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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 AFRICURE PHARMA, ROW2TECH, NIPER-G, Department of Pharmaceuticals, Ministry of Chemicals and Fertilizers, Govt. of India as ADVISOR, earlier assignment was with GLENMARK LIFE SCIENCES LTD, as CONSUlTANT, Retired from GLENMARK in Jan2022 Research Centre as Principal Scientist, Process Research (bulk actives) at Mahape, Navi Mumbai, India. Total Industry exp 32 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 Open superstar worlddrugtracker. His New Drug Approvals, Green Chemistry International, All about drugs, Eurekamoments, Organic spectroscopy international, etc in organic chemistry are some most read blogs He has hands on experience in initiation and developing novel routes for drug molecules and implementation them on commercial scale over a 32 PLUS year tenure till date Feb 2023, Around 35 plus products in his career. He has good knowledge of IPM, GMP, Regulatory aspects, he has several International patents published worldwide . He has good proficiency in Technology transfer, Spectroscopy, Stereochemistry, Synthesis, Polymorphism etc., He suffered a paralytic stroke/ Acute Transverse mylitis in Dec 2007 and is 90 %Paralysed, He is bound to a wheelchair, this seems to have injected feul in him to help chemists all around the world, he is more active than before and is pushing boundaries, He has 100 million plus hits on Google, 2.5 lakh plus connections on all networking sites, 100 Lakh plus views on dozen plus blogs, 227 countries, 7 continents, 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 38 lakh plus views on New Drug Approvals Blog in 227 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 He has total of 32 International and Indian awards

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ARIMOCLOMOL

May 30, 2022 7:42 am / 1 Comment on ARIMOCLOMOL

ARIMOCLOMOL

アリモクロモル;

Formula	C14H20ClN3O3
Exact mass	313.1193
Mol weight	313.7799

CAS 289893-25-0

289893-26-1 (Arimoclomol maleate);

INN 8300

N-[(2R)-2-hydroxy-3-piperidin-1-ylpropoxy]-1-oxidopyridin-1-ium-3-carboximidoyl chloride

BRX 220

Arimoclomol maleate is in a phase III clinical trials by Orphazyme for the treatment of Niemann-Pick disease type C (NP-C). It is also in phase II clinical studies for the treatment of amyotrophic lateral sclerosis (ALS).

Arimoclomol (INN; originally codenamed BRX-345, which is a citrate salt formulation of BRX-220) is an experimental drug developed by CytRx Corporation, a biopharmaceutical company based in Los Angeles, California. In 2011 the worldwide rights to arimoclomol were bought by Danish biotech company Orphazyme ApS.^[1] The European Medicines Agency (EMA) and U.S. Food & Drug Administration (FDA) granted orphan drug designation to arimoclomol as a potential treatment for Niemann-Pick type C in 2014 and 2015 respectively.^[2][3]


	Fig. 1 Structures of (±)-bimoclomol (1) and (R)-(+)-arimoclomol (2).

Reference：1. WO0179174A1.

Reference：1. Tetrahedron: Asymmetr. 2012, 23, 1564-1570.

PATENT

WO/2022/106614PROCESSES FOR PREPARING ARIMOCLOMOL CITRATE AND INTERMEDIATES THEREOF

https://patentscope.wipo.int/search/en/detail.jsf?docId=WO2022106614&_cid=P22-L3S8T3-04206-1

The present disclosure provides an optimized four-step process for preparing an ultra-pure composition comprising arimoclomol citrate, i.e. N-{[(2R)-2-hydroxy-3-piperidin-l-ylpropyl]oxy}pyridine-3-carboximidoyl chloride 1-oxide citrate. The optimized process comprises a plurality of optimized sub-steps, each contributing to an overall improved process, providing the ultra-pure composition comprising arimoclomol citrate. The ultra-pure composition comprising arimoclomol citrate meets the medicines agencies’ high regulatory requirements. An overview of the four-steps process is outlined below:

Step 1: Overview of process for preparing ORZY-01

Step 2: Overview of process for preparing ORZY-03

Step 4: Overview of process for preparing BRX-345 (ORZY-05)

The previously reported two-step synthesis of ORZY-01 as shown below includes a 2 hour reflux in step 1A, followed by purification of intermediate compound (V) to increase the batch quality.

PAPER

https://pubs.rsc.org/en/content/articlehtml/2017/ob/c7ob02578e

DOI: 10.1039/C7OB02578E (Communication) Org. Biomol. Chem., 2017, 15, 9794-9799

Scheme 1 Synthesis of arimoclomol (2) by reproduction of the published patent route. Reagents and conditions: (a) NH₂OH·HCl (1.2 equiv.), NaHCO₃ (1.2 equiv.), H₂O, rt, 18 h 91%; (b) piperidine (0.9 equiv.), MeOH, 65 °C, quant.; (c) 6, NaOH (1.3 equiv.), EtOH, H₂O, 70 °C, 18 h; (d) NaNO₂ (1.3 equiv.), conc. HCl, H₂O, −5 °C, 2.5 h 51% over 2 steps; (e) (−)-dibenzoyl-L-tartaric acid, EtOH then NaOH, CH₂Cl₂; (f) citric acid (1.0 equiv.), acetone; (g) supercritical fluid chromatography.


	Scheme 3 Arimoclomol (2) synthesis via chiral glycidyl nosylate synthon. Reagents and conditions: (a) (i) NaH (60% wt), DMF, 0 °C, 0.5 h; (ii) (R)-(−)-glycidyl nosylate (11) (1.06 equiv.), rt, 2 h; (iii) piperidine (1.1 equiv.), 80 °C for 4 h then rt for 18 h, 71%; (b) NaNO₂ (1.3 equiv.), conc. HCl, H₂O, −5 °C, 2.5 h, 73%.

Scheme 4 Chiral hydroxylamine route to arimoclomol (2). Reagents and conditions: (a) (i) NaH (60% wt), DMF, 0 °C, 0.5 h; (ii) (R)-(−)-glycidyl nosylate (11) (1.1 equiv.), rt, 2 h, 83%; (b) piperidine (1.05 equiv.), ⁱPrOH, 50 °C, 18 h, quant.; (c) HCl (6 M), 95 °C, 18 h; quant.; (d) Amberlyst A21, MeOH, rt, 4 h, 98%; (e) 3-cyanopyridine-N-oxide (3) (0.8 equiv.), HSCH₂CO₂H (17) (1.5 equiv.), Et₃N, EtOH, 85 °C, 24 h, 75%; (f) NaNO₂ (1.3 equiv.), conc. HCl, H₂O, −5 °C, 66%.

(R,Z)-3-(N′-(2-Hydroxy-3-(piperidine-1-yl)propoxy)carboximi-oylchloride)pyridine-1-oxide citrate (2-citrate, arimoclomol citrate) was prepared as an off-white amorphous solid (164 mg): m.p. 161–162 °C; [α]²⁰_D +8.0° (c = 1, H₂O); IR ν_max (neat): 3423, 3228, 2949, 2868, 1722, 1589, 1483, 1433, 1307, 1128, 972, 829 cm⁻¹; ¹H NMR (600 MHz, d₆-DMSO) δ: 8.54 (t, J = 1.5 Hz, 1H), 8.39–8.35 (m, 1H), 7.72–7.68 (m, 1H), 7.55 (dd, J = 8.0, 6.5 Hz, 1H), 4.28 (ddd, J = 17.6, 13.3, 7.4 Hz, 3H), 3.35 (br. s, 2H), 3.13–2.74 (m, 6H), 2.59 (d, J = 15.2 Hz, 2H), 2.56–2.51 (m, 2H), 1.77–1.61 (m, 4H), 1.48 (s, 2H); ¹³C NMR (151 MHz, d₆-DMSO) δ: 176.6, 171.3, 140.5, 136.4, 132.7, 131.5, 126.8, 123.3, 77.8, 71.4, 63.8, 58.7, 53.1, 44.0, 30.7, 23.0, 21.9; HRMS (m/z TOF MS ES+) for C₁₄H₂₀ClN₃O₃ [M + H]+ calc. 314.1271, observed 314.1263; SFC er purity R:S, >99:1.
(R,Z)-3-(N′-(2-Hydroxy-3-(piperidine-1-yl)propoxy)carboximi-oylchloride)pyridine maleate ((R)-1-maleate, bimoclomol maleate) was prepared as an off-white amorphous solid (70 mg): m.p. 137–138 °C; [α]²⁰_D +6.0° (c = 1, MeOH); IR νmax (neat): 3269, 2937, 1577, 1477, 1440, 1348, 1205, 1070, 981, 864 cm⁻¹; ¹H NMR (600 MHz, d₆-DMSO) δ: 9.09 (s, 1H), 9.01–8.98 (m, 1H), 8.73 (dd, J = 4.8, 1.5 Hz, 1H), 8.24–8.06 (m, 1H), 7.57 (ddd, J = 8.1, 4.8, 0.6 Hz, 1H), 6.02 (d, J = 4.0 Hz, 2H), 5.93 (s, 1H), 4.40–4.21 (m, 3H), 3.60–3.28 (m, 3H), 3.20 (d, J = 11.8 Hz, 1H), 3.12–3.05 (m, 1H), 3.03–2.83 (m, 2H), 1.84–1.55 (m, 5H), 1.38 (s, 1H); ¹³C NMR (151 MHz, d₆-DMSO) δ: 167.1, 151.7, 147.4, 136.0, 135.1, 134.6, 127.9, 123.9, 77.2, 63.1, 58.0, 54.1, 51.1, 22.2, 21.3; HRMS (m/z TOF MS ES+) for C₁₄H₂₀ClN₃O₂ [M + H]+ calc. 298.1322, observed 298.1319; SFC er purity R:S, 98:2.

(R,Z)-3-(N’-(2-hydroxy-3-(piperidin-1-yl)propoxy)carboximidoyl chloride)pyridine-1-oxide1 – (R)-(+)-Arimoclomol – 2 A solution of (R,Z)-3-(N’-(2-hydroxy-3-(piperidin-1-yl)propoxy)carbamimidoyl)pyridine-1-oxide 12 (205 mg, 0.70 mmol) in conc. hydrochloric acid (1.1 mL, 13.9 mmol) and water (3 mL) was cooled to -5 °C for 15 minutes. Sodium nitrite (63 mg, 0.91 mmol) in water (0.5 mL) was then added dropwise to the reaction mixture and the reaction was stirred at -5 °C for 2.5 hours. The reaction mixture was made alkaline with NaOH (7 M, 3 mL). An additional 10 mL of water was added followed by DCM (30 mL) containing EtOAc (5 mL) and the organics were dried over MgSO4, filtered and concentrated in vacuo. The residue was purified by FCC on Biotage Isolera using Biotage SNAP 10 g Si cartridge eluting with gradient elution 0-30% MeOH:DCM both containing 0.1% Et3N to afford the title compound (160 mg, 73% yield) as a colourless semi-solid. Analytical data was consistent with literature values. See ESI section SFC traces for specific enantiomeric ratios of 2 synthesised under the various methodologies quoted in the text. Optical rotation was not determined as it was determined in the ultimate product of this 2·citrate and comparative run times on SFC. 1H NMR (600 MHz, CDCl3) δ: 8.63 (t, J = 1.4 Hz, 1H), 8.16 (ddd, J = 6.4, 1.6, 0.9 Hz, 1H), 7.66 – 7.62 (m, 1H), 7.25 (dd, J = 8.0, 6.6 Hz, 1H), 4.26 (qd, J = 11.3, 5.2 Hz, 2H), 4.07 (dd, J = 9.2, 4.7 Hz, 1H), 2.62 (s, 2H), 2.47 – 2.31 (m, 4H), 1.65 – 1.51 (m, 4H), 1.42 (s, 2H); 13C NMR (151 MHz, CDCl3) δ: 140.3, 137.7, 133.1, 132.5, 125.7, 123.9, 78.7, 64.9, 60.9, 54.8, 25.8, 24.0.

(R,Z)-3-(N’-(2-hydroxy-3-(piperidin-1-yl)propoxy)carboximidoyl chloride)pyridine-1-oxide citrate

(R)-(+)- Arimoclomol citrate – 2·citrate (R,Z)-3-(N’-(2-hydroxy-3-(piperidin-1-yl)propoxy)carboximidoyl chloride)pyridine-1-oxide (159 mg, 0.51 mmol) was dissolved in acetone (3 mL) and citric acid (97 mg, 0.51 mmol) was added. The reaction mixture was left to stir at room temperature for 18 hours. After this time the mixture was sonicated and the precipitate was filtered, rinsed with cold acetone (1 mL) and dried under vacuum to afford the title compound (165 mg, 64% yield) as a white amorphous solid. Analytical data was consistent with literature values. m.p. 161-162 °C, Acetone (lit. 163-165 °C, EtOH); [α]D 20 +8.0 (c=1, H2O); IR νmax (neat): 3423, 3228, 2949, 2868, 1722, 1589, 1483, 1433, 1307, 1128, 972, 829 cm-1; 1H NMR (600 MHz, d6-DMSO) δ: 8.54 (t, J = 1.5 Hz, 1H), 8.39 – 8.35 (m, 1H), 7.72 – 7.68 (m, 1H), 7.55 (dd, J = 8.0, 6.5 Hz, 1H), 4.28 (ddd, J = 17.6, 13.3, 7.4 Hz, 3H), 3.35 (br. s, 2H), 3.13 – 2.74 (m, 6H), 2.59 (d, J = 15.2 Hz, 2H), 2.56 – 2.51 (m, 2H), 1.77 – 1.61 (m, 4H), 1.48 (s, 2H); 13C NMR (151 MHz, d6-DMSO) δ: 176.6, 171.3, 140.5, 136.4, 132.7, 131.5, 126.8, 123.3, 77.8, 71.4, 63.8, 58.7, 53.1, 44.0, 30.7, 23.0, 21.9; HRMS (m/z TOF MS ES+) for C14H20ClN3O3 [M+H]+ calc. 314.1271, observed 314.1263; SFC er purity R:S >99:1

Procedure for the conversion of (R)-(+)-Bimoclomol 1 into (R)-(+)-Arimoclomol 2 To a solution of (R)-(+)-bimoclomol (61 mg, 0.21 mmol) in acetone (2 mL) was added benzenesulfonic acid (33 mg, 0.21 mmol). The reaction mixture was stirred at room temperature for 1.5 hours. The reaction mixture was concentrated in vacuo. Separately to a suspension of hydrogen peroxide-urea adduct (39 mg, 0.41 mmol) in acetonitrile (6 mL) at -5°C (ice-salt bath) was added trifluoroacetic anhydride (58 μL, 0.41 mmol) dropwise. A suspension of (R)-(+)-bimoclomol, 1, benzenesulfonic acid salt, as made above, in acetonitrile (3 mL) was then added dropwise to this solution. The reaction mixture was stirred for 18 hours, whilst slowly warming to room temperature. Aqueous Na2S2O5 solution (0.5 M, 1 mL) was added and the reaction mixture stirred for 1 hour. The reaction mixture was made alkaline with NaOH (7 M) and extracted with DCM (2 x 30 mL). The combined organics were washed with brine, dried over MgSO4, filtered and concentrated in vacuo. The residue was purified by FCC on a Biotage Isolera using Biotage SNAP 10g Si cartridge eluting with gradient elution 0-35% MeOH in DCM to afford the title compound (35 mg, 55% yield) as a colourless semi-solid. Analytical data of the products was consistent with literature and/or previous samples synthesised above.

Click to access c7ob02578e1.pdf

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Mechanism of action

Arimoclomol is believed to function by stimulating a normal cellular protein repair pathway through the activation of molecular chaperones. Since damaged proteins, called aggregates, are thought to play a role in many diseases, CytRx believes that arimoclomol could treat a broad range of diseases.

Arimoclomol activates the heat shock response.^[4]^[5]^[6]^[7]^[8]^[9] It is believed to act at Hsp70.^[10]

History

Arimoclomol has been shown to extend life in an animal model of ALS^[11] and was well tolerated in healthy human volunteers in a Phase I study. CytRx is currently conducting a Phase II clinical trial.^[12]

Arimoclomol also has been shown to be an effective treatment in an animal model of Spinal Bulbar Muscular Atrophy (SBMA, also known as Kennedy’s Disease).^[13]

Arimoclomol was discovered by Hungarian researchers, as a drug candidate to treat insulin resistance^[14]^[15] and diabetic complications such as retinopathy, neuropathy and nephropathy. Later, the compound, along with other small molecules, was screened for further development by Hungarian firm Biorex, which was sold to CytRx Corporation, who developed it toward a different direction from 2003.

References

^ “CytRx Sells Molecular Chaperone Assets to Orphazyme in Deal Worth $120M | GEN Genetic Engineering & Biotechnology News – Biotech from Bench to Business | GEN”. GEN. 17 May 2011.
^ “European Medicines Agency – – EU/3/14/1376”. http://www.ema.europa.eu. Archived from the original on 2017-07-28. Retrieved 2022-02-15.
^ “Search Orphan Drug Designations and Approvals”. http://www.accessdata.fda.gov.
^ Kalmar B, Greensmith L (2009). “Activation of the heat shock response in a primary cellular model of motoneuron neurodegeneration-evidence for neuroprotective and neurotoxic effects”. Cell. Mol. Biol. Lett. 14 (2): 319–35. doi:10.2478/s11658-009-0002-8. PMC 6275696. PMID 19183864.
^ Kieran D, Kalmar B, Dick JR, Riddoch-Contreras J, Burnstock G, Greensmith L (April 2004). “Treatment with arimoclomol, a coinducer of heat shock proteins, delays disease progression in ALS mice”. Nat. Med. 10 (4): 402–5. doi:10.1038/nm1021. PMID 15034571. S2CID 2311751.
^ Kalmar B, Greensmith L, Malcangio M, McMahon SB, Csermely P, Burnstock G (December 2003). “The effect of treatment with BRX-220, a co-inducer of heat shock proteins, on sensory fibers of the rat following peripheral nerve injury”. Exp. Neurol. 184 (2): 636–47. doi:10.1016/S0014-4886(03)00343-1. PMID 14769355. S2CID 5316222.
^ Rakonczay Z, Iványi B, Varga I, et al. (June 2002). “Nontoxic heat shock protein coinducer BRX-220 protects against acute pancreatitis in rats”. Free Radic. Biol. Med. 32 (12): 1283–92. doi:10.1016/S0891-5849(02)00833-X. PMID 12057766.
^ Kalmar B, Burnstock G, Vrbová G, Urbanics R, Csermely P, Greensmith L (July 2002). “Upregulation of heat shock proteins rescues motoneurones from axotomy-induced cell death in neonatal rats”. Exp. Neurol. 176 (1): 87–97. doi:10.1006/exnr.2002.7945. PMID 12093085. S2CID 16071543.
^ Benn SC, Brown RH (April 2004). “Putting the heat on ALS”. Nat. Med. 10 (4): 345–7. doi:10.1038/nm0404-345. PMID 15057226. S2CID 11434434.
^ Brown IR (October 2007). “Heat shock proteins and protection of the nervous system”. Ann. N. Y. Acad. Sci. 1113 (1): 147–58. Bibcode:2007NYASA1113..147B. doi:10.1196/annals.1391.032. PMID 17656567. S2CID 36782230.
^ Kalmar B, Novoselov S, Gray A, Cheetham ME, Margulis B, Greensmith L (October 2008). “Late stage treatment with arimoclomol delays disease progression and prevents protein aggregation in the SOD1 mouse model of ALS”. J. Neurochem. 107 (2): 339–50. doi:10.1111/j.1471-4159.2008.05595.x. PMID 18673445.
^ “Phase II/III Randomized, Placebo-Controlled Trial of Arimoclomol in SOD1 Positive Familial Amyotrophic Lateral Sclerosis – Full Text View – ClinicalTrials.gov”. Archived from the original on 11 May 2009. Retrieved 2009-05-18.
^ Malik B, Nirmalananthan N, Gray A, La Spada A, Hanna M, Greensmith L (2013). “Co-induction of the heat shock response ameliorates disease progression in a mouse model of human spinal and bulbar muscular atrophy: implications for therapy”. Brain. 136 (3): 926–943. doi:10.1093/brain/aws343. PMC 3624668. PMID 23393146.
^ Kürthy M, Mogyorósi T, Nagy K, et al. (June 2002). “Effect of BRX-220 against peripheral neuropathy and insulin resistance in diabetic rat models”. Ann. N. Y. Acad. Sci. 967 (1): 482–9. Bibcode:2002NYASA.967..482K. doi:10.1111/j.1749-6632.2002.tb04306.x. PMID 12079878. S2CID 19585837.
^ Seböková E, Kürthy M, Mogyorosi T, et al. (June 2002). “Comparison of the extrapancreatic action of BRX-220 and pioglitazone in the high-fat diet-induced insulin resistance”. Ann. N. Y. Acad. Sci. 967 (1): 424–30. Bibcode:2002NYASA.967..424S. doi:10.1111/j.1749-6632.2002.tb04298.x. PMID 12079870. S2CID 23338560.

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Clinical data

Routes of administration	Oral
ATC code	N07XX17 (WHO)
Legal status
Legal status	Investigational
Identifiers
show IUPAC name
CAS Number	289893-25-0
PubChem CID	208924
ChemSpider	21106260
UNII	EUT3557RT5
KEGG	D11374
ChEMBL	ChEMBL2107726
CompTox Dashboard (EPA)	DTXSID5057701
Chemical and physical data
Formula	C₁₄H₂₀ClN₃O₃
Molar mass	313.78 g·mol⁻¹
3D model (JSmol)	Interactive image
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/////////ARIMOCLOMOL, アリモクロモル , BRX 220, INN 8300, Arimoclomol maleate, phase III, clinical, Orphazyme , Niemann-Pick disease type C, phase II, amyotrophic lateral sclerosis, (ALS)

NIDUFEXOR

March 11, 2020 11:24 am / Leave a comment

Nidufexor Chemical Structure

NIDUFEXOR

LMB763

4-[[benzyl-(8-chloro-1-methyl-4H-chromeno[4,3-c]pyrazole-3-carbonyl)amino]methyl]benzoic acid

Nidufexor is a farnesoid X receptor (FXR) agonist.

Molecular Weight	487.93
Formula	C₂₇H₂₂ClN₃O₄
CAS No.	1773489-72-7

PHASE 2 Treatment of Liver and Biliary Tract Disorders,
Agents for Diabetic Nephropathy, NOVARTIS

Nidufexor

1773489-72-7, LMB-763, UNII-CJ1PL0TE6J, CJ1PL0TE6J, BCP28929, EX-A1854

Nidufexor pound LMB-763 pound(c)

ZINC584641402

4-((N-benzyl-8-chloro-1-methyl-1,4-dihydrochromeno[4,3-c]pyrazole-3-carboxamido)methyl)benzoic acid

HY-109096

CS-0039398

https://pubs.acs.org/doi/pdf/10.1021/acs.jmedchem.9b01621

1 (7.6 g, 89% yield) as a white solid. Melting point: 232.6 °C.

1 H NMR (400 MHz, DMSO): δ 12.93 (s, 1H), 7.96−7.85 (m, 2H), 7.71 (dd, J = 7.1, 2.5 Hz, 1H), 7.42−7.20 (m, 8H), 7.06 (dd, J = 8.7, 1.9 Hz, 1H), 5.45 (d, J = 3.9 Hz, 2H), 5.25 (d, J = 9.2 Hz, 2H), 4.58 (d, J = 12.1 Hz, 2H), 4.12 (d, J = 16.6 Hz, 3H).

