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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 29 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 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 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 29 year tenure till date Aug 2016, Around 30 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, 25 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 13 lakh plus views on New Drug Approvals Blog in 212 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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PF 06821497


str1

PF 06821497

Cas 1844849-11-1

Designed to treat lymphoma

1(2H)-Isoquinolinone, 5,8-dichloro-2-[(1,2-dihydro-4-methoxy-6-methyl-2-oxo-3-pyridinyl)methyl]-3,4-dihydro-7-[(S)-methoxy-3-oxetanylmethyl]-

MF C22 H24 Cl2 N2 O5, 

MW 467.34

ChemSpider 2D Image | 5,8-Dichloro-2-[(4-methoxy-6-methyl-2-oxo-1,2-dihydro-3-pyridinyl)methyl]-7-[methoxy(3-oxetanyl)methyl]-3,4-dihydro-1(2H)-isoquinolinone | C22H24Cl2N2O5PF 06821497

5,8-Dichloro-2-[(4-methoxy-6-methyl-2-oxo-1,2-dihydro-3-pyridinyl)methyl]-7-[methoxy(3-oxetanyl)methyl]-3,4-dihydro-1(2H)-isoquinolinone

1(2H)-Isoquinolinone, 5,8-dichloro-2-[(1,2-dihydro-4-methoxy-6-methyl-2-oxo-3-pyridinyl)methyl]-3,4-dihydro-7-(methoxy-3-oxetanylmethyl)-

  • Molecular Formula C22H24Cl2N2O5
  • Average mass 467.342 Da

SCHEMBL17330377.pngPF 06821497

5,8-dichloro-2-[(4-methoxy-6-methyl-2-oxo-1H-pyridin-3-yl)methyl]-7-[(S)-methoxy(oxetan-3-yl)methyl]-3,4-dihydroisoquinolin-1-one

US2015361067

Inventors Michael Raymond Collins, Robert Steven Kania, Robert Arnold Kumpf, Pei-Pei Kung, Daniel Tyler Richter, Scott Channing Sutton, Martin James Wythes
Original Assignee Pfizer Inc.Image result
  • Epigenetic alterations play an important role in the regulation of cellular processes, including cell proliferation, cell differentiation and cell survival. The epigenetic silencing of tumor suppressor genes and activation of oncogenes may occur through alteration of CpG island methylation patterns, histone modification, and dysregulation of DNA binding protein. Polycomb genes are a set of epigenetic effectors. EZH2 (enhancer of zeste homolog 2) is the catalytic component of the Polycomb Repressor Complex 2 (PRC2), a conserved multi-subunit complex that represses gene transcription by methylating lysine 27 on Histone H3 (H3K27). EZH2 plans a key role in regulating gene expression patterns that regulate cell fate decisions, such as differentiation and self-renewal. EZH2 is overexpressed in certain cancer cells, where it has been linked to cell proliferation, cell invasion, chemoresistance and metastasis.
  • High EZH2 expression has been correlated with poor prognosis, high grade, and high stage in several cancer types, including breast, colorectal, endometrial, gastric, liver, kidney, lung, melanoma, ovarian, pancreatic, prostate, and bladder cancers. See Crea et al., Crit. Rev. Oncol. Hematol. 2012, 83:184-193, and references cited therein; see also Kleer et al., Proc. Natl. Acad. Sci. USA 2003, 100:11606-11; Mimori et al., Eur. J. Surg. Oncol. 2005, 31:376-80; Bachmann et al., J. Clin. Oncol. 2006, 24:268-273; Matsukawa et al., Cancer Sci. 2006, 97:484-491; Sasaki et al. Lab. Invest. 2008, 88:873-882; Sudo et al., Br. J. Cancer 2005, 92(9):1754-1758; Breuer et al., Neoplasia 2004, 6:736-43; Lu et al., Cancer Res. 2007, 67:1757-1768; Ougolkov et al., Clin. Cancer Res. 2008, 14:6790-6796; Varambally et al., Nature 2002, 419:624-629; Wagener et al., Int. J. Cancer 2008, 123:1545-1550; and Weikert et al., Int. J. Mol. Med. 2005, 16:349-353.
    Recurring somatic mutations in EZH2 have been identified in diffuse large B-cell lymphoma (DLBCL) and follicular lymphomas (FL). Mutations altering EZH2 tyrosine 641 (e.g., Y641C, Y641F, Y641N, Y641S, and Y641H) were reportedly observed in up to 22% of germinal center B-cell DLBCL and 7% of FL. Morin et al. Nat. Genetics 2010 February; 42(2):181-185. Mutations of alanine 677 (A677) and alanine 687 (A687) have also been reported. McCabe et al., Proc. Natl. Acad. Sci. USA 2012, 109:2989-2994; Majer et al. FEBS Letters 2012, 586:3448-3451. EZH2 activating mutations have been suggested to alter substrate specificity resulting in elevated levels of trimethylated H3K27 (H3K27me3).
    Accordingly, compounds that inhibit the activity of wild type and/or mutant forms of EZH2 may be of interest for the treatment of cancer.

SYNTHESIS

Steps

1 COUPLING, Ag2CO3

2 Alkylation, K2CO3

3 LiAlH4 REDUCTION

4 THIONYL CHLORIDE

5 N-Alkylation of Amides, t-BuOK

6 A GRIGNARD REACTION

7 AN ALKYLATION , METHYL IODIDE, t-BuOK

8 HYDROGENATION, DE BENZYLATION,  PLATINUM OXIDE

9 LAST STEP separation by chiral preparative, SFC on (R,R) Whelk O1 column, TO GET PF 06821497

PATENT

US 20150361067

///////////////PF 06821497, 1844849-11-1, PFIZER, lymphoma, Pei-Pei Kung,  @pfizer, #ACSSanFran, Michael Raymond Collins, Robert Steven Kania, Robert Arnold Kumpf, Pei-Pei Kung, Daniel Tyler Richter, Scott Channing Sutton, Martin James Wythes

Next up in #MEDI 1st time disclosures Pei-Pei Kung from @pfizer presenting a molecule designed to treat lymphoma #ACSSanFran

str0

CO[C@H](c2cc(Cl)c3CCN(CC1=C(OC)C=C(C)NC1=O)C(=O)c3c2Cl)C4COC4

CC1=CC(=C(C(=O)N1)CN2CCC3=C(C=C(C(=C3C2=O)Cl)C(C4COC4)OC)Cl)OC
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ABBV 2222


str1

ABBV 2222

Benzoic acid, 4-[(2R,4R)-4-[[[1-(2,2-difluoro-1,3-benzodioxol-5-yl)cyclopropyl]carbonyl]amino]-7-(difluoromethoxy)-3,4-dihydro-2H-1-benzopyran-2-yl]-

4-[(2R,4R)-4-({[1-(2,2-difluoro-1,3-benzodioxol-5-yl)cyclopropyl]carbonyl}- amino)-7-(difluoromethoxy)-3,4-dihydro-2H-chromen-2-yl]benzoic acid

CAS  1918143-53-9

MF C28 H21 F4 N O7
MW 559.46
1H NMR (400 MHz, CDCl.sub.3) .delta. 8.17-8.03 (m, 2H), 7.49 (d, J=8.2 Hz, 2H), 7.16-6.99 (m, 4H), 6.73-6.67 (m, 2H), 6.38 (d, J=73.6 Hz, 1H), 5.48 (td, J=10.4, 6.1 Hz, 1H), 5.36 (d, J=8.8 Hz, 1H), 5.31-5.21 (m, 1H), 2.52 (ddd, J=13.3, 6.0, 2.2 Hz, 1H), 1.86-1.71 (m, 2H), 1.68-1.60 (m, 1H), 1.10 (q, J=3.7, 2.4 Hz, 2H);
 
MS (ESI-) m/z=558 (M-H).sup.-.

Image result

DESCRIPTION

Cystic fibrosis (CF), one of the most common autosomal recessive genetic diseases in the Caucasian population, is caused by loss of function mutations in the Cystic Fibrosis Transmembrane Conductance Regulator (CFTR) gene, which is located on chromosome 7 (http://www.cff.org/AboutCF/; Rowe S. M et al. (2005); N Eng J Med. (352), 1992-2001). Approximately 1:3500 and 1:3000 infants born in the United States and in Europe, respectively, are affected by CF, resulting in ˜75,000 cases worldwide, ˜30,000 of which are in the United State. Approximately 1,000 new cases of CF are diagnosed each year, with more than 75% of patients being diagnosed by 2 years of age. Nearly half the CF population is currently 18 years of age and older. The CFTR protein (Gregory, R. J. et al. (1990) Nature 347:382-386; Rich, D. P. et al. (1990) Nature 347:358-362; Riordan, J. R. et al. (1989) Science 245:1066-1073) is a cAMP/ATP-mediated ion channel expressed in a variety of cell types, including secretory and absorptive epithelial cells. CFTR regulates chloride and bicarbonate anion flux across the cell membrane, maintaining electro neutrality and osmolarity across the epithelial membrane (Quinton, P. M. (1990), FASEB J. 4: 2709-2727). CFTR is also responsible for regulating the activity of other ion channels and proteins (Guggino, W. B. et al. (2006), Nat Revs Molecular Cell Biology 7, 426-436).

