Hz, 2H), 7.26 (s, 1 H), 6.10 (s, 1 H), 5.20 (s, 2H), 4.92 (br s, 4H), 3.77 (s, 3H), 3.76 (s, 3H), 1.45 (s, 9H), 1.38 (s, 9H). Step 2: Preparation of C103. A solution of C102 (181 mg, 0.191 mmol) in anhydrous /V,/V-dimethylformamide (2.0 mL) was treated with sulfur trioxide pyridine complex (302 mg, 1.91 mmol). The reaction mixture was allowed to stir at room temperature for 6 hours, then cooled to 0 °C and quenched with water. The resulting solid was collected by filtration and dried in vacuo to yield C103 as a white solid. Yield: 145 mg, 0.14 mmol, 74%. APCI m/z 1028.5 (M-1 ). 1H NMR (400 MHz, DMSO-d6), characteristic peaks: δ 1 1.65 (br s, 1 H), 9.37 (d, J=8.6 Hz, 1 H), 8.87 (s, 1 H), 7.49 (br d, J=8.6 Hz, 2H), 7.43 (br d, J=8.6 Hz, 2H), 7.26 (s, 1 H), 7.01 (br d, J=8.9 Hz, 2H), 7.00 (br d, J=8.8 Hz, 2H), 5.43 (s, 2H), 5.20 (dd, J=8.4, 6 Hz, 1 H), 4.01-4.07 (m, 1 H), 3.78 (s, 3H), 3.77 (s, 3H), 3.50- 3.58 (m, 1 H), 3.29-3.37 (m, 1 H), 1.44 (s, 9H), 1.37 (s, 9H). Step 3: Preparation of C104. A solution of C103 (136 mg, 0.132 mmol) in anhydrous dichloromethane (5 mL) was treated with 1 M boron trichloride in p-xylenes (0.92 mL, 0.92 mmol) and allowed to stir at room temperature for 40 minutes. The reaction mixture was cooled in an ice bath, quenched with water (0.4 mL), and transferred into a solution of methyl ferf-butyl ether: heptane (1 :2, 12 mL). The solvent was removed in vacuo and the crude product was purified via reverse phase chromatography (C-18 column; acetonitrile / water gradient with 0.1 % formic acid modifier) to yield C104 as a light yellow solid. Yield: 43 mg, 0.068 mmol, 51 %. LCMS m/z 634.4 (M+1 ). 1H NMR (400 MHz, DMSO-de), characteristic peaks: δ 9.29 (d, J=8.5 Hz, 1 H), 8.10 (s, 1 H), 7.04- 7.10 (m, 1 H), 7.00 (s, 1 H), 6.75 (s, 1 H), 5.05-5.30 (m, 3H), 4.00-4.07 (m, 1 H), 1 .42 (s, 3H), 1 .41 (s, 3H).

Step 4: Preparation of C104-Bis Na salt. A suspension of C104 (212 mg, 0.33 mmol) in water (10 mL) was cooled to 0 °C and treated with a solution of sodium bicarbonate (56.4 mg, 0.67 mmol) in water (2 mL), added dropwise. The reaction mixture was cooled to -70 °C (frozen) and lyophilized to afford C104-Bis Na salt as a white solid. Yield: 210 mg, 0.31 mmol, 93%. LCMS m/z 632.5 (M-1 ). 1H NMR (400 MHz, D20) δ 7.87 (s, 1 H), 6.94 (s, 1 H), 6.92 (s, 1 H), 5.35 (d, J=5 Hz, 1 H), 5.16 (s, 2H), 4.46-4.52 (m, 1 H), 3.71 (dd, half of ABX pattern, J=14.5, 6 Hz, 1 H), 3.55 (dd, half of ABX pattern, J=14.5, 6 Hz, 1 H), 1.43 (s, 3H), 1 .42 (s, 3H).

////////////Pfizer,  monobactam,  PF-?, 1380110-34-8, pfizer, pf, 1380110-45-1, WO 2012073138, Matthew Frank BrownSeungil HanManjinder LallMark. J. Mitton-FryMark Stephen PlummerHud Lawrence RisleyVeerabahu ShanmugasundaramJeremy T. Starr, preclinical

Design, synthesis and biological evaluation of novel 5-hydroxy-2-methyl-4H-pyran-4-one derivatives as antiglioma agents


Med. Chem. Commun., 2018, Advance Article
DOI: 10.1039/C7MD00551B, Research Article
Yi-Bin Li, Wen Hou, Hui Lin, Ping-Hua Sun, Jing Lin, Wei-Min Chen
Two series of 5-hydroxy-2-methyl-4H-pyran-4-one derivatives were synthesized and their antiglioma activities were evaluated.

Design, synthesis and biological evaluation of novel 5-hydroxy-2-methyl-4H-pyran-4-one derivatives as antiglioma agents

Author affiliations

Abstract

D-2-Hydroxyglutarate (D-2HG) is frequently found in human brain cancers. Approximately 50–80% of grade II glioma patients have a high level of D-2HG production, which can lead to cancer initiation. In this study, a series of novel 5-hydroxy-2-methyl-4H-pyran-4-one derivatives were designed and synthesized as antiglioma agents, and their related structure–activity relationships are discussed. Among these novel compounds, 4a exhibited promising anti-proliferative activity against glioma HT1080 cells and U87 cells with an IC50 of 1.43 μM and 4.6 μM, respectively. Further studies found that the most active compound (4a) shows an 86.3% inhibitory rate against the intracellular production of D-2HG at 1 μM, and dramatic inhibitory effects, even at 1 μM on the colony formation and migration of U87 and HT1080 cells.

STR1 STR2 str3 str4
6,6′-((4-(Benzyloxy)phenyl)methylene)bis(5-hydroxy-2-methyl-4H-pyran-4- one) (4a) The reaction was performed according to the general procedure C, using 1 (1.00 g, 7.90 mmol) and 4-(benzyloxy)benzaldehyde (0.84 g, 3.95 mmol).2 The crude product was recrystallized from isopropanol affording a white powder 4a (1.53 g, 87%): mp 261.4-262.1oC; 1HNMR (300 MHz, DMSO-d6)  2.22 (s, 6H, CH3), 5.08 (s, 3H, OCH2- Ph), 5.96 (s, 1H, CH-Ar), 6.25 (s, 2H, C=CH), , 7.01 (d, J = 9.0 Hz, 2H, Ar-H3’/H5’), 7.22 (d, J = 9.0 Hz, 2H, Ar-H2’/H6’), 7.31-7.45 (m, 5H, Ph-H); 13CNMR (75 MHz, DMSO-d6)  173.95, 165.08, 158.12, 151.20, 147.68, 142.19, 140.77, 137.42, 129.87, 128.91, 128.16, 127.69, 115.46, 114.97, 111.74, 69.69, 19.63; ESI-MS m/z: 447.1 [M+H]+ ; ESI-HRMS m/z: 447.1438 [M+H]+ , calcd for C26H23O7 447.1438.

Debio-1452


Image result for Debio-1452

Debio-1452, AFN 1252

AFN-1252; UNII-T3O718IKKM; API-1252; CAS 620175-39-5; CHEMBL1652621; (E)-N-methyl-N-((3-methylbenzofuran-2-yl)methyl)-3-(7-oxo-5,6,7,8-tetrahydro-1,8-naphthyridin-3-yl)acrylamide

  • MFC22 H21 N3 O3
  • 2-Propenamide, N-methyl-N-[(3-methyl-2-benzofuranyl)methyl]-3-(5,6,7,8-tetrahydro-7-oxo-1,8-naphthyridin-3-yl)-, (2E)-
  •  MW375.42
  • Phase 2, clinical trials for the oral treatment of staphylococcal infections, including hospital and community-acquired MRSA and acute bacterial skin and skin structure infections
  • Qualified Infectious Disease Product designation

GlaxoSmithKline plc INNOVATOR

Image result

Debiopharm SA,

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Image result for Affinium

Melioidosis, Enoyl ACP reductase Fabl inhibitor

Debio-1452, a novel class fatty acid biosynthesis (FAS) II pathway inhibitor, was studied in phase II clinical trials for the oral treatment of staphylococcal infections, including hospital and community-acquired MRSA and acute bacterial skin and skin structure infections. Debiopharm is developing oral and IV formulations of a prodrug of Debio-1452, Debio-1450.

Infections caused by or related to bacteria are a major cause of human illness worldwide. Unfortunately, the frequency of resistance to standard antibacterials has risen dramatically over the last decade, especially in relation to Staphylococcus aureus. For example, such resistant S. aureus includes MRSA, resistant to methicillin, vancomycin, linezolid and many other classes of antibiotics, or the newly discovered New Delhi metallo-beta-lactamase- 1 (NDM-1) type resistance that has shown to afford bacterial resistant to most known antibacterials, including penicillins, cephalosporins, carbapenems, quinolones and fluoroquinolones, macrolides, etc. Hence, there exists an urgent, unmet, medical need for new agents acting against bacterial targets..

In recent years, inhibitors of Fabl, a bacterial target involved in bacterial fatty acid synthesis, have been developed and many have been promising in regard to their potency and tolerability in humans, including a very promising Fabl inhibitor, (E)-N-methyl-N-((3-methylbenzofuran-2-yl)methyl)-3-(7-oxo-5,6,7,8-tetrahydro-l,8-naphthyridin-3-yl)acrylamide. This compound, however, has been found to be difficult or impracticable to formulate into acceptable oral and parenteral (e.g., intravenous or subcutaneous) formulations, and has marked insolubility, poor solution stability, and oral bioavailability. Much effort, over a decade or more, has been expended to design and synthesize an alternative compound that retains the significant inhibition of Fabl upon administration, but has improved physical and chemical characteristics that finally allow for practical oral and parenteral formulations. Up to now, no such compound has been identified that has adequate stability in the solid state, in aqueous solutions, together with excellent oral bioavailability that is necessary for oral and/or a parenteral administration, and is capable of being formulated into an oral and/or intravenous or intramuscular drug product using practical and commonly utilized methods of sterile formulation manufacture.

Debio-1452 is expected to have high potency against all drug-resistant phenotypes of staphylococci, including hospital and community-acquired MRSA.

Affinium obtained Debio-1452, also known as API-1252, through a licensing deal with GlaxoSmithKline. In 2014, Debiopharm acquired the product from Affinium.

In 2013, Qualified Infectious Disease Product designation was assigned to the compound for the treatment of acute bacterial skin and skin structure infections (ABSSSI).

Image result for Debio-1452

Image result for Debio-1452

AFN-1252.png

SYNTHESIS

Heck coupling of 6-bromo-3,4-dihydro-1,8-naphthyridin-2-one with t-butyl acrylate in the presence of Pd(OAc)2, DIEA and P(o-tol)3  in propionitrile/DMF or acetonitrile/DMF affords naphthyridinyl-acrylate,

Whose t-butyl ester group is then cleaved using TFA in CH2Cl2 to furnish, after treatment with HCl in dioxane, 3-(7-oxo-6,8-dihydro-5H-1,8-naphthyridin-3-yl)acrylic acid hydrochloride

SEE BELOW………

Finally, coupling of acid with N-methyl-N-(3-methylbenzofuran-2-ylmethyl)amine using EDC, HOBt and DIEA in DMF provides the target AFN-1252

Preparation of N-methyl-N-(3-methylbenzofuran-2-ylmethyl)amine :

Chlorination of 3-methylbenzofuran-2-carboxylic acid  with (COCl)2 and catalytic DMF, followed by condensation with CH3NH2 in CH2Cl2 yields the corresponding benzofuran-2-carboxamide,

Which is then reduced with LiAlH4 in THF to furnish N-methyl-N-(3-methylbenzofuran-2-ylmethyl)amine.

CONTD……..

Reduction of 2-aminonicotinic acid  with LiAlH4 in THF gives (2-amino-3-pyridinyl)methanol ,

which upon bromination with Br2 in AcOH yields (2-amino-5-bromo-3-pyridinyl)methanol hydrobromide.

Substitution of alcohol  with aqueous HBr at reflux provides the corresponding bromide,

which undergoes cyclocondensation with dimethyl malonate  in the presence of NaH in DMF/THF to furnish methyl 6-bromo-2-oxo-1,2,3,4-tetrahydro-1,8-naphthyridine-3-carboxylate.

Hydrolysis of ester with NaOH in refluxing MeOH, followed by decarboxylation in refluxing HCl leads to 6-bromo-3,4-dihydro-1,8-naphthyridin-2-one

PATENT

US-20170088822

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Aurigene Discovery Technologies Ltd

Novel co-crystalline polymorphic form of a binary enoyl-acyl carrier protein reductase (FabI) and FabI inhibitor ie AFN-1252. The FabI was isolated from Burkholderia pseudomallei (Bpm). The co-crystal is useful for identifying an inhibitor of FabI, which is useful for treating BpmFabI associated disease ie melioidosis. Appears to be the first patenting to be seen from Aurigene Discovery Technologies or its parent Dr Reddy’s that focuses on BpmFabI crystal; however, see WO2015071780, claiming alkylidine substituted heterocyclyl derivatives as FabI inhibitors, useful for treating bacterial infections. Aurigene was investigating FabI inhibitors, for treating infectious diseases, including bacterial infections such as MRSA infection, but its development had been presumed to have been discontinued since December 2015; however, publication of this application would suggest otherwise.

WO2015071780

PATENTS

US 20060142265

http://www.google.co.in/patents/US20060142265

PATENT

https://patentscope.wipo.int/search/en/detail.jsf?docId=WO2013190384&recNum=1&maxRec=&office=&prevFilter=&sortOption=&queryString=&tab=PCTDescription

Patent ID Patent Title Submitted Date Granted Date
US8901105 Prodrug derivatives of (E)-N-methyl-N-((3-M ethylbenzofuran-2-yl)methyl)-3-(7-oxo-5, 6, 7, 8-tetrahydro-1, 8-naphthyridin-3-yl)acrylamide 2013-08-26 2014-12-02
US2015065415 PRODRUG DERIVATIVES OF (E)-N-METHYL-N-((3-METHYLBENZOFURAN-2-YL)METHYL)-3-(7-OXO-5, 6, 7, 8-TETRAHYDRO-1, 8-NAPHTHYRIDIN-3-YL)ACRYLAMIDE 2014-11-06 2015-03-05
Patent ID Patent Title Submitted Date Granted Date
US7049310 Fab I inhibitors 2004-07-29 2006-05-23
US7250424 Fab I inhibitors 2006-06-01 2007-07-31
US7879872 Compositions comprising multiple bioactive agents, and methods of using the same 2006-06-29 2011-02-01
US2009042927 Salts, Prodrugs and Polymorphs of Fab I Inhibitors 2009-02-12
US7741339 Fab I Inhibitors 2009-09-03 2010-06-22
US8153652 Fab I Inhibitors 2011-04-28 2012-04-10
US2012010127 Compositions Comprising Multiple Bioactive Agents, and Methods of Using the Same 2012-01-12
US2013281442 Compounds for Treatment of Bovine Mastitis 2011-06-13 2013-10-24
US2013150400 SALTS, PRODRUGS AND POLYMORPHS OF FAB I INHIBITORS 2012-08-09 2013-06-13
US2014309191 SALTS, PRODRUGS AND POLYMORPHS OF FAB I INHIBITORS 2013-11-08 2014-10-16

////////////Debio-1452, AFN 1252,AFN-1252, UNII-T3O718IKKM, API-1252, 620175-39-5, PRECLINICAL, Phase 2, Qualified Infectious Disease Product designation

CC1=C(OC2=CC=CC=C12)CN(C)C(=O)C=CC3=CC4=C(NC(=O)CC4)N=C3

Astellas Pharma Inc. new Glucokinase Activator, ASP ? for Type 2 Diabetes


str1

ASP ?

(2R)-2-(4-cyclopropanesulfonyl-3-cyclopropylphenyl)-N-[5-(hydroxymethyl)pyrazin-2-yl]-3-[(R)-3-oxocyclopentyl]propanamide

CAS 1174229-89-0
MW C25 H29 N3 O5 S
Benzeneacetamide, 3-cyclopropyl-4-(cyclopropylsulfonyl)-N-[5-(hydroxymethyl)-2-pyrazinyl]-α-[[(1R)-3-oxocyclopentyl]methyl]-, (αR)-
Molecular Weight, 483.58
[α]D20 −128.7 (c 1.00, MeOH);
1H NMR (DMSO-d6, 400 MHz) δ 11.07 (s, 1H), 9.20 (d, J = 1.4 Hz, 1H), 8.41 (d, J = 1.4 Hz, 1H), 7.79 (d, J = 8.2 Hz, 1H), 7.41 (dd, J = 8.2, 1.8 Hz, 1H), 7.15 (d, J = 1.8 Hz, 1H), 5.52 (t, J = 5.7 Hz, 1H), 4.56 (d, J = 6.0 Hz, 2H), 4.04 (t, J = 7.6 Hz, 1H), 3.03–2.97 (m, 1H), 2.79 (tt, J = 8.4, 5.1 Hz, 1H), 2.25–1.81 (m, 8H), 1.53–1.47 (m, 1H), 1.17–1.12 (m, 2H), 1.08–1.02 (m, 4H), 0.89–0.84 (m, 2H);
13C NMR (DMSO-d6, 101 MHz) δ 218.5, 171.8, 152.1, 147.3, 145.7, 143.2, 140.3, 138.2, 134.8, 129.0, 125.3, 125.1, 62.5, 49.9, 44.4, 38.4, 38.2, 34.8, 32.1, 29.1, 12.4, 10.8, 10.7, 5.8;
FTIR (ATR, cm–1) 3544, 3257, 1727, 1692, 1546, 1507, 1363, 1285, 1149, 719;
HRMS (ESI) m/z [M + Na]+ calcd for C25H29N3O5S 506.1726, found 506.1747.
Anal. Calcd for C25H29N3O5S: C, 62.09; H, 6.04; N, 8.69. Found: C, 61.79; H, 6.19; N, 8.62.

To Astellas Pharma,Inc.

Inventors Masahiko Hayakawa, Yoshiyuki Kido, Takahiro Nigawara, Mitsuaki Okumura, Akira Kanai, Keisuke Maki, Nobuaki Amino
Applicant Astellas Pharma Inc.

Image result for Process Chemistry Labs., Astellas Pharma Inc., 160-2 Akahama, Takahagi-shi, Ibaraki 318-0001, Japan

Synthesis

contd…………………………..

PATENT

WO2009091014

https://patentscope.wipo.int/search/en/detail.jsf;jsessionid=56E9927692EF5105140FE1CD1FD14A5D.wapp1nC?docId=WO2009091014&recNum=114&maxRec=374&office=&prevFilter=&sortOption=&queryString=FP%3A%28astellas+pharma%29&tab=FullText

str1

PAPER

A Practical and Scalable Synthesis of a Glucokinase Activator via Diastereomeric Resolution and Palladium-Catalyzed C–N Coupling Reaction

Process Chemistry Labs., Astellas Pharma Inc., 160-2 Akahama, Takahagi-shi, Ibaraki 318-0001, Japan
Astellas Research Technologies Co., Ltd., 21 Miyukigaoka, Tsukuba-shi, Ibaraki 305-8585, Japan
§ Department of Applied Chemistry and Biotechnology, Graduate School of Engineering, Chiba University, 1-33 Yayoicho, Inageku, Chiba 263-8522, Japan
Org. Process Res. Dev., Article ASAP
DOI: 10.1021/acs.oprd.6b00415
 Abstract Image

Here we describe the research and development of a process for the practical synthesis of glucokinase activator (R)-1 as a potential drug for treating type-2 diabetes. The key intermediate, chiral α-arylpropionic acid (R)-2, was synthesized in high diastereomeric excess through the diasteromeric resolution of 7 without the need for a chiral resolving agent. The counterpart 2-aminopyrazine derivative 3 was synthesized using a palladium-catalyzed C–N coupling reaction. This efficient process was demonstrated at the pilot scale and yielded 19.0 kg of (R)-1. Moreover, an epimerization process to obtain (R)-7 from the undesired (S)-7 was developed.

Hayakawa, M.; Kido, Y.; Nigawara, T.; Okumura, M.; Kanai, A.; Maki, K.; Amino, N. PCT Int. Appl. WO/2009/091014 A1 20090723,2009.

https://www.astellas.com/en/ir/library/pdf/3q2017_rd_en.pdf

///////////1174229-89-0, ASTELLAS, Glucokinase Activator, TYPE 2 DIABETES, PRECLINICAL, ASP ?, WO 2009091014Masahiko Hayakawa, Yoshiyuki Kido, Takahiro Nigawara, Mitsuaki Okumura, Akira Kanai, Keisuke Maki, Nobuaki AminoWO2009091014,

O=C(Nc1cnc(cn1)CO)[C@H](C[C@@H]2CC(=O)CC2)c3ccc(c(c3)C4CC4)S(=O)(=O)C5CC5

AMG-3969


Image result for amg 3969

AMG-3969

M.Wt: 522.46
Cas : 1361224-53-4 , MF: C21H20F6N4O3S

WO 2012027261 PRODUCT PATENT

Inventors Kate Ashton, Michael David Bartberger, Yunxin Bo, Marian C. Bryan, Michael Croghan, Christopher Harold Fotsch, Clarence Henderson Hale, Roxanne Kay Kunz, Longbin Liu, Nobuko Nishimura, Mark H. Norman, Lewis Dale Pennington, Steve Fong Poon, Markian Myroslaw Stec, Jean David Joseph St., Jr., Nuria A. Tamayo, Christopher Michael Tegley, Kevin Chao Yang
Applicant Amgen Inc.

2-[4-[(2S)-4-[(6-Amino-3-pyridinyl)sulfonyl]-2-(1-propyn-1-yl)-1-piperazinyl]phenyl]-1,1,1,3,3,3-hexafluoro-2-propanol)

(S)-2-(4-(4-((6-Aminopyridin-3-yl)sulfonyl)-2-(prop-1-yn-1-yl)piperazin-1-yl)phenyl)-1,1,1,3,3,3-hexafluoropropan-2-ol,

mp 113–123 °C;
[α]D20 = +75.1 (c = 2.2, MeOH).
Agents for Type 2 Diabetes,  PRECLINICAL

AMG-3969, a novel and stable small-molecule disruptor of glucokinase (GK) and glucokinase regulatory protein (GKRP) interaction by the optimization of initial screening hit and AMG-1694. AMG-3969 potently induced the dissociation of the GK-GKRP complex and promoted GK translocation both in-vitro and in-vivo. In rodent model of diabetes, AMG-3969 reduced blood glucose levels without affecting euglycemic animals. The study represents the first successful discovery of a small molecule that targets the GK-GKRP complex as a novel pathway for managing blood glucose levels with reduced hypoglycemic risk.

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 Kate Ashton

Kate Ashton

Senior Scientist at Amgen, Inc

Amgen
Thousand Oaks, United States
Dr. Kate Ashton received a Masters in Chemistry with Industrial Experience from the University of Edinburgh. She conducted her PhD thesis research on the synthesis and structure elucidation of Reidispongiolide A with Prof. Ian Paterson at the University of Cambridge, and her postdoctoral work on SOMO catalysis with Prof. David W. C. MacMillan at both Caltech and Princeton. She has been at Amgen for 6 years and has worked on indications for cancer, Alzheimer’s and diabetes.Dr Fecke works in the area of industrial early drug discovery since 1996. He is currently Group Leader in the Primary Pharmacology department at UCB Pharma (UK) and is involved in the identification and characterization of NCE and NBE drugs in molecular interaction assays for both immunological and CNS diseases. Prior to joining UCB, he worked for Novartis and Siena Biotech in the areas of transplant rejection, neurodegeneration and oncology. He obtained his PhD at the Heinrich-Heine-University Dusseldorf in Germany in 1994.

Image result for amg 3969

(S)-2-(4-(4-((6-Aminopyridin-3-yl)sulfonyl)-2-(prop-1-yn-1-yl)piperazin-1-yl)phenyl)-1,1,1,3,3,3-hexafluoropropan-2-ol, AMG-3969

Glucokinase (GK) is a member of a family of four hexokinases that are critical in the cellular metabolism of glucose. Specifically GK, also known as hexokinase IV or hexokinase D, facilitates glucose induced insulin secretion from pancreatic β-cells as well as glucose conversion into glycogen in the liver. GK has a unique catalytic activity that enables the enzyme to be active within the physiological range of glucose (from 5mM glucose to lOmM glucose).

Genetically modified mouse models support the role of GK playing an important role in glucose homeostasis. Mice lacking both copies of the GK gene die soon after birth from severe hyperglycemia, whereas mice lacking only one copy of the GK gene present with only mild diabetes. Mice that are made to overexpress the GK gene in their livers are hypoglycemic.