13C NMR (101 MHz, DMSO-d6): δ 167.07, 162.21, 151.98, 142.65, 139.18, 132.20, 132.67, 129.70, 129.50, 129.50, 128.53, 128.53, 127.43, 127.43, 127.43, 127.43, 127.43, 125.53, 122.24, 119.0, 117.09, 116.64, 64.51, 50.68, 48.24. LC-MS m/z: 488.2/490.2 (M +H)+ ; chlorine pattern; method 3; RT = 1.41 min.

Elemental Analysis calcd for C27H22ClN3O4: C 66.46, H 4.54, N 8.61; found: C 66.43, H 4.56, N 8.62.

TRIS Salt Formation. Methanol (400 mL) was added to a mixture of 1 (4.0 g, 8.2 mmol) and 2-amino-2-hydroxymethylpropane-1,3-diol (TRIS, 1.0 g, 8.2 mmol). The mixture was heated to 70 °C for 0.5 h. After cooling to room temperature, the solvent was removed in vacuum. The residue was sonicated in dichloromethane (10 mL) and concentrated again. The resulting white solid was dried under vacuum overnight. The crude material was crystallized by slurring the solid residue in a 4:1 mixture of acetonitrile and methanol (5 mL). The mixture was stirred at room temperature for 24 h to give 4-((N-benzyl-8-chloro-1-methyl-1,4-dihydrochromeno- [4,3-c]pyrazole-3-carboxamido)methyl)benzoic acid TRIS salt as a white salt (3.7 g, 73% yield). Melting point: 195.6 °C. 1 H NMR (400 MHz, DMSO): δ 7.92−7.80 (m, 2H), 7.78−7.64 (m, 1H), 7.41− 7.19 (m, 8H), 7.13−7.00 (m, 1H), 5.44 (s, 2H), 5.25−5.14 (m, 2H), 4.61−4.48 (m, 2H), 4.18−4.03 (m, 3H), 3.39 (s, 7H). TRIS OH masked by water peak. LC-MS m/z: 488.0/490.0 (M+H)+ ; chlorine pattern, method 3. RT = 1.58 min. Elemental Analysis calc for C31H33ClN4O7: C 61.00, H 5.36, N 9.15; found: C 60.84, H 5.34, N 9.13.

https://pubs.acs.org/doi/suppl/10.1021/acs.jmedchem.9b01621/suppl_file/jm9b01621_si_001.pdf

Patent

WO 2015069666

https://patentscope.wipo.int/search/en/detail.jsf?docId=WO2015069666&tab=PCTDESCRIPTION

///////NIDUFEXOR, LMB 763, Phase II, PHASE 2, Liver and Biliary Tract Disorders, Diabetic Nephropathy, NOVARTIS

CN1C(C2=CC(Cl)=CC=C2OC3)=C3C(C(N(CC4=CC=CC=C4)CC5=CC=C(C(O)=O)C=C5)=O)=N1

LYS 228

February 24, 2020 10:24 am / Leave a comment

2D chemical structure of 1810051-96-7

LYS228

BOS-228
LYS-228

Molecular Formula, C16-H18-N6-O10-S2

Molecular Weight, 518.4783

(3S,4R)-3-((Z)-2-(2-Ammoniothiazol-4-yl)-2-((1-carboxycyclopropoxy)imino)acetamido)-2-oxo-4-((2-oxooxazolidin-3-yl)methyl)azetidine-1-sulfonate

RN: 1810051-96-7
UNII: 29H7N9XI1B

UNII-005B24W9YP

005B24W9YP

Lys-228 trihydrate

2091840-43-4

Yclopropanecarboxylic acid, 1-(((Z)-(1-(2-amino-4-thiazolyl)-2-oxo-2-(((3S,4R)-2-oxo-4-((2-oxo-3-oxazolidinyl)methyl)-1-sulfo-3-azetidinyl)amino)ethylidene)amino)oxy)-, hydrate (1:3)

1-[(Z)-[1-(2-amino-1,3-thiazol-4-yl)-2-oxo-2-[[(3S,4R)-2-oxo-4-[(2-oxo-1,3-oxazolidin-3-yl)methyl]-1-sulfoazetidin-3-yl]amino]ethylidene]amino]oxycyclopropane-1-carboxylic acid;trihydrate

BOS-228 (LYS-228) is a monobactam discovered at Novartis and currently in phase II clinical development at Boston Pharmaceuticals for the treatment of complicated urinary tract infection and complicated intraabdominal infections in adult patients.

The compound has been granted fast track and Qualified Infectious Disease Product (QIDP) designation from the FDA.

In October 2018, Novartis licensed to Boston Pharmaceuticals worldwide rights to the product.

Paper

https://pubs.acs.org/doi/10.1021/acs.oprd.9b00330

Patent

US 20150266867

PATENT

WO 2017050218

https://patentscope.wipo.int/search/en/detail.jsf?docId=WO2017050218&tab=FULLTEXT

Compound X: 1- ( ( (Z) – (1- (2-aminothiazol-4-yl) -2-oxo-2- ( ( (3S, 4R) -2-oxo-4- ( (2-oxooxazolidin-3-yl) methyl) -1-sulfoazetidin-3-yl) amino) ethylidene) amino) oxy) cyclopropanecarboxylic acid.

[0126]

Step 1: Benzhydryl 1- ( ( (Z) – (1- (2- ( (tert-butoxycarbonyl) amino) thiazol-4-yl) -2-oxo-2- ( ( (3S, 4R) -2-oxo-4- ( (2-oxooxazolidin-3-yl) methyl) azetidin-3-yl) amino) ethylidene) amino) oxy) cyclopropanecarboxylate. To a solution of (Z) -2- ( (1- ( (benzhydryloxy) carbonyl) cyclopropoxy) imino) -2- (2- ( (tert-butoxycarbonyl) amino) thiazol-4-yl) acetic acid (854 mg, 1.59 mmol) prepared according to published patent application US2011/0190254, Intermediate B (324 mg, 1.75 mmol) and HATU (785 mg, 2.07 mmol) in DMF (7.9 mL) , DIPEA was added (832 μL, 4.77 mmol) . After 1 h of stirring, it was poured into water and extracted with EtOAc. Brine was added to the aqueous layer, and it was further extracted with ethyl acetate (EtOAc) (3x) . The combined organic layers were dried over Na ₂SO ₄ and concentrated in vacuo. The crude residue was purified via silica gel chromatography (0-10％MeOH-DCM) to afford the title compound (1.09 g, 97％) as a beige foam. LCMS: R _t ＝ 0.97 min, m/z ＝705.3 (M+1) Method 2m_acidic.

[0127]

Instead of HATU, a variety of other coupling reagents can be used, such as any of the typical carbodiimides, or CDMT (2-chloro-4, 6-dimethoxy-1, 3, 5-triazine) and N-methylmorpholine to form the amide bond generated in Step 1.

[0128]

Step 2: (3S, 4R) -3- ( (Z) -2- ( (1- ( (benzhydryloxy) carbonyl) cyclopropoxy) imino) -2- (2- ( (tert-butoxycarbonyl) amino) thiazol-4-yl) acetamido) -2-oxo-4- ( (2-oxooxazolidin-3-yl) methyl) azetidine-1-sulfonic acid. Benzhydryl 1- ( ( (Z) – (1- (2- ( (tert-butoxycarbonyl) amino) thiazol-4-yl) -2-oxo-2- ( ( (3S, 4R) -2-oxo-4- ( (2-oxooxazolidin-3-yl) methyl) azetidin-3-yl) amino) ethylidene) amino) oxy) cyclopropanecarboxylate (1.00 g, 1.42 mmol) in DMF (7.0 mL) at 0 ℃ was treated with SO ₃·DMF (448 mg, 2.84 mmol) . After 2 h of stirring at rt, the solution was poured into ice-cold brine and extracted with EtOAc (3x) . The combined organic layers were dried over Na ₂SO ₄ and concentrated in vacuo, affording the title compound (assumed quantitative) as a white solid. LCMS: Rt ＝0.90 min, m/z ＝ 785.2 (M+1) Method 2m_acidic.

[0129]

Step 3: 1- ( ( (Z) – (1- (2-aminothiazol-4-yl) -2-oxo-2- ( ( (3S, 4R) -2-oxo-4- ( (2-oxooxazolidin-3-yl) methyl) -1-sulfoazetidin-3-yl) amino) ethylidene) amino) oxy) cyclopropanecarboxylic acid.

[0130]

[0131]

To a solution of (3S, 4R) -3- ( (Z) -2- ( (1- ( (benzhydryloxy) carbonyl) cyclopropoxy) imino) -2- (2- ( (tert-butoxycarbonyl) amino) thiazol-4-yl) acetamido) -2-oxo-4- ( (2-oxooxazolidin-3-yl) methyl) azetidine-1-sulfonic acid (1.10 g, 1.40 mmol) in DCM (1.5 mL) at 0℃, TFA (5.39 mL, 70.0 mmol) was added, and after 10 minutes, the ice bath was removed. Additional TFA (3.24 mL, 42.0 mmol) was added after 1 hr at rt and the solution was diluted with DCM and concentrated in vacuo after an additional 30 min. Optionally, anisole may be added to the TFA reaction to help reduce by-product formation, which may increase the yield of desired product in this step. The crude residue was purified by reverse phase prep HPLC (XSelect CSH, 30 x 100 mm, 5 μm, C18 column； ACN-water with 0.1％formic acid modifier, 60 mL/min) , affording the title compound (178 mg, 23％) as a white powder. LCMS: R _t ＝ 0.30 min, m/z ＝ 518.9 (M+1) Method 2m_acidic； ¹H NMR (400 MHz, DMSO-d ₆) δ 9.27 (d, J ＝ 9.0 Hz, 1H) 6.92 (s, 1H) 5.23 (dd, J ＝ 9.1, 5.7 Hz, 1H) 4.12-4.23 (m, 3H) 3.72-3.62 (m, 2H assumed； obscured by water) 3.61-3.52 (m, 1H assumed； obscured by water) 3.26 (dd, J ＝ 14.5, 5.9 Hz, 1H) 1.36 (s, 4H) . ¹H NMR (400 MHz, D ₂O) δ 7.23 (s, 1H) , 5.48 (d, J ＝ 5.8 Hz, 1H) , 4.71-4.65 (m, 1H) , 4.44 (t, J ＝ 8.2 Hz, 2H) , 3.89-3.73 (m, 3H) , 3.54 (dd, J ＝ 14.9, 4.9 Hz, 1H) , 1.65-1.56 (m, 2H) , 1.56-1.46 (m, 2H) . The product of this process is amorphous. Compound X can be crystallized from acetone, ethanol, citrate buffer at pH 3 (50 mM) , or acetate buffer at pH 4.5 (50 mM) , in addition to solvents discussed below.

PAPER

Bioorganic & Medicinal Chemistry Letters (2018), 28(4), 748-755.

https://www.sciencedirect.com/science/article/pii/S0960894X18300064

PATENT

WO 2019026004

https://patentscope.wipo.int/search/en/detail.jsf?docId=WO2019026004&tab=PCTDESCRIPTION

Over the past several decades, the frequency of antimicrobial resistance and its association with serious infectious diseases have increased at alarming rates. The increasing prevalence of resistance among nosocomial pathogens is particularly disconcerting. Of the over 2 million (hospital-acquired) infections occurring each year in the United States, 50 to 60% are caused by antimicrobial-resistant strains of bacteria. The high rate of resistance to commonly used antibacterial agents increases the morbidity, mortality, and costs associated with nosocomial infections. In the United States, nosocomial infections are thought to contribute to or cause more than 77,000 deaths per year and cost approximately $5 to $10 billion annually.

Important causes of Gram-negative resistance include extended-spectrum 13- lactamases (ESBLs), serine carbapenemases (KPCs) and metallo-13-lactamases (for example NDM-1 ) in Klebsiella pneumoniae, Escherichia coli, and Proteus mirabilis, high-level third-generation cephalosporin (AmpC) 13-lactamase resistance among Enterobacter species and Citrobacter freundii, and multidrug-resistance genes observed in Pseudomonas, Acinetobacter, and Stenotrophomonas. The problem of antibacterial resistance is compounded by the existence of bacterial strains resistant to multiple antibacterials. For example, Klebsiella pneumonia harboring NDM-1 metallo-13- lactamase carries frequently additional serine-13-lactamases on the same plasmid that carries the NDM-1 .

Thus there is a need for new antibacterials, particularly antibacterial compounds that are effective against existing drug-resistant microbes, or are less susceptible to development of new bacterial resistance. Monobactam antibiotic, which is referred to herein as Compound X, is primarily effective against Gram-negative bacteria, including strains that show resistance to other monobactams.

The present invention relates to a process for the preparation of monobactam antibiotic Compound X and intermediates thereof.

More particularly, the present invention relates to a process for the preparation of Compound X

Compound X

also referred to as 1 -(((Z)-(1 -(2-aminothiazol-4-yl)-2-oxo-2-(((3S,4R)-2-oxo-4-((2-oxooxazolidin-3-yl)methyl)-1 -sulfoazetidin-3-yl)amino)ethylidene)amino)oxy)cyclopropanecarboxylic acid, or a salt thereof, or a solvate including hydrate thereof.

Patent application number PCT/US2015/02201 1 describes certain monobactam antibiotics. Compound X may be prepared using the method disclosed in PCT/US2015/02201 1 , in particular example 22, and in PCT/CN2016/099482.

A drawback from these processes is that they exhibit a large number of process steps and intermediate nitrogen protection/deprotection steps, reducing the overall yield and efficiency. Furthermore, these processes require several chromatographic purification steps to be carried out in course of the processes. We have found that the preparation of Compound X, as previously prepared on a manufacturing scale, possesses a number of disadvantages, in particular poor handling characteristics.

It would thus be beneficial to develop alternative or improved processes for the production of Compound X that do not suffer from some or all of these disadvantages.

Compound ^x Compound ^x

Scheme 1

Preparation of Compound X from Intermediates 22 and 2A

Scheme 3

Examples

The Following examples are merely illustrative of the present disclosure and they should not be considered as limiting the scope of the disclosure in any way, as these examples and other equivalents thereof will become apparent to those skilled in the art in the light of the present disclosure, and the accompanying claims.

Synthesis of Compound 8 (R = benzyl)

1 .50kg oxazolidin-2-one (7b) was charged into the reactor. 7.50kg THF was charged and the stirring started. The mixture was cooled to 10~20°C. 2.18kg potassium fert-butoxide was charged intol 2.00kg THF and stirred to dissolve.

The potassium fert-butoxide solution was added dropwise into the reactor while maintaining the temperature at 10-20 °C. The reaction was stirred for 1 ~2hrs at 10-20 °C after the addition. The solution of 2.36kg methyl-2-chloroacetate (7a) in 3.00kg of THF was added to the reactor while maintaining the temperature at 10-20 °C. The reaction mixture was stirred for 16-18 h at 20-25 °C. The IPC (in process control) showed completion of the reaction. The mixture was centrifuged and the wet cake was washed with 7.50kg THF. The filtrate was concentrated and the crude 7 was provided as reddish brown liquid, which was used for the next step without further purification,

¹H NMR (400 MHz, CHLOROFORM- /) δ ppm 3.65 – 3.71 (m, 2 H) 3.74 (s, 3 H) 4.02 (s, 2 H) 4.34 – 4.45 (m, 2 H).

The dried reactor was exchanged with N2 three times. 3.71 kg LiHMDS solution in THF/Hep (1 M) and 1 .30kg THF were charged under nitrogen protection. The stirring was started and the solution was cooled to -70—60 °C. The solution of 0.71 kg benzyl acetate (6) in 5.20 kg THF was added dropwisely at -70— 60 °C, and the resulted mixture was stirred for 1 -1 .5 h after the addition. The solution of 0.65kg 7 in 3.90kg THF was added dropwise while maintaining the temperature at -70—60 °C, then stirred for 30-40 minutes. The reaction mixture was warmed to 20-25 °C and stirring was continued for 0.5-1 .0 h. IPC showed 6 was less than1 .0% (Otherwise, continue the reaction till IPC passes). The reaction mixture was poured into 13.65 kg aqueous citric acid below 10 °C. The mixture was stirred for 15-20 minutes after the addition. Phases were separated and the organic layer was collected. The aqueous layer was extracted with EA (6.50kg ^* 2). The organic layer was combined, washed by 6.50 kg 28% NaCI solution and dried with 0.65

kg anhydrous MgSC . The mixture was filtered and the wet cake was washed with 1 .30kg EA. The filtrate was concentrated under vacuum to provide crude 8. The crude 8 was stirred in 2.60 kg MTBE at 20-25 °C for 1 -1 .5 h. The mixture was cooled to 0-10 °C and stirred for 1 .5-2.0 h and filtered. The filter cake was washed with 0.65kg pre-cooled MTBE and dried under vacuum (<-0.096Mpa) at 20-25 °C for 12~16hrs till a constant weight to give 513 g of 8 as a white solid, Yield: 45%, HPLC purity 96.4%,¹ H NMR (400 MHz, CHLOROFORM-c δ ppm 3.48 – 3.55 (m, 1 H) 3.56 – 3.63 (m, 2 H) 3.66 – 3.74 (m, 1 H) 4.17 – 4.26 (m, 2 H) 4.31 – 4.44 (m, 2H) 5.12 – 5.24 (m, 2 H) 7.30 – 7.44 (m, 5 H).

Synthesis of Compound 9 (R = benzyl)

The dried reactor was charged with 3.75kg HOAc and 1 .50 kg 8. The stirring was started and the reaction mixture was cooled to 0-5 °C. 3.53kg aqueous NaN02 was added dropwise at 0-10 °C, and the reaction mixture was stirred for 15-30 minutes after the addition. IPC showed 8 was less than 0.2%. The reaction mixture was treated with 7.50kg EA and 7.50 kg water. Phases were separated and the organic layer was collected. The aqueous layer was extracted with EA (7.50kg ^* 2). The organic layers were combined, washed with 7.50 kg 28% NaCI solution, and concentrated under vacuum to provide crude 9. The crude 9 was slurried with 5.25 kg water at 10-20 °C for 3~4hrs, and filtered. The wet cake was washed with 1 .50kg water. The solid was dried under vacuum (<-0.096 Mpa) at 45-50 °C for 5-6 h till a constant weight to give 1 .44 Kg of 9, yield: 86.9%, HPLC purity 92.9%,¹H NMR (400 MHz, CHLOROFORM- /) δ ppm 3.60 – 3.76 (m, 2 H) 4.44 (t, J=8.07 Hz, 2 H) 4.60 (s, 2 H) 5.25 – 5.41 (m, 2 H) 7.30 – 7.43 (m, 5 H) 1 1 .62 (br s, 1 H).

Synthesis of Compound 9a (R = benzyl)

The dried reactor was charged with 0.58 kg Zn, 4.72kg (Βο Ο, 6.00 kg water, 1 .20 kg NH₄CI and 6.00kg THF. The reaction mixture was stirred and heated to 50-55 °C. The solution of 0.60 kg 9 in 4.20kg THF

was added dropwisely while maintaining the temperature at 50-55 °C. The reaction mixture was stirred for 0.5-1 .Ohrs after the addition. IPC showed 9 was less than 0.1 %. The reaction mixture was treated withl .50 kg ethyl acetate and stirred for 15-20 minutes. Phase was separated and the water layer was extracted by1 .50 kg ethyl acetate. The organic layers were combined, washed with 6.00 kg 28% NaCI solution and concentrated under vacuum to provide crude 9a. The crude 9a was stirred with 3.60kg^*2 n-heptane to remove excess (Βο Ο. The residue was purified by silica gel chromatography column eluted with ethyl acetate: Heptane= 1 :1 to provide crude 9a solution. The solution was concentrated under reduced pressure to obtain crude 9a. The crude 9a was slurried with 1 .80 kg MTBE for 2.0-3. Ohrs, filtered, and the wet cake was washed with MTBE. The solid was dried under vacuum (<-0.096 Mpa) at 50-55 °C for 16-18 h till a constant weight to give 392 g of 9a as a white solid, Yield: 51 %, HPLC purity 98.1 %,¹H NMR (400 MHz, DMSO-cfe) δ ppm 1.17 – 1 .57 (m, 9 H) 3.39 – 3.61 (m, 2 H) 4.20 – 4.45 (m, 3 H) 5.10 – 5.32 (m, 3 H) 5.75 (s, 1 H) 7.38 (br s, 5 H) 7.75 – 7.99 (m, 1 H).

Synthesis of compound (VII) (R = benzyl, X = CI)

9a VII

The dried reactor was charged with 13.0kg HCI in IPA and the stirring was started. 1 .33 kg 9a was charged in portions at 20-25 °C. The mixture was stirred at 20-25 °C for 3-4 h. IPC showed 9a was less than 0.1 %. The reaction solution was concentrated under vacuum 40-45 °C. The residue was treated with 21 .58kg MTBE at 20-25 °C for 3-4 h. The mixture was filtered and the wet cake was washed with 2.60kg MTBE. The solid was dried under vacuum (<-0.096 Mpa) at 45-50 °C for 5-6 h till a constant weight to give 1 .045 Kg of compound VII (R = benzyl, X = CI) as a yellow solid, Yield: 93.7%, HPLC purity 99.2%,¹ H NMR (400 MHz, DMSO-cfe) δ ppm 3.16 – 3.74 (m, 3 H) 4.10 – 4.35 (m, 4 H) 5.09 – 5.39 (m, 2 H) 7.27 – 7.60 (m, 5 H) 8.72 (br s, 2 H).

Synthesis of compound (Vile) (R = benzyl)

VII Vile

To an autoclave (3L) were added VII (R = benzyl, X = CI) (100 g, 304.2 mmol, 1 .0 equiv.), DCM (2650 g, 26.5 equiv., w/w) and (S-BINAP)RuCl2 (2.4 g, 3.04 mmol, 0.01 equiv.), successively. Air in the autoclave was replaced with N2 5 times. N2 in the autoclave was was replaced with H2 5 times. The solution was stirred with 250-260 r/min and H2 (2.1 ±0.1 MPa) at 40±5°C for 24 h. The reaction mixture was filtered, and the filter cake was washed with DCM (400 g, 4.0 equiv., w/w). The filter cake was slurried with IPA (785 g, 7.85 equiv., w/w) and H2O (40 g, 0.4 equiv., w/w) overnight (18-20 h). The mixture was filtered. The filter cake was washed with IPA (200 g, 2.0 equiv., w/w) and dried at 45±5°C overnight (18-20 h). Vile (R = benzyl) was obtained as off-white solid, 80.4 g, 79.9% yield, 95.5% purity, 97.6% de, >99.5% ee. ¹H NMR (400 MHz, DMSO-cfe) δ ppm 3.34-3.38 (m, 2 H) 3.50-3.52 (m, 1 H) 3.60-3.62 (m, 1 H) 4.18-4.24 (m, 4 H) 5.23 (s, 2H) 6.16 (s, 1 H) 7.32 (m, 5H) 8.74 (s, 1 H).