Aberrations in CFTR function result in imbalance of the airway surface liquid, leading to mucus dehydration, inflammation, recurrent bacterial infection and irreversible lung damage, which lead to premature death in affected patients. Besides respiratory disease, CF patients suffer from gastrointestinal problems and pancreatic insufficiency. The majority of males (95%) with cystic fibrosis are infertile as a result of azoospermia caused by altered vas deferens; which may be absent, atrophic, or fibrotic. Fertility is also decreased among females with cystic fibrosis due to abnormal cervical mucus.

The F508del mutation, the most common of the approximately 1900 identified polymorphisms in CFTR, results in defective processing of CFTR in the endoplasmic reticulum (ER) (http://www.cftr2.org/index.php). Approximately 90% of the CF patients carry at least one copy of the F508del mutation (deletion of a phenylalanine on position 508), and 50%-60% of the patients are homozygous for this mutation. The defective processing of CFTR results in early CFTR degradation, which leads to reduced trafficking or absence of the protein on the membrane. As there have been over 100 CF disease-causing mutations identified, they have been classified according to their phenotypic consequences and belong to synthesis, maturation, regulation, conductance, reduced number due to quantity and reduced number due to stability classifications.

Current CF drug discovery efforts focus upon developing two classes of compounds to modulate CFTR. One class, called Correctors, helps to overcome the folding defects of the mutated CFTR protein to promote its maturation resulting in higher cell surface expression. The other classes of compounds, called Potentiators, help overcome the defective regulation and/or conductance of the protein by increasing the probability of channel opening on the membrane surface.

In addition, as the modulation of CFTR protein mutations to promote proper protein folding is beneficial for CF, there are other diseases mediated by CFTR. For example, Sjögren’s Syndrome (SS), an autoimmune disorder that results in symptoms of xerostomia (dry mouth) and keratoconjunctivitis sicca (KCS, dry eyes) may result from dysregulation of moisture producing glands throughout the body. Chronic obstructive lung disease (COLD), or chronic obstructive airway disease (COAD), which is a progressive and irreversible airflow limitation in the airways is result of several physiologic abnormalities, including mucus hyper secretion and impaired mucociliary secretion. Increasing the anion secretion by CFTR potentiators have been suggested to overcome these phenotypic complexities with Sjögren’s Syndrome by increasing the corneal hydration and by overcoming the impaired mucociliary secretion in COAD (Bhowmik A, et al. (2009) Vol. 103(4), 496-502; Sloane P, et al. PLOS One (2012) Vol 7(6), 239809 (1-13)).

STEP 1

(R)-methyl 4-(7-hydroxy-4-oxochroman-2-yl)benzoate

RXN……….By reacting  7-hydroxy-4H-chromen-4-one AND  (4-(methoxycarbonyl)phenyl)boronic acid

STEP 2

(R)-methyl 4-(7-hydroxy-4-(methoxyimino)chroman-2-yl)benzoate

Reacting ABOVE compd  and O-methylhydroxylamine,

STEP 3

Methyl 4-((2R,4R)-4-amino-7-hydroxychroman-2-yl)benzoate

reacting ABOVE  compd with 5% platinum (0.05 equivalent) on carbon in acetic acid. The reaction was stirred at room temperature under hydrogen

THEN STEP 4

Methyl 4-((2R,4R)-4-amino-7-hydroxychroman-2-yl)benzoate isolated AS  trifluroroacetic acid salt

STEP 5
methyl 4-((2R,4R)-4-(1-(2,2-difluorobenzo[d][1,3]dioxol-5-yl)cyclopropanec- arboxamido)-7-hydroxychroman-2-yl)benzoate

by reacting  1-(2,2-difluorobenzo[d][1,3]dioxol-5-yl)cyclopropanecarboxylic acid  and HATU (1-[bis(dimethylamino)methylene]-1H-1,2,3-triazolo[4,5-b]pyridinium 3-oxid hexafluorophosphate, the ABOVE compound AND  N-ethyl-N-isopropylpropan-2-amine

STEP 6

Methyl 4-((2R,4R)-4-(1-(2,2-difluorobenzo[d][1,3]dioxol-5-yl)cyclopropanec- arboxamido)-7-(difluoromethoxy)chroman-2-yl)benzoate

by reacting ABOVE compound  and diethyl(bromodifluoromethyl)phosphonate

AND FINAL STEP7  is ESTER HYDROLYSIS USING lithium hydroxide to get ABBV 2222

PATENT
US 20160120841

str1

Example 122

4-[(2R,4R)-4-({[1-(2,2-difluoro-1,3-benzodioxol-5-yl)cyclopropyl]carbonyl}- amino)-7-(difluoromethoxy)-3,4-dihydro-2H-chromen-2-yl]benzoic acid

[1880] To Example 123E (130 mg, 0.227 mmol) in methanol (2 mL) and water (0.5 mL) was added lithium hydroxide (32.6 mg, 1.360 mmol). The mixture was stirred at 35.degree. C. for 4 hours, LC/MS showed the conversion was complete. Solvent was removed under reduced pressure and water (2 mL) was added. The pH of the mixture was adjusted to pH 1-2 with the addition of 2 M HCl. The precipitated white solid was collected by filtration, and dried to provide the title compound (110 mg, 0.197 mmol, 87% yield). .sup.1H NMR (400 MHz, CDCl.sub.3) .delta. 8.17-8.03 (m, 2H), 7.49 (d, J=8.2 Hz, 2H), 7.16-6.99 (m, 4H), 6.73-6.67 (m, 2H), 6.38 (d, J=73.6 Hz, 1H), 5.48 (td, J=10.4, 6.1 Hz, 1H), 5.36 (d, J=8.8 Hz, 1H), 5.31-5.21 (m, 1H), 2.52 (ddd, J=13.3, 6.0, 2.2 Hz, 1H), 1.86-1.71 (m, 2H), 1.68-1.60 (m, 1H), 1.10 (q, J=3.7, 2.4 Hz, 2H); MS (ESI-) m/z=558 (M-H).sup.-.

Example 123

methyl 4-[(2R,4R)-4-({[1-(2,2-difluoro-1,3-benzodioxol-5-yl)cyclopropyl]ca- rbonyl}amino)-7-(difluoromethoxy)-3,4-dihydro-2H-chromen-2-yl]benzoate

Example 123A

(R)-methyl 4-(7-hydroxy-4-oxochroman-2-yl)benzoate

[1881] A mixture of bis(2,2,2-trifluoroacetoxy)palladium (271 mg, 0.816 mmol), (S)-4-(tert-butyl)-2-(pyridin-2-yl)-4,5-dihydrooxazole (200 mg, 0.979 mmol), ammonium hexafluorophosphate(V) (798 mg, 4.90 mmol), (4-(methoxycarbonyl)phenyl)boronic acid (2203 mg, 12.24 mmol) and dichloroethane (8 mL) in a 20 mL vial was stirred for 5 minutes at room temperature, followed by the addition of 7-hydroxy-4H-chromen-4-one (CAS 59887-89-7, MFCD00209371, 1323 mg, 8.16 mmol) and water (256 mg, 14.19 mmol). The vial was capped and the mixture was stirred at 60.degree. C. overnight. The reaction gradually turned black, with Pd plated out on the sides of the vial. The mixture was filtered through a plug of celite and eluted with ethyl acetate to give a red solution which was washed with brine. The solvent was removed in vacuo and the crude material was chromatographed using a 100 g silica gel cartridge and eluted with a gradient of 5-40% ethyl acetate in heptane to provide the title compound (1.62 g, 66.6% yield). .sup.1H NMR (400 MHz, CDCl.sub.3) .delta. 8.15-8.04 (m, 2H), 7.87 (d, J=8.7 Hz, 1H), 7.60-7.49 (m, 2H), 6.62-6.45 (m, 2H), 5.87 (s, 1H), 5.53 (dd, J=12.8, 3.2 Hz, 1H), 3.94 (s, 3H), 3.07-2.80 (m, 2H); MS (ESI+) m/z=299 (M+H).sup.+.