Numerous human mutations in the GK gene have been identified, with the vast majority of them resulting in proteins with impaired or absent enzymatic activity. These loss-of-function mutations are thought to contribute to the hyperglycemia seen with maturity-onset diabetes of the young type II (MODY-2). A small fraction of these mutations result in a GK with increased catalytic function. These individuals present with moderate to severe hypoglycemia.

GK activity in the liver is transiently regulated by glucokinase regulatory protein (GKRP). GK catalytic activity is inhibited when GK is bound to GKRP. This interaction is antagonized by increasing concentrations of both glucose and fructose -1 -phosphate (F1P). The complex of the two proteins is localized primarily to the nuclear compartment of a cell. Post prandially as both glucose and fructose levels rise, GK released from GKRP translocates to the cytoplasm. Cytoplasmic GK is now free of the inhibitory effects of GKRP and able to kinetically respond to glucose. Evidence from the Zucker diabetic fatty rat (ZDF) indicates that their glucose intolerance may be a result of this mechanism failing to function properly.

A compound that acts directly on GKRP to disrupt its interaction with GK and hence elevate levels of cytoplasmic GK is a viable approach to modulate GK activity. Such an approach would avoid the unwanted hypoglycemic effects of over stimulation of GK catalytic activity, which has been seen in the

development of GK activators. A compound having such an effect would be useful in the treatment of diabetes and other diseases and/or conditions in which GKRP and/or GK plays a role.

CLIP

Antidiabetic effects of glucokinase regulatory protein small-molecule disruptors
Nature 2013, 504(7480): 437

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SYNTHESIS

Figure

aReagents and conditions: (a) 1-propynylmagnesium bromide, THF, 0 °C, 99%; (b) TFA, DCM, then NaBH(OAc)3 77%; (c) NH4OH, EtOH, 120 °C, 88%; (d) chiral SFC, 38%………..Nature 2013,504, 437440

PATENT

https://patentscope.wipo.int/search/en/detail.jsf?docId=WO2012027261

EXAMPLE 241 : 2-(4-(4-((6-AMINO-3-PYRIDINYL)SULFONYL)-2-(l-PROP YN- 1 – YL)- 1 -PIPERAZINYL)PHENYL)- 1,1,1 ,3 ,3 ,3 -HEXAFLUORO-2-PROPANOL

STEP 1 : 4-BENZYL 1 -TERT-BUTYL 2-0X0-1,4-PIPERAZINEDICARBOXYLATE

A 2-L Erlenmeyer flask was charged with 2-piperazinone (36.5 g, 364 mmol, Sigma- Aldrich, St. Louis, MO), sodium carbonate (116 g, 1093 mmol), 600 mL of dioxane, and 150 mL of water. To this was slowly added benzyl chloroformate (62.1 g, 364 mmol, Sigma-Aldrich, St. Louis, MO) at room temperature over 20 min. After the addition was complete, the mixture was stirred for 2 h and then diluted with water and extracted with EtOAc (2 L). The combined organic extracts were dried (MgS04), filtered, and concentrated to give a white solid. To this solid was added 500 mL of DCM, triethylamine (128 mL, 911 mmol), DMAP (4.45 g, 36.4 mmol), and di-tert-butyl dicarbonate (119 g, 546 mmol, Sigma-Aldrich, St. Louis, MO). After 1 h at room temperature, the mixture was diluted with water and the organics were separated. The organics were dried (MgS04), filtered, and concentrated to give a brown oil. To this oil was added 100 mL of DCM followed by 1 L of hexane. The resulting white solid was collected by filtration to give 4-benzyl 1-tert-butyl 2-oxo-l,4-piperazinedicarboxylate (101 g).

STEP 2: BENZYL (2-((TERT-BUTOXYCARBONYL)AMINO)ETHYL)(2-OXO-3 -PENTYN- 1 -YL)CARBAMATE

A 150-mL round-bottomed flask was charged with 4-benzyl 1-tert-butyl

2- oxo-l,4-piperazinedicarboxylate (1.41 g, 4.22 mmol) and THF (5 mL). 1-Propynylmagnesium bromide (0.5 M in THF, 20.0 mL, 10.0 mmol, Sigma-Aldrich, St. Louis, MO) was added at 0 °C slowly. The mixture was stirred at 0 °C for 2 h. Saturated aqueous NH4C1 (40 mL) was added and the aqueous phase was extracted with EtOAc (200 mL, then 2 x 100 mL). The combined organic phases were dried over sodium sulfate, filtered and concentrated in vacuo. The crude product was purified by column chromatography (50 g of silica, 0 to 50% EtOAc in hexanes) to afford benzyl (2-((tert-butoxycarbonyl)amino)ethyl)(2-oxo- 3- pentyn-l-yl)carbamate (1.55 g) as a clear oil.

STEP 3: BENZYL 3-(l-PROPYN-l-YL)-l-PIPERAZINECARBOXYLATE

A 3-L round-bottomed flask was charged with 2-((tert-butoxycarbonyl)amino)ethyl)(2-oxo-3-pentyn-l-yl)carbamate (82.2 g, 219 mmol) and 300 mL of DCM. After cooling to -10 °C, TFA (169 mL, 2195 mmol) was added and the resulting dark solution was stirred at room temperature for 15 min. Sodium triacetoxyborohydride (186 g, 878 mmol, Sigma-Aldrich, St. Louis, MO) was then added portion- wise over 10 min. After 2 h, the mixture was

concentrated, diluted with EtOAc (1 L), and neutralized with 5 N NaOH. The layers were separated and the organic extracts were washed with brine, dried (MgS04), filtered and concentrated. The resulting orange oil was purified via column chromatography (750 g of silica gel, 0 to 4.5 % MeOH/DCM) to give benzyl 3-(l-propyn-l-yl)-l-piperazinecarboxylate (43.7 g) as a brown foam.

STEP 4: BENZYL 3-(l-PROPYN-l-YL)-4-(4-(2,2,2-TRIFLUORO-l-HYDROXY- 1 -(TRIFLUOROMETHYL)ETHYL)PHENYL)- 1 -PIPERAZINECARBOXYLATE

A 150-mL reaction vessel was charged with benzyl 3-(prop-l-yn-l-yl)piperazine-l-carboxylate (2.88 g, 11.2 mmol), 2-(4-bromophenyl)-l, 1,1, 3,3,3-hexafluoropropan-2-ol (4.36 g, 13.5 mmol, Bioorg. Med. Chem. Lett. 2002, 12, 3009), dicyclohexyl(2′,6′-diisopropoxy-[ 1 , 1 ‘-biphenyl]-2-yl)phosphine, RuPhos (0.530 g, 1.14 mmol, Sigma- Aldrich, St. Louis, MO), RuPhos Palladacycle (0.417 g, 0.572 mmol, Strem Chemical Inc, Newburyport, MA), sodium tert-butoxide (2.73 g, 28.4 mmol, Strem Chemical Inc, Newburyport, MA) and toluene (35 mL). The mixture was degassed by bubbling Ar through the solution for 10 min. The vessel was sealed and heated at 100 °C for 1.5 h. The reaction mixture was cooled to room temerature and water (100 mL) was added. The aqueous phase was extracted with EtOAc (3 x 100 mL) and the combined organic phases were washed with saturated aqueous sodium chloride (150 mL). The organic extracts were dried over sodium sulfate, filtered and concentrated in vacuo. The crude product was purified by column chromatography (100 g of silica, 0 to 50% EtOAc in hexanes) to afford benzyl 3-(l-propyn-l-yl)-4-(4-(2,2,2-trifluoro- 1 -hydroxy- 1 -(trifluoromethyl)ethyl)phenyl)- 1 -piperazinecarboxylate as a yellow solid.

STEP 5: 2-(4-(4-((6-CHLORO-3-PYRIDINYL)SULFONYL)-2-(l-PROPYN-l-YL)- 1 -PIPERAZIN YL)PHENYL)- 1,1,1 ,3 ,3 ,3 -HEXAFLUORO-2-PROPANOL

A 500-mL round-bottomed flask was charged with benzyl 3-(l-propyn-l-yl)-4-(4-(2,2,2-trifluoro- 1 -hydroxy- 1 -(trifluoromethyl)ethyl)phenyl)- 1 -piperazinecarboxylate (3.13 g, 6.25 mmol) and TFA (40 mL).

Trifluoromethanesulfonic acid (1.25 mL, 14.1 mmol, Acros/Fisher Scientific, Waltham, MA) was added dropwise at room temperature. After 5 min, additional TfOH (0.45 mL, 5.1 mmol) was added. After an additional 10 min, solid

NaHC03 was carefully added in potions. Saturated aqueous NaHC03 (250 mL) was added slowly to bring pH to approximately 7. The aqueous phase was extracted with EtOAc (100 mL). At this time, more solid NaHC03 was added to the aqueous phase and extracted again with EtOAc (100 mL). The combined organic phases were washed with water (200 mL) and saturated aqueous sodium chloride (200 mL). The combined organic extracts were dried over sodium sulfate, filtered and concentrated in vacuo to afford 3.10 g of tan solid.

A 500-mL round-bottomed flask was charged with this material, triethylamine (5.00 mL, 35.9 mmol) and CH2CI2 (30 mL). 6-Chloropyridine-3-sulfonyl chloride (1.58 g, 7.43 mmol, Organic Process Research & Development 2009, 13, 875) was added in potions at 0 °C. The brown mixture was stirred at 0 °C for 10 min. The volume of the reaction mixture was reduced to approximately 10 mL in vacuo then the mixture was purified twice by column chromatography (100 g of silica, 0 to 50% EtOAc in hexanes) to afford 2-(4-(4-((6-chloro-3-pyridinyl)sulfonyl)-2-( 1 -propyn- 1 -yl)- 1 -piperazinyl)phenyl)- 1,1,1,3,3,3-hexafluoro-2-propanol (3.46 g) as an off-white solid.

STEP 6: 2-(4-(4-((6-AMINO-3-PYRIDINYL)SULFONYL)-2-(l-PROPYN-l-YL)- 1 -PIPERAZIN YL)PHENYL)- 1,1,1 ,3 ,3 ,3 -HEXAFLUORO-2-PROPANOL

A 20-mL sealed tube was charged with 2-(4-(4-((6-chloro-3-pyridinyl)sulfonyl)-2-( 1 -propyn- 1 -yl)- 1 -piperazinyl)phenyl)- 1,1,1,3,3,3-hexafluoro-2-propanol (0.340 g, 0.627 mmol), concentrated ammonium hydroxide (5.00 mL, 38.5 mmol) and EtOH (5 mL). The reaction mixture was heated in an Initiator (Biotage, AB, Uppsala, Sweden) at 120 °C for 1 h. The reaction mixture was further heated in a heating block at 110 °C for 5 h. The reaction mixture was concentrated and purified by column chromatography (25 g of silica, 30 to 80% EtOAc in hexanes) to afford 2-(4-(4-((6-amino-3-pyridinyl)sulfonyl)-2-( 1 -propyn- 1 -yl)- 1 -piperazinyl)phenyl)- 1,1,1,3,3,3-hexafluoro-2-propanol (0.289 g) as a mixture of two enantiomers.

1H NMR (400 MHz, CDC13) δ ppm 8.49 (br. s., 1 H), 7.80 (dd, J= 2.3, 8.8 Hz, 1 H), 7.59 (d, J= 8.8 Hz, 2 H), 6.97 (d, J= 9.0 Hz, 2 H), 6.55 (d, J= 8.8 Hz, 1 H), 5.05 (s, 2 H), 4.46 (br. s., 1 H), 3.85 – 3.72 (m, 2 H), 3.54 (br. s., 1 H), 3.50 – 3.34 (m, 2 H), 2.83 (dd, J= 3.3, 11.0 Hz, 1 H), 2.69 (dt, J= 3.4, 11.0 Hz, 1 H), 1.80 (s, 3 H). m/z (ESI, +ve ion) 523.1 (M+H)+. GK-GKRP IC50 (Binding) = 0.003 μΜ

The individual enantiomers were isolated using chiral SFC. The method used was as follows: Chiralpak® ADH column (21 x 250 mm, 5 μιη) using 35% methanol in supercritical C02 (total flow was 70 mL/min). This produced the two enantiomers with enantiomeric excesses greater than 98%.

2-(4-((2S)-4-((6-amino-3-pyridinyl)sulfonyl)-2-(l -propyn- 1-yl)- 1 -piperazinyl)phenyl)- 1,1,1 ,3 ,3 ,3 -hexafluoro-2-propanol and 2-(4-((2R)-4-((6-amino-3 -pyridinyl)sulfonyl)-2-( 1 -propyn- 1 -yl)- 1 -piperazinyl)phenyl)- 1,1,1,3,3,3-hexafluoro-2-propanol.

FIRST ELUTING PEAK (PEAK #1)

1H NMR (400 MHz, CDC13) δ 8.48 (d, J= 2.3 Hz, 1 H), 7.77 (dd, J= 2.5, 8.8 Hz, 1 H), 7.57 (d, J= 8.8 Hz, 2 H), 6.95 (d, J= 9.2 Hz, 2 H), 6.52 (d, J= 8.8 Hz, 1 H), 4.94 (s, 2 H), 4.44 (br. s., 1 H), 3.82 – 3.71 (m, 2 H), 3.58 – 3.33 (m, 3 H), 2.81 (dd, J= 3.2, 11.1 Hz, 1 H), 2.67 (dt, J= 3.9, 11.0 Hz, 1 H), 1.78 (d, J = 2.2 Hz, 3 H). m/z (ESI, +ve ion) 523.2 (M+H)+. GK-GKRP IC50 (Binding) = 0.002 μΜ.

SECOND ELUTING PEAK (PEAK #2)

1H NMR (400 MHz, CDC13) δ 8.49 (d, J= 1.8 Hz, 1 H), 7.78 (dd, J= 2.3, 8.8 Hz, 1 H), 7.59 (d, J= 8.6 Hz, 2 H), 6.97 (d, J= 9.0 Hz, 2 H), 6.54 (d, J= 8.8 Hz, 1 H), 4.97 (s, 2 H), 4.46 (br. s., 1 H), 3.77 (t, J= 11.7 Hz, 2 H), 3.67 (br. s., 1 H), 3.51 – 3.33 (m, 2 H), 2.82 (dd, J= 3.3, 11.0 Hz, 1 H), 2.68 (dt, J= 3.9, 11.1 Hz, 1 H), 1.79 (d, J= 2.0 Hz, 3 H). m/z (ESI, +ve ion) 523.2 (M+H)+. GK-GKRP IC50 (Binding) = 0.342 μΜ.

Alternative procedure starting after Step 4.

STEP 5 : 2-(4-(4-((6-AMINO-3-PYRIDINYL)SULFONYL)-2-(l-PROPYN-l-YL)- 1 -PIPERAZIN YL)PHENYL)- 1,1,1 ,3 ,3 ,3 -HEXAFLUORO-2-PROPANOL

Alternatively, 2-(4-(4-((6-amino-3-pyridinyl)sulfonyl)-2-( 1 -propyn- 1 -yl)-l-piperazinyl)phenyl)-l,l,l,3,3,3-hexafluoro-2-propanol was synthesized from benzyl 3-( 1 -propyn- 1 -yl)-4-(4-(2,2,2-trifluoro- 1 -hydroxy- 1 -(trifluoromethyl)ethyl)phenyl)- 1 -piperazinecarboxylate as follows.

A 2-L round-bottomed flask was charged with benzyl 3 -(1 -propyn- 1-yl)-4-(4-(2,2,2-trifluoro- 1 -hydroxy- 1 -(trifluoromethyl)ethyl)phenyl)- 1 -piperazinecarboxylate (21.8 g, 43.5 mmol, step 5) and TFA (130 mL).

Trifluoromethanesulfonic acid (11.6 mL, 131 mmol, Acros/Fisher Scientific, Waltham, MA) was added slowly at rt resulting orange cloudy mixture. After stirring at rt for 10 min, the volume of the reaction mixture was reduced to half in vacuo. Solid NaHC03 was added in potions until the mixture became sludge. Saturated aqueous NaHC03(800 mL) was added slowly until the pH was about

8. The aqueous phase was extracted with EtOAc (3 x 250 mL). The combined organic phases were washed with water (500 mL) and saturated aqueous NaCl (500 mL). The organic phase was dried over sodium sulfate, filtered and concentrated in vacuo. This material was dissolved into DCM (200 mL) and triethylamine (31.0 mL, 222 mmol) was added. Then 6-aminopyridine-3-sulfonyl chloride (9.40 g, 48.8 mmol, published PCT patent application no. WO

2009/140309) was added in potions over 10 min period. The brown mixture was stirred at room temperature for 10 min. The reaction mixture was washed with water (300 mL) and saturated aqueous NaCl (300 mL). The organic phase was dried over sodium sulfate, filtered and concentrated in vacuo. The crude product was purified by column chromatography (780 g of total silica, 30 to 90% EtOAc in hexanes) to afford 2-(4-(4-((6-amino-3-pyridinyl)sulfonyl)-2-(l-propyn-l-yl)-l-piperazinyl)phenyl)-l,l,l,3,3,3-hexafluoro-2-propanol (19.4 g) as a mixture of two enantiomers.

Paper

Nonracemic Synthesis of GK–GKRP Disruptor AMG-3969

Therapeutic Discovery, Amgen Inc., One Amgen Center Drive, Thousand Oaks, California 91320, United States
Amgen Inc. 360 Binney Street, Cambridge, Massachusetts 02142, United States
J. Org. Chem., 2014, 79 (8), pp 3684–3687

Abstract Image

A nonracemic synthesis of the glucokinase–glucokinase regulatory protein disruptor AMG-3969 (5) is reported. Key features of the synthetic approach are an asymmetric synthesis of the 2-alkynyl piperazine core via a base-promoted isomerization and a revised approach to the synthesis of the aminopyridinesulfonamide with an improved safety profile.

(S)-2-(4-(4-((6-Aminopyridin-3-yl)sulfonyl)-2-(prop-1-yn-1-yl)piperazin-1-yl)phenyl)-1,1,1,3,3,3-hexafluoropropan-2-ol, AMG-3969 (5)

(S)-2-(4-(4-((6-aminopyridin-3-yl)sulfonyl)-2-(prop-1-yn-1-yl)piperazin-1-yl)phenyl)-1,1,1,3,3,3-hexafluoropropan-2-ol (5) (64.0 g, 49% yield) as white solid. The enanatiomeric excess was found to be >99.5% by chiral SFC (see Supporting Information):
1H NMR (400 MHz, CDCl3) δ 8.47 (s, 1 H), 7.79 (d, J = 8.6 Hz, 1 H), 7.59 (d, J = 8.2 Hz, 2 H), 6.97 (d, J = 8.6 Hz, 2 H), 6.55 (d, J = 8.8 Hz, 1 H), 5.06 (br s, 2 H), 4.45 (br s, 1 H), 3.96 (br s, 1 H), 3.77 (t, J = 12.1 Hz, 2 H), 3.50–3.35 (m, 2 H), 2.82 (d, J = 11.0 Hz, 1 H), 2.68 (t, J = 10.9 Hz, 1 H), 1.79 (s, 3 H);
13C NMR (101 MHz, CD3OD) δ 163.8, 152.0, 150.1, 138.2, 129.0, 124.7 (q), 123.9, 121.1, 117.5, 109.3, 82.8, 78.3 (m), 75.5, 52.0, 47.2, 44.9, 3.2;
 
HRMS (ESI-TOF) m/z [M + H]+calcd for C21H21F6N4O3S 523.1239, found 523.1229;
 
mp 113–123 °C;
 
[α]D20 = +75.1 (c = 2.2, MeOH).
 

Clip

AMG-3969 is a disruptor of the glucokinase (GK)–glucokinase regulatory protein (GKRP) protein–protein interaction. Bourbeau and co-workers at Amgen describe their efforts towards an asymmetric synthesis of this compound ( J. Org. Chem. 2014, 79, 3684). The discovery route to this compound involved seven steps (14% overall yield), had certain safety concerns and relied upon SFC separation of the API enantiomers. The new route requires five steps (26% overall yield) and delivers the API in excellent enantiomeric excess (99% ee). A key feature of the synthetic approach was an asymmetric synthesis of the 2-alkynylpiperazine core via a base-promoted isomerization. It was found that the strongly basic conditions employed for the “alkyne-walk” did not erode the previously established stereocenter. Also, safety concerns around a late-stage amination of a 2-chloropyridine intermediate in the discovery route were alleviated by starting with a Boc-protected diaminopyridine instead.
PATENT

INTERMEDIATE A: TERT-EUTYL (5-(CHLOROSULFONYL)-2-PYRIDINYL)CARBAMATE

0,N

STEP 1 : TERT-BUTY (5-NITRO-2-PYRIDINYL)CARBAMATE

A 3-L round-bottomed flask was charged with 5-nitro-2-pyridinamine (75.0 g, 539 mmol, Alfa Aesar, Ward Hill, MA) and 500 mL of DCM. To this was added triethylamine (82 g, 810 mmol), di-tert-butyl dicarbonate (129 g, 593 mmol, Sigma-Aldrich, St. Louis, MO), and N,N-dimethylpyridin-4-amine (32.9 g, 270 mmol, Sigma-Aldrich, St. Louis, MO). After stirring at rt for 18 h, the mixture was diluted with water and the solid was collected by filtration. The yellow solid was washed with MeOH to give tert-butyl (5-nitro-2-pyridinyl)carbamate (94.6 g) as a light yellow solid.

STEP 2: TERT-BUTY (5 – AMINO-2-P YRIDINYL)C ARB AM ATE

A 3-L round-bottomed flask was charged with tert-butyl (5-nitro-2-pyridinyl)carbamate (96.4 g, 403 mmol), 500 mL of MeOH, 500 mL of THF, and 100 mL of sat aq NH4Cl. Zinc (105 g, 1610 mmol, Strem Chemical Inc, Newburyport, MA) was slowly added (over 10 min) to this solution. The mixture was stirred at room temperature for 12 h, then filtered. The filtrate was concentrated and then diluted with EtOAc and washed with water. The organic extracts were dried over MgS04, filtered, and concentrated. The resulting solid was recrystallized from MeOH to give tert-butyl(5-amino-2-pyridinyl)carbamate (38.6 g) as a light-yellow solid.

STEP 3: TERT-BUTYL (5-(CHLOROSULFONYL)-2-PYRIDINYL)CARBAMATE

A 3-L round-bottomed flask was charged with sodium nitrite (15.3 g, 221 mmol, J. T. Baker, Philipsburg, NJ), 100 mL of water and 500 mL of MeCN. After cooling to 0 °C, cone, hydrochloric acid (231 mL, 2770 mmol) was slowly added keeping the internal temperature below 10 °C. After stirring at 0 °C for 10 min, tert-butyl (5-amino-2-pyridinyl)carbamate (38.6 g, 184 mmol) was added as a suspension in MeCN (200 mL). The mixture was stirred for 30 min, then 150 mL of AcOH, copper(ii) chloride (12.4 g, 92.2 mmol, Sigma-Aldrich, St. Louis, MO), and copper(i) chloride (0.183 g, 1.85 mmol, Strem Chemical Inc,

Newburyport, MA) were added. S02 gas (Sigma-Aldrich, St. Louis, MO) was bubbled through the solution for 15 min. The mixture was stirred at 0 °C for 30 min, then about 500 mL of ice-cold water was added. The resulting precipitate was collected by filtration and dried over MgS04 to give tert-butyl (5-(chlorosulfonyl)-2-pyridinyl)carbamate (15.5 g) as a white solid.

1H NMR (400MHz, CDC13) δ ppm 8.93 (br s, 1 H), 8.63 – 8.42 (m, 1 H), 8.35 -7.94 (m, 2 H), 1.58 (s, 9 H).

INTERMEDIATE B: (3S)-l-BENZYL-3-(l-PROPYN-l-YL)PIPERAZINE

STEP 1 : (3S)-l-BENZYL-3-(2-PROPYN-l-YL)-2,5-PIPERAZINEDIONE

A 1-L round-bottoemd flask was charged with (S)-2-((tert-butoxycarbonyl)amino)pent-4-ynoic acid (42.0 g, 197 mmol, AK Scientific, Union City, CA), ethyl 2-(benzylamino)acetate (40.0 g, 207 mmol, Sigma-Aldrich, St. Louis, MO), HATU (90 g, 240 mmol, Oakwood Products, West Columbia, SC) and 200 mL of DMF. To this was added N-ethyl-N-isopropylpropan-2-amine (51.5 ml, 296 mmol, Sigma-Aldrich, St. Louis, MO). After 15 min of stirring at rt, the mixture was diluted with water 300 mL and extracted with 1 L of 20% EtOAc in diethyl ether. The layers were separated and the organic was washed with 2 M HCl, water, sat. aq. NaHC03 and brine. The extracts were dried and concentrated to give an off-white solid. To this was added 200 mL of DCM and TFA (152 ml, 1970 mmol, Sigma-Aldrich, St. Louis, MO). After stirring at rt for 30 min, the mixture was concentrated and then azetroped with 100 mL toluene (twice). To the brown oil obtained was added ammonia (2 M in MeOH, 394 ml, 789 mmol, Sigma-Aldrich, St. Louis, MO). The mixture was stirred at rt for 30 min. The mixture was concentrated, dissolved in EtOAc, and washed with water. The organics were dried (MgS04), filtered, and concentrated to give a white solid that was triturated with diethyl ether to give (S)-l-benzyl-3-(prop-2-yn-l-yl)piperazine-2,5-dione (37.3 g) as a white solid.