Alternative synthesis of compound 9a (R = benzyl)

Mg(OtBu)2

To a flask was added 5a (1 .88 g, 12.93 mmol), THF (40 mL), and CDI (2.20 g, 13.58 mmol) at 25 °C. The mixture was stirred for 3 h. To the reaction mixture was added 5b (2.00 g, 6.47 mmol), and Mg(OfBu)2 (2.21 g, 12.93 mmol). The reaction mixture was stirred at 25 °C for 24 h. The reaction mixture was concentrated under vacuum to remove most of the THF solvent. To the concentrated solution was added MTBE (40 mL), followed by addition of an aqueous solution of HCI (1 M, 60mL) to adjust to pH = 2-3. Two phases were separated, and the water phase was extracted with MTBE (20 mL). The combined organic phase was washed with aqueous NaHCC (5%, 50 mL) and brine (20%, 40 mL). The organic phase was concentrated to a weight of -19 g, and a lot of white solid was obtained in the concentration process. The suspension was cooled to 0 °C, and filtered. The filter cake was washed with cold MTBE (5 mL) and dried under vacuum to obtain product 9a (1 .6g, 63% yield).

Synthesis of compound (Vile) (R = benzyl, PG = Cbz)

Vile Vile

To a flask (5 L) were added Vile (R = benzyl) (140 g, 423.2 mmol, LOequiv.), H₂0 (1273 g, 9.09 equiv., w/w) and toluene (2206 g, 15.76 equiv., w/w). The solution was stirred and cooled to 0-5 °C with ice bath. Then NaHCOa (78.4 g, 933 mmol, 2.22 equiv.) was added and CbzCI (89.6 g, 527 mmol, 1 .24 equiv.) was dropped into the stirring solution, respectively. The solution was stirred at 30±5 °C overnight (18-20 h). Heptane (3612 g, 25.8 equiv., w/w) was added dropwise to the stirring solution over 1 h at 20-30 °C. The mixture was filtered. The filter cake was washed with heptane (280 g, 2.00 equiv., w/w) and MTBE (377 g, 2.69 equiv., w/w), respectively. The filter cake was dried at 45±5°C overnight (18-20 h). Vile (R = benzyl, PG = Cbz) was obtained as an off-white solid, 169.4 g, 93% yield, 96.7% purity, 98% de, >99.5% ee, ¹ H NMR (400 MHz, DMSO-cfe) δ ppm 3.23-3.24 (m, 1 H) 3.30 (m, 1 H) 3.51 -3.55 (m, 2 H) 3.99 (s, 1 H) 4.17-4.21 (m, 3 H) 5.02-5.03 (m, 2H) 5.12 (s, 2H) 5.46-5.48 (d, 1 H) 7.33-7.36 (m, 10H) 7.75-7.73 (d, 1 H).

Synthesis of compound (IV) (PG = Cbz)

Vile IV

Vile (R = benzyl) (220 g, 513.5 mmol, 1 .0 equiv.) was dissolved in THF (1464g, 6.65 equiv., w/w). The solution was filtered. The filter cake was washed with THF (488g, 2.22 equiv., w/w). The filtrate (Vile) was collected. To an autoclave (3L) were added the filtrate (Vile). The reactor was cooled down to -75 – -65 °C with dry-ice/EtOH bath, and bubbled with NH3 for not less than 4 h. Then the solution was stirred at 25±5 °C with NH3 (0.5-0.6 MPa) for 24 h. The autoclave was deflated to release NH3. The reaction solution was concentrated with a rotary evaporator to remove THF until the residue was around 440 g. The residue was slurried with EA (2200 g, 10 equiv., w/w) at 70±2 °C, then cooled to 25±5 °C and stirred for 16-18 h. The mixture was filtered. The filter cake was washed with EA (440 g). The filter cake was slurried with EA (1320 g, 6.00 equiv. w/w), and the temperature was raised to 70±2 °C, then cooled to 25±5 °C and stirred for 16-20 h. The mixture was filtered. The filter cake was washed with EA, and dried at 50±5 °C overnight (18-20 h). IV (PG = Cbz) was obtained as off-white solid, 141 g, 81 .5% yield, 99.1 % purity, >99.5% assay, ¹H NMR (400 MHz, DMSO-cfe) δ ppm 3.12 – 3.23 (m, 2 H) 3.31 (br s, 1 H) 3.56 (t, J=8.01 Hz, 2 H) 3.88 (quin, J=6.02 Hz, 1 H) 3.93 – 4.03 (m, 1 H) 4.20 (t, J=8.01 Hz, 2 H) 5.02 (s, 2 H) 5.27 (d, J=5.87 Hz, 1 H) 7.12 (s, 1 H) 7.22 – 7.45 (m, 5 H).

Synthesis of compound (III) (PG = Cbz, LG = S0₂CH₃)

IV III

To a flask was added IV (PG = Cbz) (14.00 g, 41 .50 mmol, 1 .00 equiv), and dry 1 , 2-dimethoxyethane (300 mL) under N2. The mixture was stirred at -5°C ~ 0°C for 1 h to obtain a good suspension. MsCI (7.89 g, 68.89 mmol, 5.33 mL, 1 .66 eq) in 1 , 2-dimethoxyethane (20.00 mL) was added dropwise during 30 min, and Et3N (12.60 g, 124.50 mmol, 17.26 mL, 3.00 eq) in 1 , 2-dimethoxyethane (20.00 mL) was added dropwise during 30 min side to side. The reaction mixture was stirred for additional 5 min at -5°C ~ 0°C, and was quenched with water (6 mL). The reaction mixture was concentrated to remove DME. The solid was slurried in water (250 mL) and MTBE (125 mL) for 1 h. The solid was collected by filtration, and then slurried in water (250 mL) for 1 hr. The solid was collected by filtration, and washed with water (25 mL) to give white solid. The solid was slurried in EA (150 mL) and dried in vacuum at 60°C for 24 h to give III (PG = Cbz, LG = SO2CH3) (15.00 g, 36.1 1 mmol, 87.01 % yield), ¹H NMR (400 MHz, DMSO-cfe) δ ppm 3.17 (s, 3 H) 3.26 (br d, J=15.04 Hz, 1 H) 3.47 – 3.57 (m, 1 H) 3.64 (br d, J=6.36 Hz, 2 H) 4.22 (br dd, J=17.79, 8.50 Hz, 2 H) 4.50 (br s, 1 H) 4.95 – 5.17 (m, 3 H) 7.21 – 7.56 (m, 5H) 7.43 (s, 1 H) 7.63 – 7.89 (m, 2 H).

Synthesis of compound II (PG = Cbz, LG = SO2CH3, M⁺ = NBu₄⁺)

O OMs o CISO3H, 2-picoline – ° O ?yO

HN Bu₄NHS0₄^< NHCbz

“Cbz

III II

To a flask was added 2-picoline (1 1 .50 g, 12.23 mL) and DMF (10 mL). The solution was cooled to 5 ^SC, followed by slow addition of chlorosulfonic acid (7.20 g, 4.14 mL). The temperature was increased to 20 ^SC. Ill (PG = Cbz, LG = SO2CH3) (5.13 g, 12.35 mmol) was added to the reaction mixture. The reaction mixture was heated to 42 ^SC for 18h. IPC (in process control) showed complete conversion of starting material. The reaction was cooled to 20 ^SC and dropwise added to a solution of tetrabutylammonium hydrogen sulfate (4.6 g, 13.6 mmol) in the mixed solvents of dichloromethane (100 mL) and water (100 mL) at 5^SC. The phases were separated and the water phase was extracted with dichloromethane (2^*50mL). The combined organic phase was washed with water (5^*100mL). The organic phase was concentrated to dryness and purified by column chromatography (dichloromethane/methanol = 15/1 v/v) to afford II (PG = Cbz, LG = SO2CH3, M⁺ = NBii4⁺) (8.4 g, 92.30%), ¹ H NMR (400 MHz, CHLOROFORM-c/) δ ppm 0.99 (t, J=7.34 Hz, 12 H) 1 .36 – 1 .50 (m, 8 H) 1 .54 – 1 .76 (m, 8 H) 3.15 (br d, J=8.31 Hz, 2 H) 3.21 – 3.35 (m, 8 H) 3.47 (br dd, J=14.73, 7.27 Hz, 1 H) 3.54 – 3.65 (m, 1 H) 3.67 – 3.81 (m, 2 H) 4.17 – 4.32 (m, 1 H) 4.39 – 4.62 (m, 1 H) 4.74 (br s, 1 H) 5.1 1 (s, 3 H) 5.32 – 5.50 (m, 1 H) 6.47 (br s, 1 H) 7.29 – 7.47 (m, 5 H) 8.69 – 8.94 (m, 1 H).

Synthesis of compound (IA)

A solution of II (PG = Cbz, LG = SO2CH3, M⁺ = NBu₄⁺) (4.0 g) in dichloromethane (38 mL) was pumped to tube A at rate of 2.0844 mL/min, and a solution of KHCO3 (3.0 g) in water (100 mL) was pumped to tube B at a rate of 1 .4156 mL/min side to side. These two streams were mixed in a cross-mixer then flowed to a tube coil that was placed in an oil bath at 100 °C. The residence time of the mixed stream in the coil was 2 min. The reaction mixture flowed through a back-pressure regulator that was set at ~ 7 bars, and was collected to a beaker. After completion of the collection, two phases was separated. The organic phase was concentrated to dryness. The residue was slurried in ethyl acetate (5 mL). The solid was filtered and the filter cake was dried to give IA (2.6 g, 75%),

¹H NMR (400 MHz, CHLOROFORM-c/) δ ppm 1.00 (t, J=7.27 Hz, 12 H) 1 .42 (sxt, J=7.31 Hz, 8 H) 1 .62 (quin, J=7.83 Hz, 8 H) 3.13 – 3.39 (m, 8 H) 3.54 – 3.69 (m, 2 H) 3.81 (dd, J=14.98, 2.51 Hz, 1 H) 3.96 – 4.13 (m, 1 H) 4.22 – 4.47 (m, 3 H) 4.99 – 5.23 (m, 3 H) 6.42 (br d, J=9.29 Hz, 1 H) 7.26 – 7.44 (m, 5 H).

Synthesis of compound 2A

Step 1

To a stirring solution of compound 16b (2 g, 10.14mmol, 1 .0 eq) in DMF (20 ml_) was added CS2CO3 (5.29g, 16.22 mmol, 1 .6 eq), then the resulting solution was stirred at room temperature for 10mins, then compound 16a (5.27g, 20.28mmol, 2eq) was added dropwise to the mixture for 2 minutes, then the resulting solution was stirred for another 2 hours. TLC showed the starting material was consumed completely. The mixture was added with water (60mL) and extracted with MTBE (20mL^*3). The combined organic layers were dried over anhydrous sodium sulfate and concentrated. The crude was slurried in heptane to give 1 .65 g 16 as a white solid (Yield: 57%), ¹H NMR (400 MHz, DMSO-cfe) δ ppm 7.48-7.28 (m, 10 H), 5.00-4.96 (t, J=6.0 Hz, 1 H), 3.81 (s, 3H), 3.44-3.42 (m, 2H), 2.40-2.37 (m, 2H).

Compound 16 (1 g, 2.66mmol, 1 eq) was dissolved in THF (20mL) under Nitrogen, and cooled to -40 °C. NaHMDS (1 .6mL, 2.0M THF solution, 1 .2 eq) was added dropwise. The reaction was stirred for 1 h at -40 °C. HPLC indicated the reaction was finished. The reaction was quenched with 10% Citric acid, extracted with MTBE (25 ml_ x 2). The combined organic layers were washed with brine (30 ml_), dried with Na2S04, filtered and concentrated to give 17 as a yellow solid, which was used for the next step without purification (assay yield: 65%); ¹H NMR (400 MHz, DMSO-cfe) δ ppm 7.27-7.13 (m, 10 H), 3.46 (s, 3H), 1 .21 -1 .17(dd, J=7.2, 10.4 Hz, 2H ); 1 .14-1 .1 1 (dd, J=7.2, 10.4 Hz, 2H).

Step 3

Compound 17 (100 mg) was dissolved in methanol (5 mL) and 2.0 M HCI IPAC solution (5 mL). The solution was heated at 45 °C for 3 days. HPLC indicated the reaction was finished. The reaction was cooled to room temperature and was diluted with 10 mL water. The reaction mixture was washed with MTBE (10 mL x 2), organic layer was discarded and the aqueous layer was concentrated to give compound 2A HCI (32 mg, 62% yield), ¹ H NMR (400 MHz, DMSO-cfe) δ ppm 3.80-3.44 (br, 4H), 1 .56 (s, 2H), 1 .38 (s, 2H).

Step 4

To a solution of 2A HCI (0.70 g, 4.57 mmol) in methanol (5 mL) was added triethylamine (1 .26 mL, 9.14 mmol) at room temperature. The solution was stirred for 20 min, and the solvent was removed under vacuum. To the residue was added IPAC (10 mL) leading to precipitation. The solid was filtered, and the filtrate was concentrated to provide 2A (0.50g, 94% yield) containing ca. 6 wt% Et3N-HCI.

Synthesis of Compound X from compound of formula (I), (IA)

Compound ^x

To a flask was charged 21 (1 .00 g, 68.43 wt%, 2.50 mmol) and DMF (10 mL). The suspension was cooled to -20 °C, to which was added diphenylphosphinic chloride (0.52 mL, 2.75 mmol). The solution was stirred at -20 °C for 30 min, followed by addition of a mixed solution of (IA) (1 .52g, 3.00 mmol) and triethylamine (0.52 mL, 3.76 mmol) in DMF (2mL). The reaction mixture was stirred at 20 °C for 20 h, followed by addition of MTBE (20 mL). The reaction mixture was adjusted to pH = 2-3 using aqueous HCI solution (37%). To the mixture was added isopropanol (100 mL). The resulting mixture was stirred for 4 h to obtain a suspension. The suspension was filtered and the filter cake was dried under vacuum to afford crude 22 (1 .17 g). The crude 22 was slurried in a combined solvent of THF/H2O (= 12 mL / 3mL), and filtered to afford 22 (0.744 g, 75 wt% by Q-NMR, 53.3% yield). ¹H NMR (400 MHz, DMSO-cfe) δ ppm 3.47 – 3.55 (m, 2 H) 3.59 – 3.63 (m, 2 H) 4.13 – 4.21 (m, 3 H ) 5.05 (dd, J=8.8, 5.6 Hz, 1 H) 8.22 (s, 1 H) 9.73 (d, J=8.7 Hz, 1 H).

To a suspension of 22 (580 mg, 75 wt%, 1 .037 mmol) in DMAC (1 .5 mL) was added 2A (214.3 mg, 85 wt%, 1 .556 mmol). The reaction was stirred at 25 °C for 3 days, and in process control showed 22, Compound X = 4/96, and Z/E = 91 /9. the mixture was slowly added into 15ml acetone to precipitate yellowish solid. The reaction mixture was filtered to afford Compound X (0.7 g, 34 wt% by QNMR, 44% yield).

Synthesis of compound 3 (R² = CH(Ph)₂)

^R2 = CH(Ph)₂

2-(2-aminothiazol-4-yl)-2-oxoacetic acid (Y) (10.00 g, 47.93 mmol) and compound W (R² = CH(Ph)₂) (13.31 g, 46.98 mmol) were suspended in DMAC (40 mL), followed by addition of triethylamine (5.01 mL, 35.95 mmol). The reaction mixture was stirred at 20 °C for 5 h. HPLC showed completion of the reaction, and Z/E

= 97/3. To the reaction mixture was added water (120 mL) with stirring. The mixture was stirred for 20 min to obtain a suspension. The suspension was filtered and the filter cake was washed with water (50 mL).

The filter cake was slurried in a combined solvent of THF/ethyl acetate (50 mL / 50 mL) at 60 °C and cooled to 20 °C. The solid was filtered and dried at 50 °C for 3 h to get 3 (R² = CH(Ph)₂) (19.5 g, 88% yield). ¹H

NMR (400 MHz, DMSO-cfe) δ ppm 1.37 -1 .42 (m, 2 H) 1 .44 – 1 .49 (m, 2 H) 6.87 (s, 1 H) 6.94 (s, 1 H) 7.22

– 7.30 (m, 6 H) 7.45 – 7.49 (m, 4 H).

Alternative Synthesis of Compound X from compound of formula (I), (IA)

Compound ^x

IA (40.14 g, 62.63 mmol) was dissolved in methanol (200 ml_), followed by addition of Pd/C (10%, 1 .1 g). The reaction mixture was maintained under hydrogen atmosphere (1 -2 bar) at 20 °C for 24 h. In process control showed completion of the reaction. The reaction mixture was filtered. The filtrate was concentrated to give an oil of IB (M⁺ = NBu₄⁺) (58.20 g, 55 wt% by Q-NMR, 100% yield). ¹ H NMR (400 MHz, DMSO-cfe) δ ppm 0.93 (t, J=7.3 Hz, 12 H) 1 .23 – 1 .36 (m, 8 H) 1 .57 (m, 8 H) 2.99 – 3.28 (m, 8 H) 3.37 (dd, J=14.3, 7.5 Hz, 1 H) 3.65 – 3.70 (m, 3 H) 3.84 – 3.88 (m, 1 H) 4.08 (d, J=5.6 Hz, 1 H) 4.18 – 4.22 (m, 2 H).

3 (R² = CH(Ph)₂) (0.95 g, 2.17 mmol) was dissolved in THF (20 ml_). To the solution was added /V-methyl morpholine (0.77 g, 7.60 mmol) and 2-chloro-4,6-dimethoxy-1 ,3,5-triazine (0.57 g, 3.26 mmol). The reaction mixture was stirred at 20 °C for 1 h followed by addition of IB (M⁺ = NBu ⁺) (2.70 g, 48.98 wt%, 2.61 mmol). The reaction was stirred at 20 °C for 5 h. In process control showed completion of the reaction. To the reaction mixture was added ethyl acetate (20 ml_). The organic phase was washed with brine (10 ml_). Solvent was removed. Acetone (40ml) was added to dissolve residue. TFA (1 .24 g, 10.86 mmol) dissolved in acetone (3 ml) was added slowly. The white solid was filtered and washed by acetone (10 ml) two times. Dried at 40 °C for 5h to get compound 4 (R² = CH(Ph)₂). ¹ H NMR (400 MHz, DMSO-cfe) δ ppm 1 .49 – 1 .55 (m, 4 H) 3.27 (dd, J=14.4, 6.2 Hz, 1 H) 3.49 – 3.65 (m, 2 H) 3.71 (dd, J=14.4, 6.2 Hz, 1 H) 4.04 – 4.10 (m, 1 H) 4.07 (dd, J=16.0, 8.6 Hz, 1 H) 4.17 (dd, J=1 1 .8, 6.0 Hz, 1 H) 5.28 (dd, J=9.0, 5.7 Hz, 1 H) 6.88 (s, 1 H) 7.03 (s, 1 H) 7.18 – 7.32 (m, 6 H) 7.43 (m, 4 H) 9.45 (d, J=9.0 Hz, 1 H).

Crude 4 (R² = CH(Ph)₂) (2.13 g) was dissolved in dichloromethane (20 ml_). The solution was cooled to 0 °C. To the solution was added anisole (0.68 ml_, 6.24 mmol) and trifluoroacetic acid (2.16 ml_, 28.08 mmol). The reaction was warmed to 20 °C, and stirred for 15 h. In process control showed completion of the

reaction. The aqueous phase was separated and added to acetone (40 mL) to obtain a suspension. The suspension was filtered to afford Compound X (0.98 g, 54.5% yield over two steps). ¹ H NMR (400 MHz, DMSO-c/e) δ ppm 1.40 (m, 4 H) 3.26 (dd, J=14.4, 6.0 Hz, 1 H) 3.54 – 3.69 (m, 3 H) 4.14 – 4.21 (m, 3 H) 5.25 (dd, J= 8.9, 5.7 Hz, 1 H) 7.02 (s, 1 H) 9.38 (d, J=9.0 Hz, 1 H).

REF

Synthesis and optimization of novel monobactams with activity against carbapenem-resistant Enterobacteriaceae – Identification of LYS228
57th Intersci Conf Antimicrob Agents Chemother (ICAAC) (June 1-5, New Orleans) 2017, Abst SATURDAY-297

//////////////LYS228, LYS 228, BOS-228, LYS-228, monobactam, Novartis, phase II, Boston Pharmaceuticals, complicated urinary tract infection, complicated intraabdominal infections, fast track, Qualified Infectious Disease Product, QIDP,

Nc1nc(cs1)\C(=N\OC2(CC2)C(=O)O)\C(=O)N[C@H]3[C@@H](CN4CCOC4=O)N(C3=O)S(=O)(=O)O

GRAPIPRANT

January 21, 2020 10:59 am / Leave a comment

ChemSpider 2D Image | grapiprant | C26H29N5O3S

Structure of GRAPIPRANT

GRAPIPRANT

Molecular FormulaC₂₆H₂₉N₅O₃S
Average mass491.605 Da

CAS 415903-37-6

UNII-J9F5ZPH7NB, CJ 023423, CJ-023423,

Phase II, Arrys Therapeutics, CANCER,

PAIN, AskAt Phase II,

N-{2-[4-(2-ethyl-4,6-dimethyl-1H-imidazo[4,5-c]pyridin-1-yl)phenyl]ethyl}-N’-[(4-methylphenyl)sulfonyl]urea

RQ-00000007, MR10A7

9763

AAT-007

Benzenesulfonamide, N-[[[2-[4-(2-ethyl-4,6-dimethyl-1H-imidazo[4,5-c]pyridin-1-yl)phenyl]ethyl]amino]carbonyl]-4-methyl-

CJ-023,423

N-[[[2-[4-(2-Ethyl-4,6-dimethyl-1H-imidazo[4,5-c]pyridin-1-yl)phenyl]ethyl]amino]carbonyl]-4-methylbenzenesulfonamide
1-[2-[4-(2-Ethyl-4,6-dimethylimidazo[4,5-c]pyridin-1-yl)phenyl]ethyl]-3-(4-methylphenyl)sulfonylurea
2-Ethyl-4,6-dimethyl-1-[4-[2-[[[[(4-methylphenyl)sulfonyl]amino]carbonyl]amino]ethyl]phenyl]-1H-imidazo[4,5-c]pyridine
AAT 007
CJ 023423

Grapiprant
MR 10A7
RQ 00000007
RQ 7

Synonyms and Mappings

415903-37-6
GRAPIPRANT [GREEN BOOK]
CJ-023
GRAPIPRANT [INN]
GRAPIPRANT [WHO-DD]
MR-10A7
AAT-007
MR10A7
RQ-00000007
RQ-7
GRAPIPRANT [USAN]
GRAPIPRANT
2-ETHYL-4,6-DIMETHYL-1-(4-(2-(((((4-METHYLPHENYL)SULFONYL)AMINO)CARBONYL)AMINO)ETHYL)PHENYL)-1H-IMIDAZO(4,5-C)PYRIDINE
N-(((2-(4-(2-ETHYL-4,6-DIMETHYL-1H-IMIDAZO(4,5-C)PYRIDIN-1-YL)PHENYL)ETHYL)AMINO)CARBONYL)-4-METHYLBENZENESULFONAMIDE
CJ 023423
BENZENESULFONAMIDE, N-(((2-(4-(2-ETHYL-4,6-DIMETHYL-1H-IMIDAZO(4,5-C)PYRIDIN-1-YL)PHENYL)ETHYL)AMINO)CARBONYL)-4-METHYL-
CJ-023,423
N-(((2-(4-(2-ETHYL-4,6-DIMETHYL-1H-IMIDAZO(4,5-C)PYRIDIN-1-YL)PHENYL)ETHYL)AMINO)CARBONYL)-4-METHYL-BENZENESULFONAMIDE
CJ-023423

SYN

Arrys Therapeutics (under license from AskAt ) and affiliate Ikena Oncology (formerly known as Kyn Therapeutics ) are developing ARY-007 , an oral formulation of grapiprant, for treating cancers; in December 2019, preliminary data were expected in 2020

Grapiprant (trade name Galliprant) is a small molecule drug that belongs in the piprant class. This analgesic and anti-inflammatory drug is primarily used as a pain relief for mild to moderate inflammation related to osteoarthritis in dogs. Grapiprant has been approved by the FDA’s Center for Veterinary Medicine and was categorized as a non-cyclooxygenase inhibiting non-steroidal anti-inflammatory drug (NSAID) in March 2016.^[1]

Preclinical studies also indicate that grapiprant is not only efficacious as a acute pain but also in chronic pain relief and inflammation drug. The effect of the drug is directly proportional to the dosage and its effects were comparable to human medication such as rofecoxib and piroxicam.^[2]

Grapiprant, a prostanoid EP4 receptor antagonist, is in phase II clinical trials at AskAt for the treatment of chronic pain. Phase I/II clinical trials are ongoing at Arrys Therapeutics in combination with pembrolizumab for the treatment of patients with microsatellite stable colorectal cancer and in patients with advanced or metastatic PD-1/L1 refractory non-small cell lung cancer (NSCLC).