Example 123B

(R)-methyl 4-(7-hydroxy-4-(methoxyimino)chroman-2-yl)benzoate

[1882] The mixture of Example 123A (960 mg, 3.22 mmol), sodium acetate (528 mg, 6.44 mmol) and O-methylhydroxylamine, hydrochloric acid (538 mg, 6.44 mmol) in methanol (10 mL) was stirred at 60.degree. C. overnight. Solvent was removed under reduced pressure. The residue was dissolved in ethyl acetate and washed with water. The organic layers was dried over MgSO.sub.4, filtered, and concentrated. The residue was washed with ether to provide the title compound (810 mg, 2.475 mmol, 77% yield). .sup.1H NMR (400 MHz, CDCl.sub.3) .delta. 8.15-8.03 (m, 2H), 7.81 (d, J=8.7 Hz, 1H), 7.58-7.43 (m, 2H), 6.50 (dd, J=8.6, 2.5 Hz, 1H), 6.45 (d, J=2.5 Hz, 1H), 5.21 (d, J=3.0 Hz, 1H), 5.12 (dd, J=12.2, 3.2 Hz, 1H), 3.95 (s, 3H), 3.93 (s, 3H), 3.45 (dd, J=17.2, 3.2 Hz, 1H), 2.63 (dd, J=17.2, 12.2 Hz, 1H); MS (ESI+) m/z 328 (M+H).sup.+.

Example 123C

Methyl 4-((2R,4R)-4-amino-7-hydroxychroman-2-yl)benzoate

[1883] A mixture of Example 123B (570 mg, 1.741 mmol) was treated with 5% platinum (0.05 equivalent) on carbon in acetic acid (5 mL). The reaction was stirred at room temperature under hydrogen (1 atmosphere) for 24 hours, LC/MS showed conversion over 95%. The mixture was filtered through a celite pad and solvent removed under reduced pressure. The residue was purified by preparative LC method TFA2 to provide the trifluroroacetic acid salt of the title compound (300 mg, 44% yield). LC/MS m/z 283 (M-NH.sub.2).sup.+.

Example 123D

methyl 4-((2R,4R)-4-(1-(2,2-difluorobenzo[d][1,3]dioxol-5-yl)cyclopropanec- arboxamido)-7-hydroxychroman-2-yl)benzoate

[1884] A mixture of 1-(2,2-difluorobenzo[d][1,3]dioxol-5-yl)cyclopropanecarboxylic acid (162 mg, 0.668 mmol) and HATU (1-[bis(dimethylamino)methylene]-1H-1,2,3-triazolo[4,5-b]pyridinium 3-oxid hexafluorophosphate, 380 mg, 1.0 mmol) in DMF (2 mL) was stirred for 5 minutes at room temperature, followed by the addition of Example 123C (200 mg, 0.334 mmol) and N-ethyl-N-isopropylpropan-2-amine (0.466 ml, 2.67 mmol). The mixture was stirred at room temperature for 2 hours, LC/MS showed reaction complete. The mixture was loaded on to a 25 g silica gel cartridge eluting with 5-50% ethyl acetate in heptane provide the title compound (204 mg, 58.3% yield). .sup.1H NMR (400 MHz, CDCl.sub.3) .delta. 8.11-7.90 (m, 2H), 7.42 (d, J=8.0 Hz, 2H), 7.16-7.02 (m, 2H), 6.94 (dd, J=37.7, 8.3 Hz, 2H), 6.49-6.32 (m, 2H), 5.67 (s, 1H), 5.36 (dt, J=15.3, 8.7 Hz, 2H), 5.18 (d, J=10.7 Hz, 1H), 3.93 (s, 3H), 2.56-2.36 (m, 1H), 1.80-1.70 (m, 2H), 1.26 (d, J=2.2 Hz, 1H), 1.10-1.04 (m, 2H); MS (ESI-) m/z=521.9 (M-H).sup.-.

Example 123E

Methyl 4-((2R,4R)-4-(1-(2,2-difluorobenzo[d][1,3]dioxol-5-yl)cyclopropanec- arboxamido)-7-(difluoromethoxy)chroman-2-yl)benzoate

[1885] To Example 123D (190 mg, 0.363 mmol) and diethyl(bromodifluoromethyl)phosphonate (0.129 ml, 0.726 mmol) in a mixture of acetonitrile (2 mL) and water (1 mL) was added 50% aqueous potassium hydroxide (244 mg, 2.178 mmol) drop wise via syringe while stirring vigorously. After the addition was completed, LC/MS showed conversion was complete with a small by-product peak. Additional water was added to the mixture and the mixture was extracted with ethyl acetate (3.times.20 mL). The combined organic extracts were washed with 1 M HCl (5 mL) and water, dried over MgSO.sub.4, filtered, and concentrated. The residue was purified by preparative LC method TFA2 to provide the title compound (150 mg, 72% yield). .sup.1H NMR (400 MHz, CDCl.sub.3) .delta. 8.09-8.00 (m, 2H), 7.49-7.41 (m, 2H), 7.15-6.99 (m, 4H), 6.75-6.66 (m, 2H), 5.50-5.40 (m, 1H), 5.33 (d, J=8.9 Hz, 1H), 5.25 (dd, J=11.3, 2.0 Hz, 1H), 3.93 (s, 3H), 2.50 (ddd, J=13.4, 6.1, 2.1 Hz, 1H), 1.84-1.71 (m, 2H), 1.65 (d, J=2.8 Hz, 1H), 1.11-1.06 (m, 2H); MS (ESI-) m/z=572 (M-H).sup.-.

REFERENCE

Next up is Xueqing Wang of @abbvie speaking about a collaboration with @GalapagosNV on a different cystic fibrosis treatment

str0

///////////ABBV 2222

O=C(O)c1ccc(cc1)[C@@H]3Oc2cc(OC(F)F)ccc2C(C3)NC(=O)C4(CC4)c5ccc6OC(F)(F)Oc6c5

BMS 986158


SCHEMBL16861831.png

str1

BMS 986158

MF C30H33N5O2, MW495.627 g/mol

CAS 1800340-40-2

5H-Pyrido[3,2-b]indole-7-methanol, 3-(1,4-dimethyl-1H-1,2,3-triazol-5-yl)-α,α-dimethyl-5-[(S)-phenyl(tetrahydro-2H-pyran-4-yl)methyl]-

MOA:Bromodomain and extraterminal domain protein inhibitor

Indication:Solid tumoursStatus:

Phase II :Bristol-Myers Squibb (Originator)

Phase I/IISolid tumours

  • Originator Bristol-Myers Squibb
  • Class Antineoplastics; Small molecules
  • Mechanism of Action Bromodomain and extraterminal domain protein inhibitors
  • 01 Jun 2015 Phase-I/II clinical trials for Solid tumours (Late-stage disease, Metastatic disease) in Canada (NCT02419417)
  • 02 Apr 2015 Bristol-Myers Squibb plans a phase I/IIa trial for Solid tumours (Late-stage disease) in USA, Australia and Canada (NCT02419417)

The genomes of eukaryotic organisms are highly organized within the nucleus of the cell. The long strands of duplex DNA are wrapped around an octomer of histone proteins to form a nucleosome. This basic unit is then further compressed by the aggregation and folding of nucleosomes to form a highly condensed chromatin structure. A range of different states of condensation are possible, and the tightness of this structure varies during the cell cycle, being most compact during the process of cell division. There has been appreciation recently that chromatin templates form a fundamentally important set of gene control mechanisms referred to as epigenetic regulation. By conferring a wide range of specific chemical modifications to histones and DNA (such as acetylation, methylation, phosphorylation, ubiquitinylation and SUMOylation) epigenetic regulators modulate the structure, function and accessibility of our genome, thereby exerting a huge impact in gene expression.

Histone acetylation is most usually associated with the activation of gene transcription, as the modification loosens the interaction of the DNA and the histone octomer by changing the electrostatics. In addition to this physical change, specific proteins bind to acetylated lysine residues within histones to read the epigenetic code. Bromodomains are small (-110 amino acid) distinct domains within proteins that bind to acetylated lysine residues commonly but not exclusively in the context of histones. There is a family of around 50 proteins known to contain bromodomains, and they have a range of functions within the cell. The BET family of bromodomain containing proteins

comprises 4 proteins (BRD2, BRD3, BRD4 and BRD-T) which contain tandem bromodomains capable of binding to two acetylated lysine residues in close proximity, increasing the specificity of the interaction.

BRD2 and BRD3 are reported to associate with histones along actively

transcribed genes and may be involved in facilitating transcriptional elongation (Leroy et al, Mol. Cell. 2008 30(1):51-60), while BRD4 appears to be involved in the recruitment of the pTEF-I3 complex to inducible genes, resulting in phosphorylation of RNA polymerase and increased transcriptional output (Hargreaves et al, Cell, 2009 138(1): 1294145). All family members have been reported to have some function in controlling or executing aspects of the cell cycle, and have been shown to remain in complex with chromosomes during cell division – suggesting a role in the maintenance of epigenetic memory. In addition some viruses make use of these proteins to tether their genomes to the host cell chromatin, as part of the process of viral replication (You et al., Cell, 2004 117(3):349-60).