STEP 2: (3S)-l-BENZYL-3-(2-PROPYN-l-YL)PIPERAZINE

A 1-L round-bottomed flask was charged with (S)-l-benzyl-3-(prop-2-yn-l-yl)piperazine-2,5-dione (37.3 g, 154 mmol) and 150 mL of THF. To this was slowly added aluminum (III) lithium hydride (1M in THF, 539 ml, 539 mmol, Sigma-Aldrich, St. Louis, MO). After the addition was complete the mixture was heated at 80 °C for 12 h. The mixture was then cooled to 0 °C and solid sodium sulfate decahydrate was added until bubbling ceased. The mixture was filtered and the filtrate was concentrated to give (S)-l-benzyl-3-(prop-2-yn-l-yl)piperazine (18.1 g) as a yellow oil.

STEP 3: (35)-l-BENZYL-3-(l-PROPYN-l-YL)PIPERAZINE

To a solution of (35)-l-benzyl-3-(2-propyn-l-yl)piperazine (2.3 g, 11 mmol) in THF (50 mL) was added potassium t-butoxide (2.41 g, 21.5 mmol, Sigma-Aldrich, St. Louis, MO). The reaction mixture was stirred at rt for 30 min, then quenched with water (200 mL) and EtOAc (300 mL) was added. The organic phase was dried over sodium sulfate, filtered and concentrated under a vacuum to give a solid that was purified by silica gel column chromatography (0 to 10% MeOH in CH2CI2) and then recrystallized from hexanes to afford (35)- 1-benzyl-3-(l-propyn-l-yl)piperazine (2.16 g) as an off-white solid.

1H NMR (400MHz, CD3OD) δ ppm 7.42 – 7.21 (m, 5 H), 3.59 – 3.49 (m, 3 H), 2.93 (td, J= 2.9, 12.4 Hz, 1 H), 2.86 – 2.73 (m, 2 H), 2.68 (d, J= 11.3 Hz, 1 H), 2.22 – 2.04 (m, 2 H), 1.80 (d, J= 2.3 Hz, 3 H).

INTERMEDIATE C: N,N-BIS(4-METHOXYBENZYL)-5-(((35)-3-(l-PROPYN- 1 – YL)- 1 -PIPERAZINYL)SULFONYL)-2-PYRIDIN AMINE

STEP 1 : (35)-l-((6-CHLORO-3-PYRIDINYL)SULFONYL)-3-(l-PROPYN-l-YL)PIPERAZINE

To a stirred solution of benzyl (35)-3-(l-propyn-l-yl)-l-piperazinecarboxylate (2.51 g, 9.71 mmol, Intermediate E) in TFA (20 mL) in 250-mL round-bottomed flask, trifluoromethanesulfonic acid (2.59 mL, 29.1 mmol, Alfa Aesar, Ward Hill, MA) was added slowly at rt. After stirring at room temperature for 3 min, the reaction mixture was concentrated to dryness under a vacuum. DCM (20 mL) was added to the residue followed by triethylamine (13.5 mL, 97 mmol). After the material went into solution, the mixture was cooled to 0 °C and 6-chloro-3-pyridinesulfonyl chloride (2.06 g, 9.73 mmol, Organic Process Research & Development 2009, 13, 875) was added portion-wise. After 5 min of stirring at 0 °C, water (40 mL) was added at that temperature and the layers were separated. The aqueous phase was extracted with DCM (2 x 50 mL). The combined organic phases were washed with saturated aqueous sodium chloride (60 mL). The organic phase was dried over sodium sulfate, filtered and concentrated under a vacuum. The crude product was purified by column chromatography (100 g of silica, 30 to 90% EtOAc in hexanes) to afford (35)- 1-((6-chloro-3-pyridinyl)sulfonyl)-3-(l-propyn-l-yl)piperazine (2.61 g) as an off-white solid.

STEP 2: N,N-BIS(4-METHOXYBENZYL)-5-(((35)-3-(l-PROPYN-l-YL)-l-PIPERAZINYL)SULFONYL)-2-PYRIDIN AMINE

A mixture of (35)-l-((6-chloro-3-pyridinyl)sulfonyl)-3-(l-propyn-l-yl)piperazine (2.6 g, 8.7 mmol), N-(4-methoxybenzyl)-l-(4-methoxyphenyl)methanamine (2.40 g, 9.33 mmol, WO2007/109810A2), and DIPEA (2.4 mL, 14 mmol) in z-BuOH (8.0 mL) was heated at 132 °C using a microwave reactor for 3 h. This reaction was run three times (total starting material amount was 7.2 g). The mixtures from the three runs were combined and partitioned between EtOAc (200 mL) and aqueous NaHC03 (half saturated, 50 mL). The organic layer was washed with aqueous NaHC03 (3 x 50 mL), dried over Na2S04, filtered, and concentrated. The residue was purified (5-times total) by chromatography on silica using MeOH:DCM:EtOAc:hexane

(4:20:20:60) as eluent to give N,N-bis(4-methoxybenzyl)-5-(((3S)-3-(l-propyn-i-yl)-l-piperazinyl)sulfonyl)-2-pyridinamine (6.6 g) as a white foam.

1H NMR (400MHz ,CDC13) δ ppm 8.55 (d, J= 2.3 Hz, 1 H), 7.64 (dd, J= 2.5, 9.0 Hz, 1 H), 7.13 (d, J= 8.6 Hz, 4 H), 6.91 – 6.81 (m, 4 H), 6.47 (d, J= 9.0 Hz, 1 H), 4.75 (s, 4 H), 3.80 (s, 6 H), 3.68 – 3.61 (m, 1 H), 3.57 (d, J= 11.2 Hz, 1 H), 3.41 (d, J= 11.3 Hz, 1 H), 3.07 (td, J= 3.3, 12.1 Hz, 1 H), 2.87 (ddd, J= 2.9, 9.7, 12.2 Hz, 1 H), 2.63 – 2.47 (m, 2 H), 1.80 (d, J= 2.2 Hz, 3 H). One exchangeable proton was not observed, m/z (ESI, +ve ion) 521.2 (M+H)+.

INTERMEDIATE D: rEi?r-BUTYL(5-(((35)-3-(l-PROPYN-l-YL)-4-(4-(2-(TRIFLUOROMETHYL)-2-OXIRANYL)PHENYL)- 1 -PIPERAZINYL)SULFONYL)-2-PYRIDINYL)CARBAMATE

step 1 step 2

STEP 1 : l-BR0M0-4-(l-(TRIFLU0R0METHYL)ETHENYL)BENZENE

To a 1-L round-bottomed flask was added methyl phenylphosphonium bromide (25.4 g, 71.1 mmol, Sigma- Aldrich, St. Louis, MO) and toluene (75 mL). The resulting mixture was stirred for 5 min then concentrated and dried under high vacuum for 30 min. To this residue was added THF (300 mL) followed by n-butyllithium (2.5 M in hexanes, 29.0 mL, 71.1 mmol, Aldrich, St. Louis, MO) dropwise via an addition funnel. After being stirred for 1 h at rt, a solution of l-(4-bromophenyl)-2,2,2-trifluoroethanone (15.0 g, 59.3 mmol, Matrix Scientific, Columbia, SC) in THF (20 mL) was added to the reaction mixture dropwise via an addition funnel. The reaction mixture was stirred at rt for 2 h. The reaction was quenched with saturated aqueous NH4C1 and the mixture was concentrated. The residue was partitioned between diethyl ether (150 mL) and saturated aqueous NH4C1 (80 mL). The organic layer was washed with water and brine, dried over MgS04, filtered, and concentrated. The resulting crude product was purified by column chromatography (330 g of silica gel, 2 to 5% EtOAc in hexanes) to afford l-bromo-4-(l-(trifluoromethyl)ethenyl)benzene (14.0 g) as a brown liquid.

STEP 2: 2-(4-BROMOPHENYL)-3,3,3-TRIFLUORO-l,2-PROPANEDIOL

To a solution of l-bromo-4-(l-(trifluoromethyl)ethenyl)benzene (13.5 g, 53.8 mmol) in acetone (100 mL) and water (100 mL) was added NMO (6.90 g, 59.2 mmol, Sigma- Aldrich, St. Louis, MO) and osmium tetroxide (0.140 mL, 2.70 mmol, Sigma-Aldrich, St. Louis, MO). The resulting mixture was stirred at rt for 6 h. The reaction mixture was filtered and the filtrate was concentrated. The residue was partitioned between EtOAc (100 mL) and water (30 mL). The aqueous layer was extracted with EtOAc (2 x 75 mL). The combined organic layers were dried over MgS04, filtered, and concentrated. The resulting product was purified by column chromatography (330 g of silica gel, 0 to 8% MeOH in DCM) to afford 2-(4-bromophenyl)-3,3,3-trifluoro-l,2-propanediol (14.5 g) as an off-white solid.

STEP 3: 4-(4-BROMOPHENYL)-2,2-DIMETHYL-4-(TRIFLUOROMETHYL)-1,3-DIOXOLANE

To a solution of 2-(4-bromophenyl)-3,3,3-trifluoro-l,2-propanediol (14.5 g, 51.0 mmol) in acetone (200 mL) was added 2,2-dimethoxypropane (19.0 mL, 153 mmol, Sigma-Aldrich, St. Louis, MO) and /?-toluenesulfonic acid (0.485 g, 2.54 mmol, Sigma-Aldrich, St. Louis, MO). The resulting mixture was stirred at rt for 20 h. Additional 2,2-dimethoxypropane (19.0 mL, 153 mmol, Sigma-Aldrich, St. Louis, MO) and /?-toluenesulfonic acid (0.485 g, 2.54 mmol, Sigma-Aldrich, St. Louis, MO) were added and the reaction was stirred for another 20 h. The reaction was quenched with saturated aqueous NaHC03 (10 mL). The reaction mixture was concentrated and the residue was partitioned between

EtOAc (100 mL) and saturated aqueous NaHC03 (60 mL). The aqueous layer was extracted with EtOAc (2 x 50 mL). The combined organic layers were dried over MgS04, filtered, and concentrated. The resulting product was purified by column chromatography (330 g of silica gel, 0 to 8% EtOAc in hexanes) to afford 4-(4-bromophenyl)-2,2-dimethyl-4-(trifluoromethyl)-l,3-dioxolane (15.7 g) as a colorless liquid.

STEP 4: BENZYL (3S)-4-(4-(2,2-DIMETHYL-4-(TRIFLUOROMETHYL)-l,3-DIOXOLAN-4-YL)PHENYL)-3-(l -PROPYN- 1 -YL)- 1 -PIPERAZINECAPvBOXYLATE

To a 20-mL vial was added benzyl (3S)-3-(l -propyn- l-yl)-l-piperazinecarboxylate (1.0 g, 3.87 mmol, Intermediate E), RuPhos Palladacycle (0.250 g, 0.310 mmol, Strem Chemical, Newburyport, MA), 4-(4-bromophenyl)-2,2-dimethyl-4-(trifluoromethyl)-l,3-dioxolane (2.50 g, 7.74 mmol), dioxane (15.0 mL), and sodium t-butoxide (0.740 g, 7.74 mmol, Sigma-Aldrich, St.

Louis, MO). The reaction mixture was degassed by bubbling N2 through the solution for 5 min, then the vial was capped. The reaction mixture was heated at 80 °C for 30 min then allowed to cool to rt and partitioned between EtOAc (70 mL) and water (40 mL). The aqueous layer was extracted with EtOAc (1 x 50 mL). The combined organic layers were dried over MgS04, filtered, and concentrated. The crude product was purified by column chromatography (80 g of silica, 5% to 30% EtOAc in hexanes) to afford benzyl (35)-4-(4-(2,2-dimethyl-4-(trifluoromethyl)- 1 ,3-dioxolan-4-yl)phenyl)-3-(l -propyn- 1 -yl)- 1 -piperazinecarboxylate (1.6 g) as a yellow foam.

STEP 5: rEi?r-BUTYL(5-(((35)-3-(l-PROPYN-l-YL)-4-(4-(2,2,2-TRIFLUORO- 1 -HYDROXY- 1 -(HYDROXYMETH YL)ETHYL)PHENYL)- 1 -PIPERAZINYL)SULFONYL)-2-PYRIDINYL)CARBAMATE

To a 150-mL round-bottomed flask was added benzyl (3S)-4-(4-(2,2-dimethyl-4-(trifluoromethyl)- 1 ,3 -dioxolan-4-yl)phenyl)-3 -( 1 -propyn- 1 -yl)- 1 -piperazinecarboxylate (1.60 g, 3.18 mmol) and TFA (20 mL, Sigma-Aldrich, St. Louis, MO). After the substrate was completely dissolved in TFA,

trifluoromethanesulfonic acid (0.850 mL, 9.55 mmol, Alfa Aesar, Ward Hill,

MA) was added and the resulting mixture was stirred at rt for 1.5 h. The reaction mixture was slowly poured into a 300-mL beaker which contained 100 mL ice water. The resulting mixture was stirred while NaOH pellets (11.0 g) were slowly added to adjust the pH to 7. The solution was extracted with EtOAc (2 x 70 mL) and 10% IPA in CHCI3 (2 x 40 mL). The combined organic layers were dried over MgS04, filtered, and concentrated. The resulting intermediate was redissolved in DCM (60 mL). Triethylamine (2.20 mL, 16.0 mmol, Sigma-Aldrich, St. Louis, MO) and tert-butyl (5-(chlorosulfonyl)-2-pyridinyl)carbamate (1.04 g, 3.60 mmol, Intermediate A) were added. The reaction mixture was stirred at rt for 1 h then partitioned between DCM (70 mL) and water (30 mL). The aqueous layer was extracted with DCM (2 x 40 mL). The combined organic layers were dried over MgS04, filtered, and concentrated. The crude product was purified by column chromatography (120 g of silica, 10% to 40% acetone in hexanes) to afford tert-butyl (5-(((35)-3-(l-propyn-l-yl)-4-(4-(2,2,2-trifiuoro-l-hydroxy- 1 -(hydroxymethyl)ethyl)phenyl)- 1 -piperazinyl)sulfonyl)-2-pyridinyl)carbamate (1.0 g) as a yellow foam.

STEP 6: rEi?r-BUTYL(5-(((35)-3-(l-PROPYN-l-YL)-4-(4-(2-(TRIFLUOROMETHYL)-2-OXIRANYL)PHENYL)- 1 -PIPERAZINYL)SULFONYL)-2-PYRIDINYL)CARBAMATE

To a solution of tert-butyl (5-(((35)-3-(l-propyn-l-yl)-4-(4-(2,2,2-trifiuoro- 1 -hydroxy- 1 -(hydroxymethyl)ethyl)phenyl)- 1 -piperazinyl)sulfonyl)-2-pyridinyl)carbamate (0.300 g, 0.513 mmol) in DCM (5 mL) was added triethylamine (0.400 mL, 2.88 mmol, Sigma-Aldrich, St. Louis, MO) and p-toluenesulfonyl chloride (0.108 g, 0.564 mmol, Sigma-Aldrich, St. Louis, MO). The resulting mixture was heated at reflux (50 °C) under N2 for 2 h. The reaction mixture was cooled to rt and partitioned between sat. NaHCOs (30 mL) and DCM (70 mL). The aqueous layer was extracted with DCM (2 x 40 mL). The combined organic layers were dried over MgS04, filtered, and concentrated. The crude product was purified by column chromatography (40 g of silica, 10 to 40%> acetone in hexanes) to afford tert-butyl (5-(((35)-3-(l-propyn-l-yl)-4-(4-(2-(trifluoromethyl)-2-oxiranyl)phenyl)- 1 -piperazinyl)sulfonyl)-2-pyridinyl)carbamate (0.240 g) as an off-white solid.

1H NMR (400MHz, CDC13) δ ppm 8.66 (dd, J= 0.6, 2.3 Hz, 1 H), 8.20 – 8.10 (m, 1 H), 8.04 (dd, J= 2.2, 8.9 Hz, 1 H), 7.63 (s, 1 H), 7.41 (d, J= 8.6 Hz, 2 H), 6.94 (d, J= 8.8 Hz, 2 H), 4.42 (d, J= 2.2 Hz, 1 H), 3.89 – 3.67 (m, 2 H), 3.38 (d, J = 5.3 Hz, 3 H), 2.97 – 2.83 (m, 2 H), 2.80 – 2.60 (m, 1 H), 1.78 (dd, J= 0.8, 2.0 Hz, 3 H), 1.55 (s, 9 H). m/z (ESI, +ve ion) 567.2 (M+H)+.

ALTERNATIVE ROUTE TO 2-(4-BROMOPHENYL)-3,3,3-TRIFLUORO-l,2-PROPANEDIOL (INTERMEDIATE D STEP 2):

F3

step 1

STEP 1 : 2-(4-BROMOPHENYL)-2-(TRIFLUOROMETHYL)OXIRANE

To a flame-dried, 50-mL, round-bottomed flask was added potassium t-butoxide (0.450 g, 4.01 mmol, Sigma- Aldrich, St. Louis, MO), DMSO (5.0 mL) and trimethylsulfoxonium iodide (1.00 g, 4.54 mmol, Sigma- Aldrich, St. Louis, MO). The resulting mixture was stirred at rt for 40 min. To this reaction mixture was added l-(4-bromophenyl)-2,2,2-trifluoroethanone (1.0 g, 4.0 mmol, Matrix Scientific, Columbia, SC) in DMSO (5.0 mL) dropwise via an addition funnel. The reaction mixture was stirred at rt for 30 min then quenched with water (1 mL) and partitioned between EtOAc (70 mL) and water (30 mL). The organic layer was washed with water (4 x 30 mL), dried over MgS04, filtered, and concentrated. The crude product was purified by column chromatography (40 g of silica, 10 to 20% acetone in hexanes) to afford 2-(4-bromophenyl)-2-(trifluoromethyl)oxirane (0.610 g) as a pale-yellow liquid.

STEP 2: 2-(4-BROMOPHENYL)-3,3,3-TRIFLUORO-l,2-PROPANEDIOL

To a 20-mL vial was added 2-(4-bromophenyl)-2-(trifluoromethyl)oxirane (0.200 g, 0.750 mmol), dioxane (2.0 mL), and water (3.0 mL). The resulting mixture was heated at 85 °C for 24 h. The reaction mixture was cooled to rt and extracted with EtOAc (3 x 50 mL). The combined organic layers were dried over MgS04, filtered and concentrated. The crude product was purified by column chromatography (40 g of silica, 10 to 30% acetone in hexanes) to afford 2-(4-bromophenyl)-3,3,3-trifluoro-l,2-propanediol (2.0 g) as a white solid.

INTERMEDIATE E: BENZYL (3S)-3-(l-PROPYN-l-YL)-l-PIPERAZINECARBOXYLATE

-Cbz

STEP 1 : 4-BENZYL 1 – TER Γ-BUT YL 2-0X0-1,4-PIPERAZINEDICARBOXYLATE

A 2-L Erlenmeyer flask was charged with 2-piperazinone (36.5 g, 364 mmol, Sigma-Aldrich, St. Louis, MO), sodium carbonate (116 g, 1090 mmol, J. T. Baker, Philipsburg, NJ), 600 mL of dioxane, and 150 mL of water. To this was slowly added benzyl chloroformate (62.1 g, 364 mmol, Sigma-Aldrich, St. Louis, MO) at rt over 20 min. After the addition was complete, the mixture was stirred for 2 h and then diluted with water and extracted with EtOAc (2 L). The combined organic extracts were dried (MgS04), filtered, and concentrated to give a white solid. To this solid was added 500 mL of DCM, triethylamine (128 mL, 911 mmol, Sigma-Aldrich, St. Louis, MO), DMAP (4.45 g, 36.4 mmol, Sigma-Aldrich, St. Louis, MO), and di-tert-butyl dicarbonate (119 g, 546 mmol, Sigma-Aldrich, St. Louis, MO). After stirring at room temperature for 1 h, the mixture was diluted with water and the organics were separated. The organics were dried (MgS04), filtered, and concentrated to give a brown oil. To this oil was added 100 mL of DCM followed by 1 L of hexane. The resulting white solid was collected by filtration to give 4-benzyl 1-tert-butyl 2-oxo-l,4-piperazinedicarboxylate (101 g).

STEP 2: BENZYL (2-((7¾’i?J,-BUTOXYCARBONYL)AMINO)ETHYL)(2-OXO-3 -PENT YN- 1 – YL)C ARB AMATE

A 150-mL round-bottomed flask was charged with 4-benzyl 1-tert-butyl 2-oxo- 1 ,4-piperazinedicarboxylate (1.41 g, 4.22 mmol) and THF (5 mL). 1-Propynylmagnesium bromide (0.5 M in THF, 20.0 mL, 10.0 mmol, Sigma-Aldrich, St. Louis, MO) was added at 0 °C slowly. The mixture was stirred at 0 °C for 2 h. Saturated aqueous NH4C1 (40 mL) was added and the aqueous phase was extracted with EtOAc (200 mL, then 2 x 100 mL). The combined organic phases were dried over sodium sulfate, filtered and concentrated under a vacuum. The crude product was purified by column chromatography (50 g of silica, 0 to 50% EtOAc in hexanes) to afford benzyl (2- tert-butoxycarbonyl)amino)ethyl)(2-oxo-3-pentyn-l-yl)carbamate (1.55 g) as a clear oil.

STEP 3: BENZYL 3-(l-PROPYN-l-YL)-l-PIPERAZINECARBOXYLATE

A 3-L round-bottomed flask was charged with 2-((tert-butoxycarbonyl)amino)ethyl)(2-oxo-3-pentyn-l-yl)carbamate (82.17 g, 219 mmol) and 300 mL of DCM. After cooling to -10 °C, TFA (169 mL, 2200

mmol) was added and the resulting dark solution was stirred at rt for 15 min.

Sodium triacetoxyborohydride (186 g, 878 mmol, Sigma- Aldrich, St. Louis, MO) was then added portion- wise over 10 min. After 2 h, the mixture was

concentrated, diluted with EtOAc (1 L), and neutralized with 5 N NaOH. The layers were separated and the organic extracts were washed with brine, dried (MgS04), filtered and concentrated. The resulting orange oil was purified via column chromatography (750 g of silica gel, 0 to 4.5 % MeOH/DCM) to give benzyl 3 -(l-propyn-l-yl)-l -piperazmecarboxylate (43.67 g) as a brown foam.

STEP 4: 4-BENZYL 1 – TER Γ-BUT YL 2-(l -PROP YN-l-YL)- 1,4-PIPERAZINEDICARBOXYLATE

A 20-mL vial was charged with benzyl 3-(l-propyn-l-yl)-l-piperazinecarboxylate (0.616 g, 2.38 mmol), di-tert-butyl dicarbonate (0.979 g, 4.49 mmol, Sigma-Aldrich, St. Louis, MO), DMAP (0.0287 g, 0.235 mmol, Sigma-Aldrich, St. Louis, MO), TEA (0.90 mL, 6.5 mmol) and DCM (8 mL). The mixture was stirred at rt for 30 min. The reaction mixture was partitioned between water (20 mL) and EtOAc (20 mL). The aqueous phase was extracted with EtOAc (20 mL). The organic phase was washed with saturated aqueous sodium chloride (40 mL), dried over sodium sulfate, filtered, and concentrated under a vacuum. The crude product was purified by column chromatography (25 g of silica, 0 to 50% EtOAc in hexanes) to afford 4-benzyl 1-tert-butyl 2-(l-propyn-l-yl)-l,4-piperazinedicarboxylate (0.488 g) as a colorless oil.

STEP 5: 4-BENZYL 1 – TER Γ-BUT YL (2S)-2-( 1 -PROP YN-l-YL)- 1,4-PIPERAZINEDICARBOXYLATE

The individual enantiomers of 4-benzyl 1-tert-butyl 2-(l-propyn-l-yl)-1 ,4-piperazinedicarboxylate were isolated using chiral SFC. The method used was as follows: Chiralpak® ADH column (Daicel Inc., Fort Lee, NJ) (30 x 250 mm, 5 μιη) using 12% ethanol in supercritical C02 (total flow was 170 mL/min).