Grapiprant is also used in humans, and was researched to be used as a pain control and inflammation associated with osteoarthritis. The effect of grapiprant could be explained through the function of prostaglandin E2, in which acts as a pro-inflammatory mediator of redness of the skin, edema and pain which are the typical signs of inflammation. The effect of PGE2 stems from its action through the four prostaglandin receptor subgroups EP1, EP2, EP3 and EP4, in which the prostaglandin EP4 receptor acts as the main intermediary of the prostaglandin-E2-driven inflammation. Grapiprant is widely accepted in veterinary medicine due to its specific and targeted approach to pain management in dogs. The serum concentration of grapiprant is increased when used in conjunction with other drugs such as acetaminophen, albendazole, and alitretinoin.

Common side effects are intestinal related effects such as mild diarrhea, appetite loss, and vomiting.^[3] Additionally, it is found that it might lead to reduced tear production due to it being a sulfa-based medication and also reduced albumin levels.

Medical uses

Grapiprant is used once a day as an oral pain relief for dogs with inflammation-related osteoarthritis. It is a non-steroidal anti-inflammatory (NSAID) that functions as a targeted action to treat osteoarthritis pain and inflammation in dogs.

Mechanism of action

Grapiprant acts as a specific antagonist that binds and blocks the prostaglandin EP4 receptor, one out of the four prostaglandin E2 (PGE2) receptor subgroups. The EP4 receptor then mediates the prostaglandin-E2-elicited response to pain, and hence grapiprant was proven to be effective in the decrease of pain in several inflammatory pain models of rats. It was also proven to be effective in reducing osteoarthritis-related pain in humans, which serves as a proof for its mechanism of action. The approximate calculation for canine efficacy dose is between the range of 1.3 and 1.7 mg/kg, in conjunction with a methylcellulose suspending agent. Based on the calculations from the comparisons of binding affinity of grapiprant to the EP4 receptors of dogs, rats, and humans, the study of plasma and serum protein binding determinations, the effective doses determined in inflammation pain models of rats, and human-related clinical studies, it is evaluated that Grapiprant should be administered just once a day. The approved dose of the commercial Grapiprant tablet by the FDA for the pain relief and inflammation associated with osteoarthritis to dogs is reported to be 2 mg/kg a day.^[4]

Absorption

Studies in animals such as horses have shown the presence of Grapiprant in serum 72 hours with a concentration >0.005 ng/ml after the initial administration of a dose of 2 mg/kg. Grapiprant is expeditiously absorbed and the reported serum concentration was reported to be 31.9 ng/ml in an amount of time of 1.5 hours. The actual body exposure to grapiprant after administration of one dose was shown to be 2000 ng.hr/ml. The degree and rate at which grapiprant is absorbed into the body, presents a mean bioavailability of 39%. A significant reduction in the bioavailability, concentration time and maximal concentration were reported to have occurred after food intake.^[1] And thus, grapiprant is usually not administered with food as it will not be as efficient.^[5]

Distribution

The volume of distribution in cat studies was reported to be 918 ml/kg.^[1]

Route of elimination

Following an oral administration, the majority of the dose was metabolized within the first 72 hours. Equine studies have shown that grapiprant is present in urine 96 hours after the first administration of a dose of 2 mg/kg and has a concentration >0.005 ng/ml. From the excreted dose conducted in horses, it is found that 55%, 15% and 19% of the orally-administered dose was excreted in bile, urine, and faeces respectively.^[1]

Toxicity

Safety studies conducted on grapiprant have demonstrated that it generally possesses an exceptional safety profile and a wide safety margin in veterinary studies.^[6] In animal studies, a research on 2.5-12 times overdose was conducted for grapiprant and the study resulted in soft-blobs and mucous-filled faeces, occasional bloody stools and emesis.

PATENT

WO-2020014465

Novel crystalline forms of grapiprant and their salts eg HCl (designated as Form A), useful for inhibiting prostaglandin EP4 receptor activity and treating cancers.

Prostaglandins are mediators of pain, fever and other symptoms associated with inflammation. Prostaglandin E₂ (PGE2) is the predominant eicosanoid detected in inflammation conditions. In addition, it is also involved in various physiological and/or pathological conditions such as hyperalgesia, uterine contraction, digestive peristalsis, awakeness, suppression of gastric acid secretion, blood pressure, platelet function, bone metabolism, angiogenesis or the like.

[0003] Four PGE2 receptor subtypes (EP1, EP2, EP3 and EP4) displaying different pharmacological properties exist. The EP4 subtype, a Gs-coupled receptor, stimulates cAMP production as well as PI3K and GSK3P signaling, and is distributed in a wide variety of tissue suggesting a major role in PGE2-mediated biological events. Various EP4 inhibitors have been described previously, for example, in WO 2002/032900, WO 2005/021508, EiS 6,710,054, and US 7,238,714, the contents of which are incorporated herein by reference in their entireties.

[0004] Accordingly, there is a need for treating, preventing, and/or reducing severity of a proliferative disorder associated with prostaglandin EP4 receptor activity. The present invention addresses such a need.

It has now been found that compounds of the present invention, and compositions thereof, are useful for treating, preventing, and/or reducing severity of a proliferative disorder associated with prostaglandin EP4 receptor activity. In general, salt forms and co-crystal forms, and pharmaceutically acceptable compositions thereof, are useful for treating or lessening the severity of proliferative disorders associated with prostaglandin EP4 receptor activity, as described in detail herein. Such compounds are represented by the chemical structure below, denoted as compound A (also known as grapiprant):

or a pharmaceutically acceptable salt thereof.

United States Patent 7,960,407, filed March 1, 2006 and issued June 14, 2011 (“the ‘407 patent,” the entirety of which is hereby incorporated herein by reference), describes certain EP4 inhibitor compounds. Such compounds include compound A:

or a pharmaceutically acceptable salt thereof.

[0037] Compound A, N-[({2-[4-(2-Ethyl-4,6-dimethyl-lH-imidazo[4,5-c]pyridin-l-yl) phenyl]ethyl}amino)carbonyl]-4-methylbenzenesulfonamide, is described in detail in the ‘407

patent, including its synthetic route. The ‘407 patent also discloses a variety of physical forms of compound A.

[0038] It would be desirable to provide a solid form of compound A (e.g., as a co-crystal thereof or salt thereof) that imparts characteristics such as improved aqueous solubility, stability and ease of formulation. Accordingly, the present invention provides both co-crystal forms and salt forms of compound A:

PATENT

WO 2002032900

PATENT

WO 2002032422

Family members of the product case ( WO0232422 ) of grapiprant have protection in most of the EU states until October 2021 and expire in the US in October 15, 2021.

PATENT

WO 2003086371

PATENT

WO2020014445 covering combinations of grapiprant and an immuno-oncology agent.

WO 2005102389

WO 2006095268

US 7960407

US 20190314390

References

^ Jump up to:^a ^b ^c ^d “Grapiprant”. http://www.drugbank.ca. Retrieved 2019-05-15.
^ PubChem. “Grapiprant”. pubchem.ncbi.nlm.nih.gov. Retrieved 2019-05-15.
^ Paul Pion, D. V. M.; Spadafori, Gina (2017-08-08). “Veterinary Partner”. VIN.com.
^ Nagahisa, A.; Okumura, T. (2017). “Pharmacology of grapiprant, a novel EP4 antagonist: receptor binding, efficacy in a rodent postoperative pain model, and a dose estimation for controlling pain in dogs”. Journal of Veterinary Pharmacology and Therapeutics. 40 (3): 285–292. doi:10.1111/jvp.12349. ISSN 1365-2885. PMID 27597397.
^ Paul Pion, D. V. M.; Spadafori, Gina (2017-08-08). “Veterinary Partner”. VIN.com.
^ Kirkby Shaw, Kristin; Rausch-Derra, Lesley C.; Rhodes, Linda (February 2016). “Grapiprant: an EP4 prostaglandin receptor antagonist and novel therapy for pain and inflammation”. Veterinary Medicine and Science. 2 (1): 3–9. doi:10.1002/vms3.13. ISSN 2053-1095. PMC 5645826. PMID 29067176.

Grapiprant
Clinical data

Trade names	Galliprant
Routes of administration	Oral
ATCvet code	QM01AX92 (WHO)
Pharmacokinetic data
Bioavailability	6.6 L/kg, high volume of distribution
Elimination half-life	5.86 hours in horses
Excretion	Urine
Identifiers
IUPAC name [show]
CAS Number	415903-37-6
PubChem CID	11677589
DrugBank	DB12836
ChemSpider	9852318
UNII	J9F5ZPH7NB
CompTox Dashboard (EPA)	DTXSID60194419
Chemical and physical data
Formula	C₂₆H₂₉N₅O₃S
Molar mass	491.61 g·mol⁻¹
3D model (JSmol)	Interactive image
SMILES [hide] CCC1=NC2=C(C)N=C(C)C=C2N1C1=CC=C(CCNC(=O)NS(=O)(=O)C2=CC=C(C)C=C2)C=C1

//////////////GRAPIPRANT, 415903-37-6, UNII-J9F5ZPH7NB, CJ 023423, CJ-023423, RQ-00000007, MR10A7, Galliprant, Phase II, Arrys Therapeutics, CANCER, PAIN, AskAt

CCC1=NC2=C(N1C3=CC=C(C=C3)CCNC(=O)NS(=O)(=O)C4=CC=C(C=C4)C)C=C(N=C2C)C

CK-101

August 30, 2019 2:30 pm / Leave a comment

N-[3-[2-[2,3-Difluoro-4-[4-(2-hydroxyethyl)piperazin-1-yl]anilino]quinazolin-8-yl]phenyl]prop-2-enamide.png

CK-101, RX-518

CAS 1660963-42-7

MF C₂₉ H₂₈ F₂ N₆ O₂

MW 530.57

2-Propenamide, N-[3-[2-[[2,3-difluoro-4-[4-(2-hydroxyethyl)-1-piperazinyl]phenyl]amino]-8-quinazolinyl]phenyl]-

N-[3-[2-[[2,3-Difluoro-4-[4-(2-hydroxyethyl)piperazin-1-yl]phenyl]amino]quinazolin-8-yl]phenyl]acrylamide

N-(3-(2-((2,3-Difluoro-4-(4-(2-hydroxyethyl)piperazin-1-yl)phenyl)amino)quinazolin-8-yl)phenyl)acrylamide

EGFR-IN-3

UNII-708TLB8J3Y

708TLB8J3Y

AK543910

Suzhou NeuPharma (Originator)
Checkpoint Therapeutics

Non-Small Cell Lung Cancer Therapy
Solid Tumors Therapy

PHASE 2 Checkpoint Therapeutics, Cancer, lung (non-small cell) (NSCLC), solid tumour

RX518(CK-101) is an orally available third-generation and selective inhibitor of certain epidermal growth factor receptor (EGFR) activating mutations, including the resistance mutation T790M, and the L858R and exon 19 deletion (del 19) mutations, with potential antineoplastic activity.

In August 2019, Suzhou Neupharma and its licensee Checkpoint Therapeutics are developing CK-101 (phase II clinical trial), a novel third-generation, covalent, EGFR inhibitor, as a capsule formulation, for the treatment of cancers including NSCLC and other advanced solid tumors. In September 2017, the FDA granted Orphan Drug designation to this compound, for the treatment of EGFR mutation-positive NSCLC; in January 2018, the capsule was being developed as a class 1 chemical drug in China.

CK-101 (RX-518), a small-molecule inhibitor of epidermal growth factor receptor (EGFR), is in early clinical development at Checkpoint Therapeutics and Suzhou NeuPharma for the potential treatment of EGFR-mutated non-small cell lung cancer (NSCLC) and other advanced solid malignancies.

In 2015, Suzhou NeuPharma granted a global development and commercialization license to its EGFR inhibitor program, excluding certain Asian countries, to Coronado Biosciences (now Fortress Biotech). Subsequently, Coronado assigned the newly acquired program to its subsidiary Checkpoint Therapeutics.

In 2017, the product was granted orphan drug designation in the U.S. for the treatment of EGFR mutation-positive NSCLC.

There are at least 400 enzymes identified as protein kinases. These enzymes catalyze the phosphorylation of target protein substrates. The phosphorylation is usually a transfer reaction of a phosphate group from ATP to the protein substrate. The specific structure in the target substrate to which the phosphate is transferred is a tyrosine, serine or threonine residue. Since these amino acid residues are the target structures for the phosphoryl transfer, these protein kinase enzymes are commonly referred to as tyrosine kinases or serine/threonine kinases.

[0003] The phosphorylation reactions, and counteracting phosphatase reactions, at the tyrosine, serine and threonine residues are involved in countless cellular processes that underlie responses to diverse intracellular signals (typically mediated through cellular receptors), regulation of cellular functions, and activation or deactivation of cellular processes. A cascade of protein kinases often participate in intracellular signal transduction and are necessary for the realization of these cellular processes. Because of their ubiquity in these processes, the protein kinases can be found as an integral part of the plasma membrane or as cytoplasmic enzymes or localized in the nucleus, often as components of enzyme complexes. In many instances, these protein kinases are an essential element of enzyme and structural protein complexes that determine where and when a cellular process occurs within a cell.

[0004] The identification of effective small compounds which specifically inhibit signal transduction and cellular proliferation by modulating the activity of tyrosine and serine/threonine kinases to regulate and modulate abnormal or inappropriate cell proliferation, differentiation, or metabolism is therefore desirable. In particular, the identification of compounds that specifically inhibit the function of a kinase which is essential for processes leading to cancer would be beneficial.

[0005] While such compounds are often initially evaluated for their activity when dissolved in solution, solid state characteristics such as polymorphism are also important. Polymorphic forms of a drug substance, such as a kinase inhibitor, can have different physical properties, including melting point, apparent solubility, dissolution rate, optical and mechanical properties, vapor pressure, and density. These properties can have a direct effect on the ability to process or manufacture a drug substance and the drug product. Moreover, differences in these properties

can and often lead to different pharmacokinetics profiles for different polymorphic forms of a drug. Therefore, polymorphism is often an important factor under regulatory review of the ‘sameness’ of drug products from various manufacturers. For example, polymorphism has been evaluated in many multi-million dollar and even multi-billion dollar drugs, such as warfarin sodium, famotidine, and ranitidine. Polymorphism can affect the quality, safety, and/or efficacy of a drug product, such as a kinase inhibitor. Thus, there still remains a need for polymorphs of kinase inhibitors. The present disclosure addresses this need and provides related advantages as well.

PATENT

WO2015027222

https://patentscope.wipo.int/search/en/detail.jsf?docId=WO2015027222

PATENT

WO-2019157225

https://patentscope.wipo.int/search/en/detail.jsf?docId=WO2019157225&tab=PCTDESCRIPTION&_cid=P10-JZNKMN-12945-1

Crystalline form II-VIII of the compound presumed to be CK-101 (first disclosed in WO2015027222 ), for treating a disorder mediated by epidermal growth factor receptor (EGFR) eg cancer.

SCHEME A

Scheme B

General Procedures

Example 1: Preparation of the compound of Formula I (N-(3-(2-((2,3-difluoro-4-(4-(2-hydroxyethyl)piperazin-l-yl)phenyl)amino)quinazolin-8-yl)phenyl)acrylamide)

[0253] To a solution of l,2,3-trifluoro-4-nitrobenzene (2.5 g, 14 mmol, 1.0 eq.) in DMF (20 mL) was added K₂C0₃ (3.8 g, 28 mmol, 2.0 eq.) followed by 2-(piperazin-l-yl)ethanol (1.8 g, 14 mmol, 1.0 eq.) at 0 °C and the mixture was stirred at r.t. overnight. The mixture was poured into ice-water (200 mL), filtered and dried in vacuo to afford 2-(4-(2,3-difluoro-4-nitrophenyl)piperazin-l-yl)ethanol (2.7 g, 67.5%).

[0254] To a solution of 2-(4-(2,3-difluoro-4-nitrophenyl)piperazin-l-yl)ethanol (2.7 g, 9.0 mmol) in MeOH (30 mL) was added Pd/C (270 mg) and the resulting mixture was stirred at r.t.

overnight. The Pd/C was removed by filtration and the filtrate was concentrated to afford 2-(4-(4-amino-2,3-difluorophenyl)piperazin-l-yl)ethanol (2.39 g, 99% yield) as off-white solid.

[0255] To a solution of 8-bromo-2-chloroquinazoline (15.4 g, 63.6 mmol, 1 eq. ) and (3-aminophenyl)boronic acid (8.7 g, 63.6 mmol, 1 eq.) in dioxane/H₂0 (200 mL/20 mL) was added Na₂C0₃ (13.5 g, 127.2 mmol, 2 eq.), followed by Pd(dppf)Cl₂ (2.6 g, 3.2 mmol, 0.05 eq.) under N₂, then the mixture was stirred at 80 °C for 12 h. Then the solution was cooled to r.t.,

concentrated and the residue was purified via column chromatography (PE/EA=3 :2, v/v) to afford 3-(2-chloroquinazolin-8-yl)aniline as yellow solid (8.7 g, 53.7% yield).

[0256] To a solution of 3-(2-chloroquinazolin-8-yl)aniline (8.7 g, 34 mmol, 1 eq.) in DCM ( 200 mL ) cooled in ice-bath was added TEA (9.5 mL, 68 mmol, 2 eq. ), followed by acryloyl chloride (4.1 mL, 51 mmol, 1.5 eq.) dropwise. The resulting mixture was stirred at r.t. for 1 h, then washed with brine, dried over anhydrous N₂S0₄ concentrated and the residue was purified via column chromatography (PE/EA=l : 1, v:v) to afford N-(3-(2-chloroquinazolin-8-yl)phenyl)acryl amide as yellow solid(6.6 g, 65% yield).

[0257] To a suspension of 2-(4-(4-amino-2,3-difluorophenyl)piperazin-l-yl)ethanol (83 mg,

0.32 mmol, 1 eq.) and N-(3-(2-chloroquinazolin-8-yl)phenyl)acrylamide (100 mg, 0.32 mmol, 1 eq.) in n-BuOH (5 mL) was added TFA (68 mg, 0.64 mmol, 2 eq.) and the resulting mixture was stirred at 90 °C overnight. The mixture was concentrated, diluted with DCM (20 mL) , washed with Na₂C0₃ solution (20 mL), dried over anhydrous Na₂S0₄, concentrated and the residue was purified via column chromatography (MeOH/DCM=l/30, v:v) to afford N-(3-(2-((2,3-difluoro-4-(4-(2-hydroxyethyl)piperazin-l-yl)phenyl)amino)quinazolin-8-yl)phenyl)acrylamide as a yellow solid(l6.3 mg, 9.5% yield). LRMS (M+H⁺) m/z calculated 531.2, found 531.2. 1H NMR

(CD₃OD, 400 MHz) d 9.21 (s, 1 H), 7.19-8.01 (m, 10 H), 8.90 (s, 1 H), 6.41-6.49 (m, 3 H), 5.86 (m, 1 H), 3.98-4.01 (m, 3 H), 3.70-3.76 (m, 3 H), 3.40-3.49 (m, 2 H), 3.37-3.39 (m, 4 H), 3.18 (m, 2H).

Example 2. Preparation of Form I of the compound of Formula I

[0258] Crude compound of Formula I (~30 g, 75% of weight based assay) was dissolved in ethyl acetate (3 L) at 55-65 °C under nitrogen. The resulting solution was filtered via silica gel pad and washed with ethyl acetate (3 L><2) at 55-65 °C. The filtrate was concentrated via vacuum at 30-40 °C to ~2.4 L. The mixture was heated up to 75-85 °C and maintained about 1 hour.

Then cooled down to 50-60 °C and maintained about 2 hours. The heat-cooling operation was repeated again and the mixture was then cooled down to 20-30 °C and stirred for 3 hours. The resulting mixture was filtered and washed with ethyl acetate (60 mL><2). The wet cake was dried via vacuum at 30-40 °C to get (about 16 g) of the purified Form I of the compound of Formula I.

Example 3. Preparation of Form III of the compound of Formula I

[0259] The compound of Formula I (2 g) was dissolved in EtOH (40 mL) at 75-85 °C under nitrogen. n-Heptane (40 mL) was added dropwise into reaction at 75-85 °C. The mixture was stirred at 75-85 °C for 1 hour. Then cooled down to 50-60 °C and maintained about 2 hours. The heat-cooling operation was repeated again and continued to cool the mixture down to 20-30 °C and stirred for 3 hours. The resulting mixture was filtered and washed with EtOH/n-Heptane (1/1, 5 mL><2). The wet cake was dried via vacuum at 30-40 °C to get the purified Form III of the compound of Formula I (1.7 g).

Example 4. Preparation of Form IV of the compound of Formula I The crude compound of Formula I (15 g) was dissolved in ethyl acetate (600 mL) at 75-85 °C under nitrogen and treated with anhydrous Na₂S0₄, activated carbon, silica metal scavenger for 1 hour. The resulting mixture was filtered via neutral Al₂0₃ and washed with ethyl acetate (300 mL><2) at 75-85 °C. The filtrate was concentrated under vacuum at 30-40 °C and swapped with DCM (150 mL). n-Heptane (75 mL) was added into this DCM solution at 35-45 °C, and then the mixture was cooled down to 20-30 °C slowly. The resulting mixture was filtered and washed with DCM/n-Heptane (2/1, 10 mL><3). The wet cake was dried via vacuum at 35-40 °C to get the purified Form IV of the compound of Formula I (9.6 g).

Example 5. Preparation of Form V of the compound of Formula I

[0260] Polymorph Form III of the compound of Formula I was dried in oven at 80 °C for 2 days to obtain the polymorph Form V.

Example 6. Preparation of Form VI of the compound of Formula I

[0261] The compound of Formula I (1 g) was dissolved in IPA (20 mL) at 75-85 °C under nitrogen. n-Heptane (20 mL) was added dropwise into reaction at 75-85 °C. The mixture was stirred at 45-55 °C for 16 hours. Then heated up to 75-85 °C and maintained about 0.5 hour.