Recent articles relating to this target include Prinjha et al., Trends in

Pharmacological Sciences, March 2012, Vol. 33, No. 3, pp. 146-153; Conway, ACS Med. Chem. Lett., 2012, 3, 691-694 and Hewings et al, J. Med. Chem., 2012, 55, 9393-9413.

Small molecule BET inhibitors that are reported to be in development include GSK-525762A, OTX-015, TEN-010 as well as others from the University of Oxford and Constellation Pharmaceuticals Inc.

Hundreds of epigenetic effectors have been identified, many of which are chromatin-binding proteins or chromatin-modifying enzymes. These proteins have been associated with a variety of disorders such as neurodegenerative disorders, metabolic diseases, inflammation and cancer. Thus, these compounds which inhibit the binding of a bromodomain with its cognate acetylated proteins, promise new approaches in the treatment of a range of autoimmune and inflammatory diseases or conditions and in the treatment of various types of cancer.

 
Inventors Derek J. Norris, George V. Delucca, Ashvinikumar V. Gavai, Claude A. Quesnelle, Patrice Gill, Daniel O’MALLEY, Wayne Vaccaro, Francis Y. Lee, Mikkel V. DEBENEDETTO, Andrew P. Degnan, Haiquan Fang, Matthew D. Hill, Hong Huang, William D. Schmitz, JR John E. STARRETT, Wen-Ching Han, John S. Tokarski, Sunil Kumar MANDAL
Applicant Bristol-Myers Squibb Company

PATENT

WO 2015100282

Examples 54 & 55

2-[3-(Dimethyl-lH-l,2,3-triazol-5-yl)-5-[oxan-4-yl(phenyl)methyl]-5H-pyrido[3,2- b] indol-7-yl] pr opan-2-ol

Enantiomer A, Example 54 Enantiomer B, Example 55

Step 1 : 2-C hloro-5-(l ,4-dimethyl- 1H- 1 ,2,3-triazol-5-yl)pyridin-3-amine

To a 100 mL round bottom flask containing 5-bromo-2-chloropyridin-3-amine (2.90 g, 14.0 mmol), l,4-dimethyl-5-(tributylstannyl)-lH-l,2,3-triazole (2.70 g, 6.99 mmol) [Seefeld, M.A. et al. PCT Int. AppL, 2008, WO2008098104] and Pd(PPh3)4 (0.61 g, 0.52 mmol) in DMF (20 mL) was added cuprous iodide (0.20 g, 1.05 mmol) and Et3N (1.9 mL, 14.0 mmol). The reaction mixture was purged with N2 for 3 min and then heated at 100 °C for 1 h. After cooling to room temperature, the mixture was diluted withl0% LiCl solution and extracted with EtOAc (2x). The combined organics were washed with sat. NaCl, dried over MgS04, filtered and concentrated. CH2C12 was added, and the resulting precipitate was collected by filtration. The mother liquor was concentrated and purified using ISCO silica gel chromatography (40 g column, gradient from 0% to 100% EtOAc/CH2Cl2). The resulting solid was combined with the precipitate and triturated with cold EtOAc to give the title compound (740 mg, 47%) as a light tan solid. LCMS (M+H) = 224.1; HPLC RT = 1.03 min (Column: Chromolith ODS S5 4.6 x 50 mm; Mobile Phase A: 10:90 MeOH: water with 0.1% TFA; Mobile Phase B: 90: 10 MeOH:water with 0.1% TFA; Temperature: 40 °C; Gradient: 0-100% B over 4 min; Flow: 4 mL/min).

Step 2: Methyl 3-((2-chloro-5-(l,4-dimethyl-lH-l,2,3-triazol-5-yl)pyridin-3-yl)amino)benzoate

Following a procedure analogous to that described in Step 2 of Example 1, 2-chloro-5-(l ,4-dimethyl-lH-l,2,3-triazol-5-yl)pyridin-3-amine (740 mg, 3.31 mmol) was converted to the title compound (644 mg, 54%). 1H NMR (400 MHz, CDC13) δ 7.94 (t, J=1.9 Hz, 1H), 7.88 (d, J=2.1 Hz, 1H), 7.83 (dt, J=7.8, 1.3 Hz, 1H), 7.49 (t, J=7.9 Hz, 1H), 7.40 (d, J=2.1 Hz, 1H), 7.36 (ddd, J=8.0, 2.3, 0.9 Hz, 1H), 6.38 (s, 1H), 3.99 (s, 3H), 3.93 (s, 3H), 2.34 (s, 3H); LCMS (M+H) = 358.2; HPLC RT = 2.34 min (Column:

Chromolith ODS S5 4.6 x 50 mm; Mobile Phase A: 10:90 MeOH:water with 0.1% TFA; Mobile Phase B: 90: 10 MeOH:water with 0.1% TFA; Temperature: 40 °C; Gradient: 0-100% B over 4 min; Flow: 4 mL/min).

Step 3: Methyl 3-(l,4-dimethyl-lH-l,2,3-triazol-5-yl)-5H-pyrido[3,2-6]indole-7-carboxylate

Following a procedure analogous to that described in Step 3 of Example 1 , methyl 3-((2-chloro-5-(l,4-dimethyl-lH-l,2,3-triazol-5-yl)pyridin-3-yl)amino)benzoate (2.82 g, 7.88 mmol) was converted to the title compound (1.58 g, 62%). 1H NMR (500 MHz, DMSO-de) δ 11.93 (s, 1H), 8.62 (d, J=1.8 Hz, 1H), 8.36 (dd, J=8.2, 0.6 Hz, 1H), 8.29 -8.22 (m, 1H), 8.16 (d, J=1.8 Hz, 1H), 7.91 (dd, J=8.2, 1.4 Hz, 1H), 4.02 (s, 3H), 3.94 (s, 3H), 2.31 (s, 3H); LCMS (M+H) = 322.3; HPLC RT = 1.98 min (Column: Chromolith ODS S5 4.6 x 50 mm; Mobile Phase A: 10:90 MeOH:water with 0.1% TFA; Mobile Phase B: 90: 10 MeOH:water with 0.1% TFA; Temperature: 40 °C; Gradient: 0-100% B over 4 min; Flow: 4 mL/min).

Alternate synthesis of Methyl 3-(l,4-dimethyl-lH-l,2,3-triazol-5-yl)-5H-pyrido[3,2-b] indole-7-carboxylate

A mixture of methyl 3-bromo-5H-pyrido[3,2-b]indole-7-carboxylate (Step 2 of Example 40, 3.000 g, 9.83 mmol), l,4-dimethyl-5-(tributylstannyl)-lH-l,2,3-triazole (4.18 g, 10.82 mmol), copper (I) iodide (0.281 g, 1.475 mmol), Pd(Ph3P)4 (0.738 g, 0.639 mmol) and triethylamine (2.74 mL, 19.66 mmol) in DMF (25 mL) was purged under a nitrogen stream and then heated in a heating block at 95 °C for 2 hours. After cooling to room temperature the reaction mixture was diluted with water and extracted into ethyl acetate. Washed with water, NH4OH, brine and concentrated. The residue was triturated with 100 mL CHC13, filtered off the solid and rinsed with CHC13 to give. 1.6 g of product. The filtrate was loaded unto the ISCO column (330 g column, A: DCM; B:

10%MeOH/DCM, 0 to 100% gradient) and chromatographed to give an additional 0.7 g. of methyl 3 -( 1 ,4-dimethyl- 1 H- 1 ,2,3 -triazol-5 -yl)-5H-pyrido [3 ,2-b]indole-7-carboxylate (2.30 g total, 7.16 mmol, 72.8 % yield).

Step 4: Methyl 3-(l,4-dimethyl-lH-l,2,3-triazol-5-yl)-5-(phenyl(tetrahydro-2H-pyran-4-yl)methyl)-5H-pyrido[3,2-b]indole-7-carboxylate

Following a procedure analogous to that described in Step 4 of Example 1 , methyl 3-(l,4-dimethyl-lH-l,2,3-triazol-5-yl)-5H-pyrido[3,2-¾]indole-7-carboxylate (80 mg, 0.25 mmol) was converted to the title compound (65 mg, 53%) after purification by prep HPLC (Column: Phen Luna C 18, 30 x 100 mm, 5 μιη particles; Mobile Phase A: 5:95 acetonitrile: water with 0.1% TFA; Mobile Phase B : 95 : 5 acetonitrile: water with 0.1% TFA; Gradient: 10-100% B over 14 min, then a 2-min hold at 100% B; Flow: 40 mL/min). 1H NMR (400 MHz, CDC13) δ 8.51 (d, J=1.8 Hz, 1H), 8.50 (s, 1H), 8.47 (d, J=8.1 Hz, 1H), 8.10 (dd, J=8.1, 1.1 Hz, 1H), 7.63 (d, J=1.8 Hz, 1H), 7.46 (d, J=7.3 Hz, 2H), 7.40 – 7.30 (m, 3H), 5.62 (d, J=10.6 Hz, 1H), 4.11 – 4.03 (m, 4H), 3.92 – 3.83 (m, 4H), 3.56 (td, J=l 1.9, 1.8 Hz, 1H), 3.35 (td, J=l 1.9, 1.9 Hz, 1H), 3.18 – 3.05 (m, 1H), 2.30 (s, 3H), 2.04 (d, J=13.0 Hz, 1H), 1.71 – 1.58 (m, 1H), 1.50 – 1.37 (m, 1H), 1.09 (d, J=12.8 Hz, 1H); LCMS (M+H) = 496.3; HPLC RT = 2.93 min (Column: Chromolith ODS S5 4.6 x 50 mm; Mobile Phase A: 10:90 MeOH:water with 0.1% TFA; Mobile Phase B: 90: 10 MeOH:water with 0.1% TFA; Temperature: 40 °C; Gradient: 0-100% B over 4 min; Flow: 4 mL/min).