This separated the two enantiomers with enantiomeric excesses greater than 98%. The first eluting peak was subsequently identified as 4-benzyl 1-tert-butyl (2S)-2-(l-propyn-l-yl)-l,4-piperazinedicarboxylate and used in the next step.

STEP 6: BENZYL (3S)-3-(l-PROPY -l-YL)-l-PIPERAZINECAPvBOXYLATE

A 100-mL round-bottomed flask was charged with 4-benzyl 1-tert-butyl (25)-2-(l-propyn-l-yl)-l,4-piperazinedicarboxylate (0.145 g, 0.405 mmol), TFA (1.0 mL, 13 mmol) and DCM (2 mL). The mixture was stirred at rt for 40 min. The mixture was concentrated and solid NaHC03 was added followed by saturated aqueous NaHC03. The aqueous phase was extracted with EtOAc (2 x 20 mL). The combined organic phases were washed with IN NaOH (40 mL), saturated aqueous NaHC03 (40 mL), water (40 mL) and saturated aqueous sodium chloride (40 mL). The organic phase was dried over sodium sulfate, filtered, and concentrated under a vacuum to afford benzyl (35)-3-(l-propyn-l-yl)-l-piperazinecarboxylate (0.100 g) as a pale yellow clear oil which solidified upon standing to give a pale yellow solid.

1H NMR (400MHz, MeOD) δ ppm 7.47 – 7.13 (m, 5 H), 5.27 – 5.00 (m, 2 H), 3.88 – 3.58 (m, 3 H), 3.48 – 3.33 (m, 2 H), 3.22 – 3.02 (m, 1 H), 2.89 – 2.63 (m, 1 H), 1.80 (s, 3 H). m/z (ESI, +ve ion) 259.1 (M+H)+.

XAMPLE 23: 5-(((3S)-3-(l-PROPYN-l-YL)-4-(4-(l,2,2,2-TETRAFLUORO-1 -(TRIFLUOROMETHYL)ETHYL)PHENYL)- 1 -PIPERAZINYL)SULFONYL)-2-PYRIDIN AMINE

STEP 1 : 2-(4-((2S)-4-BENZYL-2-(l-PROPYN-l-YL)-l-PIPERAZINYL)PHENYL)-1 , 1 ,1 ,3,3,3-HEXAFLUORO-2-PROPANOL

A 20-mL vial was charged with (3S)-l-benzyl-3-(l-propyn-l-yl)piperazine (2.143 g, 10 mmol, Intermediate B), 2-(4-bromophenyl)-1,1,1, 3,3, 3-hexafluoropropan-2-ol (3.09 g, 11.5 mmol, Bioorg. Med. Chem. Lett. 2002, 12, 3009), sodium 2-methylpropan-2-olate (1.92 g, 20.0 mmol, Sigma-Aldrich, St. Louis, MO), dioxane (5 mL), RuPhos palladacycle (0.364 g, 0.500 mmol, Strem Chemical Inc., Newburyport, MA), and RuPhos (0.233 g, 0.500 mmol, Strem Chemical Inc., Newburyport, MA). The vial was sealed and heated at 100 °C for 1 h. The mixture was allowed to cool to rt, and diluted with water and extracted with EtOAc. The combined organic phases were dried over sodium sulfate, filtered and concentrated under a vacuum to give a solid that was purified by silica gel column chromatography (0 to 40% EtOAc in hexanes) to afford 2-(4-((2S)-4-benzyl-2-( 1 -propyn- 1 -yl)- 1 -piperazinyl)phenyl)- 1,1,1,3,3,3-hexafluoro-2-propanol (1.75 g) as a slightly yellow oil.

STEP 2: l,l,l,3,3,3-HEXAFLUORO-2-(4-((2S)-2-(l-PROPYN-l-YL)-l-PIPERAZINYL)PHENYL)-2-PROPANOL

A 250 mL round-bottomed flask was charged with 2-(4-((2S)-4-benzyl-2-( 1 -propyn- 1 -yl)- 1 -piperazinyl)phenyl)- 1,1,1 ,3 ,3 ,3-hexafluoro-2-propanol (1.75 g, 4.35 mmol), potassium carbonate (2.40 g, 17.4 mmol, Sigma-Aldrich, St. Louis, MO), CH2CI2 (25 mL), and 1-chloroethyl chlorocarbonate (1.88 mL, 17.4 mmol, Sigma-Aldrich, St. Louis, MO). After 30 min at rt, the reaction was filtered and the filtrate was concentrated. To the resulting oil was added MeOH (25 mL). This mixture was heated at 75 °C for 1.5 h then concentrated. The residue was triturated with diethyl ether to give l,l,l,3,3,3-hexafluoro-2-(4-((2S)-2-(l-propyn-l-yl)-l-piperazinyl)phenyl)-2-propanol (1.44 g) as a white solid.

STEP 3: TERT-BUTYL (5-(((3S)-3-(l-PROPYN-l-YL)-4-(4-(2,2,2-TRIFLUORO- 1 -HYDROXY- 1 -(TRIFLUOROMETHYL)ETHYL)PHENYL)- 1 -PIPERAZINYL)SULFONYL)-2-PYRIDINYL)CARBAMATE

A 250-mL round-bottomed flask was charged with 1,1,1,3,3,3-hexaf uoro-2-(4-((2S)-2-( 1 -propyn- 1 -yl)- 1 -piperazinyl)phenyl)-2-propanol (18.9 g, 51.6 mmol) and DCM (150 mL) and cooled to 0 °C. TEA was added (14.4 mL, 103 mmol, Sigma-Aldrich, St. Louis, MO) followed by tert-butyl (5- (chlorosulfonyl)pyridin-2-yl)carbamate (15.9 g, 54.2 mmol, Intermediate A) portionwise. After 10 min, the reaction mixture was diluted with water (100 mL) and the organic layer was separated, dried over Na2S04, filtered and concentrated under a vacuum to give a solid that was purified by silica gel column

chromatography (0 to 50% EtO Ac in hexanes) to afford tert-butyl (5 -(((3 S)-3 -( 1 -propyn- 1 -yl)-4-(4-(2,2,2-trifluoro- 1 -hydroxy- 1 -(trifluoromethyl)ethyl)phenyl)- 1 -piperazinyl)sulfonyl)-2-pyridinyl)carbamate (19.9 g) as a tan foam.

STEP 4: 5-(((3S)-3-(l-PROPYN-l-YL)-4-(4-(l,2,2,2-TETRAFLUORO-l- (TRIFLUOROMETHYL)ETHYL)PHENYL)- 1 -PIPERAZINYL)SULFONYL)-2-PYRIDIN AMINE

A 500-mL round-bottomed flask was charged with tert-butyl (5-(((3S)-3-(1 -propyn- 1 -yl)-4-(4-(2,2,2-trifluoro- 1 -hydroxy- 1 – (trifluoromethyl)ethyl)phenyl)-l-piperazinyl)sulfonyl)-2-pyridinyl)carbamate (19.7 g, 31.6 mmol) and DCM (300 mL) and cooled to 0 °C.

(Diethylamino)sulfur trifluoride (4.18 mL, 31.6 mmol, Matrix Scientific, Columbia, SC) was added, and after 10 min, the reaction was diluted with water (250 mL) and DCM (200 mL). The organic layer was separated, dried over

Na2S04, filtered and concentrated under a vacuum. The resultant foam was taken up in DCM (200 mL) and cooled to 0 °C. TFA (100 mL, 1298 mmol) was added and the reaction mixture was warmed to rt for 1.5 h. The reaction was then re-cooled to 0 °C and solid sodium bicarbonate was added slowly until gas evolution ceased. The mixture was diluted with water (250 mL) and DCM (300 mL) and the organic layer was separated, dried over Na2S04, filtered and concentrated under a vacuum to give a solid that was purified by silica gel column chromatography (0 to 100% EtOAc in hexanes) to afford 5-(((3S)-3-(l-propyn- 1 -yl)-4-(4-( 1 ,2,2,2-tetrafluoro- 1 -(trifluoromethyl)ethyl)phenyl)- 1 -piperazinyl)sulfonyl)-2-pyridinamine (11.05 g) as a single enantiomer.

1H NMR (400MHz, CD3OD) δ ppm 8.31 (d, J= 2.2 Hz, 1 H), 7.74 (dd, J= 2.4, 8.9 Hz, 1 H), 7.47 (d, J = 8.8 Hz, 2 H), 7.12 (d, J = 9.0 Hz, 2 H), 6.63 (d, J= 8.8 Hz, 1 H), 4.76-4.70 (m, 1 H), 3.76 (dd, J= 1.9, 11.2 Hz, 2 H), 3.66 – 3.52 (m, 1 H), 3.29 – 3.20 (m, 1 H), 2.79 – 2.72 (m, 1 H), 2.66 – 2.53 (m, 1 H), 1.76 (d, J = 2.2 Hz, 3 H). m/z (ESI, +ve ion) 525.2 (M+H)+. GK-GKRP IC50 (Binding) = 0.187 μΜ.

PAPER

Small Molecule Disruptors of the Glucokinase–Glucokinase Regulatory Protein Interaction: 2. Leveraging Structure-Based Drug Design to Identify Analogues with Improved Pharmacokinetic Profiles

Department of Therapeutic Discovery—Medicinal Chemistry, Department of Therapeutic Discovery—Molecular Structure and Characterization, §Department of Metabolic Disorders, Department of Pharmacokinetics and Drug Metabolism, Department of Pathology, #Department of Pharmaceutics Amgen, Inc., One Amgen Center Drive, Thousand Oaks, California, 91320 and 360 Binney Street, Cambridge, Massachusetts, 02142, United States
J. Med. Chem., 2014, 57 (2), pp 325–338
DOI: 10.1021/jm4016747
Abstract Image

In the previous report, we described the discovery and optimization of novel small molecule disruptors of the GK-GKRP interaction culminating in the identification of 1 (AMG-1694). Although this analogue possessed excellent in vitro potency and was a useful tool compound in initial proof-of-concept experiments, high metabolic turnover limited its advancement. Guided by a combination of metabolite identification and structure-based design, we have successfully discovered a potent and metabolically stable GK-GKRP disruptor (27, AMG-3969). When administered to db/db mice, this compound demonstrated a robust pharmacodynamic response (GK translocation) as well as statistically significant dose-dependent reductions in fed blood glucose levels.

2-(4-((2S)-4-((6-Amino-3-pyridinyl)sulfonyl)-2-(1-propyn-1-yl)-1-piperazinyl)phenyl)-1,1,1,3,3,3-hexafluoro-2-propanol (27)

1H NMR (400 MHz, CDCl3) δ 8.48 (d, J = 2.3 Hz, 1 H), 7.77 (dd, J = 2.5, 8.8 Hz, 1 H), 7.57 (d, J = 8.8 Hz, 2 H), 6.95 (d, J = 9.2 Hz, 2 H), 6.52 (d, J = 8.8 Hz, 1 H), 4.94 (s, 2 H), 4.44 (br s, 1 H), 3.82–3.71 (m, 2 H), 3.58–3.33 (m, 3 H), 2.81 (dd, J = 3.2, 11.1 Hz, 1 H), 2.67 (dt, J = 3.9, 11.0 Hz, 1 H), 1.78 (d, J = 2.2 Hz, 3 H).
m/z (ESI, +ve ion) 523.2 (M + H)+.
REFERENCES
St Jean, D.J. Jr.; Ashton, K.; Andrews, K.; et al.
Small molecule disruptors of the glucokinase-glucokinase regulatory protein (GK-GKRP) interaction
34th Natl Med Chem Symp (May 18-21, Charleston) 2014, Abst 4
Small molecule disruptors of the GK-GKRP interaction as potential antidiabetics
247th Am Chem Soc (ACS) Natl Meet (March 16-20, Dallas) 2014, Abst MEDI 214
Use of non-traditional conformational restriction in the design of a novel, potent, and metabolically stable series of GK-GKRP inhibitors
248th Am Chem Soc (ACS) Natl Meet (August 10-14, San Francisco) 2014, Abst MEDI 267
Small molecule inhibitors for glucokinase-glucokinase regulatory protein (GK-GKRP) binding: Optimization for in vivo target assessment of type II diabetes
248th Am Chem Soc (ACS) Natl Meet (August 10-14, San Francisco) 2014, Abst MEDI 268

MAKING CONNECTIONS Aleksandra Baranczak (right), a fourth-year grad student in Gary A. Sulikowski’s lab at Vanderbilt University, discusses her efforts to synthesize the core of the diazo-containing natural product lomaiviticin A with Kate Ashton, a medicinal chemist at Amgen
Dr. Kate Ashton

Mark Norman

Mark Norman

Michael Bartberger

Michael Bartberger

Chris Fotsch

Chris Fotsch

David St. Jean

David St. Jean

Klaus Michelsen

Klaus Michelsen

///////////1361224-53-4, AMGEN, AMG 3969, Type 2 Diabetes,  PRECLINICAL
O=S(=O)(c1ccc(N)nc1)N2C[C@H](C#CC)N(CC2)c3ccc(cc3)C(O)(C(F)(F)F)C(F)(F)F

BMS-960


Figure imgf000099_0001

str1

BMS-960

PRECLINICAL

(S)-1-((S)-2-Hydroxy-2-(4-(5-(3-phenyl-4-(trifluoromethyl)isoxazol-5-yl)-1,2,4-oxadiazol-3-yl)phenyl)ethyl)piperidine-3-carboxylic Acid

3-Piperidinecarboxylic acid, 1-[(2S)-2-hydroxy-2-[4-[5-[3-phenyl-4-(trifluoromethyl)-5-isoxazolyl]-1,2,4-oxadiazol-3-yl]phenyl]ethyl]-, (3S)-

(S)-1-((S)-2-Hydroxy-2-(4-(5-(3-phenyl-4-(trifluoromethyl)isoxazol-5-yl)-1,2,4-oxadiazol-3-yl)phenyl)ethyl)piperidine-3-carboxylic Acid

CAS 1265321-86-5 FREE FORM

FREE FORM 528.48, C26 H23 F3 N4 O5

CAS 1265323-40-7 HCL SALT

BASIC PATENT WO201117578, 2011, (US Patent 8399451)

Inventors John L. Gilmore, James E. Sheppeck
Applicant Bristol-Myers Squibb Company

Image result for Bristol-Myers Squibb Company

Sphingosine-1-phosphate (S1P) is the endogenous ligand for the sphingosine-1-phophate receptors (S1P1–5) and triggers a number of cellular responses through their stimulation. S1P and its interaction with the S1P receptors play a significant role in a variety of biological processes including vascular stabilization, heart development, lymphocyte homing, and cancer angiogenesis. Agonism of S1P1, especially, has been shown to play an important role in lymphocyte trafficking from the thymus and secondary lymphoid organs, inducing immunosuppression, which has been established as a novel mechanism of treatment for immune diseases and vascular diseases

Sphingosine-1 -phosphate (SlP) has been demonstrated to induce many cellular effects, including those that result in platelet aggregation, cell proliferation, cell morphology, tumor cell invasion, endothelial cell and leukocyte chemotaxis, endothelial cell in vitro angiogenesis, and lymphocyte trafficking. SlP receptors are therefore good targets for a wide variety of therapeutic applications such as tumor growth inhibition, vascular disease, and autoimmune diseases. SlP signals cells in part via a set of G protein-coupled receptors named SlPi or SlPl, SlP2 or S1P2, SlP3 or S1P3, SlP4 Or S1P4, and SlP5 or S1P5 (formerly called EDG-I, EDG-5, EDG-3, EDG-6, and EDG-8, respectively).

SlP is important in the entire human body as it is also a major regulator of the vascular and immune systems. In the vascular system, SlP regulates angiogenesis, vascular stability, and permeability. In the immune system, SlP is recognized as a major regulator of trafficking of T- and B-cells. SlP interaction with its receptor SlPi is needed for the egress of immune cells from the lymphoid organs (such as thymus and lymph nodes) into the lymphatic vessels. Therefore, modulation of SlP receptors was shown to be critical for immunomodulation, and SlP receptor modulators are novel immunosuppressive agents.

The SlPi receptor is expressed in a number of tissues. It is the predominant family member expressed on lymphocytes and plays an important role in lymphocyte trafficking. Downregulation of the SlPi receptor disrupts lymphocyte migration and homing to various tissues. This results in sequestration of the lymphocytes in lymph organs thereby decreasing the number of circulating lymphocytes that are capable of migration to the affected tissues. Thus, development of an SlPi receptor agent that suppresses lymphocyte migration to the target sites associated with autoimmune and aberrant inflammatory processes could be efficacious in a number of autoimmune

Among the five SlP receptors, SlPi has a widespread distribution and is highly abundant on endothelial cells where it works in concert with SIP3 to regulate cell migration, differentiation, and barrier function. Inhibition of lymphocyte recirculation by non-selective SlP receptor modulation produces clinical immunosuppression preventing transplant rejection, but such modulation also results in transient bradycardia. Studies have shown that SlPi activity is significantly correlated with depletion of circulating lymphocytes. In contrast, Sl P3 receptor agonism is not required for efficacy. Instead, SIP3 activity plays a significant role in the observed acute toxicity of nonselective SlP receptor agonists, resulting in the undesirable cardiovascular effects, such as bradycardia and hypertension. (See, e.g., Hale et al, Bioorg. Med. Chem. Lett., 14:3501 (2004); Sanna et al., J. Biol. Chem., 279: 13839 (2004); Anliker et al., J. Biol. Chem., 279:20555 (2004); Mandala et al., J. Pharmacol. Exp. Ther., 309:758 (2004).)

An example of an SlPi agonist is FTY720. This immunosuppressive compound FTY720 (JPI 1080026-A) has been shown to reduce circulating lymphocytes in animals and humans, and to have disease modulating activity in animal models of organ rejection and immune disorders. The use of FTY720 in humans has been effective in reducing the rate of organ rejection in human renal transplantation and increasing the remission rates in relapsing remitting multiple sclerosis (see Brinkman et al., J. Biol. Chem., 277:21453 (2002); Mandala et al., Science, 296:346 (2002); Fujino et al., J.

Pharmacol. Exp. Ther., 305:45658 (2003); Brinkman et al, Am. J. Transplant., 4: 1019 (2004); Webb et al., J. Neuroimmunol, 153: 108 (2004); Morris et al., Eur. J. Immunol, 35:3570 (2005); Chiba, Pharmacology & Therapeutics, 108:308 (2005); Kahan et al., Transplantation, 76: 1079 (2003); and Kappos et al., N. Engl. J. Med., 335: 1124 (2006)). Subsequent to its discovery, it has been established that FTY720 is a prodrug, which is phosphorylated in vivo by sphingosine kinases to a more biologically active agent that has agonist activity at the SlPi, SIP3, SlP4, and SIP5 receptors. It is this activity on the SlP family of receptors that is largely responsible for the pharmacological effects of FTY720 in animals and humans. [0007] Clinical studies have demonstrated that treatment with FTY720 results in bradycardia in the first 24 hours of treatment (Kappos et al, N. Engl. J. Med., 335: 1124 (2006)). The observed bradycardia is commonly thought to be due to agonism at the SIP3 receptor. This conclusion is based on a number of cell based and animal experiments. These include the use of SIP3 knockout animals which, unlike wild type mice, do not demonstrate bradycardia following FTY720 administration and the use of SlPi selective compounds. (Hale et al., Bioorg. Med. Chem. Lett., 14:3501 (2004); Sanna et al., J. Biol. Chem., 279: 13839 (2004); and Koyrakh et al., Am. J. Transplant, 5:529 (2005)).

The following applications have described compounds as SlPi agonists: WO 03/061567 (U.S. Patent Publication No. 2005/0070506), WO 03/062248 (U.S. Patent No. 7,351,725), WO 03/062252 (U.S. Patent No. 7,479,504), WO 03/073986 (U.S. Patent No. 7,309,721), WO 03/105771, WO 05/058848, WO 05/000833, WO 05/082089 (U.S. Patent Publication No. 2007/0203100), WO 06/047195, WO 06/100633, WO 06/115188, WO 06/131336, WO 2007/024922, WO 07/109330, WO 07/116866, WO 08/023783 (U.S. Patent Publication No. 2008/0200535), WO 08/029370, WO 08/114157, WO 08/074820, WO 09/043889, WO 09/057079, and U.S. Patent No. 6,069,143. Also see Hale et al., J. Med. Chem., 47:6662 (2004).

There still remains a need for compounds useful as SlPi agonists and yet having selectivity over Sl P3.

Applicants have found potent compounds that have activity as SlPi agonists. Further, applicants have found compounds that have activity as SlPi agonists and are selective over SIP3. These compounds are provided to be useful as pharmaceuticals with desirable stability, bioavailability, therapeutic index, and toxicity values that are important to their drugability.

SYNTHESIS

Figure

(S)-1-((S)-2-Hydroxy-2-(4-(5-(3-phenyl-4-(trifluoromethyl)isoxazol-5-yl)-1,2,4-oxadiazol-3-yl)phenyl)ethyl)piperidine-3-carboxylic acid, HCl (BMS-960). CAS 1265323-40-7

(S)-1-((S)-2-hydroxy-2-(4-(5-(3-phenyl-4-(trifluoromethyl)isoxazol-5-yl)-1,2,4-oxadiazol-3-yl)phenyl)ethyl)piperidine-3-carboxylic acid, HCl (BMS-960)

1H NMR (400 MHz, DMSO-d6) δ 12.88 (br. s, 1H), 10.5 (br. s, 1H), 8.14 (d, J = 8.6 Hz, 2H), 7.72 (d, J = 8.4 Hz, 2H), 7.69–7.57 (m, 5H), 6.43 (br. s., 1H), 5.37 (d, J = 10.8 Hz, 1H), 3.89–3.60 (m, 2H), 3.50–2.82 (m, 6H), 2.14–1.99 (m, 1H), 1.97–1.75 (m, 1H), 1.63–1.35 (m, 1H);

13C NMR (101 MHz, CDCl3) δ 172.8, 168.5, 164.0, 161.6, 155.4, 156.2, 131.2, 129.0, 128.9, 127.4, 127.2, 125.5, 124.3, 122.2, 111.6, 66.6. 63.0, 52.9, 52.2, 38.8, 25.0, 21.7;

19F NMR (376 MHz, DMSO-d6) δ −54.16;

Anal. calcd for C26H23F3N4O5·HCl: C, 54.71; H, 4.36; N, 9.80. Found: C, 54.76; H, 3.94; N, 9.76;

HRMS (ESI) m/e 529.17040 [(M + H)+, calcd for C26 H24 N4 O5 F3 529.16933].

PATENT

WO 2011017578

Example 14

(S)-l-((S)-2-Hydroxy-2-(4-(5-(3-phenyl-4-(trifluoromethyl)isoxazol-5-yl)-l,2,4- oxadiazol-3-yl)phenyl)ethyl)piperidine-3-carboxylic acid

Figure imgf000099_0001

Preparation 14A: (3S)-Ethyl l-(2-(4-cyanophenyl)-2-hydroxyethyl)piperidine-3- carboxylate

Figure imgf000099_0002

(14A)-isomer A (14A)-isomer B [00210] To a mixture of (S)-ethyl piperidine-3-carboxylate (1.3 g, 8.27 mmol) in toluene (50 mL) was added 4-(2-bromoacetyl)benzonitrile (2.4 g, 10.71 mmol). The reaction mixture was stirred overnight. LCMS indicated completion of reaction. MeOH (10 mL) was added to the mixture, followed by the portionwise addition of sodium borohydride (0.313 g, 8.27 mmol). After 1 hour, LCMS show complete reduction to the desired alcohol. The reaction was quenched with water. The reaction mixture was diluted with ethyl acetate and washed with saturated NaCl. The organic layer was dried with MgSO4, filtered, concentrated, and purified on a silica gel cartridge using an EtOAc/hexanes gradient to yield 2.0 g of solid product. The product was separated by chiral HPLC (Berger SFC MGIII instrument equipped with a CHIRALCEL® OJ (25 x 3 cm, 5 μM). Temp: 30 0C; Flow rate: 130 mL/min; Mobile phase: C(V(MeOH +

0.1%DEA) in 9: 1 ratio isocratic:

[00211] Peak 1 (Isomer A): RT = 2.9 min. for (S)-ethyl l-((S)-2-(4-cyanophenyl)-2- hydroxyethyl)piperidine-3-carboxylate (>99% d.e.). The absolute and relative stereochemistry of compound 14A-isomer A was assigned (S,S) by X-ray crystal structure (see Alternative Route data). 1H NMR (400 MHz, CDCl3) δ ppm 7.63 (2 H, m, J=8.35 Hz), 7.49 (2 H, m, J=8.35 Hz), 4.77 (1 H, dd, J=10.55, 3.52 Hz), 4.17 (2 H, q, J=7.03 Hz), 3.13 (1 H, d, J=9.23 Hz), 2.53-2.67 (3 H, m), 2.44 (2 H, dd, J=18.68, 9.89 Hz), 2.35 (1 H, dd, J=12.74, 10.55 Hz), 1.87-2.01 (1 H, m), 1.71-1.82 (1 H, m), 1.52-1.70 (2 H, m), 1.28 (3 H, t, J=7.03 Hz).