Then cooled down to 45-55 °C for 0.5 hour and continued to cool the mixture down to 20-30 °C and stirred for 3 hours. Filtered and washed with IPA/n-Heptane (1/1, 3 mL><2). The wet cake was dried via vacuum at 75-80 °C for 2 hours to get the purified Form VI of the compound of Formula I.

Example 7. Preparation of Form VIII of the compound of Formula I

[0262] The polymorph Form VI of the compound of Formula I was dried in oven at 80 °C for 2 days to obtain the polymorph Form VIII.

Example 8. X-ray powder diffraction (XRD)

[0263] X-ray powder diffraction (XRD) patterns were obtained on a Bruker D8 Advance. A CuK source (=1.54056 angstrom) operating minimally at 40 kV and 40 mA scans each sample between 4 and 40 degrees 2-theta. The step size is 0.05°C and scan speed is 0.5 second per step.

Example 9. Thermogravimetric Analyses (TGA)

[0264] Thermogravimetric analyses were carried out on a TA Instrument TGA unit (Model TGA 500). Samples were heated in platinum pans from ambient to 300 °C at 10 °C/min with a nitrogen purge of 60mL/min (sample purge) and 40mL/min (balance purge). The TGA temperature was calibrated with nickel standard, MP=354.4 °C. The weight calibration was performed with manufacturer-supplied standards and verified against sodium citrate dihydrate desolvation.

Example 10. Differential scanning calorimetry (DSC)

[0265] Differential scanning calorimetry analyses were carried out on a TA Instrument DSC unit (Model DSC 1000 or 2000). Samples were heated in non-hermetic aluminum pans from ambient to 300 °C at 10 °C/min with a nitrogen purge of 50mL/min. The DSC temperature was calibrated with indium standard, onset of l56-l58°C, enthalpy of 25-29J/g.

Example 11. Hygroscopicity (DVS)

[0266] The moisture sorption profile was generated at 25°C using a DVS Moisture Balance Flow System (Model Advantage) with the following conditions: sample size approximately 5 to 10 mg, drying 25°C for 60 minutes, adsorption range 0% to 95% RH, desorption range 95% to 0% RH, and step interval 5%. The equilibrium criterion was <0.01% weight change in 5 minutes for a maximum of 120 minutes.

Example 12: Microscopy

[0267] Microscopy was performed using a Leica DMLP polarized light microscope equipped with 2.5X, 10X and 20X objectives and a digital camera to capture images showing particle shape, size, and crystallinity. Crossed polars were used to show birefringence and crystal habit for the samples dispersed in immersion oil.

Example 13: HPLC

[0256] HPLCs were preformed using the following instrument and/or conditions.

///////////////CK-101 , CK 101 , CK101 , phase II , Suzhou Neupharma, Checkpoint Therapeutics , Orphan Drug designation, EGFR mutation-positive NSCLC, NSCLC, CANCER, SOLID TUMOUR, China, RX-518, AK543910

OCCN1CCN(CC1)c5ccc(Nc2nc3c(cccc3cn2)c4cccc(NC(=O)C=C)c4)c(F)c5F

Fluazolepali, 氟唑帕利 , Fluzoparib

July 25, 2019 1:31 pm / Leave a comment

Fluazolepali

CAS 2170504-09-1

Fluzoparib; SHR-3162, (HS10160)

HS 10160
SHR 3162

An orally available inhibitor of poly(ADP-ribose) polymerase 1 and 2 (PARP-1/2) for treatment of solid tumors (Jiangsu Hengrui Medicine Co. Ltd., Lianyungang, China)

Fluazolepali, developed by Hengrui and Howson, is intended for the treatment of recurrent ovarian cancer, triple-negative breast cancer, advanced gastric cancer and other advanced solid tumors. Currently, the drug has been introduced into China for recurrent ovarian cancer. Clinical stage.

In February 2019, a randomized, double-blind, controlled, multicenter, phase III clinical study (CTR20190294) of flazopril capsule versus placebo for maintenance of recurrent ovarian cancer was initiated in China and was sponsored by Hengrui Medicine.

Jiangsu Hansoh Pharmaceutical , in collaboration with Jiangsu Hengrui Medicine , is developing an oral capsule formulation of fluazolepali (fluzoparib; SHR-3162), a small molecule inhibitor to PARP-1 and PARP-2, for the treatment of solid tumors including epithelial ovarian, fallopian tube or primary peritoneal, breast and gastric cancer.

Originator Jiangsu Hengrui Medicine Co.
Class Antineoplastics
Mechanism of Action Poly(ADP-ribose) polymerase 1 inhibitors; Poly(ADP-ribose) polymerase 2 inhibitors

Phase II Ovarian cancer

Phase I Breast cancer; Fallopian tube cancer; Gastric cancer; Peritoneal cancer; Solid tumours

09 Jul 2019 Jiangsu HengRui Medicine initiates a phase I trial in Solid tumors in China (NCT04013048) [14C]-Fluzoparib
01 Jul 2019 Jiangsu HengRui Medicine plans a phase I drug-drug interaction trial (In volunteers) in China (PO) (NCT04011124)
12 Jun 2019 Jiangsu HengRui Medicine completes a phase I trial in Gastric cancer (Combination therapy, Recurrent, Metastatic disease, Second-line therapy or greater, Late-stage disease) in China (PO) (NCT03026881)

Fluzoparib (SHR 3162) is a selective poly [ADP-ribose] polymerase 1 (PARP1) and poly [ADP-ribose] polymerase 2 inhibitor (PARP2), being developed by Jiangsu HengRui Medicine, for the treatment of cancer. PARP enzymes play a vital role in repair of DNA damage and maintaining genomic stability. Fluzoparib inhibits PARP enzymes and induces DNA-double strands breaks, G₂/M arrest and apoptosis in homologous recombination repair (HR)-deficient cells. Clinical development for ovarian cancer, breast cancer, fallopian tube cancer, peritoneal cancer, gastric cancer and solid tumours is underway in China and Australia.

An orally available inhibitor of poly (ADP-ribose) polymerase (PARP) types 1 and 2, with potential antineoplastic activity. Upon oral administration, fluzoparib inhibits PARP 1 and 2 activity, which inhibits PARP-mediated repair of damaged DNA via the base excision repair (BER) pathway, enhances the accumulation of DNA strand breaks, promotes genomic instability, and leads to an induction of apoptosis. The PARP family of proteins catalyze post-translational ADP-ribosylation of nuclear proteins, which then transduce signals to recruit other proteins to repair damaged DNA. PARP inhibition may enhance the cytotoxicity of DNA-damaging agents and may reverse tumor cell chemoresistance and radioresistance. Check for active clinical trials using this agent. (NCI Thesaurus)

PATENT

WO-2019137358

https://patentscope.wipo.int/search/en/detail.jsf?docId=WO2019137358&tab=FULLTEXT&_cid=P20-JYI5A2-54836-1

Process for preparing heterocyclic compounds (presumed to be fluazolepali ) and its intermediates as PARP inhibitors useful for treating cancer.

Example 1

The compound and 5.0kg of 10% palladium on carbon 250g, 80L of methanol was added to the kettle at 0.4MPa, 24h 25 ℃ hydrogenation reaction. The palladium carbon was removed by filtration, the filter cake was washed with methanol, and the filtrate was collected, evaporated to dryness under reduced pressure, and ethyl acetate (20 L) was added to the concentrate, and the mixture was stirred and evaporated, and then cooled to 0° C. ～3, stirring, filtration, filter cake and then adding 20 L of ethyl acetate, pulping at room temperature for 3 to 4 h, filtration, vacuum drying at 45 ° C for 6-8 h to obtain 5.5 kg of compound 3 solid, yield 91.7%, HPLC purity 99.69%.

Example 2

According to the method of Example 19 of CN102686591A, 2 g of the compound 3 and 2.79 g of the compound 4 were charged to obtain 3.6 g of the compound of the formula I in a yield of 87.8%.

Example 3

At room temperature, 2.0 g of compound 2 (prepared according to the method disclosed in WO2009025784) was dissolved in 30 mL of isopropanol, and concentrated sulfuric acid was added dropwise with stirring to adjust the pH to 3, and stirred at room temperature without solid precipitation; the reaction solution was poured into 150 mL of n-hexane. After stirring at room temperature, no solid precipitated, and the sulfate solid of Compound 2 could not be obtained.

Example 4

1. At room temperature, 1.11 g of compound 2 was dissolved in 10 mL of isopropanol, and 15% phosphoric acid/isopropanol solution was added dropwise with stirring to adjust the pH to 3, stirred at room temperature, filtered, and the filter cake was washed with isopropyl alcohol and dried under vacuum. Compound 2 phosphate solid 1.46 g, yield 87.1%, HPLC purity 99.72%.

Example 5

At room temperature, 1.28 g of compound 2 was dissolved in 10 mL of isopropanol, and 20% acetic acid/isopropanol solution was added dropwise with stirring to adjust the pH to 3, and stirred at room temperature without solid precipitation; the reaction solution was poured into 100 mL of n-hexane, and continued. After stirring at room temperature, no solid precipitated, and the acetate solid of Compound 2 could not be obtained.

Example 6

1.05g of compound 2 was dissolved in 10mL of isopropanol at room temperature, and the pH was adjusted to 3 by adding 15% citric acid/isopropanol solution while stirring. At room temperature, no solid precipitated; the reaction solution was poured into 100 mL of n-hexane. After stirring at room temperature, no solid precipitated, and the citrate solid of Compound 2 could not be obtained.

Example 7

1.12 g of compound 2 was dissolved in 10 mL of isopropanol at room temperature, and 0.74 g of maleic acid was added thereto with stirring. The mixture was stirred at room temperature, filtered, and the filter cake was washed with isopropyl alcohol and dried in vacuo to obtain the maleate salt of compound 2. 1.51 g, yield 84.6%.

PATENT

WO2019109938

claiming synergistic combination comprising PARP inhibitor fluazolepali and apatinib mesylate .

PATENT

WO 2018005818

WO 2018129553

WO 2018129559

WO 2018208968

WO 2018213732

WO 2018191277

WO 2018201096

WO 2018085469

WO 2018085468

WO 2019090227

WO 2019133697

WO 2019067978

WO 2019071123

WO 2019090141

///////////Fluazolepali, Jiangsu Hansoh Pharmaceutical, Jiangsu Hengrui Medicine, fluzoparib, SHR-3162, 氟唑帕利 , Phase II, Ovarian cancer, HS10160, CHINA, HS 10160

https://med.sina.com/article_detail_103_2_64751.html

Tanzisertib

July 22, 2019 1:30 pm / 1 Comment on Tanzisertib

ChemSpider 2D Image | Tanzisertib | C21H23F3N6O2

Tanzisertib

CAS 899805-25-5

trans-4-((9-((3S)-Tetrahydrofuran-3-yl)-8-((2,4,6-trifluorophenyl)amino)-9H-purin-2-yl)amino)cyclohexanol

4-[[9-[(3S)-oxolan-3-yl]-8-(2,4,6-trifluoroanilino)purin-2-yl]amino]cyclohexan-1-ol

C21-H23-F3-N6-O2, 448.4467

9557

Cyclohexanol, 4-[[9-[(3S)-tetrahydro-3-furanyl]-8-[(2,4,6-trifluorophenyl)amino]-9H-purin-2-yl]amino]-, trans-

CC 930
CC-930
Tanzisertib
UNII-M5O06306UO
A c-Jun amino-terminal kinase inhibitor.UNII, M5O06306UO

Treatment of Idiopathic Pulmonary Fibrosis (IPF)

Originator Celgene Corporation
Class Antifibrotics; Small molecules
Mechanism of ActionJ NK mitogen-activated protein kinase inhibitors

Orphan Drug Status Yes – Idiopathic pulmonary fibrosis

Discontinued Discoid lupus erythematosus; Idiopathic pulmonary fibrosis

16 Jul 2012 Celgene Corporation terminates a phase II trial in Discoid lupus erythematosus in USA (NCT01466725)
23 Feb 2012 Celgene initiates enrolment in a phase II trial for Discoid lupus erythematosus in the USA (NCT01466725)
08 Nov 2011The Committee for Orphan Medicinal Products (COMP) recommends orphan drug designation for tanzisertib in European Union for Idiopathic pulmonary fibrosis

Tanzisertib has been granted orphan drug status by the FDA for the treatment of idiopathic pulmonary fibrosis. A positive opinion has been received from the EU Committee for Orphan Medicinal Products (COMP

Tanzisertib has been used in trials studying the treatment of Fibrosis, Discoid Lupus, Pulmonary Fibrosis, Interstitial Lung Disease, and Lung Diseases, Interstitial, among others.

PATENT

https://patents.google.com/patent/US20090048275A1/de

Image result for US 20090048275

PATENT

WO 2006076595

US 20070060598

WO 2008057252

US 20080021048

US 20140094456

WO 2014055548

PATENT

WO 2015153683

https://patentscope.wipo.int/search/en/detail.jsf?docId=WO2015153683

/////////Tanzisertib, CC 930, Idiopathic Pulmonary Fibrosis, Orphan Drug, phase II, CELGENE

c1c(c(c(cc1F)F)Nc2n(c3nc(ncc3n2)N[C@H]4CC[C@@H](CC4)O)[C@@H]5COCC5)F

Reldesemtiv

July 11, 2019 10:13 am / Leave a comment

Image result for Reldesemtiv

Reldesemtiv

CK-2127107

CAS 1345410-31-2

UNII-4S0HBYW6QE, 4S0HBYW6QE

MW 384.4 g/mol, MF C₁₉H₁₈F₂N₆O

1-[2-({[trans-3-fluoro-1-(3-fluoropyridin-2-yl)cyclobutyl]methyl}amino)pyrimidin-5-yl]-1H-pyrrole-3- carboxamide

1-[2-[[3-fluoro-1-(3-fluoropyridin-2-yl)cyclobutyl]methylamino]pyrimidin-5-yl]pyrrole-3-carboxamide

Reldesemtiv, also known as CK-2127107, is a skeletal muscle troponin activator (FSTA) and is a potential treatment for people living with debilitating diseases and conditions associated with neuromuscular or non-neuromuscular dysfunction, muscular weakness, and/or muscle fatigue such as SMA, COPD, and ALS.

Cytokinetics , in collaboration with Astellas , is developing reldesemtiv, the lead from a program of selective fast skeletal muscle troponin activators, in an oral suspension formulation, for the treatment of indications associated with neuromuscular dysfunction, including spinal muscular atrophy and amyotrophic lateral sclerosis.

Originator Cytokinetics
Developer Astellas Pharma; Cytokinetics
Class Pyridines; Pyrimidines; Pyrroles; Small molecules
Mechanism of Action Troponin stimulants

Orphan Drug Status Yes – Spinal muscular atrophy

Phase II Amyotrophic lateral sclerosis; Chronic obstructive pulmonary disease; Spinal muscular atrophy
Suspended Muscle fatigue
No development reported Muscular atrophy

05 May 2019 Safety and efficacy data from the phase II FORTITUDE-ALS trial in Amyotrophic lateral sclerosis presented at the American Academy of Neurology Annual Meeting (AAN-2019)
07 Mar 2019 Cytokinetics completes the phase III FORTITUDE-ALS trial for Amyotrophic lateral sclerosis in USA, Australia, Canada, Spain, Ireland and Netherlands (PO) (NCT03160898)
22 Jan 2019 Cytokinetics plans a phase I trial in Healthy volunteers in the first quarter of 2019

Reldesemtiv, a next-generation, orally-available, highly specific small-molecule is being developed by Cytokinetics, in collaboration with Astellas Pharma, for the improvement of skeletal muscle function associated with neuromuscular dysfunction, muscle weakness and/or muscle fatigue in spinal muscular atrophy (SMA), chronic obstructive pulmonary disease (COPD) and amyotrophic lateral sclerosis (ALS). The drug candidate is a fast skeletal muscle troponin activator (FSTA) or troponin stimulant intended to slow the rate of calcium release from the regulatory troponin complex of fast skeletal muscle fibers. Clinical development for ALS, COPD and SMA is underway in the US, Australia, Canada, Ireland, Netherlands and Spain. No recent reports of development had been identified for phase I development for muscular atrophy in the US. Due to lack of of efficacy determined at interim analysis Cytokinetics suspended phase I trial in muscle fatigue in the elderly.

The cytoskeleton of skeletal and cardiac muscle cells is unique compared to that of all other cells. It consists of a nearly crystalline array of closely packed cytoskeletal proteins called the sarcomere. The sarcomere is elegantly organized as an interdigitating array of thin and thick filaments. The thick filaments are composed of myosin, the motor protein responsible for transducing the chemical energy of ATP hydrolysis into force and directed movement. The thin filaments are composed of actin monomers arranged in a helical array. There are four regulatory proteins bound to the actin filaments, which allows the contraction to be modulated by calcium ions. An influx of intracellular calcium initiates muscle contraction; thick and thin filaments slide past each other driven by repetitive interactions of the myosin motor domains with the thin actin filaments.

[0003] Of the thirteen distinct classes of myosin in human cells, the myosin-II class is responsible for contraction of skeletal, cardiac, and smooth muscle. This class of myosin is significantly different in amino acid composition and in overall structure from myosin in the other twelve distinct classes. Myosin-II forms homo-dimers resulting in two globular head domains linked together by a long alpha-helical coiled-coiled tail to form the core of the sarcomere’s thick filament. The globular heads have a catalytic domain where the actin binding and ATPase functions of myosin take place. Once bound to an actin filament, the release of phosphate (cf. ADP-Pi to ADP) signals a change in structural conformation of the catalytic domain that in turn alters the orientation of the light-chain binding lever arm domain that extends from the globular head; this movement is termed the powerstroke. This change in orientation of the myosin head in relationship to actin causes the thick filament of which it is a part to move with respect to the thin actin filament to which it is bound. Un-binding of the globular head from the actin filament (Ca²⁺ regulated) coupled with return of the catalytic domain and light chain to their starting conformation/orientation completes the catalytic cycle, responsible for intracellular movement and muscle contraction.

Tropomyosin and troponin mediate the calcium effect on the interaction on actin and myosin. The troponin complex is comprised of three polypeptide chains: troponin C, which binds calcium ions; troponin I, which binds to actin; and troponin T, which binds to tropomyosin. The skeletal troponin-tropomyosin complex regulates the myosin binding sites extending over several actin units at once.

Troponin, a complex of the three polypeptides described above, is an accessory protein that is closely associated with actin filaments in vertebrate muscle. The troponin complex acts in conjunction with the muscle form of tropomyosin to mediate the

Ca²⁺ dependency of myosin ATPase activity and thereby regulate muscle contraction. The troponin polypeptides T, I, and C, are named for their tropomyosin binding, inhibitory, and calcium binding activities, respectively. Troponin T binds to tropomyosin and is believed to be responsible for positioning the troponin complex on the muscle thin filament. Troponin I binds to actin, and the complex formed by troponins I and T, and tropomyosin inhibits the interaction of actin and myosin. Skeletal troponin C is capable of binding up to four calcium molecules. Studies suggest that when the level of calcium in the muscle is raised, troponin C exposes a binding site for troponin I, recruiting it away from actin. This causes the tropomyosin molecule to shift its position as well, thereby exposing the myosin binding sites on actin and stimulating myosin ATPase activity.

U.S. Patent No. 8962632 discloses l-(2-((((trans)-3-fluoro-l-(3-fluoropyridin-2-yl)cyclobutyl)methyl)amino)pyrimidin-5-yl)-lH-pyrrole-3-carboxamide, a next-generation fast skeletal muscle troponin activator (FSTA) as a potential treatment for people living with debilitating diseases and conditions associated with neuromuscular or non-neuromuscular dysfunction, muscular weakness, and/or muscle fatigue.

PATENT

WO 2011133888

https://patentscope.wipo.int/search/en/detail.jsf?docId=WO2011133888&recNum=202&docAn=US2011033614&queryString=&maxRec=57668

PATENT

WO2016039367 ,

https://patentscope.wipo.int/search/en/detail.jsf?docId=WO2016039367&tab=FULLTEXT

claiming the use of a similar compound for treating stress urinary incontinence.

Compound A is 1- [2-({[trans-3-fluoro-1- (3-fluoropyridin-2-yl) cyclobutyl] methyl} amino) pyrimidin-5-yl] -1H Pyrrole-3-carboxamide, which is the compound described in Example 14 of the aforementioned US Pat. The chemical structure is as shown below.

[Chemical formula 1]

PATENT

WO-2019133605

https://patentscope.wipo.int/search/en/detail.jsf?docId=WO2019133605&tab=PCTDESCRIPTION&_cid=P11-JXY4C3-99085-1

Process for preparing reldesemtiv , a myosin, actin, tropomyosin, troponin C, troponin I, troponin T modulator, useful for treating neuromuscular disorders, muscle wasting, claudication and metabolic syndrome.

Scheme 1

[0091] Scheme 1 illustrates a scheme of synthesizing the compound of Formula (1C).

Scheme 2

[0092] Scheme 2 illustrates an alternative scheme of synthesizing the compound of Formula (1C).

TFAA DS, toluene

HCI, H₂0

50°C

Scheme 3

[0093] Scheme 3 illustrates a scheme of converting the compound of Formula (1C) to the compound of Formula (II).

H₂

Ni Raney

NH₃

Scheme 4

[0094] Scheme 4 illustrates a scheme of converting the compound of Formula (II) to the compound of Formula (1).

Examples

[0095] To a flask was added N-methylpyrrolidone (30 mL), tert-butyl cyanoacetate (8.08 g) at room temperature. To a resulting solution was added potassium tert-butoxide (7.71 g), l,3-dibromo-2,2-dimethoxy propane (5.00 g) at 0 °C. To another flask, potassium iodide (158 mg), 2,6-di-tert-butyl-p-cresol (42 mg), N-methylpyrrolidone (25 mL) were added at room temperature and then resulting solution was heated to 165 °C. To this solution, previously prepared mixture was added dropwise at 140-165 °C, then stirred for 2 hours at 165 °C. To the reaction mixture, water (65 mL) was added. A resulting solution was extracted with toluene (40 mL, three times) and then combined organic layer was washed with water (20 mL, three times) and 1N NaOH aq. (20 mL). A resulting organic layer was concentrated below 50 °C under reduced pressure to give 3, 3 -dimethoxy cyclobutane- l-carbonitrile (66% yield,

GC assay) as toluene solution. 1H MR (CDCl₃, 400 MHz) d 3.17 (s, 3H), 3.15 (s, 3H), 2.93-2.84 (m, 1H), 2.63-2.57 (m, 2H), 2.52-2.45 (m, 2H).

Example 2 Synthesis of methyl 3,3-dimethoxycyclobutane-l-carboxylate

[0096] A reactor was vacuumed to 0.02 MPa and less and then inerted with nitrogen to atmosphere for three times. MeOH (339.00 kg), 3-oxocyclobutanecarboxylic acid (85.19 kg, 746.6 mol, 1.0 eq.), Amberlyst-l5 ion exchange resin (8.90 kg, 10% w/w), and

trimethoxymethane (196.00 kg, 1847.3 mol, 2.5 eq.) were charged into the reactor and the resulting mixture was heated to 55±5°C and reacted for 6 hours to give methyl 3,3-dimethoxycyclobutane-l-carboxylate solution in MeOH. 1H NMR (CDCl₃, 400 MHz) d 3.70 (s, 3H), 3.17 (s, 3H), 3.15 (s, 3H), 2.94-2.85 (m, 1H), 2.47-2.36 (m, 4H).