Step 5 : 2- [3-(Dimethyl- lH-1 ,2,3-triazol-5-yl)-5- [oxan-4-yl(phenyl)methyl] -5H-pyrido [3,2-6] indol-7-yl] pr opan-2-ol,

Following a procedure analogous to that described in Step 5 of Example 1 , methyl 3-(l ,4-dimethyl- IH- 1 ,2,3-triazol-5-yl)-5-(phenyl(tetrahydro-2H-pyran-4-yl)methyl)-5H-pyrido[3,2-b]indole-7-carboxylate (65 mg, 0.13 mmol) was converted to racemic 2-[3-(dimethyl-lH-l,2,3-triazol-5-yl)-5-[oxan-4-yl(phenyl)methyl]-5H-pyrido[3,2-¾]indol-7-yl]propan-2-ol, which was separated by chiral prep SFC (Column: Chiralpak IB 25 x 2 cm, 5 μιη; Mobile Phase: 70/30 C02/MeOH; Flow: 50 mL/min);to give Enantiomer A (24 mg, 36%) and Enantiomer B (26 mg, 38%). Enantiomer A: 1H NMR (500 MHz, CDC13) 5 8.44 (d, J=1.8 Hz, IH), 8.36 (d, J=8.2 Hz, IH), 7.98 (s, IH), 7.56 (d, J=1.7 Hz, IH), 7.47 – 7.41 (m, 3H), 7.37 – 7.32 (m, 2H), 7.31 – 7.28 (m, IH), 5.59 (d, J=10.5 Hz, IH), 4.06 (dd, J=11.8, 2.8 Hz, IH), 3.90 – 3.84 (m, 4H), 3.55 (td, J=11.9, 2.0 Hz, IH), 3.35 (td, J=11.9, 2.0 Hz, IH), 3.15 – 3.04 (m, IH), 2.30 (s, 3H), 2.04 (d, J=13.6 Hz, IH), 1.92 (s, IH), 1.75 (s, 6H), 1.69 – 1.58 (m, IH), 1.47 – 1.38 (m, IH), 1.12 (d, J=13.4 Hz, IH); LCMS (M+H) = 496.4; HPLC RT = 2.46 min (Column: Chromolith ODS S5 4.6 x 50 mm; Mobile Phase A: 10:90 MeOH:water with 0.1% TFA; Mobile Phase B: 90: 10 MeOH:water with 0.1% TFA; Temperature: 40 °C; Gradient: 0- 100% B over 4 min; Flow: 4 mL/min). SFC RT = 5.50 min (Column: Chiralpak IB 250 x 4.6 mm, 5 μιη; Mobile Phase: 70/30 C02/MeOH; Flow: 2 mL/min); SFC RT = 1.06 min (Column:

Chiralcel OD-H 250 x 4.6 mm, 5 μιη; Mobile Phase: 50/50 C02/(1 : 1 MeOH/CH3CN); Flow: 2 mL/min); [a]D2° = -117.23 (c = 0.08, CHC13). Enantiomer B: 1H NMR (500 MHz, CDC13) δ 8.44 (d, J=l .8 Hz, IH), 8.36 (d, J=8.2 Hz, IH), 7.98 (s, IH), 7.56 (d, J=1.7 Hz, IH), 7.47 – 7.41 (m, 3H), 7.37 – 7.32 (m, 2H), 7.31 – 7.28 (m, IH), 5.59 (d, J=10.5 Hz, IH), 4.06 (dd, J=11.8, 2.8 Hz, IH), 3.90 – 3.84 (m, 4H), 3.55 (td, J=11.9, 2.0 Hz, IH), 3.35 (td, J=l 1.9, 2.0 Hz, IH), 3.15 – 3.04 (m, IH), 2.30 (s, 3H), 2.04 (d, J=13.6 Hz, IH), 1.92 (s, IH), 1.75 (s, 6H), 1.69 – 1.58 (m, IH), 1.47 – 1.38 (m, IH), 1.12 (d, J=13.4 Hz, IH); LCMS (M+H) = 496.4; HPLC RT = 2.46 min (Column: Chromolith ODS S5 4.6 x 50 mm; Mobile Phase A: 10:90 MeOH:water with 0.1% TFA; Mobile Phase B: 90: 10 MeOH:water with 0.1% TFA; Temperature: 40 °C; Gradient: 0-100% B over 4 min; Flow: 4 mL/min). SFC RT = 8.30 min (Column: Chiralpak IB 250 x 4.6 mm, 5 μιη; Mobile Phase: 70/30 C02/MeOH; Flow: 2 mL/min); SFC RT = 2.83 min (Column: Chiralcel OD-H 250 x 4.6 mm, 5 μιη; Mobile Phase: 50/50 C02/(1 : 1 MeOH/CH3CN); Flow: 2 mL/min); [a]D2° = +88.78 (c = 0.10, CHC13).

Alternate Synthesis of Examples 54

2-[3-(Dimethyl-lH-l,2,3-triazol-5-yl)-5-[oxan-4-yl(phenyl)methyl]-5H-pyrido[3,2- b] indol-7-yl] propan-2-ol.

Enantiomer A, Example 54

Step 1: (S)-methyl 3-(l,4-dimethyl-lH-l,2,3-triazol-5-yl)-5-(phenyl(tetrahydro-2H-pyran-4-yl)methyl)-5H-pyrido[3,2-b]indole-7-carboxylate

The enantiomers of phenyl(tetrahydro-2H-pyran-4-yl)methanol ( 2.0 g, 10.4 mmol) [Orjales, A. et al. J. Med. Chem. 2003, 46, 5512-5532], were separated on preperative SFC. (Column: Chiralpak AD 5 x 25 cm, 5 μιη; Mobile Phase: 74/26

C02/MeOH; Flow: 270 mL/min; Temperature 30°C). The separated peaks were concentrated and dried under vacuum to give white solids. Enantiomer A: (S)-phenyl(tetrahydro-2H-pyran-4-yl)methanol: (0.91 g, 45.5%) SFC RT = 2.32 min

(Column: Chiralpac AD 250 x 4.6 mm, 5 μιη; Mobile Phase: 70/30 C02/MeOH; Flow: 3 mL/min); Temperature 40°C. Enantiomer B: (R)-phenyl(tetrahydro-2H-pyran-4-yl)methanol. (0.92 g, 46%) SFC RT = 3.09 min (Column: Chiralpac AD 250 x 4.6 mm, 5 μιη; Mobile Phase: 70/30 C02/MeOH; Flow: 3 mL/min); Temperature 40°C.

Following a procedure analogous to that described in Step 4 of Example 1 except using toluene (120mL) as the solvent, methyl 3-(l ,4-dimethyl-lH-l,2,3-triazol-5-yl)-5H-pyrido[3,2-b]indole-7-carboxylate (4 g, 12.45 mmol) and (R)-phenyl(tetrahydro-2H-pyran-4-yl)methanol (Enantiomer B above, 5.86 g, 30.5 mmol) was converted to the title compound (5.0 g, 81%). HPLC RT = 2.91 min (Column: Chromolith ODS S5 4.6 x 50 mm; Mobile Phase A: 10:90 MeOFLwater with 0.1% TFA; Mobile Phase B: 90: 10 MeOFLwater with 0.1% TFA; Temperature: 40 °C; Gradient: 0- 100% B over 4 min; Flow: 4 mL/min).