[00212] Peak 2 (Isomer B): RT = 3.8 min for (S)-ethyl l-((R)-2-(4-cyanophenyl)-2- hydroxyethyl)piperidine-3-carboxylate (>99% d.e.). The absolute and relative stereochemistry of 14A-isomer B was assigned (S,R) based on the crystal structure of 14A-isomer A. 1H NMR (400 MHz, CDCl3) δ ppm 7.63 (2 H, m, J=8.35 Hz), 7.49 (2 H, m, J=8.35 Hz), 4.79 (1 H, dd, J=10.55, 3.52 Hz), 4.16 (2 H, q, J=7.03 Hz), 2.69-2.91 (3 H, m), 2.60-2.68 (1 H, m), 2.56 (1 H, dd, J=12.30, 3.52 Hz), 2.36 (1 H, dd, J=12.52, 10.77 Hz), 2.25 (1 H, t, J=8.79 Hz), 1.65-1.90 (3 H, m), 1.52-1.64 (1 H, m, J=12.69, 8.49, 8.49, 4.17 Hz), 1.27 (3 H, t, J=7.25 Hz).

[00213] (S)-Ethyl l-((S)-2-(4-cyanophenyl)-2-hydroxyethyl)piperidine-3-carboxylate (14A-isomer A) was carried forward to make Example 14 and (S)-ethyl l-((R)-2-(4- cyanophenyl)-2-hydroxyethyl)piperidine-3-carboxylate (14A-isomer B) was carried forward to make Example 15.

Preparation 14B: (S)-Ethyl l-((S)-2-hydroxy-2-(4-((Z)-N’-hydroxycarbamimidoyl) phenyl)ethyl)piperidine-3 -carboxylate

Figure imgf000100_0001

[00214] To a mixture of ((S)-ethyl l-((S)-2-hydroxy-2-(4-((Z)-N’- hydroxycarbamimidoyl) phenyl)ethyl)piperidine-3 -carboxylate (14A-Isomer A) (58 mg, 0.192 mmol) and hydroxylamine hydrochloride (26.7 mg, 0.384 mmol) in 2-propanol (10 mL) was added sodium bicarbonate (64.5 mg, 0.767 mmol). The reaction mixture was heated at 85 0C. The reaction mixture was diluted with ethyl acetate and washed with sat NaCl. The organic layer was dried with MgSO4, filtered, and concentrated to yield 56 mg. MS (M+l) = 464. HPLC Peak RT = 1.50 minutes.

Preparation 14C: (S)-Ethyl l-((S)-2-hydroxy-2-(4-(5-(3-phenyl-4-(trifluoromethyl) isoxazol-5-yl)-l,2,4-oxadiazol-3-yl)phenyl)ethyl)piperidine-3-carboxylate

Figure imgf000101_0001

[00215] 3-Phenyl-4-(trifluoromethyl)isoxazole-5-carbonyl fluoride, InM-G (214 mg, 0.78 mmol) was dissolved in acetonitrile (5.00 mL). DIEA (0.272 mL, 1.555 mmol) and (S)-ethyl- 1 -((S)-2-hydroxy-2-(4-((Z)-N’-hydroxycarbamimidoyl) phenyl)ethyl)- piperidine-3-carboxylate (261 mg, 0.778 mmol) were added. The reaction mixture was stirred for 2 hours, then IM TBAF in THF (0.778 mL, 0.778 mmol) was added. The reaction mixture was stirred overnight at room temperature. The reaction mixture was filtered and purified by HPLC in three batches. HPLC conditions: PHENOMENEX® Luna C18 5 micron column (250 x 30mm); 25-100% CH3CN/water (0.1% TFA); 25 minute gradient; 30 mL/min. Isolated fractions with correct mass were partitioned between EtOAc and saturated NaHCO3 with back extracting aqueous layer once. The organic layer was dried with MgSO4, filtered, and concentrated to give 155mg of (S)- ethyl l-((S)-2-hydroxy-2-(4-(5-(3-phenyl-4-(trifluoromethyl)isoxazol-5-yl)-l,2,4- oxadiazol-3-yl)phenyl)ethyl) piperidine-3-carboxylate. 1H NMR (400 MHz, MeOH-d3) δ ppm 8.04 (2 H, d, J=8.13 Hz), 7.55-7.60 (2 H, m), 7.41-7.54 (5 H, m), 4.81 (1 H, ddd, J=8.35, 4.06, 3.84 Hz), 3.96-4.10 (2 H, m), 2.82-3.08 (1 H, m), 2.67-2.82 (1 H, m), 2.36- 2.61 (3 H, m), 2.08-2.33 (2 H, m), 1.73-1.87 (1 H, m, J=8.54, 8.54, 4.45, 4.17 Hz), 1.32- 1.70 (3 H, m), 1.09-1.19 (3 H, m). MS (m+l) = 557. HPLC Peak RT = 3.36 minutes. Purity = 99%.

Example 14: [00216] (S)-Ethyl l-((S)-2-hydroxy-2-(4-(5-(3-phenyl-4-(trifluoromethyl)isoxazol-5- yl)-l,2,4-oxadiazol-3-yl)phenyl)ethyl)piperidine-3-carboxylate (89 mg, 0.16 mmol) was heated at 50 0C in 6N HCl (5 mL) in acetonitrile (5 mL). The reaction mixture was stirred overnight and then filtered and purified by HPLC. HPLC conditions:

PHENOMENEX® Luna C 18 5 micron column (250 x 30mm); 25-100% CH3CN/water (0.1% TFA); 25 minute gradient; 30 mL/min. Isolated fractions with correct mass were freeze-dried overnight to yield 36 mg of (S)-l-((S)-2-hydroxy-2-(4-(5-(3-phenyl-4- (trifluoromethyl)isoxazol-5-yl)-l,2,4-oxadiazol-3-yl)phenyl)ethyl) piperidine-3- carboxylic acid as a TFA salt. 1H NMR (400 MHz, MeOH-d3) δ ppm 8.23 (2 H, d, J=8.35 Hz), 7.65-7.74 (4 H, m), 7.54-7.65 (3 H, m), 5.29 (1 H, t, J=7.03 Hz), 4.00 (1 H, br. s.), 3.43-3.75 (1 H, m), 3.34-3.41 (2 H, m), 2.82-3.24 (2 H, m), 2.26 (1 H, d, J=I 1.86 Hz), 1.84-2.14 (2 H, m), 1.52-1.75 (1 H, m). MS (m+1) = 529. HPLC Peak RT = 3.24 minutes. Purity = 98%. Example 14-Alternate Synthesis Route 1

Preparation 14D (Alternate Synthesis Route 1): (S)-4-(Oxiran-2-yl)benzonitrile

Figure imgf000102_0001

[00217] To 800 mL of 0.2M, pH 6.0 sodium phosphate buffer in a 2 L flask equipped with an overhead stirrer was added D-glucose (38.6 g, 1.2 eq), β-nicotinamide adenine dinucleotide, free acid (1.6 g, mmol), glucose dehydrogenase (36 mg, 3.2 kU,

CODEXIS® GDH- 102, 90 U/mg), and enzyme KRED-NADH-110 (200 mg,

CODEXIS®, 25 U/mg). The vessels containing the reagents above were rinsed with 200 mL of fresh sodium phosphate buffer and added to the reaction which was stirred to dissolution and then heated to 40 0C. To this mixture was added a solution of 2-bromo- 4′-cyanoacetophenone (40 g, 178.5 mmol) in 100 mL DMSO through an addition funnel in about 30 min. The container was rinsed with 20 mL DMSO and the rinse was added to the reactor. A pH of 5.5-6.0 was maintained by adding 1 M NaOH through a fresh addition funnel (total volume of 200 mL over 6h) after which HPLC showed complete consumption of the starting material. The reaction mixture was extracted with 800 mL MTBE x 2 and the combined extracts were washed with 300 mL of 25% brine. The crude alcohol was transferred to a 3L 3-neck flask and treated with solid NaOtBu (34.3 g, 357 mmol) stirring for 1 h and then additional NaOtBu (6.9 g, 357 mmol) and stirring for 30 min. The reaction mixture was filtered and the solution was washed with 300 mL 0.2 M pH 6.0 sodium phosphate buffer, brine, and then the solvent was removed in vacuo and the resulting white solid was dried in a vacuum oven to give (S)-4-(oxiran-2- yl)benzonitrile (23 g, 90% yield, 100% e.e.). 1H NMR (400 MHz, CDCl3) δ ppm 7.62 (2 H, d), 7.35 (2 H, d), 3.88 (1 H, dd), 3.18 (1 H, app t), 2.73 (1 H, dd) Purity = 99%.

[00218] Chiral HPLC was done on a CHIRALP AK® AD-RH 4.6x150mm (Daicel Chemical Industries Ltd.) column using gradient of solvent A (10 mM NH4OAc in water/acetonitrile, 90: 10) and solvent B (10 mM NH4OAc in water/acetonitrile, 10:90) with 70% to 90% in 40 min at a flow rate of 0.5 ml/min at ambient temperature. The detection employed UV at 235 nm. The retention times are as follows:

[00219] Peak 1 (Isomer A): RT = 16.7 min. for (S)-4-(oxiran-2-yl)benzonitrile

[00220] Peak 2 (Isomer B): RT = 14.0 min. for (R)-4-(oxiran-2-yl)benzonitrile Preparation of 14A-isomer A (Alternate Synthesis Route 1): (S)-Ethyl l-((S)-2-(4- cyanophenyl)-2 -hydroxy ethyl)piperidine-3-carboxylate

Figure imgf000103_0001

(14A)-isomer A

[00221] (S)-4-(Oxiran-2-yl)benzonitrile (10.00 g, 68.9 mmol), (S)-ethyl piperidine-3- carboxylate (10.83 g, 68.9 mmol) and iPrOH (100 mL) was charged into a round bottom flask under N2. After heating at 55 0C for 4 hours, 4-dimethylaminopyridine (1.683 g, 13.78 mmol) was then added. The reaction mixture was then heated to 50 0C for an additional 12 hours. At this time HPLC indicated the starting material was completely converted to the desired product. The reaction mixture was then cooled to room temperature. EtOAc (120 ml) was added, followed by 100 ml of water. The organic layer was separated, extracted with EtOAc (2x 100 mL) and concentrated under vacuo to give a crude product. The crude product was recrystallized from EtOH/EtOAc/H2O (3/2/2) (8ml/lg) to give a crystalline off-white solid 14A-alt (15 g, 72% yield, 99.6% e.e.). The absolute and relative stereochemistry was determined by single X-ray crystallography employing a wavelength of 1.54184 A. The crystalline material had an orthorhombic crystal system and unit cell parameters approximately equal to the following:

a = 5.57 A α = 90.0°

b = 9.7l A β = 90.0°

c = 30.04 A γ = 90.0°

Space group: P212121

Molecules/asymmetric unit: 2

Volume/Number of molecules in the unit cell = 1625 A3

Density (calculated) = 1.236 g/cm3

Temperature 298 K.

Preparation 14E (Alternate Route 1): (S)-Ethyl l-((S)-2-(tert-butyldimethylsilyloxy)-2- (4-cyanophenyl)ethyl)piperidine-3-carboxylate

Figure imgf000104_0001

[00222] To a mixture of (S)-ethyl 1 -((S)-2-(4-cyanophenyl)-2-hydroxy ethyl) piperidine-3-carboxylate (17.0 g, 56.2 mmol) and DIPEA (17.68 ml, 101 mmol) in CH2Cl2 (187 mL) was added tert-butyldimethylsilyl trifluoromethanesulfonate (16 ml, 69.6 mmol) slowly. The reaction was monitored with HPLC. The reaction completed in 2 hours. The reaction mixture (a light brown solution) was quenched with water, the aqueous layer was extracted with DCM. The organic phase was combined and dried with Na2SO4. After concentration, the crude material was further purified on a silica gel cartridge (33Og silica, 10-30% EtOAc/hexanes gradient) to afford a purified product (S)- ethyl 1 -((S)-2-(tert-butyldimethylsilyloxy)-2-(4-cyanophenyl)ethyl) piperidine-3 – carboxylate (22.25 g, 53.4 mmol, 95 % yield). 1H NMR (400 MHz, CDCl3) δ ppm 7.61 (2 H, d), 7.45 (2 H, d), 4.79 (1 H, m), 4.15 (2 H, m), 2.88 (1 H, m), 2.75 (1 H, m), 2.60 (1 H, dd), 2.48 (1 H, m), 2.40 (1 H, dd), 2.33 (1 H, tt), 2.12 (1 H, tt), 1.90 (1 H, m), 1.68 (1 H, dt), 1.52 (1 H, m), 1.48 (1 H, m), 1.27 (3 H, t), 0.89 (9 H, s), 0.08 (3 H, s), -0.07 (3 H, s).

Preparation 14F (Alternate Route 1): (S)-Ethyl l-((S)-2-(tert-butyldimethylsilyloxy)-2- (4-((Z)-N’-hydroxycarbamimidoyl)phenyl)ethyl)piperidine-3-carboxylate

Figure imgf000105_0001

[00223] (S)-Ethyl- 1 -((S)-2-(tert-butyldimethylsilyloxy)-2-(4-cyanophenyl)ethyl) piperidine-3-carboxylate (31.0 g, 74.4 mmol) was dissolved in EtOH (248 mL).

Hydroxylamine (50% aq) (6.84 ml, 112 mmol) was added and stirred at room temperature overnight. Then all volatiles were removed with ROTA VAPOR®. The residue was purified with on a silica gel cartridge (33Og silica, 0-50% EtOAc/hexanes gradient) to give (S)-ethyl l-((S)-2-(tert-butyldimethylsilyloxy)-2-(4-((Z)-N’- hydroxycarbamimidoyl)phenyl)ethyl)piperidine-3-carboxylate (31 g, 68.9 mmol, 93 % yield) as a white foam. 1H NMR (400 MHz, CDCl3) δ ppm 8.38 (1 H, br s), 7.58 (2 H, d), 7.37 (2 H, d), 4.88 (2 H, br s), 4.81 (1 H, m), 4.13 (2 H, m), 2.96 (1 H, m), 2.82 (1 H, m), 2.61 (1 H, dd), 2.51 (1 H, m), 2.42 (1 H, dd), 2.32 (1 H, tt), 2.13 (1 H, dt), 1.91 (1 H, m), 1.66 (1 H, dt), 1.58 (1 H, m), 1.48 (1 H, m), 1.27 (3 H, t), 0.89 (9 H, s), 0.08 (3 H, s), -0.09 (3 H, s). Preparation 14G (Alternate Route 1): (S)-Ethyl l-((S)-2-(tert-butyldimethylsilyloxy)-2- (4-(5-(3-phenyl-4-(trifluoromethyl)isoxazol-5-yl)-l,2,4-oxadiazol-3- yl)phenyl)ethyl)piperidine-3-carboxylate

Figure imgf000105_0002

[00224] (S)-Ethyl- 1 -((S)-2-(tert-butyldimethylsilyloxy)-2-(4-((Z)-N’- hydroxycarbamimidoyl)phenyl)ethyl)piperidine-3-carboxylate (32.6g, 72.5 mmol) was dissolved in acetonitrile (145 ml) (anhydrous) and cooled to ~3 0C with ice-bath. 3- phenyl-4-(trifluoromethyl)isoxazole-5-carbonyl chloride (19.98 g, 72.5 mmol) was dissolved in 5OmL anhydrous acetonitrile and added dropwise. The internal temperature was kept below 10 0C during addition. After addition, the reaction mixture was allowed to warm to room temperature. At 30 minutes, HPLC showed completion of the first reaction step. The reaction mixture was re-cooled to below 10 0C. DIEA (18.99 ml, 109 mmol) was added slowly. After the addition, the reaction mixture was heated up to 55 0C for 17 hr s. HPLC/LCMS showed completion of the reaction. The solvents were removed by ROTA VAPOR®. The residue was stirred in 25OmL 20% EtOAc/hexanes and the DIPEA HCl salt precipitated from solution and was removed via filtration. The filtrate was concentrated and purified using a silica gel cartridge (3X33Og silica, 0-50%

EtOAc/hexanes gradient). (S)-ethyl l-((S)-2-(tert-butyldimethylsilyloxy)-2-(4-(5-(3- phenyl-4-(trifluoromethyl)isoxazol-5-yl)-l,2,4-oxadiazol-3-yl)phenyl)ethyl)piperidine-3- carboxylate (43g, 64.1 mmol, 88 % yield) was obtained a light yellow oil. 1H NMR (400 MHz, CDCl3) δ ppm 8.16 (2 H, d), 7.68 (2 H, d), 7.57 (5 H, m), 4.85 (1 H, m), 4.14 (2 H, m), 2.95 (1 H, m), 2.82 (1 H, m), 2.64 (1 H, dd), 2.51 (1 H, m), 2.49 (1 H, dd), 2.35 (1 H, tt), 2.14 (1 H, dt), 1.91 (1 H, m), 1.66 (1 H, dt), 1.57 (1 H, m), 1.48 (1 H, m), 1.27 (3 H, t), 0.92 (9 H, s), 0.11 (3 H, s), -0.05 (3 H, s).

Example 14 (Alternate Route 1): (S)-l-((S)-2-Hydroxy-2-(4-(5-(3-phenyl-4- (trifluoromethyl)isoxazol-5-yl)-l,2,4-oxadiazol-3-yl)phenyl)ethyl)piperidine-3- carboxylic acid

Figure imgf000106_0001

[00225] (S)-Ethyl l-((S)-2-(tert-butyldimethylsilyloxy)-2-(4-(5-(3-phenyl-4- (trifluoromethyl)isoxazol-5-yl)-l,2,4-oxadiazol-3-yl)phenyl)ethyl)piperidine-3- carboxylate (42g, 62.6 mmol) was dissolved in dioxane (150 ml) and treated with 6M HCl (150 ml). The reaction mixture was heated to 65 0C for 6 hours (the reaction was monitored with HPLC, EtOH was distilled out to push the equilibrium forward). Dioxane was removed and the residue was redissolved in ACN/water and lyophilized separately to give crude (S)-l-((S)-2-hydroxy-2-(4-(5-(3-phenyl-4-(trifluoromethyl) isoxazol-5-yl)- l,2,4-oxadiazol-3-yl)phenyl)ethyl)piperidine-3-carboxylic acid, HCl, (37g crude foamy solid). The crude solid (36 g, 63.7 mmol) was suspended in acetonitrile (720 mL) and heated to 60 0C and water (14.4 mL) was added dropwise. A clear solution was obtained, which was cooled to room temperature and concentrated to a viscous oil, treated with ethyl acetate (1.44 L) with vigorously stirring, heated to 60 0C, and cooled to room temperature. (S)-l-((S)-2-hydroxy-2-(4-(5-(3-phenyl-4-(trifluoromethyl)isoxazol-5-yl)- l,2,4-oxadiazol-3-yl)phenyl)ethyl) piperidine-3-carboxylic acid, HCl (28g, 49.3 mmol, 77 % yield) was collected and vacuum dried. Characterization of product by 1H NMR and chiral HPLC matched Example 14 prepared in previous synthesis.

Preparation of Intermediate (14A)-isomer A-Alternate Route 2; 2-Steps: (S)-Ethyl 1- ((S)-2-(4-cyanophenyl)-2-hydroxyethyl)piperidine-3-carboxylate

Figure imgf000107_0001

(14A)-isomer A

Step 1 : Preparation (14D) (Alternate Route 2): (S)-Ethyl l-(2-(4-cyanophenyl)-2- oxoethyl)piperidine-3-carboxylate hydrobromide

Figure imgf000107_0002

(14D)-isomer A

[00226] To a solution of commercially available (S)-ethyl piperidine-3-carboxylate (10 g, 63.6 mmol) in 200 mL toluene was added 4-(2-bromoacetyl)benzonitrile (17g, 76 mmol). The reaction mixture was stirred overnight. The next day, the precipitated solid was collected by filtration and washed with ethyl acetate (x3) and dried under vacuum to give 15.2g of (S)-ethyl l-(2-(4-cyanophenyl)-2-oxoethyl)piperidine-3-carboxylate hydrobromide. MS (M+ 1) = 301. HPLC Peak RT = 1.51 minutes.

Step 2: Preparation of 14 A-isomer A (Alternate Route 2): (S)-Ethyl l-((S)-2-(4- cyanophenyl)-2-hydroxyethyl)piperidine-3 -carboxylate

[00227] Phosphate buffer (1100 mL, BF045, pH 7.0, 0. IM) was added into two liter jacketed glass reactor. The temperature of the reactor was adjusted to 20 0C with the help of a circulator and the reaction mixture was stirred with a magnetic stirrer. Dithiothretol (185.2 mg, 1 mM), magnesium sulfate (288.9 mg, 2 mM), and D-glucose (11.343 g, 62.95 m moles) were added into the reactor. (5*)-Ethyl l-(2-(4-cyanophenyl)-2-oxoethyl) piperidine-3 -carboxylate HBr salt (12 g, 31.47 m moles dissolved in 60 mL DMSO) was added into the reactor slowly with continuous stirring, β-nicotinamide adenine dinucleotide phosphate sodium salt (NADP), 918.47 mg, glucose dehydrogenase, 240 mg (total 18360 U, 76.5 U/mg, ~ 15U/mL, Amano Lot. GDHY1050601) and KRED-114, 1.2 g (CODEXIS® assay 7.8 U/mg of solid), were dissolved in 2.0 mL, 2.0 mL and 10 ml of the same buffer, respectively. Next, NADP, GDH and KRED-114 were added to the reactor in that order. The remaining 26 mL of same buffer was used to wash the NADP, GDH and KRED-114 containers and buffer was added into the same reactor. The starting pH of the reaction was 7.0 which decreased with the progress of the reaction and was maintained at pH 6.5 during the course of the reaction (used pH stat, maintained with IM NaOH). The reaction was run for 4.5 hours and immediately stopped and extracted with ethyl acetate. The ethyl acetate solution was evaporated under reduced pressure and weight of the dark brown residue was 12.14 g. The product was precipitated with dichloromethane and heptane to give 9 g of crude product which was further purified by dissolving it in minimum amount of dichloromethane and re-precipitating by the addition of excess amount of heptane to give 5.22 g. The process was repeated to give an additional 2.82 g of highly pure product for a total of 8.02 g of de > 99.5%.

[00228] Chiral HPLC was done on a CHIRALP AK® AD-RH 4.6x150mm (Daicel Chemical Industries Ltd.) column using gradient of solvent A (10 mM NH4OAc in water/acetonitrile, 90: 10) and solvent B (IO mM NH4OAc in water/acetonitrile, 10:90) with 70% to 90% in 40 min at a flow rate of 0.5 ml/min at ambient temperature. The detection was done by UV at 235 nm. The retention times are as follows: [00229] Peak 1 (14A-isomer A): RT = 20.7 min. for (S)-ethyl l-((S)-2-(4- cyanophenyl)-2-hydroxyethyl)piperidine-3-carboxylate.

[00230] Peak 2 (14B-isomer B): RT = 30.4 min. for (S)-ethyl l-((R)-2-(4- cyanophenyl)-2-hydroxyethyl)piperidine-3-carboxylate.

[00231] Compound 14A-isomer A prepared using this asymmetric method was unambiguously assigned since it was identical to the 14A-isomer A (by 1H NMR and chiral HPLC retention time) that was prepared above and determined by X-ray crystallography. Synthesis of Example 14 from this material followed the same route as described above.

paper

Regioselective Epoxide Ring Opening for the Stereospecific Scale-Up Synthesis of BMS-960, A Potent and Selective Isoxazole-Containing S1P1Receptor Agonist

Discovery Chemistry, Bristol-Myers Squibb, Princeton, New Jersey 08540, United States
Chemical & Synthetic Development, Bristol-Myers Squibb, New Brunswick, New Jersey 08903, United States
Org. Process Res. Dev., Article ASAP
DOI: 10.1021/acs.oprd.6b00366
Abstract Image

This article presents a stereospecific scale-up synthesis of (S)-1-((S)-2-hydroxy-2-(4-(5-(3-phenyl-4-(trifluoromethyl)isoxazol-5-yl)-1,2,4-oxadiazol-3-yl)phenyl)ethyl)piperidine-3-carboxylic acid (BMS-960), a potent and selective isoxazole-containing S1P1 receptor agonist. The process highlights an enzymatic reduction of α-bromoketone toward the preparation of (S)-bromo alcohol, a key precursor of (S)-4-(oxiran-2-yl)benzonitrile. A regioselective and stereospecific epoxide ring-opening reaction was also optimized along with improvements to 1,2,4-oxadiazole formation, hydrolysis, and crystallization. The improved process was utilized to synthesize batches of BMS-960 for Ames testing and other toxicological studies.