Example 3 Synthesis of 3, 3-dimethoxycyclobutane-l -carboxamide

[0097] The methyl 3, 3 -dimethoxy cyclobutane- l-carboxylate solution in MeOH prepared as described in Example 2 was cooled to below 25°C and centrifuged. The filter cake was washed with MeOH(7.00 kg) and the filtrate was pumped to the reactor. The solution was concentrated under vacuum below 55°C until the system had no more than 2 volumes. MeOH

(139.40 kg) was charged to the reactor and the solution was concentrated under vacuum below 55°C until the system had no more than 2 volumes. MeOH (130.00 kg) was charged to the reactor and the solution was concentrated under vacuum below 55°C until the system had no more than 2 volumes. Half of the resulting solution was diluted with MeOH (435.00 kg) and cooled to below 30°C. NH₃ gas (133.80 kg) was injected into the reactor below 35°C for

24 hours. The mixture was stirred at 40±5°C for 72 hours. The resulting solution was

concentrated under vacuum below 50°C until the system had no more than 2 volumes.

MTBE(l8l.OO kg) was charged into the reactor. The resulting solution was concentrated under vacuum below 50°C until the system had no more than 2 volumes. PE (318.00 kg) was charged into the reactor. The resulting mixture was cooled to 5±5°C, stirred for 4 hours at 5±5°C, and centrifuged. The filter cake was washed with PE (42.00 kg) and the wet filter cake was put into a vacuum oven. The filter cake was dried at 30±5°C for at least 8 hours to give 3,3-dimethoxycyclobutane-l-carboxamide as off-white solid (112.63 kg, 94.7% yield). 1H NMR (CDCf, 400 MHz) d 5.76 (bs, 1H), 5.64 (bs, 1H), 3.18 (s, 3H), 3.17 (s, 3H), 2.84-2.76 (m, 1H), 2.45-2.38 (m, 4H).

[0098] A reactor was vacuumed to 0.02 MPa and less and then inerted with nitrogen to atmosphere for three times. Toluene (500.00 kg), 3,3-dimethoxycyclobutane-l-carboxamide (112.54kg, 706.9 mol, 1.0 eq.), and TEA (158.00 kg, 1561.3 mol, 2.20 eq) were charged into the reactor and the resulting mixture was cooled to 0+ 5°C. TFAA (164.00 kg, 781 mol, 1.10 eq.) was added dropwise at 0±5°C. The resulting mixture was stirred for 10 hours at 20±5°C and cooled below 5±5°C. H₂0 (110.00 kg) was charged into the reactor at below 15 °C. The resulting mixture was stirred for 30 minutes and the water phase was separated. The aqueous phase was extracted with toluene (190.00 kg) twice. The organic phases were combined and washed with H₂0 (111.00 kg). H₂0 was removed by azeotrope until the water content was no more than 0.03%. The resulting solution was cooled to below 20°C to give 3,3-dimethoxycyclobutane-l-carbonitrile solution in toluene (492.00 kg with 17.83% assay content, 87.9% yield).

Example 5 Synthesis of l-(3-fluoropyridin-2-yl)-3,3-dimethoxycyclobutane-l-carbonitrile

[0099] A reactor was vacuumed to 0.02 MPa and less and then inerted with nitrogen to atmosphere for three times. The 3,3-dimethoxycyclobutane-l-carbonitrile solution in toluene prepared as described in Example 4 (246.00 kg of a 17.8% solution of 3,3-dimethoxycyclobutane-l-carbonitrile in toluene, 1.05 eq.) and 2-chloro-3-fluoropyridine (39.17 kg, 297.9 mol, 1.00 eq.) were charged into the reactor. The reactor was vacuumed to 0.02 MPa and less and then inerted with nitrogen to atmosphere for three times. The mixture was slowly cooled to -20±5°C. NaHDMS (2M in THF) (165.71 kg, 1.20 eq) was added

dropwise at -20±5°C. The resulting mixture was stirred at -l5±5°C for 1 hour. The mixture was stirred until the content of 2-chloro-3-fluoropyridine is no more than 2% as measured by HPLC. Soft water (16.00 kg) was added dropwise at below 0°C while maintaining the reactor temperature. The resulting solution was transferred to another reactor. Aq. NH₄Cl (10% w/w, 88.60 Kg) was added dropwise at below 0°C while maintaining the reactor temperature. Soft water (112.00 kg) was charged into the reactor and the aqueous phase was separated and collected. The aqueous phase was extracted with ethyl acetate (70.00 kg) and an organic phase was collected. The organic phase was washed with sat. NaCl (106.00 kg) and collected. The above steps were repeated to obtain another batch of organic phase. The two batches of organic phase were concentrated under vacuum below 70°C until the system had no more than 2 volumes. The resulting solution was cooled to below 30°C to give a l-(3-fluoropyridin-2-yl)-3, 3 -dimethoxy cyclobutane- l-carbonitrile solution. 1H NMR (CDC1₃, 400 MHz) d 8.42-8.38 (m, 1H), 7.50-7.45 (m, 1H), 7.38-7.33 (m, 1H), 3.28 (s, 3 H), 3.13 (s, 3H), 3.09-3.05 (m, 4H).

Example 6 Synthesis of I-(3-fluoropyridin-2-yl)-3-oxocyclohutanecarhonitrile

[0100] A reactor was vacuumed to 0.02 MPa and less and then inerted with nitrogen to atmosphere for three times. Water (603.00 kg) was added to the reactor and was stirred.

Concentrated HC1 (157.30 kg) was charged into the reactor at below 35°C. The l-(3-fluoropyridin-2-yl)-3, 3 -dimethoxy cyclobutane- l-carbonitrile solution prepared as described in Example 5 (206.00 kg) was charged into the reactor and the resulting mixture was heated to 50±5°C and reacted for 3 hours at 50±5°C. The mixture was reacted until the content of 1-(3 -fluoropyridin-2-yl)-3, 3 -dimethoxycyclobutane- l-carbonitrile was no more than 2.0% as measured by HPLC. The reaction mixture was cooled to below 30°C and extracted with ethyl acetate (771.00 kg). An aqueous phase was collected and extracted with ethyl acetate (770.00 kg). The organic phases were combined and the combined organic phase was washed with soft water (290.00 kg) and brine (385.30 kg). The organic phase was concentrated under vacuum at below 60°C until the system had no more than 2 volumes. Propan-2-ol (218.00 kg) was charged into the reactor. The organic phase was concentrated under vacuum at below

60°C until the system had no more than 1 volume. PE (191.00 kg) was charged into the reactor at 40±5 °C and the resulting mixture was heated to 60±5 °C and stirred for 1 hour at 60±5 °C. The mixture was then slowly cooled to 5±5 °C and stirred for 5 hours at 5±5 °C. The mixture was centrifuged and the filter cake was washed with PE (48.00 kg) and the wet filter cake was collected. Water (80.00 kg), concentrated HC1 (2.20 kg), propan-2-ol (65.00 kg), and the wet filter cake were charged in this order into a drum. The resulting mixture was stirred for 10 minutes at 20±5 °C. The mixture was centrifuged and the filter cake was washed with a mixture solution containing 18.00 kg of propan-2-ol, 22.50 kg of soft water, and 0.60 kg of concentrated HC1. The filter cake was put into a vacuum oven and dried at 30±5°C for at least 10 hours. The filter cake was dried until the weight did not change to give l-(3-fluoropyridin-2-yl)-3-oxocyclobutanecarbonitrile as off-white solid (77.15 kg, 68.0% yield). 1H NMR (CDCl₃, 400 MHz) d 8.45-8.42 (m, 1H), 7.60-7.54 (m, 1H), 7.47-7.41 (m, 1H), 4.18-4.09 (m, 2H), 4.02-3.94 (m, 2H).

Example 7 Synthesis of I-(3-fhtoropyridin-2-yl)-3-hydroxycyclobulanecarbonilrile

[0101] To a solution of l-(3-fluoropyridin-2-yl)-3-oxocyclobutanecarbonitrile (231 g,

1.22 mol) in a mixture ofDCM (2 L) and MeOH (200 mL) was added NaBH₄ portionwise at -78° C. The reaction mixture was stirred at -78°C. for 1 hour and quenched with a mixture of methanol and water (1 : 1). The organic layer was washed with water (500 mL><3), dried over Na₂S0₄, and concentrated. The residue was purified on silica gel (50% EtO Ac/hexanes) to provide the title compound as an amber oil (185.8 g, 77.5%). Low Resolution Mass

Spectrometry (LRMS) (M+H) m/z 193.2.

Example 8 Synthesis of (ls,3s)-3-fluoro-l-(3-fluoropyridin-2-yl)cyclobutane-l-carbonitrile

[0102] To a solution of 1 -(3 -fluoropyridin-2-yl)-3 -hydroxy cyclobutanecarbonitrile (185 g, 0.96 mol) in DCM (1 L) was added DAST portionwise at 0-10 °C. Upon the completion of addition, the reaction was refluxed for 6 hours. The reaction was cooled to rt and poured onto sat. NaHCCf solution. The mixture was separated and the organic layer was washed with water, dried over Na₂S0₄, and concentrated. The residue was purified on silica gel (100% DCM) to provide the title compound as a brown oil (116g) in a 8: 1 transxis mixture. The above brown oil (107 g) was dissolved in toluene (110 mL) and hexanes (330mL) at 70 °C. The solution was cooled to 0 °C and stirred at 0 °C overnight. The precipitate was filtered and washed with hexanes to provide the trans isomer as a white solid (87.3 g). LRMS (M+H) m/z 195.1.

Example 9 Synthesis of ((lr,3r)-3-fluoro-l-(3-fluoropyridin-2-yl)cyclobutyl)methanamine

[0103] A mixture of ( 1.v,3.v)-3-fluoro- 1 -(3-fluoropyridin-2-yl)cyclobutane- 1 -carbonitrile (71 g, 0.37 mol) and Raney nickel (~7 g) in 7N ammonia in methanol (700 mL) was charged with hydrogen (60 psi) for 2 days. The reaction was filtered through a celite pad and washed with methanol. The filtrate was concentrated under high vacuum to provide the title compound as a light green oil (70 g, 97.6%). LRMS (M+H) m/z 199.2.

Example 10 Synthesis of t-butyl 5-bromopyrimidin-2-yl((trans-3-fluoro-l-(3-fluoropyridin-2-yl)cyclobutyl)methyl) carbamate

[0104] A mixture of ((lr,3r)-3-fluoro-l-(3-fluoropyridin-2-yl)cyclobutyl)methanamine (37.6 g, 190 mmol), 5-bromo-2-fluoropyrimidine (32.0 g, 181 mmol), DIPEA (71 mL, 407 mmol), and NMP (200 mL) was stirred at rt overnight. The reaction mixture was then diluted with EtOAc (1500 mL) and washed with saturated sodium bicarbonate (500 mL). The

organic layer was separated, dried over Na₂S0₄, and concentrated. The resultant solid was dissolved in THF (600 mL), followed by the slow addition of DMAP (14 g, 90 mmol) and Boc₂0 (117.3 g, 542 mmol). The reaction was heated to 60° C. and stirred for 3 h. The reaction mixture was then concentrated and purified by silica gel chromatography

(EtO Ac/hex) to give 59.7 g oft-butyl 5-bromopyrimidin-2-yl((trans-3-fluoro-l-(3-fluoropyridin-2-yl)cyclobutyl)methyl)carbamate as a white solid.

Example 11 Synthesis of t-butyl 5-(3-cyano- 1 H -pyrrol- 1 -yl)pyrimidin-2-yl(((lrans)-3-fhtoro-l-(3-fluoropyridin-2-yl)cyclohutyl)methyl)carhamate

[0105] To a solution oft-butyl 5-bromopyrimidin-2-yl((trans-3-fluoro-l-(3-fluoropyridin-2-yl)cyclobutyl)methyl) carbamate (1.0 g, 2.8 mmol) in 15 mL of toluene (degassed with nitrogen) was added copper iodide (100 mg, 0.6 mmol), potassium phosphate (1.31 g, 6.2 mmol), trans-N,N’-dimethylcyclohexane-l, 2-diamine (320 mg, 2.2 mmol), and 3-cyanopyrrole (310 mg, 3.6 mmol). The reaction was heated to 100 °C and stirred for 2 h. The reaction was then concentrated and purified by silica gel chromatography (EtOAc/hexanes) to afford 1.1 g of t-butyl 5-(3-cyano-lH-pyrrol-l-yl)pyrimidin-2-yl(((trans)-3-fluoro-l-(3-fluoropyridin-2-yl)cyclobutyl)methyl)carbamate as a clear oil.

Example 12 Synthesis of l-(2-((((trans)-3-fluoro-l-(3-fluoropyridin-2-yl)cyclobutyl)methyl)amino)pyrimidin-5-yl)-lH-pyrrole-3-carboxamide

[0106] To a solution oft-butyl 5-(3-cyano-lH-pyrrol-l-yl)pyrimidin-2-yl(((trans)-3-fluoro-l-(3-fluoropyridin-2-yl)cyclobutyl)methyl)carbamate (1.1 g, 3.1 mmol) in DMSO (10 mL) was added potassium carbonate (1.3 g, 9.3 mmol). The mixture was cooled to 0 °C and hydrogen peroxide (3 mL) was slowly added. The reaction was warmed to rt and stirred for 90 min. The reaction was diluted with EtO Ac (75 mL) and washed three times with brine (50 mL). The organic layer was then dried over Na₂S0₄, filtered, and concentrated to give a crude solid that was purified by silica gel chromatography (10% MeOH/CH₂Cl2) to afford 1.07 g of a white solid compound. This compound was dissolved in 25% TFA/CH2CI2 and stirred for 1 hour. The reaction was then concentrated, dissolved in ethyl acetate (75 mL), and washed three times with saturated potassium carbonate solution. The organic layer was then dried over Na₂S04, filtered, and concentrated to give a crude solid that was triturated with 75% ethyl acetate/hexanes. The resultant slurry was sonicated and filtered to give 500 mg of l-(2-((((trans)-3-fluoro-l-(3-fluoropyridin-2-yl)cyclobutyl)methyl)amino)pyrimidin-5-yl)-lH-pyrrole-3 -carboxamide as a white solid. LRMS (M+H=385).

REFERENCES

1: Andrews JA, Miller TM, Vijayakumar V, Stoltz R, James JK, Meng L, Wolff AA, Malik FI. CK-2127107 amplifies skeletal muscle response to nerve activation in humans. Muscle Nerve. 2018 May;57(5):729-734. doi: 10.1002/mus.26017. Epub 2017 Dec 11. PubMed PMID: 29150952.

2: Gross N. The COPD Pipeline XXXII. Chronic Obstr Pulm Dis. 2016 Jul 14;3(3):688-692. doi: 10.15326/jcopdf.3.3.2016.0150. PubMed PMID: 28848893; PubMed Central PMCID: PMC5556764.

//////////////CK-2127107, CK 2127107, CK2127107, Reldesemtiv, Cytokinetics, Astellas, neuromuscular disorders, muscle wasting, claudication, metabolic syndrome, spinal muscular atrophy, amyotrophic lateral sclerosis, Orphan Drug Status, Spinal muscular atrophy, Phase II

C1C(CC1(CNC2=NC=C(C=N2)N3C=CC(=C3)C(=O)N)C4=C(C=CC=N4)F)F

Ceralasertib, AZD 6738

July 8, 2019 12:39 pm / 1 Comment on Ceralasertib, AZD 6738

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AZD-6738, Ceralasertib

Molecular Formula C₂₀H₂₄N₆O₂S
Average mass 412.509 Da

CAS 1352226-88-0 [RN]

1H-Pyrrolo[2,3-c]pyridine, 4-[4-[(3R)-3-methyl-4-morpholinyl]-6-[1-(S-methylsulfonimidoyl)cyclopropyl]-2-pyrimidinyl]-

4-{4-[(3R)-3-Methyl-4-morpholinyl]-6-[1-(S-methylsulfonimidoyl)cyclopropyl]-2-pyrimidinyl}-1H-pyrrolo[2,3-c]pyridine

1H-Pyrrolo(2,3-b)pyridine, 4-(4-(1-((S(R))-S-methylsulfonimidoyl)cyclopropyl)-6-((3R)-3-methyl-4-morpholinyl)-2-pyrimidinyl)-

imino-methyl-[1-[6-[(3R)-3-methylmorpholin-4-yl]-2-(1H-pyrrolo[2,3-b]pyridin-4-yl)pyrimidin-4-yl]cyclopropyl]-oxo-λ6-sulfane

85RE35306Z

AZD-6738

UNII:85RE35306Z

CAS : 1352226-88-0 (free base) 1352280-98-8 (formic acid) 1352226-97-1 (racemic)

4-[4-[1-[[S(R)]-S-Methylsulfonimidoyl]cyclopropyl]-6-[(3R)-3-methyl-4-morpholinyl]-2-pyrimidinyl]-1H-pyrrolo[2,3-b]pyridine
AZD 6738
Ceralasertib

Originator AstraZeneca; University of Pennsylvania
Class Antineoplastics; Morpholines; Pyrimidines; Small molecules
Mechanism of Action ATR protein inhibitors

Phase II Breast cancer; Gastric cancer; Non-small cell lung cancer; Ovarian cancer
Phase I/II Chronic lymphocytic leukaemia; Solid tumours
Phase I Non-Hodgkin’s lymphoma
Preclinical Diffuse large B cell lymphoma
No development reported B-cell lymphoma; Lymphoid leukaemia

26 Mar 2019 National Cancer Institute plans a phase II trial for Cholangiocarcinoma (Combination therapy, Second-line therapy or greater) and Solid tumours (Combination therapy, Second-line therapy or greater) in March 2019 (NCT03878095)
18 Mar 2019 Royal Marsden NHS Foundation Trust and AstraZeneca re-initiate the phase I PATRIOT trial in Solid tumours (Second-line therapy or greater) in United Kingdom (NCT02223923)
25 Dec 2018 University of Michigan Cancer Center plans the phase II TRAP trial for Prostate cancer (Combination therapy; Metastatic disease; Second-line therapy or greater) in February 2019 (NCT03787680)

Inhibits ATR kinase.

Ceralasertib, also known as AZD6738, is an orally available morpholino-pyrimidine-based inhibitor of ataxia telangiectasia and rad3 related (ATR) kinase, with potential antineoplastic activity. Upon oral administration, ATR kinase inhibitor Ceralasertib selectively inhibits ATR activity by blocking the downstream phosphorylation of the serine/threonine protein kinase CHK1. This prevents ATR-mediated signaling, and results in the inhibition of DNA damage checkpoint activation, disruption of DNA damage repair, and the induction of tumor cell apoptosis.

ATR (also known as FRAP-Related Protein 1; FRP1; MEC1; SCKL; SECKL1) protein kinase is a member of the PI3 -Kinase like kinase (PIKK) family of proteins that are involved in repair and maintenance of the genome and its stability (reviewed in Cimprich K.A. and Cortez D. 2008, Nature Rev. Mol. Cell Biol. 9:616-627). These proteins co-ordinate response to DNA damage, stress and cell-cycle perturbation. Indeed ATM and ATR, two members of the family of proteins, share a number of downstream substrates that are themselves recognised components of the cell cycle and DNA-repair machinery e.g. Chkl, BRCAl, p53 (Lakin ND et al,1999, Oncogene; Tibbets RS et al, 2000, Genes & Dev.). Whilst the substrates of ATM and ATR are to an extent shared, the trigger to activate the signalling cascade is not shared and ATR primarily responds to stalled replication forks (Nyberg K.A. et al., 2002, Ann. Rev.

Genet. 36:617-656; Shechter D. et al. 2004, DNA Repair 3:901-908) and bulky DNA damage lesions such as those formed by ultraviolet (UV) radiation (Wright J. A. et al, 1998, Proc. Natl. Acad. Sci. USA, 23:7445-7450) or the UV mimetic agent, 4-nitroquinoline-1-oxi-e, 4NQO (Ikenaga M. et al. 1975, Basic Life Sci. 5b, 763-771). However, double strand breaks (DSB) detected by ATM can be processed into single strand breaks (SSB) recruiting ATR; similarly SSB, detected by ATR can generate DSB, activating ATM. There is therefore a significant interplay between ATM and ATR.

Mutations of the ATR gene that result in complete loss of expression of the ATR protein are rare and in general are not viable. Viability may only result under heterozygous or hypomorphic conditions. The only clear link between ATR gene mutations and disease exists in a few patients with Seckel syndrome which is characterized by growth retardation and microcephaly (O’Driscoll M et al, 2003 Nature Genet. Vol3, 497-501). Cells from patients with hypomorphic germline mutations of ATR (seckel syndrome) present a greater susceptibility to chromosome breakage at fragile sites in presence of replication stress compared to wild type cells (Casper 2004). Disruption of the ATR pathway leads to genomic instability. Patients with Seckel syndrome also present an increased incidence of cancer,suggestive of the role of ATR in this disease in the maintenance of genome stability .

Moreover, duplication of the ATR gene has been described as a risk factor in rhabdomyosarcomas (Smith L et al, 1998, Nature Genetics 19, 39-46). Oncogene-driven tumorigenesis may be associated with ATM loss-of- function and therefore increased reliance on ATR signalling (Gilad 2010). Evidence of replication stress has also been reported in several tumour types such as colon and ovarian cancer, and more recently in glioblastoma, bladder, prostate and breast (Gorgoulis et al, 2005; Bartkova et al. 2005a; Fan et al., 2006; Tort et al, 2006; Nuciforo et al, 2007; Bartkova et al., 2007a). Loss of Gl checkpoint is also frequently observed during tumourigenesis. Tumour cells that are deficient in Gl checkpoint controls, in particular p53 deficiency, are susceptible to inhibition of ATR activity and present with premature chromatin condensation (PCC) and cell death (Ngheim et al, PNAS, 98, 9092-9097).

ATR is essential to the viability of replicating cells and is activated during S-phase to regulate firing of replication origins and to repair damaged replication forks (Shechter D et al, 2004, Nature cell Biology Vol 6 (7) 648-655). Damage to replication forks may arise due to exposure of cells to clinically relevant cytotoxic agents such as hydroxyurea (HU) and platinums (O’Connell and Cimprich 2005; 118, 1-6). ATR is activated by most cancer chemotherapies (Wilsker D et al, 2007, Mol. Cancer Ther. 6(4) 1406-1413). Biological assessment of the ability of ATR inhibitors to sensitise to a wide range of chemotherapies have been evaluated. Sensitisation of tumour cells to chemotherapeutic agents in cell growth assays has been noted and used to assess how well weak ATR inhibitors (such as Caffeine) will sensitise tumour cell lines to cytotoxic agents. (Wilsker D .et al, 2007, Mol Cancer Ther. 6 (4)1406-1413; Sarkaria J.N. et al, 1999, Cancer Res. 59, 4375-4382). Moreover, a reduction of ATR activity by siRNA or ATR knock-in using a dominant negative form of ATR in cancer cells has resulted in the sensitisation of tumour cells to the effects of a number of therapeutic or experimental agents such as antimetabolites (5-FU, Gemcitabine, Hydroxyurea, Metotrexate, Tomudex), alkylating agents (Cisplatin, Mitomycin C, Cyclophosphamide, MMS) or double-strand break inducers (Doxorubicin, Ionizing radiation) (Cortez D. et al. 2001, Science, 294:1713-1716; Collis S.J. et al, 2003, Cancer Res. 63:1550-1554; Cliby W.A. et al, 1998, EMBO J. 2:159-169) suggesting that the combination of ATR inhibitors with some cytotoxic agents might be therapeutically beneficial.