Step 2. (S)-2-[3-(Dimethyl-lH-l,2,3-triazol-5-yl)-5-[oxan-4-yl(phenyl)methyl]-5H-pyrido [3,2-b] indol-7-yl] propan-2-ol

A 500 mL round bottom flask containing (S)-methyl 3-(l,4-dimethyl-lH-l,2,3-triazol-5-yl)-5-(phenyl(tetrahydro-2H-pyran-4-yl)methyl)-5H-pyrido[3,2-b]indole-7-carboxylate (5.0 g, 10.09 mmol) in THF (150 mL) was cooled in an ice/MeOH bath. MeMgBr, (3M in Et20, 17.0 mL, 51.0 mmol) was added slowly over 4 min. The resulting solution was stirred for 2 h and then quenched carefully with sat. NH4C1. The reaction mixture was diluted with 10% LiCl solution extracted with EtOAc. The organic layer was dried over MgS04, filtered and concentrated. The crude material was purified using ISCO silica gel chromatography (120 g column, gradient from 0%> to 6%>

MeOH/CH2Cl2). The product was collected and concentrated then dissolved in hot MeOH(35mL). To the mixture was added 15mL water and the mixture was cooled to room temperature. The resulting white precipitate was collected by filtration with 2: 1 MeOH/water rinse then dried under vacuum to give the title compound (3.2 g, 62%>). 1H

NMR (500 MHz, CDC13) δ 8.40 (d, J=1.8 Hz, 1H), 8.33 (d, J=8.2 Hz, 1H), 7.93 (s, 1H), 7.53 (d, J=l .8 Hz, 1H), 7.46 (d, J=7.3 Hz, 2H), 7.42 (dd, J=8.2, 1.4 Hz, 1H), 7.37 – 7.31 (m, 2H), 7.30 – 7.28 (m, 1H), 5.56 (d, J=10.5 Hz, 1H), 4.06 (d, J=8.9 Hz, 1H), 3.89 – 3.83 (m, 1H), 3.55 (td, J=11.9, 2.1 Hz, 1H), 3.35 (td, J=11.9, 2.1 Hz, 1H), 3.10 (q, J=10.8 Hz, 1H), 2.39 (s, 3H), 2.23 (s, 3H), 2.03 (d, J=14.2 Hz, 1H), 1.89 (s, 1H), 1.74 (s, 6H), 1.68 -1.59 (m, 1H), 1.46 – 1.36 (m, 1H), 1.12 (d, J=12.2 Hz, 1H); LCMS (M+H) = 496.3; HPLC RT = 2.44 min (Column: Chromolith ODS S5 4.6 x 50 mm; Mobile Phase A: 10:90 MeOH: water with 0.1% TFA; Mobile Phase B: 90: 10 MeOH: water with 0.1%

TFA; Temperature: 40 °C; Gradient: 0-100% B over 4 min; Flow: 4 mL/min); SFC RT = 2.01 min (Column: Chiralcel OD-H 250 x 4.6 mm, 5 μιη; Mobile Phase: 60/40 C02/(1 : 1 MeOH/CH3CN); Flow: 2 mL/min). SFC RT = 1.06 min (Column: Chiralcel OD-H 250 x 4.6 mm, 5 μιη; Mobile Phase: 50/50 C02/(1 : 1 MeOH/CH3CN); Flow: 2 mL/min).

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US9458156 Tricyclic compounds as anticancer agents 2014-12-23 2016-10-04

3rd speaker at 1st time disclosures is Ashvin Gavai of @bmsnews talking about an oral BET inhibitor to treat cancer

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CC(C)(O)c2cc3n(c1cc(cnc1c3cc2)c4c(C)nnn4C)[C@@H](C5CCOCC5)c6ccccc6

Process Development and Good Manufacturing Practice Production of a Tyrosinase Inhibitor via Titanium-Mediated Coupling between Unprotected Resorcinols and Ketones


(S)-4-(2,4-Dihydroxyphenyl)-N-(1-phenylethyl)piperidine-1-carboxamide (1)

In a………………….. to yield crude 1 (3.51 kg, 77%, 97.7 A% purity). Recrystallization: In a 100 L double jacketed reactor were charged crude 1 (3.51 kg, 10.31 mol, 1.0 equiv), iPrOH (27.0 L, 7.5 vol), AcOH (74.1 g), and water (27.0 L, 7.5 vol). The suspension was warmed to reflux and turned to a solution after 30 min of reflux. Heating was stopped, and the reaction medium was allowed to cool to 23 °C over 20 h. The suspension was filtered through a 25 μm filter medium; the cake was washed with a mixture of water (3.6 L) and AcOH (7.3 g) and the solid collected and dried under vacuum at 45 °C for 48 h to yield 1 (2.86 kg, 81%, 98.5 A% purity).
1H NMR (400 MHz, DMSO-d6): δ 9.11 (s, 1H), 8.96 (s, 1H), 7.30–7.31 (m, 4), 7.19–7.20 (m, 1H), 6.79 (d, J = 8.3 Hz, 2H), 6.7 (d, J = 7.9 Hz, 2H), 6.28 (d, J = 2.4 Hz, 1H), 6.16 (dd, J = 8.3, 2.4 Hz, 1H), 4.85–4.87 (m, 1 H), 4.13 (d, J = 12.9 Hz, 2H), 2.85 (t, J = 11.9 Hz, 1H), 2.70 (t, J = 12.7 Hz, 2H), 1.64 (d, J = 12.1 Hz, 2H), 1.40–1.41 (m, 5H).
13C NMR (101 MHz, DMSO-d6) δ 156.6, 156.0, 155.2, 146.3, 127.9, 126.7, 126.1, 125.9, 122.5, 106.0, 102.4, 49.3, 44.4, 34.7, 31.8, 31.7, 22.9;
mp: 200–201 °C;
HRMS (m/z, ES+) for C20H25N2O3 (M + H)+ calcd. 341.1865, measd. 341.1859.

Process Development and Good Manufacturing Practice Production of a Tyrosinase Inhibitor via Titanium-Mediated Coupling between Unprotected Resorcinols and Ketones

Nestlé Skin Health R&D, 2400 Route des colles BP 87, 06902 Sophia-Antipolis Cedex, France
Org. Process Res. Dev., Article ASAP
DOI: 10.1021/acs.oprd.7b00036

ACS Editors’ Choice – This is an open access article published under an ACS AuthorChoice License, which permits copying and redistribution of the article or any adaptations for non-commercial purposes.

Thibaud Gerfaud

Thibaud Gerfaud

Team Leader Process Chemistry

Nestlé Skin Health Logo

Boiteau Jean-Guy

Boiteau Jean-Guy

Head of Process Research & Development

Nestlé Skin Health

Nestlé Skin Health Logo

Abstract

Abstract Image

A concise and economically attractive process for the synthesis of a novel tyrosinase inhibitor has been developed and implemented on a multikilogram scale under GMP. A major achievement to the success of the process is the development of a direct coupling between free resorcinol and ketone. First developed under basic conditions, this coupling has been turned to a novel titanium(IV) mediated process allowing good selectivity, easy isolation, and high atom efficiency. Other key steps feature an alkene reduction by palladium catalyzed transfer hydrogenation and a urea formation using N,N′-disuccinimidyl carbonate as the carbonyl source. This route allowed us to produce kilogram batches of the candidate to support preclinical and clinical studies.

Figure

Boiteau, J.-G.; Bouquet, K.; Talano, S.; Millois-Barbuis, C. Patent WO 2010/063774 A1, 2010.

More………………

str1

Cas 1228342-28-6
MF C20 H24 N2 O3,
MW  340.42
1-Piperidinecarboxamide, 4-(2,4-dihydroxyphenyl)-N-[(1S)-1-phenylethyl]-
  • 4-(2,4-Dihydroxyphenyl)-N-[(1S)-1-phenylethyl]-1-piperidinecarboxamide
  • 4-(2,4-Dihydroxyphenyl)piperidine-1-carboxylic acid N-((S)-1-phenylethyl)amide
Inventors Jean-Guy Boiteau , Karine Bouquet , Sandrine Talano , Barbuis Corinne Millois
Applicant Galderma Research & Development

Hyperpigmentation disorders such as melasma are characterized by an increase in melanin synthesis which accumulates in the epidermis and is responsible for a darkening of the skin. Melanogenesis occurs in the basal layer of the epidermis into specific organelles of the melanocytes called melanosomes.

A detailed analysis of the biosynthetic pathway reveals that tyrosinase is a key enzyme in melanogenesis and is responsible for the oxidation of tyrosine into DOPA (3,4-dihydroxyphenylalanine) and DOPA quinone.

It is a melanogenesis inhibitor working through the inhibition of tyrosinase (IC50 = 0.1 μM on normal human epidermal melanocytes) currently under development at Nestlé Skin Health R&D for the topical treatment of hyperpigmentation disorders. REF 1-5

WO 2010063774

Novel 4- (azacycloalkyl)benzene-l ,3-diol compounds as tyrosinase inhibitors, process for the preparation thereof and use thereof in human medicine and in cosmetics

The invention relates to novel 4- (azacycloalkyl) benzene-1, 3-diol compounds as industrial and useful products. It also relates to the process for the preparation thereof and to the use thereof, as tyrosinase inhibitors, in pharmaceutical or cosmetic compositions for use in the treatment or prevention of pigmentary disorders.