PAPER

Journal of Medicinal Chemistry (2016), 59(13), 6248-6264.

Discovery and Structure–Activity Relationship (SAR) of a Series of Ethanolamine-Based Direct-Acting Agonists of Sphingosine-1-phosphate (S1P1)

Abstract

Abstract Image

Sphingosine-1-phosphate (S1P) is a bioactive sphingolipid metabolite that regulates a multitude of physiological processes such as lymphocyte trafficking, cardiac function, vascular development, and inflammation. Because of the ability of S1P1 receptor agonists to suppress lymphocyte egress, they have great potential as therapeutic agents in a variety of autoimmune diseases. In this article, the discovery of selective, direct acting S1P1 agonists utilizing an ethanolamine scaffold containing a terminal carboxylic acid is described. Potent S1P1 agonists such as compounds 18a and 19a which have greater than 1000-fold selectivity over S1P3 are described. These compounds efficiently reduce blood lymphocyte counts in rats through 24 h after single doses of 1 and 0.3 mpk, respectively. Pharmacodynamic properties of both compounds are discussed. Compound 19a was further studied in two preclinical models of disease, exhibiting good efficacy in both the rat adjuvant arthritis model (AA) and the mouse experimental autoimmune encephalomyelitis model (EAE).

BASE

(S)-1-((S)-2-Hydroxy-2-(4-(5-(3-phenyl-4-(trifluoromethyl) isoxazol-5-yl)-1,2,4-oxadiazol-3-yl)phenyl)ethyl)piperidine-3-carboxylic Acid (18a)

(S)-ethyl 1-((S)-2-hydroxy-2-(4-(5-(3-phenyl-4-(trifluoromethyl)isoxazol-5-yl)-1,2,4-oxadiazol-3-yl)phenyl)ethyl)piperidine-3-carboxylate (36%).

1H NMR (400 MHz, MeOH-d3) δ ppm 8.04 (2 H, d, J = 8.13 Hz), 7.55–7.60 (2 H, m), 7.41–7.54 (5 H, m), 4.81 (1 H, ddd, J = 8.35, 4.06, 3.84 Hz), 3.96–4.10 (2 H, m), 2.82–3.08 (1 H, m), 2.67–2.82 (1 H, m), 2.36–2.61 (3 H, m), 2.08–2.33 (2 H, m), 1.73–1.87 (1 H, m, J = 8.54, 8.54, 4.45, 4.17 Hz), 1.32–1.70 (3 H, m), 1.09–1.19 (3 H, m).

MS (M + H)+ at m/z 557. HPLC purity: 99%, tr = 3.36 min (method B).

TFA salt

(S)-1-((S)-2-hydroxy-2-(4-(5-(3-phenyl-4-(trifluoromethyl)isoxazol-5-yl)-1,2,4-oxadiazol-3-yl)phenyl)ethyl)piperidine-3-carboxylic acid, TFA salt (18a, 61%) as a white solid.

1H NMR (400 MHz, MeOH-d3) δ ppm 8.23 (2 H, d, J = 8.35 Hz), 7.65–7.74 (4 H, m), 7.54–7.65 (3 H, m), 5.29 (1 H, t, J = 7.03 Hz), 4.00 (1 H, br s), 3.43–3.75 (1 H, m), 3.34–3.41 (2 H, m), 2.82–3.24 (2 H, m), 2.26 (1 H, d, J = 11.86 Hz), 1.84–2.14 (2 H, m), 1.52–1.75 (1 H, m).

MS (M + H)+ at m/z 529.

HPLC tr = 3.27 min (method B). HPLC purity: 99.4%, tr = 8.78 min (method E); 99.0%, tr = 7.29 min (method F).

HCL SALT

This material was converted to the HCl salt for the following analyses: mp: 219.2 °C. Anal. Calcd for C26H23N4O5F3·HCl: 0.14% water: C, 55.2; H, 4.31; N, 9.87; Cl, 6.25. Found: C, 55.39; H, 4.10; N, 9.88; Cl, 6.34. [α]D20 + 30.47 (c 0.336, MeOH). HPLC with chiral stationary phase (A linear gradient using CO2 (solvent A) and IPA with 0.1% DEA (solvent B); t = 0 min, 30% B, t = 10 min, 55% B was employed on a Chiralcel AD-H 250 mm × 4.6 mm ID, 5 μm column; flow rate was 2.0 mL/min): tr = 5.38 min with >99% ee.

References

Gilmore, J. L.; Sheppeck, J. E.; Watterson, S. H.; Haque, L.; Mukhopadhyay, P.; Tebben, A. J.; Galella, M. A.; Shen, D. R.; Yarde, M.; Cvijic, M. E.; Borowski, V.; Gillooly, K.; Taylor, T.; McIntyre, K. W.; Warrack, B.; Levesque, P. C.; Li, J. P.; Cornelius, G.; D’Arienzo, C.; Marino, A.; Balimane, P.; Salter-Cid, L.; Barrish, J. C.; Pitts, W. J.; Carter, P. H.; Xie, J.; Dyckman, A. J.Discovery and Structure Activity Relationship (SAR) of a Series of Ethanolamine-Based Direct-Acting Agonists of Sphingosine-1-Phosphate (S1P1) J. Med. Chem. 2016, 59, 62486264, DOI: 10.1021/acs.jmedchem.6b00373
Gilmore, J. L.; Sheppeck, J. E. Preparation of 3-(4-(1-hydroxyethyl)phenyl)-1,2,4-oxadiazole derivatives as sphingosine-1-phosphate receptor agonists for the treatment of autoimmune disease and inflammation. PCT Int. Appl. 2011, WO 2011017578.

//////BMS-960, PRECLINICAL, BMS 960

Cl.O=C(O)[C@H]1CCCN(C1)C[C@@H](O)c2ccc(cc2)c3nc(on3)c5onc(c4ccccc4)c5C(F)(F)F

DNDI-VL-2098


str0

DNDI-VL-2098

CAS 681492-17-1

(R)-2-Methyl-6-nitro-2-(4-trifluoromethoxyphenoxymethyl)-2,3-dihydroimidazo[2,1-b]oxazole

Watch this post, will be updated………..

MF C14 H12 F3 N3 O5,
MW 359.26
Imidazo[2,1-b]oxazole, 2,3-dihydro-2-methyl-6-nitro-2-[[4-(trifluoromethoxy)phenoxy]methyl]-, (2R)-
Image result for OTSUKA
Medicinal Chemistry Research Institute, Otsuka Pharmaceutical Co., Ltd., 463-10 Kagasuno, Kawauchi-cho, Tokushima 771-0192, Japan, and Microbiological Research Institute, Otsuka Pharmaceutical Co., Ltd., 463-10 Kagasuno, Kawauchi-cho, Tokushima 771-0192, Japan
Image result for OTSUKA Hidetsugu Tsubouchi
(left to right) Hidetsugu Tsubouchi, Ph.D., Compliance & Ethics Department, manager; Hirofumi Sasaki, Medicinal Chemistry Research Laboratories, associate head and project OPC; Makoto Matsumoto, Ph.D, Pharmaceutical Business Division, senior director; Hiroyuki Hashizume, Pharmaceutical Marketing Headquarters, Product Planning and Management Group, product management manager; Masanori Kawasaki, TB Projects, associate director
Melting Point: 176-178 °C , Condition: Solvent ethyl acetate; isopropanol

(2R)-2-Methyl-6-nitro-2-(4-trifluoromethoxyphenoxymethyl)-2,3-dihydroimidazo[2,1-b]oxazole

Mp: 169–171 °C; Org. Process Res. Dev., Article ASAP, DOI: 10.1021/acs.oprd.6b00331

HPLC (area %): 99.52%; HPLC (chiral): 99.8% (a/a);

1H NMR (400 MHz, CDCl3): δ 7.57 (s, 1H), 7.14–7.16 (d, 2H, J = 10.0 Hz), 6.83–6.86 (d, 2H, J = 7.2 Hz), 4.48–4.50 (d, 1H, J = 10.0 Hz), 4.22–4.24 (d, 1H, J = 10.0 Hz), 4.05–4.10 (t, 2H, J = 9.6 and 10.4 Hz), 1.79 (s, 3H);

13C NMR (100 MHz, CDCl3): δ 156.0, 155.8, 147.1, 143.5, 122.6, 115.5, 112.6, 122.6, 121.7, and 119.1 (JC–F = 255.1 Hz), 116.6, 92.9, 71.8, 51.3, 23.0;

19F NMR (CDCl3, 376 MHz): δ −58.4;

IR (KBr, cm–1): 3155, 2996, 1607, 1456, 1281, 1106, 978, 921, 834,783, 708;

mass (m/z): 360.3 (M + 1)+;

[α]25589 = (+)8.445 (c 1.00 g/100 mL, CHCl3).

Visceral leishmaniasis (VL), infamously known as kala-azar (black fever) in the Indian subcontinent, is the most lethal form of leishmaniasis and is caused by protozoan parasites. This deadly disease is the second largest parasitic killer in the world, surpassed only by malaria, with a worldwide distribution in Asia, East Africa, South America, and the Mediterranean region. In the search for effective treatments for visceral leishmaniasis, the Drugs for Neglected Diseases initiative (DNDi) recently evaluated fexinidazole a nitroimidazole being developed as a treatment for Human African Trypanosomiasis. Fexinidazole  showed potential as a safe and effective oral drug for the treatment of visceral leishmaniasis and is now in clinical trials.

Figure

fexinidazole (1) and DNDI-VL-2098 (2).

Earlier, through an agreement with TB Alliance and in association with the ACSRC at the University of Auckland (NZ), DNDi screened about 70 other nitroimidazole analogues belonging to four chemical subclasses and investigated them for antileishmanial activity

Image result for DNDI-VL-2098

Image result for DNDI-VL-2098

Paper

http://pubs.acs.org/doi/abs/10.1021/acs.jmedchem.5b01699

Repositioning Antitubercular 6-Nitro-2,3-dihydroimidazo[2,1-b][1,3]oxazoles for Neglected Tropical Diseases: Structure–Activity Studies on a Preclinical Candidate for Visceral Leishmaniasis

Auckland Cancer Society Research Centre, School of Medical Sciences, The University of Auckland, Private Bag 92019, Auckland 1142, New Zealand
Faculty of Infectious & Tropical Diseases, London School of Hygiene & Tropical Medicine, Keppel Street, London WC1E 7HT, United Kingdom
§ Laboratory for Microbiology, Parasitology and Hygiene, Faculty of Pharmaceutical, Biomedical and Veterinary Sciences, University of Antwerp, Universiteitsplein 1, B-2610 Antwerp, Belgium
Division of Parasitology, CSIR-Central Drug Research Institute, Lucknow 226031, India
Drugs for Neglected Diseases Initiative, 15 Chemin Louis Dunant, 1202 Geneva, Switzerland
# Institute for Tuberculosis Research, College of Pharmacy, University of Illinois at Chicago, 833 South Wood Street, Chicago, Illinois 60612, United States
Global Alliance for TB Drug Development, 40 Wall Street, New York 10005, United States
J. Med. Chem., 2016, 59 (6), pp 2530–2550
DOI: 10.1021/acs.jmedchem.5b01699
*Phone: (+649) 923-6145. Fax: (+649) 373-7502. E-mail: am.thompson@auckland.ac.nz.

Abstract

Abstract Image

6-Nitro-2,3-dihydroimidazo[2,1-b][1,3]oxazole derivatives were initially studied for tuberculosis within a backup program for the clinical trial agent pretomanid (PA-824). Phenotypic screening of representative examples against kinetoplastid diseases unexpectedly led to the identification of DNDI-VL-2098 as a potential first-in-class drug candidate for visceral leishmaniasis (VL). Additional work was then conducted to delineate its essential structural features, aiming to improve solubility and safety without compromising activity against VL. While the 4-nitroimidazole portion was specifically required, several modifications to the aryloxy side chain were well-tolerated e.g., exchange of the linking oxygen for nitrogen (or piperazine), biaryl extension, and replacement of phenyl rings by pyridine. Several less lipophilic analogues displayed improved aqueous solubility, particularly at low pH, although stability toward liver microsomes was highly variable. Upon evaluation in a mouse model of acute Leishmania donovani infection, one phenylpyridine derivative (37) stood out, providing efficacy surpassing that of the original preclinical lead.

Figure

Structures of various antileishmanial or antitubercular agents.

 str1 str0

CLICK ON IMAGE

2-Methyl-6-nitro-2-{[4-(trifluoromethoxy)phenoxy]methyl}-2,3-dihydroimidazo[2,1- b][1,3]oxazole (7).

Method A (Scheme 1B): Reaction of alcohol 88 with NaH, using procedure C, followed by chromatography of the product on silica gel, eluting with CH2Cl2, gave 71 (87%) as a pale yellow solid: mp (CH2Cl2/hexane) 122-124 C (lit.1 mp 126.8-127.9 C); 1 H NMR (CDCl3)  7.56 (s, 1 H), 7.16 (br d, J = 9.1 Hz, 2 H), 6.85 (br d, J = 9.2 Hz, 2 H), 4.48 (d, J = 10.2 Hz, 1 H), 4.23 (d, J = 10.1 Hz, 1 H), 4.09 (d, J = 10.1 Hz, 1 H), 4.05 (d, J = 10.2 Hz, 1 H), 1.79 (s, 3 H); 13C NMR (CDCl3)  156.3 (C-1’), 156.1 (C-7a), 147.4 (C- 6), 143.9 (q, JC-F = 2.1 Hz, C-4’), 122.8 (2 C, C-3’,5’), 120.7 (q, JC-F = 256.5 Hz, 4’-OCF3), 115.8 (2 C, C-2’,6’), 112.8 (C-5), 93.1 (C-2), 72.2 (2-CH2O), 51.6 (C-3), 23.3 (2-CH3). Anal. (C14H12F3N3O5) C, H, N.

Method B (Scheme 2B): Reaction of 2-bromo-1-[(2-methyloxiran-2-yl)methyl]-4-nitro-1Himidazole2 (98) with 4-(trifluoromethoxy)phenol (0.95 equiv) and NaH (1.2 equiv), using procedure I, followed by chromatography of the product on silica gel, eluting with 2:1 and 3:1 CH2Cl2/petroleum ether (foreruns) and then with 3:1 CH2Cl2/petroleum ether and CH2Cl2, S8 gave a crude product, which was crystallized from CH2Cl2/hexane (and the mother liquors further purified by chromatography on silica gel, eluting as before), to give 71 (55%) as a pale yellow solid (see data above). Method C (Scheme 2D): Reaction of 2-chloro-1-[(2-methyloxiran-2-yl)methyl]-4-nitro-1Himidazole1 (109) with 4-(trifluoromethoxy)phenol (1.0 equiv) and NaH, using procedure I, followed by chromatography of the product on silica gel, eluting with 1:1 and 3:2 CH2Cl2/petroleum ether (foreruns) and then with 3:1 CH2Cl2/petroleum ether and CH2Cl2, gave a crude product, which was crystallized from CH2Cl2/hexane (and the mother liquors further purified by chromatography on silica gel, eluting with 1:1 and 3:1 Et2O/petroleum ether and then with Et2O and CH2Cl2), to give 71 (51%) as a pale yellow solid (see data above).

Synthesis of 9 (Scheme 2A): (2R)-2-Methyl-6-nitro-2-{[4-(trifluoromethoxy)phenoxy]methyl}-2,3-dihydroimidazo- [2,1-b][1,3]oxazole (9). Reaction of 2-chloro-1-{[(2R)-2-methyloxiran-2-yl]methyl}-4-nitro- 1H-imidazole3 (96) with 4-(trifluoromethoxy)phenol and NaH, using procedure H, gave 91,3 (36%) as a pale brown solid: mp 170-171 C (lit.1 mp 176.5-178 C); 1 H NMR (CDCl3)  7.56 (s, 1 H), 7.16 (br d, J = 8.8 Hz, 2 H), 6.85 (br d, J = 9.0 Hz, 2 H), 4.48 (d, J = 10.2 Hz, 1 H), 4.23 (d, J = 10.0 Hz, 1 H), 4.09 (d, J = 10.2 Hz, 1 H), 4.05 (d, J = 10.3 Hz, 1 H), 1.79 (s, 3 H); [α] 25 D 9.0 (c 1.002, CHCl3) [lit.1 [α] 28 D 7.67 (c 1.030, CHCl3)]. Anal. (C14H12F3N3O5) C, H, N. HPLC purity: 100%. Chiral HPLC (using a CHIRALPAK AD-H analytical column and eluting with 15% EtOH/hexane at 1 mL/min) determined that the ee of 9 was 98.7%.

Paper

Sasaki, Hirofumi; Journal of Medicinal Chemistry 2006, VOL 49(26), Pg 7854-7860

Synthesis and Antituberculosis Activity of a Novel Series of Optically Active 6-Nitro-2,3-dihydroimidazo[2,1-b]oxazoles

Medicinal Chemistry Research Institute, Otsuka Pharmaceutical Co., Ltd., 463-10 Kagasuno, Kawauchi-cho, Tokushima 771-0192, Japan, and Microbiological Research Institute, Otsuka Pharmaceutical Co., Ltd., 463-10 Kagasuno, Kawauchi-cho, Tokushima 771-0192, Japan
J. Med. Chem., 2006, 49 (26), pp 7854–7860
DOI: 10.1021/jm060957y

Abstract

Abstract Image

In an effort to develop potent new antituberculosis agents that would be effective against both drug-susceptible and drug-resistant strains of Mycobacterium tuberculosis, we prepared a novel series of optically active 6-nitro-2,3-dihydroimidazo[2,1-b]oxazoles substituted at the 2-position with various phenoxymethyl groups and a methyl group and investigated the in vitro and in vivo activity of these compounds. Several of these derivatives showed potent in vitro and in vivo activity, and compound 19 (OPC-67683) in particular displayed excellent in vitro activity against both drug-susceptible and drug-resistant strains of M. tuberculosis H37Rv (MIC = 0.006 μg/mL) and dose-dependent and significant in vivo efficacy at lower oral doses than rifampicin in mouse models infected with M. tuberculosis Kurono. The synthesis and structure−activity relationships of these new compounds are presented.

(R)-2-Methyl-6-nitro-2-(4-trifluoromethoxyphenoxymethyl)-2,3-dihydroimidazo[2,1-b]oxazole (8). Mp 176−178 °C.

1H NMR (CDCl3) δ 1.79 (3H, s), 4.06 (1H, d, J = 6.8 Hz), 4.10 (1H, d, J = 6.8 Hz), 4.23 (1H, d, J = 10.1 Hz), 4.49 (1H, d, J = 10.1 Hz), 6.84 (2H, d, J = 9.0 Hz), 7.13 (2H, d, J = 9.0 Hz), 7.56 (1H, s).

MS (DI) m/z 359 (M+). Anal. (C14H12F3N3O5) C, H, N.

PAPER

Abstract Image

A process suitable for kilogram-scale synthesis of (2R)-2-methyl-6-nitro-2-{[4-(trifluoromethoxy)phenoxy]methyl}-2,3-dihydroimidazo[2,1-b][1,3]oxazole (DNDI-VL-2098, 2), a preclinical drug candidate for the treatment of visceral leishmaniasis, is described. The four-step synthesis of the target compound involves the Sharpless asymmetric epoxidation of 2-methyl-2-propen-1-ol, 8. Identification of a suitable synthetic route using retrosynthetic analysis and development of a scalable process to access several kilograms of 2 are illustrated. The process was simplified by employing in situ synthesis of some intermediates, reducing safety hazards, and eliminating the need for column chromatography. The improved reactions were carried out on the kilogram scale to produce 2 in good yield, high optical purity, and high quality.

http://pubs.acs.org/doi/abs/10.1021/acs.oprd.6b00331

Development of a Scalable Process for the Synthesis of DNDI-VL-2098: A Potential Preclinical Drug Candidate for the Treatment of Visceral Leishmaniasis

Process Chemistry Division, Advinus Therapeutics Ltd., 21 & 22, Phase II, Peenya Industrial Area, Bangalore 560058, Karnataka, India
Auckland Cancer Society Research Centre, School of Medical Sciences, The University of Auckland, Private Bag 92019, Auckland 1142, New Zealand
Drugs for Neglected Diseases initiative (DNDi), 15 Chemin Louis Dunant, 1202 Geneva, Switzerland
Org. Process Res. Dev., Article ASAP
DOI: 10.1021/acs.oprd.6b00331
*Process Chemistry Division, Advinus Therapeutics Ltd., 21 & 22, Phase II, Peenya Industrial Area, Bangalore -560058, Karnataka, India. E-mail: hari.pati@advinus.com. Tel. No.: (+91)9900212096.
 
Hiroyuki Fujiki, Ph.D, New Drug Research Division, Biology and Translational Research Unit, senior research scientist; Yoshitaka Yamamura, Pharmaceutical Business Division, senior director; Youichi Yabuuchi, Ph.D, Otsuka Pharmaceutical Factory, Inc., corporate adviser; Hidenori Ogawa, Ph.D, Medicinal Chemistry Research Laboratories
/////////////preclinical, DNDI-VL-2098, 681492-17-1, Visceral Leishmaniasis

CEP 33779


img

CEP-33779, CEP33779
CAS 1257704-57-6
Chemical Formula: C24H26N6O2S
Molecular Weight: 462.57
Elemental Analysis: C, 62.32; H, 5.67; N, 18.17; O, 6.92; S, 6.93

N-(3-(4-methylpiperazin-1-yl)phenyl)-8-(4-(methylsulfonyl)phenyl)-[1,2,4]triazolo[1,5-a]pyridin-2-amine

PRECLINICAL Treatment of Rheumatoid Arthritis, Agents for Colorectal Cancer Therapy Systemic Lupus Erythematosus,

Jak2 Inhibitors

Image result for teva logo

Matthew A. Curry, Bruce D. Dorsey, Benjamin J. Dugan, Diane E. Gingrich, Eugen F. Mesaros, Karen L. Milkiewicz,
Applicant Cephalon, Inc.

Worldwide Discovery Research, Cephalon, Inc., 145 Brandywine Parkway, West Chester, Pennsylvania 19380, United States

Image result for Cephalon, Inc.

Matt Curry

 Matthew A. Curry

Bruce Dorsey

Bruce Dorsey

Image result for Cephalon, Inc. Benjamin J. Dugan

Benjamin Dugan

Benjamin J. Dugan received a B.S. degree in Chemistry from the University of Delaware in 1993 under the tutelage of the late Dr. Cynthia McClure. He began his career at FMC Corporation in the agricultural products division. In 2006, he moved to Cephalon, Inc., acquired by Teva Pharmaceutical Industries Ltd. in 2011, and engaged in oncology research focused on small molecule, ATP competitive, kinase inhibitors culminating with the discovery of CEP-33779. He is currently a Research Scientist focused on the development of novel, bioactive small molecules for treatment of central nervous system disorders.

Cephalon Inc.
Malvern, United States

Image result for Cephalon, Inc. Diane E. Gingrich

Members of the Cephalon research team that discovered CEP-5214 and CEP-7055 include (from left) Hudkins, Thelma S. Angeles, Bruce A. Ruggeri, and Diane E. Gingrich. CEPHALON PHOTO

Eugen F. Mesaros

Cephalon Inc.
Malvern, United States
Image result for cephalon Karen L. Milkiewicz

Lupus (systemic lupus erythematosus, SLE) is a chronic autoimmune disease characterized by the presence of activated T and B cells, autoantibodies and chronic inflammation that attacks various parts of the body including the joints, skin, kidneys, CNS, cardiac tissue and blood vessels. In severe cases, antibodies are deposited in the cells (glomeruli) of the kidneys, leading to inflammation and possibly kidney failure, a condition known as lupus nephritis.

Although the cause of lupus remains unknown, manifestations of the disease have been linked to genetic polymorphisms, environmental toxins and pathogens (Morel;

Fairhurst, Wandstrat et al. 2006). In addition, gender, hormonal influences and cytokine dysregulation have been tightly linked to the development of lupus (Aringer and Smolen 2004; Smith-Bouvier, Divekar et al. 2008). Lupus affects nine times as many women as men. It may occur at any age, but appears most often in people between the ages of 10 and 50 years. African Americans and Asians are affected more often than people from other races.