An additional phenotypic assay has been described to define the activity of specific ATR inhibitory compounds is the cell cycle profile (PJ Hurley, D Wilsker and F Bunz, Oncogene, 2007, 26, 2535-2542). Cells deficient in ATR have been shown to have defective cell cycle regulation and distinct characteristic profiles, particularly following a cytotoxic cellular insult. Furthermore, there are proposed to be differential responses between tumour and normal tissues in response to modulation of the ATR axis and this provides further potential for therapeutic intervention by ATR inhibitor molecules (Rodnguez-Bravo V et al, Cancer Res., 2007, 67, 11648-11656).

Another compelling utility of ATR-specific phenotypes is aligned with the concept of synthetic lethality and the observation that tumour cells that are deficient in G1 checkpoint controls, in particular p53 deficiency, are susceptible to inhibition of ATR activity resulting in premature chromatin condensation (PCC) and cell death (Ngheim et al, PNAS, 98, 9092-9097). In this situation, S-phase replication of DNA occurs but is not completed prior to M-phase initiation due to failure in the intervening checkpoints resulting in cell death from a lack of ATR signalling. The G2/M checkpoint is a key regulatory control involving ATR (Brown E. J. and Baltimore D., 2003, Genes Dev. 17, 615-628) and it is the compromise of this checkpoint and the prevention of ATR signalling to its downstream partners which results in PCC. Consequently, the genome of the daughter cells is compromised and viability of the cells is lost (Ngheim et al, PNAS, 98, 9092-9097).

It has thus been proposed that inhibition of ATR may prove to be an efficacious approach to future cancer therapy (Collins I. and Garret M.D., 2005, Curr. Opin. Pharmacol., 5:366-373; Kaelin W.G. 2005, Nature Rev. Cancer, 5:689-698) in the appropriate genetic context such as tumours with defects in ATM function or other S-phase checkpoints. Until recently, There is currently no clinical precedent for agents targeting ATR, although agents targeting the downstream signalling axis i.e. Chk1 are currently undergoing clinical evaluation (reviewed in Janetka J.W. et al. Curr Opin Drug Discov Devel, 2007, 10:473-486). However, inhibitors targeting ATR kinase have recently been described (Reaper 2011, Charrier 2011).

In summary ATR inhibitors have the potential to sensitise tumour cells to ionising radiation or DNA-damage inducing chemotherapeutic agents, have the potential to induce selective tumour cell killing as well as to induce synthetic lethality in subsets of tumour cells with defects in DNA damage response.

PAPER

Discovery and Characterization of AZD6738, a Potent Inhibitor of Ataxia Telangiectasia Mutated and Rad3 Related (ATR) Kinase with Application as an Anticancer Agent

Kevin M. Foote

Cite This:J. Med. Chem.201861229889-9907

Publication Date:October 22, 2018

https://doi.org/10.1021/acs.jmedchem.8b01187

The kinase ataxia telangiectasia mutated and rad3 related (ATR) is a key regulator of the DNA-damage response and the apical kinase which orchestrates the cellular processes that repair stalled replication forks (replication stress) and associated DNA double-strand breaks. Inhibition of repair pathways mediated by ATR in a context where alternative pathways are less active is expected to aid clinical response by increasing replication stress. Here we describe the development of the clinical candidate 2(AZD6738), a potent and selective sulfoximine morpholinopyrimidine ATR inhibitor with excellent preclinical physicochemical and pharmacokinetic (PK) characteristics. Compound 2 was developed improving aqueous solubility and eliminating CYP3A4 time-dependent inhibition starting from the earlier described inhibitor 1 (AZ20). The clinical candidate 2 has favorable human PK suitable for once or twice daily dosing and achieves biologically effective exposure at moderate doses. Compound 2 is currently being tested in multiple phase I/II trials as an anticancer agent.

ATR Inhibitors

4-{4-[(3R)-3-Methylmorpholin-4-yl]-6-[1-((R)-S-methylsulfonimidoyl)cyclopropyl]pyrimidin-2-yl}-1H-pyrrolo[2,3-b]pyridine (2)

2 (139 g, 42%) as a white crystalline solid.

¹H NMR (400 MHz, DMSO-d₆): 1.19 (3H, d), 1.29–1.50 (3H, m), 1.61–1.72 (1H, m), 3.01 (3H, s), 3.22 (1H, d), 3.43 (1H, td), 3.58 (1H, dd), 3.68–3.76 (2H, m), 3.87–3.96 (1H, m), 4.17 (1H, d), 4.60 (1H, s), 6.98 (1H, s), 7.20 (1H, dd), 7.55–7.58 (1H, m), 7.92 (1H, d), 8.60 (1H, d), 11.67 (1H, s).

¹³C NMR (176 MHz, DMSO-d₆) 11.29, 12.22, 13.39, 38.92, 41.14, 46.48, 47.81, 65.97, 70.19, 101.54, 102.82, 114.58, 117.71, 127.21, 136.70, 142.21, 150.12, 161.88, 162.63, 163.20.

HRMS-ESI m/z 413.17529 [MH⁺]; C₂₀H₂₄N₆O₂S requires 413.1760.

Chiral HPLC: (HP1100 system 4, 5 μm Chiralpak AS-H (250 mm × 4.6 mm) column, eluting with isohexane/EtOH/MeOH/TEA 50/25/25/0.1) R_f = 8.252, >99%. Anal. Found (% w/w): C, 58.36; H, 5.87; N, 20.20; S, 7.55; H₂O, <0.14. C₂₀H₂₄N₆O₂S requires C, 58.23; H, 5.86; N, 20.37; S, 7.77.

Patent

WO 2011154737

https://patentscope.wipo.int/search/en/detail.jsf;jsessionid=CF8CA857FDD8BF59DA9F336056132BB7.wapp2nA?docId=WO2011154737&tab=PCTDESCRIPTION

Example 1.01

4-{4-[(3R)-3-Methylmorpholin-4-yl]-6-[((R)-S-methylsulfonimidoyl)methyl]pyrimidin-2-yl}-1H-pyrrolo[2,3-b]pyridine

(R)-3-Methyl-4-(6-((R)-S-methylsulfonimidoylmethyl)-2-(1-tosyl-1H-pyrrolo[2,3-b]pyridin-4-yl)pyrimidin-4-yl)morpholine (98 mg, 0.18 mmol) was dissolved in MeOH (10 ml) and DCM (10 ml) and heated to 50 °C. Sodium hydroxide, 2M aqueous solution (0.159 ml, 0.32 mmol) was then added and heating continued for 5 hours. The reaction mixture was evaporated and the residue dissolved in DME: water :MeCN 2: 1 : 1 (4 ml) and then purified by preparative HPLC using decreasingly polar mixtures of water (containing 1% NH₃) and MeCN as eluents. Fractions containing the desired compound were evaporated and the residue trituated with Et₂O

(1 ml) to afford the title compound (34.6 mg, 49%); ¹HNMR (400 MHz, CDCl₃) 1.40 (3H, d), 3.17 (3H, s), 3.39 (1H, tt), 3.62 (1H, td), 3.77 (1H, dd), 3.85 (1H, d), 4.08 (1H, dd), 4.18 (1H, d), 4.37 – 4.48 (2H, q), 4.51 (1H, s), 6.59 (1H, s), 7.35 (1H, t), 7.46 (1H, d), 8.06 (1H, d), 8.42 (1H, d), 10.16 (1H, s); m/z: (ES+) MH⁺, 387.19.

The (R)-3-methyl-4-(6-((R)-S-methylsulfonimidoylmethyl)-2-(1-tosyl-1H-pyrrolo[2,3-b]pyridin-4-yl)pyrimidin-4-yl)morpholine, used as starting material, can be prepared as follows:

a) (R)-3-methylmorpholine (7.18 g, 71.01 mmol) and triethylamine (12.87 ml, 92.31 mmol) were added to methyl 2,4-dichloropyrimidine-6-carboxylate (14.70 g, 71.01 mmol) in DCM (100 ml). The resulting mixture was stirred at RT for 18 hours. Water (100 ml) was added, the layers separated and extracted with DCM (3 × 75 ml). The combined organics were

dried over MgSO₄, concentrated in vacuo and the residue triturated with Et₂O to yield (R)-methyl 2-chloro-6-(3-methylmorpholino)pyrimidine-4-carboxylate (14.77 g, 77%); ¹H NMR (400 MHz, CDCl₃) 1.35 (3H, d), 3.34 (1H, td), 3.55 (1H, td), 3.70 (1H, dd), 3.81 (1H, d), 3.97 (3H, s), 4.03 (1H, dd), 4.12 (1H, br s), 4.37 (1H, br s), 7.15 (1H, s); m/z: (ESI+) MH⁺, 272.43. The liquors were concentrated onto silica and purified by chromatography on silica eluting with a gradient of 20 to 40% EtOAc in isohexane. Fractions containing product were combined and evaporated to afford (R)-methyl 2-chloro-6-(3-methylmorpholino)pyrimidine-4-carboxylate (1.659 g, 9%); ¹H NMR (400 MHz, CDCl₃) 1.35 (3H, d), 3.33 (1H, td), 3.55 (1H, td), 3.69 (1H, dd), 3.80 (1H, d), 3.97 (3H, s), 4.03 (1H, dd), 4.12 (1H, br s), 4.36 (1H, br s), 7.15 (1H, s); m/z: (ESI+) MH⁺, 272.43.

b) Lithium borohydride, 2M in THF (18 ml, 36.00 mmol) was added dropwise to (R)-methyl 2-chloro-6-(3-methylmorpholino)pyrimidine-4-carboxylate (16.28 g, 59.92 mmol) in THF (200 ml) at 0°C over a period of 20 minutes under nitrogen. The resulting solution was stirred at 0 °C for 30 minutes and then allowed to warm to RT and stirred for a further 18 hours. Water (200 ml) was added and the THF evaporated. The aqueous layer was extracted with EtOAc (2 × 100 ml) and the organic phases combined, dried over MgSO4 and then evaporated to afford (R)-(2-chloro-6-(3-methylmorpholino)pyrimidin-4-yl)methanol (14.54 g, 100%) which was used in the next step without purification; ¹HNMR (400 MHz, CDCl₃) 1.32 (3H, d), 2.65 (1H, br s), 3.25 – 3.32 (1H, m), 3.51 – 3.57 (1H, m), 3.67 – 3.70 (1H, m), 3.78 (1H, d), 3.98 – 4.09 (2H, m), 4.32 (1H, br s), 4.59 (2H, s), 6.44 (1H, s); m/z: (ESI+) MH⁺, 244.40.

c) Methanesulfonyl chloride (4.62 ml, 59.67 mmol) was added dropwise to (R)-(2-chloro-6-(3-methylmorpholino)pyrimidin-4-yl)methanol (14.54 g, 59.67 mmol) and triethylamine (8.32 ml, 59.67 mmol) in DCM (250 ml) at 25 °C over a period of 5 minutes. The resulting solution was stirred at 25 °C for 90 minutes. The reaction mixture was quenched with water (100 ml) and extracted with DCM (2 × 100 ml). The organic phases were combined, dried over MgSO₄, filtered and evaporated to afford (R)-(2-chloro-6-(3-methylmorpholino)pyrimidin-4-yl)methyl methanesulfonate (20.14 g, 105%) which was used in the next step without further purification; ¹H NMR (400 MHz, CDCl₃) 1.33 (3H, d), 3.13 (3H, s), 3.27 – 3.34 (1H, m), 3.51 -3.57 (1H, m), 3.66 – 3.70 (1H, m), 3.79 (1H, d), 3.99 – 4.03 (2H, m), 4.34 (1H, br s), 5.09 (2H, d) , 6.52 (1H, s); m/z: (ESI+) MH⁺, 322.83.

Alternatively, this step can be carried out as follows:

In a 3 L fixed reaction vessel with a Huber 360 heater / chiller attached, under a nitrogen atmosphere, triethylamine (0.120 L, 858.88 mmol) was added in one go to a stirred solution of (R)-(2-chloro-6-(3-methylmorpholino)pyrimidin-4-yl)methanol (161 g, 660.68 mmol) in DCM (7.5vol) (1.2 L) at 20°C (3°C exotherm seen). The mixture was cooled to 5°C and then methanesulfonyl chloride (0.062 L, 792.81 mmol) was added dropwise over 15 minutes, not allowing the internal temperature to exceed 15°C. The reaction mixture was stirred at 15°C for 2 hours and then held (not stirring) overnight at RT under a nitrogen atmosphere. Water (1.6 L, 10 vol) was added and the aqueous layer was separated and then extracted with DCM (2 × 1.6 L, 2 × 10 vol). The organics were combined, washed with 50% brine / water (1.6 L, 10 vol), dried over magnesium sulphate, filtered and then evaporated to afford a mixture of

approximately two thirds (R)-(2-chloro-6-(3-methylmorpholino)pyrimidin-4-yl)methyl methanesulfonate and one third (R)-4-(2-chloro-6-(chloromethyl)pyrimidin-4-yl)-3-methylmorpholine (216 g) which was used in the next step without further purification, d) Lithium iodide (17.57 g, 131.27 mmol) was added to (R)-(2-chloro-6-(3-methylmorpholino)pyrimidin-4-yl)methyl methanesulfonate (19.2 g, 59.67 mmol) in dioxane (300 ml) and heated to 100 °C for 2 hours under nitrogen. The reaction mixture was quenched with water (200 ml) and extracted with EtOAc (3 × 200 ml). The organic layers were combined and washed with 2M sodium bisulfite solution (400 ml), water (400 ml), brine (400 ml) dried over MgSO₄ and then evaporated. The residue was triturated with Et₂O to afford (R)-4-(2-chloro-6-(iodomethyl)pyrimidin-4-yl)-3-methylmorpholine (13.89 g, 66%); ¹H NMR (400 MHz, CDCl₃) 1.32 (3H, d), 3.28 (1H, td), 3.54 (1H, td), 3.69 (1H, dd), 3.78 (1H, d), 3.98 -4.02 (2H, m), 4.21 (2H, s), 4.29 (1H, br s), 6.41 (1H, s); m/z: (ESI+) MH⁺ 354.31.

The mother liquors were concentrated down and triturated with Et₂O to afford a further crop of (R)-4-(2-chloro-6-(iodomethyl)pyrimidin-4-yl)-3-methylmorpholine (2.46 g, 12%); ¹HNMR (400 MHz, CDCI₃) 1.32 (3H, d), 3.28 (1H, td), 3.54 (1H, td), 3.69 (1H, dd), 3.78 (1H, d), 3.98 – 4.02 (2H, m), 4.21 (2H, s), 4.30 (1H, s), 6.41 (1H, s); m/z: (ESI+) MH⁺, 354.31.

Alternatively, this step can be carried out as follows:

(R)-(2-Chloro-6-(3-methylmorpholino)pyrimidin-4-yl)methyl methanesulfonate (80 g, 248.62 mmol) and lithium iodide (83 g, 621.54 mmol) were dissolved in dioxane (300 ml) and then heated at 107 °C for 1 hour. The reaction mixture was quenched with water (250 ml), extracted with EtOAc (3 × 250 ml), the organic layer was dried over MgSO4, filtered and evaporated. The residue was dissolved in DCM and Et2O was added, the mixture was passed through silica (4 inches) and eluted with Et₂O. Fractions containing product were evaporated and the residue was then triturated with Et₂O to give a solid which was collected by filtration and dried under vacuum to afford (R)-4-(2-chloro-6-(iodomethyl)pyrimidin-4-yl)-3-methylmorpholine (75 g, 86%) ; m/z: (ESI+) MH⁺, 354.27.

e) (R)-4-(2-Chloro-6-(iodomethyl)pyrimidin-4-yl)-3-methylmorpholine (17.0 g, 48.08 mmol) was dissolved in DMF (150 ml), to this was added sodium methanethiolate (3.37 g, 48.08 mmol) and the reaction was stirred for 1 hour at 25 °C. The reaction mixture was quenched with water (50 ml) and then extracted with Et₂O (3 × 50 ml). The organic layer was dried over MgSO₄, filtered and then evaporated. The residue was purified by flash

chromatography on silica, eluting with a gradient of 50 to 100% EtOAc in iso-hexane. Pure fractions were evaporated to afford (R)-4-(2-chloro-6-(methylthiomethyl)pyrimidin-4-yl)-3-methylmorpholine (12.63 g, 96%); m/z: (ES+) MH⁺, 274.35.

Alternatively, (R)-4-(2-chloro-6-(methylthiomethyl)pyrimidin-4-yl)-3-methylmorpholine, may be prepared as follows:

In a 3 L fixed vessel, sodium thiomethoxide (21% in water) (216 g, 646.69 mmol) was added dropwise over 5 minutes to a stirred solution of a mixture of approximately two thirds (R)-(2-chloro-6-(3-methylmorpholino)pyrimidin-4-yl)methyl methanesulfonate and one third (R)-4-(2-chloro-6-(chloromethyl)pyrimidin-4-yl)-3-methylmorpholine (130.2 g, 431 mmol) and sodium iodide (1.762 ml, 43.11 mmol) in MeCN (1 L) at RT (temperature dropped from 20 °C to 18 °C over the addition and then in the next 5 minutes rose to 30 °C). The reaction mixture was stirred for 16 hours and then diluted with EtOAc (2 L), and washed sequentially with water (750 ml) and saturated brine (1 L). The organic layer was dried over MgSO₄, filtered and then evaporated to afford (R)-4-(2-chloro-6-(methylthiomethyl)pyrimidin-4-yl)-3-methylmorpholine (108 g, 91%); ¹H NMR (400 MHz, DMSO- d₆) 1.20 (3H, d), 2.07 (3H, s), 3.11 – 3.26 (1H, m), 3.44 (1H, td), 3.53 (2H, s), 3.59 (1H, dd), 3.71 (1H, d), 3.92 (1H, dd), 3.92 – 4.04 (1H, br s), 4.33 (1H, s), 6.77 (1H, s); m/z: (ES+) MH⁺, 274.36.

f) (R)-4-(2-Chloro-6-(methylthiomethyl)pyrimidin-4-yl)-3-methylmorpholine (12.63 g, 46.13 mmol) was dissolved in DCM (100 ml), to this was added mCPBA (7.96 g, 46.13 mmol) in one portion and the reaction mixture was stirred for 10 minutes at 25 °C. An additional portion of mCPBA (0.180 g) was added. The reaction mixture was quenched with saturated Na₂CO₃ solution (50 ml) and extracted with DCM (3 × 50 ml). The organic layer was dried over MgSO₄, filtered and then evaporated. The residue was dissolved in DCM (80 ml) in a 150

ml conical flask which was placed into a beaker containing Et₂O (200 ml) and the system covered with laboratory film and then left for 3 days. The obtained crystals were filtered, crushed and sonicated with Et₂O. The crystallisation procedure was repeated to afford (R)-4-(2-chloro-6-((R)-methylsulfinylmethyl)pyrimidin-4-yl)-3-methylmorpholine as white needles (3.87 g, 29%); ¹HNMR (400 MHz, CDCl₃) 1.33 (3H, d), 2.62 (3H, s), 3.30 (1H, td), 3.53 (1H, td), 3.68 (1H, dd), 3.76 (2H, dd), 3.95 (1H, d), 4.00 (1H, dd), 4.02 (1H, s), 4.32 (1H, s), 6.42 (1H, s).

The remaining liquour from the first vapour diffusion was purified by flash chromatography on silica, eluting with a gradient of 0 to 5% MeOH in DCM. Pure fractions were evaporated to afford (R)-4-(2-chloro-6-((S)-methylsulfinylmethyl)pyrimidin-4-yl)-3-methylmorpholine as an orange gum (5.70 g, 43%); ¹ HNMR (400 MHz, CDCl₃) 1.33 (3H, d), 2.62 (3H, d), 3.29 (1H, td), 3.54 (1H, td), 3.68 (1H, dd), 3.73 – 3.82 (2H, m), 3.94 (1H, dd), 4.00 (2H, dd), 4.33 (1H, s), 6.42 (1H, s).

Alternatively, this step can be carried out as follows:

Sodium meta-periodate (64.7 g, 302.69 mmol) was added in one portion to (R)-4-(2-chloro-6-(methylthiomethyl)pyrimidin-4-yl)-3-methylmorpholine (82.87 g, 302.69 mmol) in water (500 ml), EtOAc (1000 ml) and MeOH (500 ml). The resulting solution was stirred at 20 °C for 16 hours. Sodium metabisulfite (50 g) was added and the mixture stirred for 30 minutes. The reaction mixture was filtered and then partially evaporated to remove the MeOH. The organic layer was separated, dried over MgSO₄, filtered and then evaporated. The aqueous layer was washed with DCM (3 x 500 ml). The organic layers were combined, dried over MgSO₄, filtered and then evaporated. The residues were combined and dissolved in DCM (400 ml) and purified by flash chromatography on silica, eluting with a gradient of 0 to 5% MeOH in DCM. Fractions containing product were evaporated and the residue was dissolved in DCM (400 ml) and then divided into four 450 ml bottles. An aluminium foil cap was placed over the top of each bottle and a few holes made in each cap. The bottles were placed in pairs in a large dish containing Et₂O (1000 ml), and then covered and sealed with a second glass dish and left for 11 days. The resultant white needles were collected by filtration and dried under vacuum. The crystals were dissolved in DCM (200 ml) and placed into a 450 ml bottle. An aluminium foil cap was placed over the top of the bottle and a few holes made in the cap. The bottle was placed in a large dish containing Et₂O (1500 ml) and then covered and sealed with a second glass dish and left for 6 days. The resultant crystals were collected by filtration and dried under vacuum to afford (R)-4-(2-chloro-6-((R)-methylsulfinylmethyl)pyrimidin-4-yl)-3-methylmorpholine (16.53 g, 19%); ¹H NMR (400 MHz, CDCl₃) 1.33 (3H, d), 2.61 (3H, s),

3.29 (1H, td), 3.53 (1H, td), 3.68 (1H, dd), 3.76 (2H, dd), 3.95 (1H, d), 3.99 (1H, dd), 4.02 (1H, s), 4.31 (1H, s), 6.41 (1H, s). Chiral HPLC: (HP1100 System 5, 20μm Chiralpak AD-H (250 mm × 4.6 mm) column eluting with Hexane/EtOH/TEA 50/50/0.1) Rf, 12.192 98.2%.

The filtrate from the first vapour diffusion was concentrated in vacuo to afford an approximate

5:2 mixture of (R)-4-(2-chloro-6-((S)-methylsulfinylmethyl)pyrimidin-4-yl)-3-methylmorpholine and (R)-4-(2-chloro-6-((R)-methylsulfinylmethyl)pyrimidin-4-yl)-3-methylmorpholine (54.7 g, 62%).