Skin pigmentation, in particular human skin pigmentation, is the result of melanin synthesis by dendritic cells, melanocytes. Melanocytes contain organelles called melanosomes which transfer melanin into the upper layers of keratinocytes which are then transported to the surface of the skin through differentiation of the epidermis (Gilchrest BA, Park HY, Eller MS, Yaar M, Mechanisms of ultraviolet light-induced pigmentation. Photochem Photobiol 1996; 63: 1-10; Hearing VJ, Tsukamoto K, Enzymatic control of pigmentation in mammals. FASEB J 1991; 5: 2902-2909) .

Among the enzymes of melanogenesis, tyrosinase is a key enzyme which catalyses the first two steps of melanin synthesis. Homozygous mutations of tyrosinase cause oculocutaneous albinism type I characterized by a complete lack of melanin synthesis (Toyofuku K, Wada I, Spritz RA, Hearing VJ, The molecular basis of oculocutaneous albinism type 1 (OCAl) : sorting failure and degradation of mutant tyrosinases results in a lack of pigmentation. Biochem J 2001; 355: 259-269) .

In order to treat pigmentation disorders resulting from an increase in melanin production, for which there is no treatment that meets all the expectations of patients and dermatologists, it is important to develop new therapeutic approaches.

Most of the skin-lightening compounds that are already known are phenols or hydroquinone derivatives.

These compounds inhibit tyrosinase, but the majority of them are cytotoxic to melanocytes owing to the formation of quinones. There is a risk of this toxic effect causing a permanent depigmentation of the skin. The obtaining of compounds that can inhibit melanogenesis while at the same time being very weakly cytotoxic or devoid of toxicity to melanocytes is most particularly sought.

Among the compounds already described in the literature, patent application WO 99/15148 discloses the use of 4-cycloalkyl resorcinols as depigmenting agents .

Patent FR2704428 discloses the use of 4-halo-resorcinols as depigmenting agents.

Patent applications WO 2006/097224 and WO 2006/097223 disclose the use of 4-cycloalkylmethyl resorcinols as depigmenting agents.

Patent application WO 2005/085169 discloses the use of alkyl 3- (2, 4-dihydroxyphenyl) propionate as a depigmenting agent.

Patent application WO 2004/017936 discloses the use of 3- (2, 4-dihydroxyphenyl) acrylamide as a depigmenting agent.

Patent application WO 2004/052330 discloses the use of 4- [ 1, 3] dithian-2-ylresorcinols as depigmenting agents .

More particularly, patent EP0341664 discloses the use of 4-alkyl resorcinols as depigmenting agents, among which 4-n-butyl resorcinol, also known as rucinol, is part of the composition of a depigmenting cream sold under the name Iklen®.

The applicant has now discovered, unexpectedly and surprisingly, that novel compounds of 4- (azacycloalkyl) benzene-1, 3-diol structure have a very good tyrosinase enzyme-inhibiting activity and a very low cytotoxicity. Furthermore, these compounds have a tyrosinase enzyme-inhibiting activity that is greater than that of rucinol while at the same time being less cytotoxic with respect to melanocytes than rucinol.

These compounds find uses in human medicine, in particular in dermatology, and in the cosmetics field.

FR 2939135

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  1. Briganti, S.; Camera, E.; Picardo, M. Pigm. Cell Res. 2003, 16, 101, DOI: 10.1034/j.1600-0749.2003.00029.x

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    Brenner, M.; Hearing, V. J. Photochem. Photobiol. 2008, 84, 539, DOI: 10.1111/j.1751-1097.2007.00226.x

  3. 3.

    (a) Schallreuter, K. U.; Kothari, S.; Chavan, B.; Spencer, J. D. Exp. Dermatol. 2008, 17, 395, DOI: 10.1111/j.1600-0625.2007.00675.x

    (b) Cooksey, C. J.; Garratt, P. J.;Land, E. J.; Pavel, S.; Ramsden, C. A.; Riley, P. A.; Smit, N. P.J. Biol. Chem. 1997, 272, 26226, DOI: 10.1074/jbc.272.42.26226

    (c) Stratford, M. R. L.; Ramsden, C. A.; Riley, P. A.Bioorg. Med. Chem. 2013, 21, 1166, DOI: 10.1016/j.bmc.2012.12.031

  4. 4.

    Chang, T. S. Int. J. Mol. Sci. 2009, 10, 2440, DOI: 10.3390/ijms10062440

  5. 5.

    Hypopigmentation effect have already been demonstrated for resorcinols; see:

    (a) Kim, D. S.; Kim, S. Y.;Park, S. H.; Choi, Y. G.; Kwon, S. B.; Kim, M. K.; Na, J. I.; Youn, S. W.; Park, K. C. Biol. Pharm. Bull. 2005,28, 2216, DOI: 10.1248/bpb.28.2216

    (b) Khemis, A.; Kaiafa, A.;Queille-Roussel, C.; Duteil, L.; Ortonne, J. P. Br. J. Dermatol.2007, 156, 997, DOI: 10.1111/j.1365-2133.2007.07814.x

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

O=C(N[C@@H](C)c1ccccc1)N2CCC(CC2)c3ccc(O)cc3O

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,

Image result for DEBIOPHARM

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

Image result for Aurigene Discovery Technologies Ltd

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

SEN 826


figure

SEN 826
CAS 1160833-51-1
C25 H31 N5 O, 417.55
Methanone, [1-[3-(1-methyl-1H-benzimidazol-2-yl)phenyl]-4-piperidinyl](4-methyl-1-piperazinyl)-
CAS HBr SALT 1612250-71-1

WO2009074300 product patent

Russell John Thomas, Mohr Gal.La Pericot, Giacomo Minetto, Annette Cornelia Bekker, Pietro Ferruzzi
Applicant Siena Biotech S.P.A.
Image result for Siena Biotech S.P.A.
Siena Biotech S.p.A. operates as a drug discovery and development company which develops a portfolio of disease modifying small molecule therapeutics for oncology and neurodegenerative diseases. Its products include blood-brain barrier penetrant compounds, which are in pipeline, for the treatment of brain cancers and peripheral tumors capable of metastasizing to the brain; clinical candidates for Alzheimer’s disease; and SEN196, a Sirtuin 1 inhibitor against Huntington disease. The company also provides contract research services, drug discovery, integrated chemistry, in-vitro technologies, and preclinical technologies. Siena Biotech S.p.A. has a strategic partnership with Aptuit Inc. The company was founded in 2000 and is based in Siena, Italy. Siena Biotech S.p.A operates as a subsidiary of THERAMetrics holding AG
Russell Thomas

Russell Thomas

https://www.linkedin.com/in/russell-thomas-0317464/

PLEASE MAIL ME AT amcrasto@gmail.com if picture is a mistake or cal +919323115463

The SMO receptor mediates Hedgehog (Hh) signaling critical to development, differentiation, growth, and cell migration. In normal conditions, activation of the pathway is induced by binding of specific endogenous ligands (i.e., Sonic Hh) to its receptor Patched (Ptch), which in turns reverts the Ptch inhibitory effect on SMO. SMO activation ultimately determines specific target genes activation through a family of three transcription factors, Gli1, Gli2 and Gli3.
Although Hh signaling is significantly curtailed in adults, it retains functional roles in stem cell maintenance, and aberrant Hh signaling has been described in a range of tumours.
Mutational inactivation of the inhibitory pathway components results in a constitutive ligand-independent activation seen in tumours such as basal cell carcinoma (BCC) and medulloblastoma. Ligand-dependent activation is seen in tumours such as prostate cancer, pancreatic cancer, gastrointestinal malignancies, melanoma, gliomas, breast cancer, ovarian cancer, leukemia, and B-cell lymphomas. A significant body of evidence supports the conclusion that SMO receptor antagonism will block the downstream signaling events.
As part of a program to address unmet medical need with regard to tumours in the CNS, Siena Biotech has designed and investigated selective antagonists of the SMO receptor. The newly designed API development candidate SEN826 1  is part of a group of potent antagonists of the Hedgehog pathway.
SYNTHESIS

PATENT

WO 2009074300

Figure imgf000025_0001

Figure imgf000019_0002

Figure

The synthesis starts with the formation of the 2-arylbenzimidazole derivative 6 which can be carried out starting from N-methylphenylenediamine 2 (Method A; blue path in Scheme 1) or employing o-phenylenediamine 4 in the ring closure reaction followed by N-methylation (Method B; orange path in Scheme 1). Sodium hydrogen sulfite is used to promote the condensation of the corresponding o-phenylenediamine with the Br-aromatic aldehyde 3.(6b) The next step is the coupling of the aryl bromide with isonipecotic ethyl ester in Buchwald conditions. After acidic hydrolysis with HCl under microwave irradiation, the final amide 1 was synthesized with CDI as coupling agent.