There is no cure for lupus. Current treatments for lupus are aimed at controlling symptoms and are limited to toxic and immunosuppressive agents with severe side-effects such as high dose glucocorticoids and/or hydroxchloroquine. Severe disease (e.g., patients that have signs of renal involvement) require more aggressive drugs including

mycophenolate mofetil (MMF), azathioprine (AZA) and/or cyclophosphamide (CTX) (Bertsias and Boumpas 2008). CTX, AZA and MMF are very toxic and

immunosuppressive, and only 50% of treated patients enter complete remission, with relapse rates up to 30% over a 2-year period.

Memory B cells, and more important, long-lived plasma cells (LL-PCs) which differentiate from memory B cells, are key cell types involved in lupus (Neubert, Meister et al. 2008; Sanz and Lee 2010). Long-lived plasma cells synthesize and secrete large quantities of high-affinity isotype switched antibodies (Meister, Schubert et al. 2007;

Muller, Dieker et al. 2008). Circulating antinuclear antibodies (ANAs) increase the chances of antibody depositing onto self tissues, forming immune-complexes and eventually leading to tissue destruction, epitope spreading and involvement of other organ systems. LL-PCs are commonly found to be chemo- and radio-resistant, over expressing various heat shock proteins and drug pumps (Obeng, Carlson et al. 2006; Neubert, Meister et al. 2008). In addition, LL-PCs primarily reside in the bone marrow where they are protected from current lupus therapies such as cyclophosphamide and glucocorticoids.

A need exists for new treatments for lupus, including lupus nephritis. A need particularly exists for lupus treatments that can target and reduce LL-PCs.

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CEP-33779 is a highly selective, orally active, small-molecule inhibitor of JAK2. CEP-33779 induced regression of established colorectal tumors, reduced angiogenesis, and reduced proliferation of tumor cells. Tumor regression correlated with inhibition of STAT3 and NF-κB (RelA/p65) activation in a CEP-33779 dose-dependent manner. The ability of CEP-33779 to suppress growth of colorectal tumors by inhibiting the IL-6/JAK2/STAT3 signaling suggests a potential therapeutic utility of JAK2 inhibitors in multiple tumors types, particularly those with a strong inflammatory component.

str0

{[8-(4-Methanesulfonyl-phenyl)-[1,2,4]triazolo[1,5-a]pyridin-2-yl]-[3-(4-methyl-piperazin-1-yl)-phenyl]-amine} (1)

LC/MS: (M+H+)+ = 463.2;
1H NMR (DMSO, 400 MHz) δ 9.61 (s, 1H), 8.85 (d, J = 6.8 Hz, 1H), 8.43 (d, J = 6.8 Hz, 2H), 8.06 (d, J = 6.8 Hz, 2H), 7.96 (d, J = 7.5 Hz, 1H), 7.59 (s, 1H), 7.17 (t, J = 6.8 Hz, 1H), 7.11 (t, J = 8.0 Hz, 1H), 7.05 (d, J = 8.6 Hz 1H), 6.49 (d, J = 8.0 Hz, 1H), 3.30 (s, 3H), 3.13 (m, 4H), 2.48 (m, 4H), 2.24 (s, 3H).
CEP-33779 Diglycolate Salt
1H NMR (DMSO, 400 MHz) δ 9.61 (s, 1H), 8.85 (d, J = 6.7 Hz, 1H), 8.43 (d, J = 6.7 Hz, 2H), 8.06 (d, J = 6.7 Hz, 2H), 7.97 (d, J = 7.5 Hz, 1H), 7.59 (s, 1H), 7.18 (d, J = 6.7 Hz, 1H), 7.11 (m, 1H), 7.05 (d, J = 8.6 Hz, 1H), 6.50 (d, J = 8.0 Hz, 1H), 3.89 (s, 4H), 3.30 (s, 3H), 3.13 (m, 4H), 2.48 (m, 4H), 2.24 (s, 3H).
DSC: Endotherm onset at 153.0 °C; Peak at 155.8 °C.

PATENT

WO 2010141796

https://www.google.com/patents/WO2010141796A3?cl=en

Example 35 [8-(4-Methanesulfonyl-phenyl)-[ 1 ,2,4]triazolo[ 1 ,5-a]pyridin-2-yl]-[3-(4-methyl-piperazin-

1 -yl)-phenyl]-amine

Figure imgf000156_0001

35 a) l-(3-Bromo-phenyl)-4-methyl-piperazine was prepared from l-(3-bromo-phenyl)- piperazine (1.33 g, 5.52 mmol) in a manner analogous to Step 32a. The reaction product was isolated as a pale yellow oil (1.4 g, 100%). 1H NMR (400 MHz, CDCl3, δ, ppm): 7.10 (dd, J=8.2, 8.2 Hz, IH), 7.04 (dd, J=2.1, 2.1 Hz, IH), 6.95 (ddd, J=I. S, 1.7, 0.7 Hz, IH), 6.83 (ddd, J=8.3, 2.4, 0.6 Hz, IH), 3.23-3.18 (m, 4H), 2.58-2.54 (m, 4H), 2.35 (s, 3H). MS = 255, 257 (MH)+. 35b) [8-(4-Methanesulfonyl-phenyl)-[ 1 ,2,4]triazolo[ 1 ,5-a]pyridin-2-yl]-[3-(4-methyl- piperazin-l-yl)-phenyl]-amine was prepared from 8-(4-methanesulfonyl-phenyl)- [l,2,4]triazolo[l,5-a]pyridin-2-ylamine (75.0 mg, 0.260 mmol) and l-(3-bromo-phenyl)-4- methyl-piperazine (80.0 mg, 0.314 mmol) with 2,2′-bis-dicyclohexylphosphanyl-biphenyl (30.0 mg, 0.0549 mmol) as the ligand in a manner analogous to Step 2d and was isolated as a yellow solid (0.072 g, 60%).

MP = 232-234 0C.

1H NMR (400 MHz, CDCl3, δ, ppm): 8.49 (d, J=I 2 Hz, IH), 8.25 (d, J=I .5 Hz, 2H), 8.08 (d, J=I .9 Hz, 2H), 7.65 (d, J=I .1 Hz, IH), 7.38 (s, IH), 7.27-7.20 (m, IH), 7.04-6.95 (m, 2H), 6.84 (s, IH), 6.60 (d, J=8.0 Hz, IH), 3.30-3.25 (m, 4H), 3.10 (s, 3H), 2.63-2.58 (m, 4H), 2.38 (s, 3H).

MS = 463 (MH)+.

PATENT

WO 2012078504

PATENT

WO 2012078574

https://google.com/patents/WO2012078574A2?cl=da

COMPOUND A is a JAK2 inhibitor with the chemical name [8-(4-methanesulfonyl-phenyl)-[1,2,4]triazolo[1,5-a]pyridin-2-yl]-[3-(4-methyl-piperazin-1-yl)-phenyl]-amine. COMPOUND A has the following structure:

COMPOUND A

COMPOUND A was prepared in a manner analogous to the five-step method described below (see Example 35 of International Application No. PCT/US10/37363):

Step 1 : To a solution of 1-(3-bromo-phenyl)-piperazine (about 1 g) and acetic acid (about 0.4 mL) in methanol (about 25 mL) is added 37% formaldehyde in water/methanol (about 56.7:37:6.3, water:formaldehyde:methanol; about 5 mL). The mixture is stirred at room temperature for about 18 hours. The suspension is cooled to about 5°C in an ice/water bath and sodium cyanoborohydride (about 5 g) is added in small portions. The mixture is stirred and warmed to room temperature for about 18 hours. The mixture is slowly poured into saturated aqueous ammonium chloride (about 200 mL) and stirred for about 1 hour. The mixture is extracted with dichloromethane (3 x about 75 mL). The combined organic layers are dried over magnesium sulfate, filtered and evaporated. The material is placed under high vacuum for about 18 hours to yield 1-(3-bromo-phenyl)-4-methyl-piperazine as a pale yellow oil (about 1 g). 1H NMR (400 MHz, CDCl3, δ, ppm): 7.10 (dd, J=8.2, 8.2 Hz, 1H), 7.04 (dd, J=2.1, 2.1 Hz, 1H), 6.95 (ddd, J=7.8, 1.7, 0.7 Hz, 1H), 6.83 (ddd, J=8.3, 2.4, 0.6 Hz, 1H), 3.23-3.18 (m, 4H), 2.58-2.54 (m, 4H), 2.35 (s, 3H). MS = 255, 257 (MH)+.

Step 2: To a solution of 3-bromo-pyridin-2-ylamine (about 10 g) in 1,4-dioxane (about 100 mL) is added dropwise ethoxycarbonyl isothiocyanate (about 7 mL). The mixture is stirred under an atmosphere of nitrogen for about 18 hours. The volatiles are evaporated to yield a waxy solid. The recovered material is triturated with hexane (about 250 mL). N-(3-bromo-2-pyridinyl)-N’-carboethoxy-thiourea is isolated and used without further purification. 1H NMR (400 MHz, (D3C)2SO, δ, ppm): 11.46 (s, 1H), 11.43 (s, 1H), 8.49 (dd, J=4.6, 1.5 Hz, 1H), 8.18 (dd, J=8.0, 1.5 Hz, 1H), 7.33 (dd, J=8.0, 4.7 Hz, 1H), 4.23 (q, J=7.1 Hz, 2H), 1.27 (t, J=7.2 Hz, 3H). MS = 215 (MH)+.

Step 3: To a stirred suspension of hydroxylamine hydrochloride (about 17 g) and Ν,Ν-diisopropylethylamine (about 26 mL) in a mixture of methanol (about 70 mL) and

ethanol (about 70 mL) is added N-(3-bromo-2-pyridinyl)-N’-carboethoxy-thiourea. The mixture is stirred for about 2 hours at room temperature then heated to about 60°C for about 18 hours. The suspension is cooled to room temperature, filtered and rinsed with methanol, water then methanol. 8-Bromo-[1,2,4]triazolo[1,5-a]pyridin-2-ylamine is isolated as an off-white solid (about 8 g). 1H NMR (400 MHz, (D3C)2SO, δ, ppm): 8.58 (d, J=6.4 Hz, 1H), 7.73 (d, J=7.6 Hz, 1H), 6.80 (t, J=7.0 Hz, 1H), 6.25 (s, 2H). MS = 213, 215 (MH)+.

Step 4: An oven dried tube is charged with palladium acetate (about 0.2 g) and triphenylphosphine (about 0.6 g). The tube is evacuated under high vacuum and backflushed under a stream of nitrogen for about 5 minutes. A suitable solvent such as

1,4-dioxane (about 10 mL) is added and the mixture is stirred under nitrogen for a suitable time (e.g., for about 10 minutes). 8-Bromo-[1,2,4]triazolo[1,5-a]pyridin-2-ylamine (about 0.75 g), (4-methylsulfonylphenyl)boronic acid (about 1 g), a suitable solvent, such as N,N-dimethylformamide (about 10 mL) and a suitable base, such as about 1.5 M of sodium carbonate in water (about 10 mL) are added. The mixture is stirred for about 2 minutes at room temperature under nitrogen then the tube is sealed and heated at about 80°C for about 18 hours. The mixture is transferred to a round bottom flask and the volatiles are evaporated under reduced pressure. The product is isolated in a suitable manner. For example, water (about 100 mL) may be added and the mixture stirred. The solid may then be collected by filtration, and optionally rinsed with water, air dried, triturated with ether/dichloromethane (about 4: 1; about 10 mL), filtered and rinsed with ether. 8-(4-methanesulfonyl-phenyl)-[1,2,4]triazolo[1,5-a]pyridin-2-ylamine is isolated as a tan solid (about 0.6 g). MP = 236-239 °C. 1H NMR (400 MHz, (D3C)2SO, δ, ppm): 8.63 (d, J=6.3 Hz, 1H), 8.38 (d, J=7.9 Hz, 2H), 8.03 (d, J=7.9 Hz, 2H), 7.84 (d, J= 7.3 Hz, 1H), 7.03 (t, J=7.0 Hz, 1H), 6.21 (br s, 2H), 3.28 (s, 3H). MS = 289 (MH)+.

Step 5: To an oven dried tube is added palladium acetate (about 10 mg) and 2,2′-bis-dicyclohexylphosphanyl-biphenyl (about 30 mg), 8-(4-methanesulfonyl-phenyl)-[1,2,4]triazolo[1,5-a]pyridin-2-ylamine (about 75 mg), 1-(3-bromo-phenyl)-4-methyl-piperazine (about 80 mg), a suitable base, such as cesium carbonate (about 270 mg) and a suitable solvent, such as 1,4-dioxane (about 5 mL). The tube is evacuated and backflushed with nitrogen three times. The tube is sealed and heated at about 80°C for about 72 hours. The mixture is cooled to room temperature and the product isolated in a suitable manner.

For example, the cooled mixture may be diluted with dichloromethane (about 10 mL), filtered through a plug of diatomaceous earth, rinsed with dichloromethane and evaporated. The material may then be purified, e.g., via chromatography, e.g., utilizing an ISCO automated purification apparatus (e.g., amine modified silica gel column 5%→100% ethyl acetate in hexanes). [8-(4-Methanesulfonyl-phenyl)-[1,2,4]triazolo[1,5-a]pyridin-2-yl]-[3-(4-methyl-piperazin-1-yl)-phenyl]-amine (i.e., COMPOUND A) is isolated as a yellow solid (about 0.07 g). MP = 232-234 °C. 1H NMR (400 MHz, CDCl3, δ, ppm): 8.49 (d, J=7.2 Hz, 1H), 8.25 (d, J=7.5 Hz, 2H), 8.08 (d, J=7.9 Hz, 2H), 7.65 (d, J=7.7 Hz, 1H), 7.38 (s, 1H), 7.27-7.20 (m, 1H), 7.04-6.95 (m, 2H), 6.84 (s, 1H), 6.60 (d, J=8.0 Hz, 1H), 3.30-3.25 (m, 4H), 3.10 (s, 3H), 2.63-2.58 (m, 4H), 2.38 (s, 3H). MS = 463 (MH)+.

PATENT

WO 2015089153

https://www.google.com/patents/WO2015089153A1?cl=un

This disclosure relates to a l,2,4 riazolo[l,5a]pyridine derivative, [8-(4 methanesulfonyl-phenyl)-[ 1 ,2,4]triazoio[1 ,5-a]pyridin-2-yl]-[3-(4-methyl-piperazin- 1 -yl phenyl] -amine, re g structure:

or a pharmaceutical salt thereof, and its use in the treatment of multiple sclerosis.

Compound A is a potent, orally active, small molecule inhibitor of JA 2. See, e.g..International Application No. PCT/USlO/37363, U.S. Patent Nos. 8,501,936 and ,633,173, and U.S. Published Patent Application Nos. 2013/0267535 and 2014/0024655, each of which is incorporated by reference herein. Compound A can be prepared, for example, using methods analogous to Example 35 of International Application No.PCT/US 10/37363.

PAPER

A Selective, Orally Bioavailable 1,2,4-Triazolo[1,5-a]pyridine-Based Inhibitor of Janus Kinase 2 for Use in Anticancer Therapy: Discovery of CEP-33779

Worldwide Discovery Research, Cephalon, Inc., 145 Brandywine Parkway, West Chester, Pennsylvania 19380, United States
J. Med. Chem., 2012, 55 (11), pp 5243–5254
DOI: 10.1021/jm300248q
Publication Date (Web): May 10, 2012
Copyright © 2012 American Chemical Society
*Phone: 610-738-6733. Fax: 610-738-6643. E-Mail: bdugan@cephalon.com.

Abstract

Abstract Image

Members of the JAK family of nonreceptor tyrosine kinases play a critical role in the growth and progression of many cancers and in inflammatory diseases. JAK2 has emerged as a leading therapeutic target for oncology, providing a rationale for the development of a selective JAK2 inhibitor. A program to optimize selective JAK2 inhibitors to combat cancer while reducing the risk of immune suppression associated with JAK3 inhibition was undertaken. The structure–activity relationships and biological evaluation of a novel series of compounds based on a 1,2,4-triazolo[1,5-a]pyridine scaffold are reported. Para substitution on the aryl at the C8 position of the core was optimum for JAK2 potency (17). Substitution at the C2 nitrogen position was required for cell potency (21). Interestingly, meta substitution of C2-NH-aryl moiety provided exceptional selectivity for JAK2 over JAK3 (23). These efforts led to the discovery of CEP-33779 (29), a novel, selective, and orally bioavailable inhibitor of JAK2.

[8-(4-Methanesulfonyl-phenyl)-[1,2,4]triazolo[1,5-a]pyridin-2-yl]-[3-(4-methyl-piperazin-1-yl)-phenyl]-amine (29)

 1H NMR (CDCl3) δ 8.49 (dd, J = 6.6, 1.0 Hz, 1H), 8.25 (d, J = 8.4 Hz, 2H), 8.08 (d, J = 8.4 Hz, 2H), 7.66 (dd, J = 7.5, 0.9 Hz, 1H), 7.39–7.36 (m, 1H), 7.23 (t, J = 8.2 Hz, 1H), 7.02 (t, J = 7.1 Hz, 1H), 6.97 (dd, J = 7.8, 1.4 Hz, 1H), 6.88 (s, 1H), 6.60 (dd, J = 8.3, 1.8 Hz, 1H), 3.30–3.25 (m, 4H), 3.10 (s, 3H), 2.63–2.58 (m, 4H), 2.38 (s, 3H).
13C NMR (CDCl3) δ 162.65, 152.28, 148.87, 141.00, 140.91, 140.05, 129.64, 129.29, 128.18, 127.85, 127.76, 124.77, 112.03, 109.40, 108.59, 104.80, 55.19, 49.02, 46.19, 44.59;
mp 208–211 °C.
High resolution mass spectrum (ESI+) m/z 463.1925 [(M + H)+calcd for C24H26N6O2S: 463.1916]. HPLC: 95 A%.

PAPER

An Improved Synthesis of the Free Base and Diglycolate Salt of CEP-33779; A Janus Kinase 2 Inhibitor

Chemical Process Research and Development, Teva Branded Pharmaceutical Products R&D Inc., 383 Phoenixville Pike, Malvern, Pennsylvania 19355, United States
Org. Process Res. Dev., Article ASAP
DOI: 10.1021/acs.oprd.6b00311
Publication Date (Web): November 30, 2016
Copyright © 2016 American Chemical Society

Abstract

Abstract Image

CEP-33779 is a triazole that has been reported to show highly selective inhibition of Janus kinase 2 (JAK2). An efficient process to form CEP-33779 will be presented that uses multiple palladium couplings to provide the drug substance in a convergent manner. The existing medicinal chemistry route was modified to avoid chromatographic purification, improve safety, and utilize palladium ligands which are available in quantities amenable to scale-up. Challenges faced during the development of the new process included optimization of conditions for Buchwald–Hartwig and Suzuki couplings, control of homocoupled impurities and removal of residual palladium. In addition, a screen of conditions to form a diglycolate salt of the parent compound are also presented.

REFERENCES

1: Dugan BJ, Gingrich DE, Mesaros EF, Milkiewicz KL, Curry MA, Zulli AL, Dobrzanski P, Serdikoff C, Jan M, Angeles TS, Albom MS, Mason JL, Aimone LD, Meyer SL, Huang Z, Wells-Knecht KJ, Ator MA, Ruggeri BA, Dorsey BD. A selective, orally bioavailable 1,2,4-triazolo[1,5-a]pyridine-based inhibitor of Janus kinase 2 for use in anticancer therapy: discovery of CEP-33779. J Med Chem. 2012 Jun 14;55(11):5243-54. doi: 10.1021/jm300248q. Epub 2012 May 18. PubMed PMID: 22594690.

2: Tagoe C, Putterman C. JAK2 inhibition in murine systemic lupus erythematosus. Immunotherapy. 2012 Apr;4(4):369-72. doi: 10.2217/imt.12.20. PubMed PMID: 22512630.

3: Seavey MM, Lu LD, Stump KL, Wallace NH, Hockeimer W, O’Kane TM, Ruggeri BA, Dobrzanski P. Therapeutic efficacy of CEP-33779, a novel selective JAK2 inhibitor, in a mouse model of colitis-induced colorectal cancer. Mol Cancer Ther. 2012 Apr;11(4):984-93. doi: 10.1158/1535-7163.MCT-11-0951. Epub 2012 Feb 14. PubMed PMID: 22334590.

4: Lu LD, Stump KL, Wallace NH, Dobrzanski P, Serdikoff C, Gingrich DE, Dugan BJ, Angeles TS, Albom MS, Mason JL, Ator MA, Dorsey BD, Ruggeri BA, Seavey MM. Depletion of autoreactive plasma cells and treatment of lupus nephritis in mice using CEP-33779, a novel, orally active, selective inhibitor of JAK2. J Immunol. 2011 Oct 1;187(7):3840-53. doi: 10.4049/jimmunol.1101228. Epub 2011 Aug 31. PubMed PMID: 21880982.

5: Stump KL, Lu LD, Dobrzanski P, Serdikoff C, Gingrich DE, Dugan BJ, Angeles TS, Albom MS, Ator MA, Dorsey BD, Ruggeri BA, Seavey MM. A highly selective, orally active inhibitor of Janus kinase 2, CEP-33779, ablates disease in two mouse models of rheumatoid arthritis. Arthritis Res Ther. 2011 Apr 21;13(2):R68. doi: 10.1186/ar3329. PubMed PMID: 21510883; PubMed Central PMCID: PMC3132063.

/////////////CEP-33779, CEP33779, CEP 33779, 1257704-57-6, PRECLINICAL, TEVA,  Rheumatoid Arthritis, Colorectal Cancer Therapy, Systemic Lupus Erythematosus,

Jak2 Inhibitors

O=S(C1=CC=C(C2=CC=CN3C2=NC(NC4=CC=CC(N5CCN(C)CC5)=C4)=N3)C=C1)(C)=O

str1 str2

str0

Novel Autotaxin Inhibitors for the Treatment of Osteoarthritis Pain from Lilly Research Laboratories


SCHEMBL15875396.png

str1Figure imgf000023_0002

2-(2-(1H-1,2,3-triazol-5-yl)ethoxy)-1-(2-((2,3-dihydro-1H-inden-2-yl)amino)-5,7-dihydro-6Hpyrrolo[3,4-d]pyrimidin-6-yl)ethan-1-one

l-[2-(2,3-dihydro- lH-inden-2-ylamino)-5,7-dihydro-6H-pyrrolo[3,4- d]pyrimidin-6-yl]-2-[2-(lH- l ,2,3-triazol-4-yl)ethoxy]ethanone.

CAS 1619971-30-0

1-[2-(2,3-dihydro-1H-inden-2-ylamino)-5,7-dihydro-6H-pyrrolo[3,4-d]pyrimidin-6-yl]-2-[2-(1H-1,2,3-triazol-4-yl)ethoxy]ethanone;
Molecular Formula: C21H23N7O2
Molecular Weight: 405.45302 g/mol

US2014200231

Scheme A

Scheme B

Scheme C

VI

Scheme E

Autotaxin is an enzyme reported to be the source of lysophosphatidic acid (LPA) which up-regulates pain-related proteins through one if its cognate receptors, LPAi. LPA is an intracellular lipid mediator which influences a multiplicity of biological and biochemical processes. Targeted inhibition of autotaxin-mediated LPA biosynthesis may provide a novel mechanism to prevent nerve injury-induced neuropathic pain.

Compounds that inhibit autotaxin are desired to offer a potential treatment option for patients in need of treatment for pain.

Pain associated with osteoarthritis (OA) is reported to be the primary symptom leading to lower extremity disability in OA patients. Over 20 million Americans have been diagnosed with OA, the most common of the arthropathies. The currently approved treatments for OA pain may be invasive, lose efficacy with long term use, and may not be appropriate for treating all patients. Additional treatment options for patients suffering from pain associated with OA are desired. Compounds that inhibit autotaxin represent another possible treatment option for patients with pain associated with OA.

U.S. Patent 7,524,852 (‘852) discloses substituted bicyclic pyrimidine derivatives as anti-inflammatory agents.

PCT/US2011/048477 discloses indole compounds as autotoxin inhibitors.

There is a need for novel compounds that provide autotaxin inhibition. The present invention provides novel compounds which are autotaxin inhibitors. The present invention provides certain novel compounds that inhibit the production of LPA.

Autotaxin inhibitor compounds are desired to provide treatments for autotaxin mediated conditions, such as pain and pain associated with OA.

PAPER

Abstract Image

In an effort to develop a novel therapeutic agent aimed at addressing the unmet need of patients with osteoarthritis pain, we set out to develop an inhibitor for autotaxin with excellent potency and physical properties to allow for the clinical investigation of autotaxin-induced nociceptive and neuropathic pain. An initial hit identification campaign led to an aminopyrimidine series with an autotaxin IC50 of 500 nM. X-ray crystallography enabled the optimization to a lead compound that demonstrated favorable potency (IC50 = 2 nM), PK properties, and a robust PK/PD relationship.

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Novel Autotaxin Inhibitors for the Treatment of Osteoarthritis Pain: Lead Optimization via Structure-Based Drug Design

Lilly Research Laboratories, A Division of Eli Lilly and Company, Indianapolis, Indiana 46285, United States
ACS Med. Chem. Lett., 2016, 7 (9), pp 857–861
DOI: 10.1021/acsmedchemlett.6b00207
*E-mail: jonessp@lilly.com. Tel: +1-317-277-5543.

http://pubs.acs.org/doi/abs/10.1021/acsmedchemlett.6b00207

Spencer Jones

Spencer Jones

Senior Research Scientist at Eli Lilly and Company

2-(2-(1H-1,2,3-triazol-5-yl)ethoxy)-1-(2-((2,3-dihydro-1H-inden-2-yl)amino)-5,7-dihydro-6Hpyrrolo[3,4-d]pyrimidin-6-yl)ethan-1-one (9)

………… Purified the resulting residue by silica gel chromatography (gradient elution: 0-9% methanol in ethyl acetate ) to give the title compound……..

1H NMR (400 MHz, CDCl3): 60:40 mixure of rotamers * indicates minor rotamer δ 8.18 (bs, 0.6H), *8.13 (bs, 0.4H), 7.49 (s, 1H), 7.21-7.09 (m, 4 H), 5.70-5.50 (m, 1H), 4.87-4.78 (m, 1H), 4.75 (s, 1.2H), *4.67 (s, 0.8H), 4.64 (s, 1.2H) *4.53 (s, 0.8H), *4.30 (s, 0.8H), 4.28 (s, 1.2H), 3.93 (t, J = 5.6 Hz, 2H), 3.43 (dd, J = 16.2, 7.1 Hz, 2H), 3.10 (t, J = 5.6 Hz, 2H), 2.89 (dd, J = 16.2, 4.9 Hz, 2H).

13C NMR (400 MHz, CDCl3): * indicates minor δ *169.3, 16 169.2, 167.0, *166.8, *162.4, 162.2, 152.8, *152.3, 141.1, 137.8, 130.9, 126.7, 124.9, 115.9, 69.8, 69.3, *69.0, 52.7, *52.5, 51.2, 49.0, *47.9, 40.1, 24.7.

LC/MS (ESI+ ): (m/z) 406 (C21H24N7O2 = (M+1)+ ).

PATENT

WO-2014110000-A1

Example 2

Synthesis of l-[2-(2,3-dihydro- lH-inden-2-ylamino)-5,7-dihydro-6H-pyrrolo[3,4- d]pyrimidin-6-yl]-2-[2-(lH- l ,2,3-triazol-4-yl)ethoxy]ethanone.

Figure imgf000023_0002

Stir a mixture of 2-[2-(lH-triazol-5-yl)ethoxy]acetic acid 2,2,2-trifluoroacetic acid

(20.22 g; 70.90 mmol), N-(2,3-dihydro- lH-inden-2-yl)-6,7-dihydro-5H-pyrrolo[3,4- d]pyrimidin-2-amine dihydrochloride hydrate (27.99 g; 81.54 mmol) and triethylamine (98.83 mL; 709.03 mmol) in dimethylformamide (404.40 mL) at 0°C. Add a solution of 1-propanephosphonic acid cyclic anhydride (50% solution in DMF; 51.89 mL; 81.54 mmol) over 30 minutes, and stir the mixture at room temperature for 18 hours.

Concentrate the reaction mixture under reduced pressure to give a residue. Add water (200 mL) and extract the mixture with ethyl acetate (4 x 250 mL) and

dichloromethane (4 x 250 mL). Wash the combined organic layers with saturated aqueous sodium bicarbonate (2 x 100 mL) and brine (100 mL), then dry over anhydrous sodium sulfate. Filter the mixture and concentrate the solution under reduced pressure to give a red solid (25.70 g) that is slurried in ethyl acetate/methanol (9: 1 mixture; 200 mL) for 2 hours at room temperature. Filter the resulting solid and wash with cold ethyl acetate (50 mL) to give a solid (ca.18.2 g) that is re-slurried in ethyl acetate (200 mL) at reflux for 1 hour. On cooling to room temperature, stir the mixture for 1 hour and filter the resulting light pink solid.

Slurry the light pink solid in water/methanol (1 : 1 mixture; 200 mL) and heat the mixture at 50°C for 30 minutes. Add ammonium hydroxide solution (32% ; 50 mL) and continue to heat the mixture at 50°C for 30 minutes. Upon cooling to room temperature, add additional ammonium hydroxide solution (32% ; 50 mL) and continue stirring for 1 hour at room temperature. Filter the resulting light gray solid, dry and slurry again in ethyl acetate (200 mL) for 1 hour to afford a light gray solid that is filtered, washed with ethyl acetate (25 mL), and dried to give the title compound (12.42 g; 43%) as a gray solid. MS (m/z): 406 (M+l).

PATENT

US-20140200231-A1

https://www.google.com/patents/US20140200231

Scheme E

Figure imgf000014_0001

Preparation 7

Synthesis of 2-[2-(lH-triazol-5-yl)ethoxy]acetic acid.

Figure imgf000018_0001

Pressurize 1 atmosphere of hydrogen (g) to a flask containing [2-(l-benzyl-lH- l,2,3-triazol-5-yl)ethoxy]acetic acid (10.1 g; 1.00 equiv; 38.66 mmoles) and palladium (II) chloride (3 g; 16.92 mmoles; 3.00 g) in isopropyl alcohol (300 mL) and water (60 mL). Maintain the flask under a hydrogen atmosphere for 3 h, then filter through Celite™ and concentrate. Add toluene (2×50 mL) and concentrate to afford the title compound (7.96 g, 100%). ]H NMR (d6-DMSO): 2.86 (t, / = 7 Hz, 2 H), 3.65 (t, / = 7 Hz, 2 H), 3.98 (s, 2 H), 7,77 (s, 1 H), 13.4 – 13.6 (br s, 2 H).

Example 1

Synthesis of l-[2-(2,3-dihydro-lH-inden-2-ylamino)-7,8-dihydropyrido[4,3-d]pyrimidin- 6(5H)-yl]-2-[2-(lH-l,2,3-triazol-4- l)ethoxy]ethanone.

Figure imgf000018_0002

Add N-indan-2-yl-5,6,7,8-tetrahydropyrido[4,3-d]pyrimidin-2-amine (4.2 g, 15.8 mmol) to a mixture of 2-[2-(lH-triazol-5-yl)ethoxy]acetic acid (2.7 g, 15.8 mmol), 1-hydroxybenzotriazole (3.20 g, 23.7 mmol), and dimethylaminopropyl)-3- ethylcarbodiimide hydrochloride (5.44 g, 28.4 mmol) in dichloromethane (40 mL) at 25 °C. Add triethylamine (4.40 mL, 31.6 mmol) to the reaction mixture and stir for 16 h. Wash with water (2 x 50 mL) and concentrate the organic layer. Purify by silica gel column chromatography, eluting with ethyl acetate/methanol, to give the title compound (4.0 g, 60%) as a solid. MS (m/z): 420 (M + Η). Preparation 8

Synthesis of 2-chloro-l-[2-(2,3-dihydro-lH-inden-2-ylamino)-7,8-dihydropyrido[4,3- d]pyrimidin-6(5H)-yl]ethanone.

Figure imgf000019_0001

To N-indan-2-yl-5,6,7,8-tetrahydropyrido[4,3-d]pyrimidin-2-amine (11.0 g, 41.3 mmol) and triethylamine (7.48 mL, 53.7 mmol) in dichloromethane (200 mL), add 2- chloroacetyl chloride (3.61 mL, 5.13 g, 45.4 mmol) dropwise over five minutes at 23 °C. Stir for 30 minutes and pour the reaction mixture into 1 : 1 50% saturated aqueous sodium bicarbonate: dichloromethane (75 mL). Separate the organic layer from the aqueous layer and further extract the aqueous layer with dichloromethane (2 x 25 mL). Combine the organic extracts and dry over anhydrous sodium sulfate, filter, and concentrate. Dissolve the residue in chloroform (10 mL) and purify via silica gel column chromatography (gradient elution: 25% ethyl acetate in hexanes to 100% ethyl acetate) to give the title compound (9.75 g, 69%). ]H NMR (CDC13, * = minor amide rotamer) δ 2.77* (t, 2H), 2.84 (dd, 2H), 2.87 (t, 2H), 3.35 (dd, 2H), 3.76 (t, 2H), 3.85* (t, 2H), 4.12 (s, 2H), 4.52* (s, 2H), 4.57 (s, 2H), 4.72-4.82 (m, IH), 5.48-5.64 (m, IH), 7.12-7.21 (m, 4H), 8.03-8.10 (m, IH).

Preparation 9

Synthesis of 2-(but-3-yn-l-yloxy)-l-[2-(2,3-dihydro-lH-inden-2-ylamino)-7,8- dihydropyrido[4,3-d]p rimidin-6(5H)-yl]ethanone.

Figure imgf000019_0002

To sodium hydride (60 wt% in mineral oil, 1.58 g, 39.6 mmol) in tetrahydrofuran (50 mL) at 23 °C, add 3-butyn-l-ol (7.93 g, 8.59 mL, 113.2 mmol) dropwise, then stir at 23 °C for 20 minutes. Add this solution to 2-chloro-l-[2-(2,3-dihydro-lH-inden-2- ylamino)-7,8-dihydropyrido[4,3-d]pyrimidin-6(5H)-yl]ethanone (9.70 g, 28.3 mmol) in tetrahydrofuran (150 mL) at 23 °C and stir for one hour. Pour the reaction mixture into 50% saturated aqueous sodium bicarbonate solution. Separate the organic layer and further extract the aqueous layer with ethyl ether (x 2) and ethyl acetate (x 2). Combine the organic extracts and wash with brine, then dry over anhydrous sodium sulfate, filter, and concentrate. Purify the resulting crude product by silica gel column chromatography (gradient elution: 20% ethyl acetate in hexanes to 100% ethyl acetate) to give the title compound (8.16 g, 77%). MS (m/z): 377 (M + 1).

Example la

Alternative synthesis of l-[2-(2,3-dihydro- lH-inden-2-ylamino)-7,8-dihydropyrido[4,3- d]pyrimidin-6(5H)-yl]-2-[2-(lH- l,2,3-triazol-4- l)ethoxy]ethanone.

Figure imgf000020_0001

Sparge a solution of 2-(but-3-yn- l-yloxy)-l-[2-(2,3-dihydro-lH-inden-2- ylamino)-7,8-dihydropyrido[4,3-d]pyrimidin-6(5H)-yl]ethanone (8.15 g, 21.7 mmol) and L-ascorbic acid sodium salt (8.58 g, 43.3 mmol) in dimethylformamide (60 mL) and water (60 mL) with nitrogen for ten minutes, then evacuate and backfill with nitrogen three times. Add copper (II) sulfate pentahydrate (1.08 g, 4.33 mmol) and heat to 90 °C, then add azidotrimethylsilane (23.1 mL, 20.0 g, 173 mmol) dropwise and stir for one hour. Cool reaction mixture to 23 °C and pour into water (50 mL). Extract this mixture with ethyl acetate (4 x 50 mL). Combine the organic extracts and wash with saturated aqueous sodium chloride, dry over anhydrous sodium sulfate, filter, and concentrate.

Purify the resulting crude product by silica gel column chromatography (gradient elution: 0 to 10% methanol in ethyl acetate) to give the title compound (3.60 g, 40%). MS (m/z): 420 (M + 1). Preparation 10

Synthesis of tert-butyl-2-(2,3-dihydro-lH-inden-2-ylamino)-5,7-dihydro-6H-pyrrolo[3,4- d]pyrimidine-6-carboxylate.

Figure imgf000021_0001

Charge 450 rriL (2.58 mol) of N-ethyl-N-isopropylpropan-2-amine into a 15 °C solution of tert-butyl 2-chloro-5,7-dihydro-6H-pyrrolo[3,4-d]pyrimidine-6-carboxylate (220 g, 860.37 mmol) and 2,3-dihydro-lH-inden-2-amine (137.7 g, 1.03 mol) in 1- methylpyrrolidin-2-one (3.6 L). Heat the resulting mixture to 80 °C for 16 h, then cool to 30 °C and transfer the resulting mixture into 5 L of water at 25 °C. Filter the resulting solid and rinse the filter cake with water (2 x 300 rriL). Reslurry the solid in ethyl acetate (350 iriL) for 45 min at 15 °C. Filter the slurry, rinsing with 15 °C ethyl acetate ( 2 x 250 rriL), and dry to give the title compound (226 g, 75%) as an off-white solid. ‘H NMR (d6-DMSO) 1.45 (s, 9 H), 2.87 (dd, /= 7.2, 15.8 Hz, 2 H), 3.24 (dd, /= 7.2, 15.8 Hz, 2 H), 4.36 (d, 10.4 Hz, 2 H), 4.44 (d, /= 12.8 Hz, 2 H), 4.60 (m, 1 H), 7.14 (m, 2 H), 7.20 (m, 2 H), 7.55 (d, /= 6.8 Hz, 1 H), 8.27 (d, /= 7.2 Hz, 1 H).

Preparation 11

Synthesis of N-(2,3-dihydro-lH-inden-2-yl)-6,7-dihydro-5H-pyrrolo[3,4-d]pyrimidin-2- amine dihydrochloride hydrate.

Figure imgf000021_0002

Charge 670 rriL of 5 M hydrochloric acid (3.35 mol) to a solution of tert-butyl 2-

(2,3-dihydro-lH-inden-2-ylamino)-5,7-dihydro-6H pyrrolo[3,4-d]pyrimidine-6- carboxylate (226 g, 641.25 mmol) in tetrahydrofuran (2.0 L) at 17 °C, maintaining the internal temperature below 26 °C during the addition. Heat the resulting solution to 50 °C for 16 h, cool to 25 °C and dilute with 500 rriL of water and 500 mL of tert- butylmethylether. Separate the resulting layers and extract with tert-butylmethylether (3 x 1 L). Concentrate the water phase down to a reaction volume of ca. 200 mL, and filter the resulting slurry. Rinse the cake with tert-butylmethylether (2 x 200 mL) and dry to give the title product (177 g, 80%) as a light brown solid. MS (m/z): 253.2 (M-2HC1- H20+1).

Preparation 12

Syntheis of tert-butyl 2-but-3-ynox acetate.

Figure imgf000022_0001

Stir a mixture of but-3-yn-l-ol (6.00 g; 85.60 mmol), tetrabutylammonium sulfate (2.07 g; 8.54 mmol) and sodium hydroxide (40% wt/wt; 150 mL) in dichloromethane (150 mL) at 0°C. Add tert-butyl bromoacetate (19.34 mL; 128.40 mmol) dropwise and stir the mixture for 2.5 hours at room temperature. Dilute the reaction mixture with dichloromethane (200 mL) and water (100 mL), separate the layers, and further extract the aqueous layer with dichloromethane (2 x 100 mL). Wash the combined organic layers with brine (100 mL), dry over anhydrous sodium sulfate, and concentrate to afford the crude title compound as a brown oil (11.93 g). Purify the oil by silica gel column chromatography, eluting with hexane: ethyl acetate (0% to 10% mixtures) to give the title compound (11.35 g; 72%) as a colorless oil. ]H NMR (CDCI3) δ 1.48 (s, 9H), 2.00 (m, 1H), 2.52 (m, 2H), 3.67 (m, 2H), 4.01 (bs, 2H).

Preparation 13

Synthesis of tert-butyl 2-[2-(lH-triazol-5- l)ethoxy]acetate.

Figure imgf000022_0002

Stir tert-Butyl 2-but-3-ynoxyacetate (11.34 g; 61.55 mmol) and copper(I)iodide (584 mg; 3.07 mmol) in a mixture of dimethylformamide (56.70 mL) and methanol (11.34 mL) at 0°C. Add azido(trimethyl)silane (12.33 mL; 86.47 mmol) dropwise and heat the mixture at 90°C for 18 hours.

In a second batch, stir tert-butyl 2-but-3-ynoxyacetate (4.38 g; 23.77 mmol) and copper(I)iodide (226 mg; 1.19 mmol) in a mixture of dimethylformamide (22 mL) and methanol (6 mL) at 0°C. Add azido(trimethyl)silane (4.8 mL; 33.66 mmol) dropwise and the mixture heated at 90°C for 18 hours.

Upon cooling to room temperature, combine the crude products from both batches and concentrate the mixture to afford a greenish residue. Purify the crude product by filtration through a plug of silica eluting with dichloromethane: ethyl acetate (75% to 100% mixtures) to afford the title compound (14.15 g, 73%) as a colorless oil. MS (m/z): 228.15 (M+l).

Preparation 14

Synthesis of 2-[2-(lH-triazol-5-yl)ethoxy]acetic acid 2,2,2-trifluoroacetic acid.

Figure imgf000023_0001

Stir a mixture of ieri-butyl 2-[2-(lH-triazol-5-yl)ethoxy]acetate (14.15 g; 62.26 mmol) and trifluoroacetic acid (70.75 mL, 935.69 mmol) in dichloromethane (70.75 mL) for 2 hours at room temperature. Concentrate the reaction mixture under reduced pressure to provide the title compound containing additional trifluoroacetic acid (20.22 g, >100%) as a brown solid. MS (m/z): 172.05 (M+l).

Example 2

Synthesis of l-[2-(2,3-dihydro- lH-inden-2-ylamino)-5,7-dihydro-6H-pyrrolo[3,4- d]pyrimidin-6-yl]-2-[2-(lH- l ,2,3-triazol-4-yl)ethoxy]ethanone.

Figure imgf000023_0002

Stir a mixture of 2-[2-(lH-triazol-5-yl)ethoxy]acetic acid 2,2,2-trifluoroacetic acid

(20.22 g; 70.90 mmol), N-(2,3-dihydro- lH-inden-2-yl)-6,7-dihydro-5H-pyrrolo[3,4- d]pyrimidin-2-amine dihydrochloride hydrate (27.99 g; 81.54 mmol) and triethylamine (98.83 mL; 709.03 mmol) in dimethylformamide (404.40 mL) at 0°C. Add a solution of 1-propanephosphonic acid cyclic anhydride (50% solution in DMF; 51.89 mL; 81.54 mmol) over 30 minutes, and stir the mixture at room temperature for 18 hours.

Concentrate the reaction mixture under reduced pressure to give a residue. Add water (200 mL) and extract the mixture with ethyl acetate (4 x 250 mL) and

dichloromethane (4 x 250 mL). Wash the combined organic layers with saturated aqueous sodium bicarbonate (2 x 100 mL) and brine (100 mL), then dry over anhydrous sodium sulfate. Filter the mixture and concentrate the solution under reduced pressure to give a red solid (25.70 g) that is slurried in ethyl acetate/methanol (9: 1 mixture; 200 mL) for 2 hours at room temperature. Filter the resulting solid and wash with cold ethyl acetate (50 mL) to give a solid (ca.18.2 g) that is re-slurried in ethyl acetate (200 mL) at reflux for 1 hour. On cooling to room temperature, stir the mixture for 1 hour and filter the resulting light pink solid.

Slurry the light pink solid in water/methanol (1 : 1 mixture; 200 mL) and heat the mixture at 50°C for 30 minutes. Add ammonium hydroxide solution (32% ; 50 mL) and continue to heat the mixture at 50°C for 30 minutes. Upon cooling to room temperature, add additional ammonium hydroxide solution (32% ; 50 mL) and continue stirring for 1 hour at room temperature. Filter the resulting light gray solid, dry and slurry again in ethyl acetate (200 mL) for 1 hour to afford a light gray solid that is filtered, washed with ethyl acetate (25 mL), and dried to give the title compound (12.42 g; 43%) as a gray solid. MS (m/z): 406 (M+l).

Preparation 15

Synthesis of 2-chloro- l-[2-(2,3-dihydro- lH-inden-2-ylamino)-5,7-dihydro-6H- pyrrolo[3,4-d]pyrimidin-6-yl]ethanone.

Figure imgf000024_0001

Stir a suspension of N-(2,3-dihydro-lH-inden-2-yl)-6,7-dihydro-5H-pyrrolo[3,4- d]pyrimidin-2-amine dihydrochloride hydrate (14.4 g, 41.9 mmol) and triethylamine (14.3 g, 19.7 mL, 141.4 mmol) in dichloromethane (200 mL) at 23 °C for 10 minutes, then cool to -30 °C. Add 2-chloroacetyl chloride (5.49 g, 3.86 mL, 48.6 mmol) over two minutes and warm to 23 °C over 10 minutes. Add methanol (5 mL) and remove the solvent in vacuo. Slurry the crude reaction mixture in methanol (30 mL), add 50 g silica gel and remove solvent in vacuo. Load the resulting residue onto a loading column and purify via silica gel column chromatography (gradient elution: 50% ethyl acetate in hexanes to ethyl acetate to 10% methanol in ethyl acetate) to give the title compound (11.5 g, 84%). MS (m/z): 329(M+1).

Preparation 16

Synthesis of 2-(but-3-yn-l-yloxy)-l-[2-(2,3-dihydro-lH-inden-2-ylamino)-5,7-dihydro- 6H-pyrrolo[3,4-d]pyrimidin-6-yl]ethanone.

Figure imgf000025_0001

To sodium hydride (60 wt% in mineral oil, 2.06 g, 51.4 mmol) in tetrahydrofuran (86 mL) at 0 °C, add 3-butyn-l-ol (4.64 g, 5.03 mL, 64.3 mmol), then stir at 23 °C for 15 minutes. Add this solution to 2-chloro-l-[2-(2,3-dihydro-lH-inden-2-ylamino)-5,7- dihydro-6H-pyrrolo[3,4-d]pyrimidin-6-yl]ethanone (8.45 g, 25.7 mmol) in

tetrahydrofuran (86 mL) at 0 °C and stir for five minutes. Pour reaction mixture into 50% saturated aqueous sodium bicarbonate solution. Separate the organic layer and further extract the aqueous layer with ethyl ether and ethyl acetate (2 x 50 mL each). Combine the organic extracts and wash with brine, then dry over anhydrous sodium sulfate, filter, and concentrate. Combine the crude product with the crude product from a second reaction (run reaction under identical conditions and stoichiometry employing 2-chloro- 1- [2-(indan-2-ylamino)-5,7-dihydropyrrolo[3,4-d]pyrimidin-6-yl]ethanone (3.0 g, 9.1 mmol)) and purify by silica gel column chromatography (gradient elution: 25% ethyl acetate in hexanes to 100% ethyl acetate) to give the title compound (2.90 g, 23%). MS

(m/z): 363(M+1). Example 2a

Alternative synthesis of l-[2-(2,3-dihydro-lH-inden-2-ylamino)-5,7-dihydro- pyrrolo[3,4-d]pyrimidin-6-yl]-2-[2-(lH-l,2,3-triazol-4-yl)ethoxy]ethanone.

Figure imgf000026_0001

Add dimethylformamide (27 mL) and water (27 mL) to a flask containing 2-(but- 3-yn-l-yloxy)-l-[2-(2,3-dihydro-lH-inden-2-ylamino)-5,7-dihydro-6H-pyrrolo[3,4- d]pyrimidin-6-yl]ethanone (2.90 g, 8.00 mmol). Add copper (II) sulfate pentahydrate (400 mg, 1.60 mmol) and L-ascorbic acid sodium salt (3.17 g, 16.0 mmol). Evacuate flask and backfill with nitrogen (x 2), then add azidotrimethylsilane (7.37 g, 8.53 mL, 64.0 mmol) and heat the reaction to 90 °C for 70 minutes. Cool the reaction mixture to 23 °C and remove all solvent in vacuo. Suspend the residue in methanol/dichloromethane and then add silica gel and remove solvent in vacuo. Load this material onto a loading column and purify via silica gel column chromatography (gradient elution: 0-9% methanol in ethyl acetate) to give the title compound (980 mg, 30%). MS (m/z):

406(M+1).

/////////Autotaxin LPA osteoarthritis tool molecule, lily, Spencer Jones, PRECLINICAL

N1(Cc2cnc(nc2C1)NC3Cc4ccccc4C3)C(=O)COCCc5cnnn5

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