PAPER

A Scalable Route to the SMO Receptor Antagonist SEN826: Benzimidazole Synthesis via Enhanced in Situ Formation of the Bisulfite–Aldehyde Complex

Process Chemistry Unit, Siena Biotech SpA, 53100 Siena, Italy
Compound Management & Analysis Unit, Siena Biotech SpA, 53100 Siena, Italy
Org. Process Res. Dev., 2014, 18 (6), pp 699–708
Abstract Image

A practical and scalable route to the SMO antagonist SEN826 1 is described herein, including the discussion of an alternative approach to the synthesis of the target molecule. The optimized route consists of five chemical steps. A new and efficient access to the key intermediate 6 via the bisulfite–aldehyde complex was developed, significantly enhancing the yields and reducing costs. As a result, a synthetic procedure for preparation of multihundred gram quantities of the final product has been developed.

1 as hydrobromide salt. Yield: 71%.
UPLC–MS: tR = 1.24 min; m/z = 418 [M + 1]+.
HRMS calcd for C25H33N5O [M + 1]+ 418.26069, found 418.26075.
HPLC: tR = 5.99 min; purity 99.1%.
1H NMR (400 MHz DMSO-d6): δ 9.80 (broad, 1H), 7.89 (m, 1H), 7.77 (m, 1H), 7.55–7.45 (m, 3H), 7.38 (s, 1H), 7.24 (m, 2H), 4.48–4.15 (m, 2H), 3.96 (s, 3H), 3.86 (m, 2H), 3.55–3.15 (m, 3H), 3.10–2.82 (m, 6H), 2.81 (s, 3H), 1.76–1.57 (m, 4H).
13C NMR (100 MHz DMSO-d6): δ 173.5, 152.3, 151.5, 135.1, 135.0, 130.5, 126.2, 125.6, 125.3, 119.9, 119.1, 117.1, 116.5, 113.0, 53.2, 48.2, 42.7, 38.8, 37.4, 33.1, 28.2.
Water content (KF): 3.5 wt %.
Pd content (ICP-MS): 128 ppm.
Bromine content (ionic exchange LC): 20 wt % (1.2 equiv).
str1 str2
/////////////////

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.

Image result for AMGEN

 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

Image result for Antidiabetic effects of glucokinase regulatory protein small-molecule disruptors.

Image result for Antidiabetic effects of glucokinase regulatory protein small-molecule disruptors.

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

Hoshinolactam, A new antitrypanosomal lactam


Abstract Image
Tropical diseases caused by parasitic protozoa are a threat to human health, mainly in developing countries. Trypanosomiasis (Chagas disease and sleeping sickness) and leishmaniasis, inter alia, are classified as neglected tropical diseases, and over 400 million people are at risk of contracting these diseases.

In addition, a parasite of the Trypanosoma genus, Trypanosoma brucei brucei, is the causative agent of Nagana disease in wild and domestic animals, and this disease is a major obstacle to the economic development of affected rural areas.

Although some therapeutic agents for these diseases exist, they have limitations, such as serious side effects and the emergence of drug resistance. Thus, new and more effective antiprotozoal medicines are needed

Marine natural products have recently been considered to be good sources for drug leads. In particular, secondary metabolites produced by marine cyanobacteria have unique structures and versatile biological activities, and some of these compounds show antiprotozoal activities. For example, coibacin A isolated from cf. Oscillatoria sp. exhibited potent antileishmanial activity, and viridamide A isolated from Oscillatoria nigro-viridis showed antileishmanial and antitrypanosomal activities.

constituents of marine cyanobacteria and reported an antitrypanosomal cyclodepsipeptide, janadolide.

The marine cyanobacterium was collected at the coast near Hoshino, Okinawa.

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Okinawa
沖縄市
Uchinaa
City
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EARLIER MERCK TEAM HAD REPORTED

CAS 159153-15-8
MF C20 H33 N O5
MW 367.48
2-Pyrrolidinone, 3,4-dihydroxy-5-(hydroxymethyl)-3-[3-(2-nonylcyclopropyl)-1-oxo-2-propenyl]-, [3S-[3α,3[E(1S*,2S*)],4β,5α]]-
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Antitrypanosomal
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Marine cyanobacterium
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Human fetal lung fibroblast MRC-5 cells
Majusculoic acid.png
Majusculoic acid
Image result for malyngamide A.
Malyngamide A.

PAPER

http://pubs.acs.org/doi/suppl/10.1021/acs.orglett.7b00047

Recently, we isolated a new antitrypanosomal lactam, hoshinolactam (1), from a marine cyanobacterium.Structurally, 1 contains a cyclopropane ring and a γ-lactam ring. So far, some metabolites possessing either a cyclopropane ring or a γ-lactam ring have been discovered from marine cyanobacteria, such as majusculoic acid and malyngamide A. To the best of our knowledge, on the other hand, hoshinolactam (1) is the first compound discovered in marine cyanobacteria that possesses both of these ring systems. In addition, we clarified that 1 exhibited potent antitrypanosomal activity without cytotoxicity against human fetal lung fibroblast MRC-5 cells. Here, we report the isolation, structure elucidation, first total synthesis, and preliminary biological characterization of hoshinolactam (1).

Isolation and Total Synthesis of Hoshinolactam, an Antitrypanosomal Lactam from a Marine Cyanobacterium

Department of Chemistry, Keio University, 3-14-1 Hiyoshi, Kohoku-ku, Yokohama, Kanagawa 223-8522, Japan
Research Center for Tropical Diseases, Kitasato Institute for Life Sciences, and §Graduate School of Infection Control Sciences, Kitasato University, 5-9-1, Shirokane, Minato-ku, Tokyo 108-8641, Japan
Org. Lett., Article ASAP
DOI: 10.1021/acs.orglett.7b00047

Abstract Image

In the search for new antiprotozoal substances, hoshinolactam, an antitrypanosomal lactam, was isolated from a marine cyanobacterium. The gross structure was elucidated by spectroscopic analyses, and the absolute configuration was determined by the first total synthesis. Hoshinolactam showed potent antitrypanosomal activity with an IC50 value of 3.9 nM without cytotoxicity against human fetal lung fibroblast MRC-5 cells (IC50 > 25 μM).

Table 1. 1H and 13C NMR Data for 1 in C6D6
unit position δCa δHb (J in Hz)
HIMP 1 177.8, C
2 44.1, CH 2.51, dq (5.2, 7.6)
3 80.8, CH 4.94, dd (4.6, 5.2)
4 57.3, CH 3.49, ddd (4.6, 4.7, 9.4)
5a 44.6, CH2 1.21, m
5b 1.36, m
6 25.0, CH 1.61, m
7 21.7, CH3 0.74, d (6.2)
8 23.2, CH3 0.76, d (6.3)
9 15.0, CH3 1.33, d (7.6)
NH 7.65, s
PCPA 1 166.0, C
2 117.4, CH 5.88, d (15.5)
3 155.0, CH 6.59, dd (10.3, 15.5)
4 22.4, CH 0.91, m
5 23.3, CH 0.59, m
6 35.7, CH2 0.96, m
7 22.5, CH2 1.20, tq (7.1, 7.3)
8 14.0, CH3 0.78, t (7.3)
9a 16.1, CH2 0.35, ddd (4.5, 6.0, 8.2)
9b 0.42, ddd (4.5, 4.5, 8.8)
aMeasured at 100 MHz.
bMeasured at 400 MHz.
Positive HRESIMS data (m/z 308.2228, calcd for C18H30NO3 [M + H]+ 308.2225). Table 1 shows the NMR data for 1.
An analysis of the 1H NMR spectrum indicated the presence of four methyl groups (δH 0.74, 0.76, 0.78 and 1.33), four protons of the cyclopropane ring (δH 0.35, 0.42, 0.59 and 0.91), and two olefinic protons (δH 5.88 and 6.59).
The 13C NMR and HMQC spectra revealed the existence of two carbonyl groups (δC 166.0 and 177.8) and two sp2 methines (δC 117.4 and 155.0).
Examination of the COSY and HMBC spectra established the presence of two fragments derived from 4-hydroxy-5-isobutyl-3-methylpyrrolidin-2-one (HIMP) and 3-(2-propylcyclopropyl) acrylic acid (PCPA), respectively. The configuration of the C-2–C-3 olefinic bond in the PCPA was determined to be trans on the basis of the coupling constant (3JH2–H3 = 15.5 Hz). The connectivity of the two partial structures was determined from the HMBC correlation (H-3 of HIMP/C-1 of PCPA).
1H, 13C, COSY, HMQC, HMBC, and NOESY NMR spectra in C6D6 and 1H and 13C NMR spectra in CD3OD for hoshinolactam (1)
1H, 13C, COSY, HMQC, HMBC, and NOESY NMR spectra in C6D6

1H and 13C NMR spectra in CD3OD

1H NMR PREDICT

13 C NMR PREDICT

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OKINAWA

///////////Hoshinolactam

CC(C)C[C@@H]2NC(=O)[C@H](C)C2OC(=O)/C=C/[C@H]1C[C@@H]1CCC

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

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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

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