Alternatively, this step can be carried out as follows:

Sodium meta-periodate (2.87 g, 13.44 mmol) was added in one portion to (R)-4-(2-chloro-6-(methylthiomethyl)pyrimidin-4-yl)-3-methylmorpholine (3.68 g, 13.44 mmol) in water (10.00 ml), EtOAc (20 ml) and MeOH (10.00 ml). The resulting solution was stirred at 20 °C for 16 hours. The reaction mixture was diluted with DCM (60 ml) and then filtered. The DCM layer was separated and the aqueous layer washed with DCM (3 × 40 ml). The organics were combined, dried over MgSO₄, filtered and then evaporated. The residue was purified by flash chromatography on silica, eluting with a gradient of 0 to 7% MeOH in DCM. Pure fractions were evaporated to afford (R)-4-(2-chloro-6-(methylsulfinylmethyl)pyrimidin-4-yl)-3-methylmorpholine (2.72 g, 70%); ¹H NMR (400 MHz, DMSO-d₆) 1.22 (3H, d), 2.64 (3H, d), 3.14 – 3.26 (1H, m), 3.45 (1H, td), 3.59 (1H, dd), 3.73 (1H, d), 3.88 – 3.96 (2H, m), 4.00 (1H, d), 4.07 (1H, dt), 4.33 (1H, s), 6.81 (1H, s); m/z: (ESI+) MH⁺, 290.43.

The (3R)-4-(2-chloro-6-(methylsulfinylmethyl)pyrimidin-4-yl)-3-methylmorpholine (2.7 g, 9.32 mmol) was purified by preparative chiral chromatography on a Merck 100 mm 20 μm Chiralpak AD column, eluting isocratically with a 50:50:0.1 mixture of iso-Hexane:EtOH:TEA as eluent. The fractions containing product were evaporated to afford (R)-4-(2-chloro-6-((S)-methylsulfinylmethyl)pyrimidin-4-yl)-3-methylmorpholine (1.38 g, 51%) as the first eluting compound; ¹HNMR (400 MHz, CDCl₃) 1.29 (3H, dd), 2.56 (3H, s), 3.15 – 3.33 (1H, m), 3.46 (1H, tt), 3.55 – 3.83 (3H, m), 3.85 – 4.06 (3H, m), 4.31 (1H, s), 6.37 (1H, s). Chiral HPLC: (HP1100 System 6, 20μm Chiralpak AD (250 mm × 4.6 mm) column eluting with iso-Hexane/EtOH/TEA 50/50/0.1) Rf, 7.197 >99%.

and (R)-4-(2-chloro-6-((R)-methylsulfinylmethyl)pyrimidin-4-yl)-3-methylmorpholine (1.27 g, 47 %) as the second eluting compound; ¹H NMR (400 MHz, CDCl₃) 1.28 (3H, d), 2.58 (3H, s),

3.26 (1H, td), 3.48 (1H, td), 3.62 (1H, dt), 3.77 (2H, dd), 3.88 – 4.13 (3H, m), 4.28 (1H, s), 6.37 (1H, s). Chiral HPLC: (HP1100 System 6, 20μm Chiralpak AD (250 mm × 4.6 mm) column eluting with iso-Hexane/EtOH/TEA 50/50/0.1) Rf, 16.897 >99%.

g) Iodobenzene diacetate (18.98 g, 58.94 mmol) was added to (R)-4-(2-chloro-6-((R)-methylsulfinylmethyl)pyrimidin-4-yl)-3-methylmorpholine (17.08 g, 58.94 mmol), 2,2,2-trifluoroacetamide (13.33 g, 117.88 mmol), magnesium oxide (9.50 g, 235.76 mmol) and rhodium(II) acetate dimer (0.651 g, 1.47 mmol) in DCM (589 ml) under air. The resulting suspension was stirred at 20 °C for 24 hours. Further 2,2,2-trifluoroacetamide (13.33 g, 117.88 mmol), magnesium oxide (9.50 g, 235.76 mmol), iodobenzene diacetate (18.98 g, 58.94 mmol) and rhodium(II) acetate dimer (0.651 g, 1.47 mmol) were added and the suspension was stirred at 20 °C for 3 days. The reaction mixture was filtered and then silica gel (100 g) added to the filtrate and the solvent removed in vacuo. The resulting powder was purified by flash chromatography on silica, eluting with a gradient of 20 to 50% EtOAc in isohexane. Pure fractions were evaporated to afford N-[({2-chloro-6-[(3R)-3-methylmorpholin-4-yl]pyrimidin-4-yl}methyl)(methyl)oxido-λ6-(R)-sulfanylidene]-2,2,2-trifluoroacetamide (19.39 g, 82%); ¹H NMR (400 MHz, DMSO-d₆) 1.22 (3H, d), 3.17 – 3.27 (1H, m), 3.44 (1H, td), 3.59 (1H, dd), 3.62 (3H, s), 3.74 (1H, d), 3.95 (1H, dd), 4.04 (1H, br s), 4.28 (1H, s), 5.08 (2H, q), 6.96 (1H, s); m/z: (ESI+) MH⁺, 401.12 and 403.13.

h) Dichlorobis(triphenylphosphine)palladium(II) (8.10 mg, 0.01 mmol) was added in one portion to N-[({2-chloro-6-[(3R)-3-methylmorpholin-4-yl]pyrimidin-4-yl}methyl)(methyl)oxido-λ6-(R)-sulfanylidene]-2,2,2-trifluoroacetamide (185 mg, 0.46 mmol), 2M aqueous Na2CO3 solution (0.277 ml, 0.55 mmol) and 4-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)-1-tosyl-1H-pyrrolo[2,3-b]pyridine (193 mg, 0.48 mmol) in DME:water 4: 1 (5 ml) at RT. The reaction mixture was stirred at 90 °C for 1 hour, filtered and then purified by preparative HPLC using decreasingly polar mixtures of water (containing 1% NH₃) and MeCN as eluents. Fractions containing the desired compound were evaporated to afford (R)-3-methyl-4-(6-((R)-S-methylsulfonimidoylmethyl)-2-(1-tosyl-1H-pyrrolo[2,3-b]pyridin-4-yl)pyrimidin-4-yl)morpholine (102 mg, 41%); ¹HNMR (400 MHz, CDCl₃) 1.33 (3H, d), 3.21 – 3.38 (1H, m), 3.42 (3H, d), 3.45 – 3.57 (1H, m), 3.61 – 3.70 (1H, m), 3.78 (1H, d), 4.01 (1H, dd), 3.90 -4.15 (1H, br s), 4.30 (1H, s), 4.64 (1H, dd), 4.84 (1H, dd), 6.49 (1H, d); m/z: (ESI+) MH⁺, 541.35

The 4-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)-1-tosyl-1H-pyrrolo[2,3-b]pyridine, used as starting material, can be prepared as follows:

a) To a 3L fixed vessel was charged 3-chlorobenzoperoxoic acid (324 g, 1444.67 mmol) portionwise to 1H-pyrrolo[2,3-b]pyridine (150 g, 1244.33 mmol) in DME (750 ml) and heptane (1500 ml) at 20°C over a period of 1 hour under nitrogen. The resulting slurry was stirred at 20 °C for 18 hours. The precipitate was collected by filtration, washed with DME / heptane (1/2 5 vol) (750 ml) and dried under vacuum at 40°C to afford 1H-pyrrolo[2,3-b] pyridine 7-oxide 3-chlorobenzoate (353 g, 97%) as a cream solid, which was used without further purification; ¹H NMR (400 MHz, DMSO-d₆) 6.59 (1H, d), 7.07 (1H, dd), 7.45 (1H, d), 7.55 (1H, t), 7.65 (1H, dd), 7.70 (1H, ddd), 7.87 – 7.93 (2H, m), 8.13 (1H, d), 12.42 (1H, s), 13.32 (1H, s).

b) A 2M solution of potassium carbonate (910 ml, 1819.39 mmol) was added dropwise to a stirred slurry of 1H-pyrrolo[2,3-b]pyridine 7-oxide 3-chlorobenzoate (352.6 g, 1212.93 mmol) in water (4.2 vol) (1481 ml) at 20°C, over a period of 1 hour adjusting the pH to 10. To the resulting slurry was charged water (2 vol) (705 ml) stirred at 20 °C for 1 hour. The slurry was cooled to 0°C for 1 hour and the slurry filtered, the solid was washed with water (3 vol 1050ml) and dried in a vacuum oven at 40°C over P₂O₅ overnight to afford 1H-pyrrolo[2,3-b] pyridine 7-oxide (118 g, 73%); ¹H NMR (400 MHz, DMSO-d₆) 6.58 (1H, d), 7.06 (1H, dd), 7.45 (1H, d), 7.64 (1H, d), 8.13 (1H, d), 12.44 (1H, s); m/z: (ES+) (MH+MeCN)⁺, 176.03. c) To a 3L fixed vessel under an atmosphere of nitrogen was charged methanesulfonic anhydride (363 g, 2042.71 mmol) portionwise to 1H-pyrrolo[2,3-b]pyridine 7-oxide (137 g, 1021.36 mmol), and tetramethylammonium bromide (236 g, 1532.03 mmol) in DMF (10 vol) (1370 ml) cooled to 0°C over a period of 30 minutes under nitrogen. The resulting suspension was stirred at 20 °C for 24 hours. The reaction mixture was quenched with water (20 vol, 2740 ml) and the reaction mixture was adjusted to pH 7 with 50% sodium hydroxide (approx 200 ml). Water (40 vol, 5480 ml) was charged and the mixture cooled to 10°C for 30 minutes. The solid was filtered, washed with water (20 vol, 2740 ml) and the solid disssolved into

DCM/methanol (4: 1, 2000 ml), dried over MgSO₄ and evaporated to provide a light brown solid. The solid was taken up in hot methanol (2000 ml) and water added dropwise until the solution went turbid and left overnight. The solid was filtered off and discarded, the solution was evaporated and the solid recrystallised from MeCN (4000 ml). The solid was filtered and washed with MeCN to afford 4-bromo-1H-pyrrolo[2,3-b]pyridine (68.4 g, 34%) as a pink

solid; ¹H NMR (400 MHz, OMSO-d₆) 6.40 – 6.45 (1H, m), 7.33 (1H, d), 7.57 – 7.63 (1H, m), 8.09 (1H, t), 12.02 (1H, s); m/z: (ES+) MH⁺, 198.92. The crude mother liquors were purified by Companion RF (reverse phase CI 8, 415g column), using decreasingly polar mixtures of water (containing 1% NH₃) and MeCN as eluents (starting at 26% upto 46% MeCN). Fractions containing the desired compound were evaporated to afford 4-bromo-1H-pyrrolo[2,3-b]pyridine (5.4 g, 3%) as a pink solid; ¹H NMR (400 MHz, DMSO-d₆) 6.43 (1H, dd), 7.33 (1H, d), 7.55 – 7.66 (1H, m), 8.09 (1H, d), 12.03 (1H, s); m/z: (ES+) MH⁺, 199.22.

d) Sodium hydroxide (31.4 ml, 188.35 mmol) was added to 4-bromo-1H-pyrrolo[2,3-b]pyridine (10.03 g, 50.91 mmol), tosyl chloride (19.41 g, 101.81 mmol) and

tetrabutylammonium hydrogensulfate (0.519 g, 1.53 mmol) in DCM (250 ml) at RT. The resulting mixture was stirred at RT for 1 hour. The reaction was quenched through the addition of saturated aqueous NH₄Cl, the organic layer removed and the aqueous layer further extracted with DCM (3 × 25 ml). The combinbed organics were washed with brine (100 ml), dried over Na₂SO₄ and then concentrated under reduced pressure. The residue was purified by flash chromatography on silica, eluting with a gradient of 0 to 20% EtOAc in isohexane. Pure fractions were evaporated to afford 4-bromo-1-tosyl-1H-pyrrolo[2,3-b]pyridine (14.50 g, 81%); ¹H NMR (400 MHz, CDCl₃) 2.38 (3H, s), 6.64 (1H, d), 7.28 (2H, d), 7.36 (1H, d), 7.78 (1H, d), 8.06 (2H, d), 8.22 (1H, d); m/z: (ES+) MH⁺, 353.23.

e) 1,1′-Bis(diphenylphosphino)ferrocenedichloropalladium(II) (3.37 g, 4.13 mmol) was added in one portion to 4-bromo-1-tosyl-1H-pyrrolo[2,3-b]pyridine (14.5 g, 41.28 mmol), bis(pinacolato)diboron (20.97 g, 82.57 mmol) and potassium acetate (12.16 g, 123.85 mmol) in anhydrous DMF (300 ml) at RT. The resulting mixture was stirred under nitrogen at 90 °C for 24 hours. After cooling to RT, 1N aqueous NaOH was added untill the aqueous layer was taken to pH 10. The aqueous layer was washed with DCM (1L), carefully acidified to pH 4 with 1 N aqueous HCl, and then extracted with DCM (3 × 300 ml). The organic layer was concentrated under reduced pressure to afford a dark brown solid. The solid was triturated with diethyl ether, filtered and dried to afford 4-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)-1-tosyl-1H-pyrrolo[2,3-b]pyridine (7.058 g, 43%); ¹H NMR (400 MHz, CDCl₃) 1.36 (12H, s), 2.35 (3H, s), 7.01 (1H, d), 7.22 (2H, d), 7.52 (1H, d), 7.74 (1H, d), 8.03 (2H, m), 8.42 (1H, d); m/z: (ES+) MH⁺, 399.40. The mother liquors were concentrated in vacuo and the residue triturated in isohexane, filtered and dried to afford a further sample of 4-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)-1-tosyl-1H-pyrrolo[2,3-b]pyridine (3.173 g, 19%); ¹H NMR (400 MHz,

CDCI₃) 1.36 (12H, s), 2.35 (3H, s), 7.01 (1H, d), 7.23 (2H, d), 7.52 (1H, d), 7.74 (1H, d), 8.03 (2H, d), 8.42 (1H, d); m/z: (ES+) MH⁺, 399.40.

Example 2.01 and example 2.02

4-{4-[(3R)-3-Methylmorpholin-4-yl]-6-[1-((S)-S-methylsulfonimidoyl)cyclopropyl]pyrimidin-2-yl}-1H-pyrrolo[2,3-blpyridine, and

4-{4-[(3R)-3-Methylmorpholin-4-yl]-6-[1-((R)-S-methylsulfonimidoyl)cyclopropyl]pyrimidin-2-yl}-1H-pyrrolo[2,3-blpyridine

(3R)-3-Methyl-4-(6-(1-(S-methylsulfonimidoyl)cyclopropyl)-2-(1-tosyl-1H-pyrrolo[2,3-b]pyridin-4-yl)pyrimidin-4-yl)morpholine (1.67 g, 2.95 mmol) was dissolved in DME:water 4: 1 (60 ml) and heated to 50 °C. Sodium hydroxide, 2M aqueous solution (2.58 ml, 5.16 mmol) was then added and heating continued for 18 hours. The reaction mixture was acidified with 2M H Cl (~2 ml) to pH5. The reaction mixture was evaporated to dryness and the residue dissolved in EtOAc (250 ml), and washed with water (200 ml). The organic layer was dried over MgSO₄, filtered and evaporated onto silica gel (10 g). The resulting powder was purified by flash chromatography on silica, eluting with a gradient of 0 to 7% MeOH in DCM. Pure fractions were evaporated and the residue was purified by preparative chiral chromatography on a Merck 50mm, 20μm ChiralCel OJ column, eluting isocratically with 50% isohexane in EtOH/MeOH (1 : 1) (modified with TEA) as eluent. The fractions containing the desired compound were evaporated to dryness to afford the title compound: 4-{4-[(3R)-3-methylmorpholin-4-yl]-6-[1-((R)-S-methylsulfonimidoyl)cyclopropyl]pyrimidin-2-yl}-1H-pyrrolo[2,3-b]pyridine (0.538g, 44%) as the first eluting compound; ¹H NMR (400 MHz,

DMSO-d⁶) 1.29 (3H, d), 1.51 (3H, m), 1.70 – 1.82 (1H, m), 3.11 (3H, s), 3.28 (1H, m, obscured by water peak), 3.48 – 3.60 (1H, m), 3.68 (1H, dd), 3.75 – 3.87 (2H, m), 4.02 (1H, dd), 4.19 (1H, d), 4.60 (1H, s), 7.01 (1H, s), 7.23 (1H, dd), 7.51 – 7.67 (1H, m), 7.95 (1H, d), 8.34 (1H, d), 11.76 (1H, s); m/z: (ES+) MH⁺, 413.12. Chiral HPLC: (HP1100 System 4, 5μm Chiralcel OJ-H (250 mm × 4.6 mm) column eluting with iso-Hexane/EtOH/MeOH/TEA 50/25/25/0.1) Rf, 9.013 >99%. Crystals were grown and isolated by slow evaporation to dryness in air from EtOAc. These crystals were used to obtain the structure shown in Fig 1 by X-Ray diffraction (see below). Example 2.02: 4-{4-[(3R)-3-methylmorpholin-4-yl]-6-[1-((R)-S-methylsulfonimidoyl)cyclopropyl]pyrimidin-2-yl}-1H-pyrrolo[2,3-b]pyridine (326 mg, 0.79 mmol) was dissolved in DCM (3 ml). Silica gel (0.5 g) was added and the mixture concentrated in vacuo. The resulting powder was purified by flash chromatography on silica, eluting with a gradient of 0 to 5% MeOH in DCM. Pure fractions were evaporated to dryness and the residue was crystallized from EtOAc/n-heptane to afford 4-{4-[(3R)-3-methylmorpholin-4-yl]-6-[1-((R)-S-methylsulfonimidoyl)cyclopropyl]pyrimidin-2-yl}-1H-pyrrolo[2,3-b]pyridine (256 mg, 79%) as a white crystalline solid; ¹H NMR (400 MHz, DMSO-d⁶) 1.29 (3H, d), 1.39 – 1.60 (3H, m), 1.71 – 1.81 (1H, m), 3.10 (3H, d), 3.21 – 3.29 (1H, m), 3.52 (1H, td), 3.67 (1H, dd), 3.80 (2H, t), 4.01 (1H, dd), 4.19 (1H, d), 4.59 (1H, s), 7.01 (1H, s), 7.23 (1H, dd), 7.54 – 7.62 (1H, m), 7.95 (1H, d), 8.34 (1H, d), 11.75 (1H, s). DSC (Mettler-Toledo DSC 820, sample run at a heating rate of 10°C per minute from 30°C to 350°C in a pierced aluminium pan) peak, 224.1 FC.

and the title compound: 4-{4-[(3R)-3-methylmorpholin-4-yl]-6-[1-((S)-S-methylsulfonimidoyl)cyclopropyl]pyrimidin-2-yl}-1H-pyrrolo[2,3-b]pyridine (0.441 g, 36%) as the second eluting compound; ¹H NMR (400 MHz, DMSO-d⁶) 1.28 (3H, d), 1.40 – 1.58 (3H, m), 1.70 – 1.80 (1H, m), 3.10 (3H, d), 3.23 – 3.27 (1H, m), 3.51 (1H, dt), 3.66 (1H, dd), 3.80 (2H, d), 4.01 (1H, dd), 4.21 (1H, d), 4.56 (1H, s), 6.99 (1H, s), 7.22 (1H, dd), 7.54 – 7.61 (1H, m), 7.94 (1H, d), 8.33 (1H, d), 11.75 (1H, s); m/z: (ES+) MH⁺, 413.12. Chiral HPLC: (HP1100 System 4, 5μm Chiralcel OJ-H (250 mm × 4.6 mm) column eluting with iso-Hexane/EtOH/MeOH/TEA 50/25/25/0.1) Rf, 15.685 >99%. Example 2.01 : 4-{4-[(3R)-3-methylmorpholin-4-yl]-6-[1-((S)-S-methylsulfonimidoyl)cyclopropyl]pyrimidin-2-yl}-1H-pyrrolo[2,3-b]pyridine (66.5 mg) was purified by crystallisation from EtOH/water to afford 4-{4-[(3R)-3-methylmorpholin-4-yl]-6-[1-((S)-S-methylsulfonimidoyl)cyclopropyl]pyrimidin-2-yl}-1H-pyrrolo[2,3-b]pyridine (0.050 g); ¹H NMR (400 MHz, CDCl₃) 1.40 (3H, d), 1.59 (2H, s), 1.81 (2H, s), 2.41 (1H, s), 3.16 (3H, s), 3.39 (1H, td), 3.59 – 3.67 (1H, m), 3.77 (1H, dd), 3.86 (1H, d), 4.07 (1H, dd), 4.17 (1H, d), 4.54 (1H, s), 6.91 (1H, s), 7.34 (1H, t), 7.43 (1H, t), 8.05 (1H, d), 8.41 (1H, d), 9.14 (1H, s).

Scheme 1. Medicinal Chemistry Route to AZD6738

Reagent and conditions:

(a) (3R)-3-methylmorpholine, TEA, DCM, 77%;

(b) LiBH₄, THF, 100%;

(d) LiI, dioxane, 78%;

(e) NaSMe, DMF, 96%;

(f) m-CPBA, DCM;

(g) crystallization or chromatography, 40% (two steps);

(h) IBDA, trifluoroacetamide, MgO, DCM, Rh₂(OAc)₄ 82%;

(i) 1,2-dibromoethane, sodium hydroxide, TOAB, 2-MeTHF, 47%;

(j) TsCl, tetrabutylammonium hydrogen sulfate, sodium hydroxide, DCM, 92%;

(k) bis(pinacolato)diboron, potassium acetate, 1,1′-bis(diphenylphosphino)ferrocene dichloro palladium(II), DMF, 62%;

(l) Pd(II)Cl₂(PPh₃)₂, Na₂CO₃, DME, water, 80%;

(m) 2 N NaOH, DME, water, 92%.

Foote, K. M. N.; Johannes, W. M.; Turner, P.. Morpholino Pyrimidines and their use in therapy. WO 2011/154737 A1, 15 December 2011.

PAPER

Development and Scale-up of a Route to ATR Inhibitor AZD6738

William R. F. Goundry et al

Cite This:Org. Process Res. Dev.2019XXXXXXXXXX-XXX

Publication Date:June 21, 2019

https://doi.org/10.1021/acs.oprd.9b00075

AZD6738 is currently being tested in multiple phase I/II trials for the treatment of cancer. Its structure, comprising a pyrimidine core decorated with a chiral morpholine, a cyclopropyl sulfoximine, and an azaindole, make it a challenging molecule to synthesize on a large scale. We describe the evolution of the chemical processes, following the manufacture of AZD6738 from the initial scale-up through to multikilos on plant scale. During this evolution, we developed a biocatalytic process to install the sulfoxide with high enantioselectivity, followed by introduction of the cyclopropyl group first in batch, then in a continuous flow plate reactor, and finally through a series of continuous stirred tank reactors. The final plant scale process to form AZD6738 was operated on 46 kg scale with an overall yield of 18%. We discuss the impurities formed throughout the process and highlight the limitations of this route for further scale-up.

imino-methyl-[1-[6-[(3R)-3-methylmorpholin-4-yl]-2-(1H-pyrrolo[2,3-b]pyridin-4-yl)pyrimidin-4-yl]cyclopropyl]-oxo-λ6-sulfane (1) (30.0 g) were added at 75 °C, and the reaction mixture was held for 2 h. The mixture was cooled to 20 °C, and n-heptane (141.9 kg) was added at the rate of 40 kg/h. The solid was collected by filtration, washed with a mixture of 1-butanol and n-heptane (9.3 and 22.4 kg respectively), and then given a further wash with n-heptane (32.2 kg). The solid was dried at 40 °C to give imino-methyl-[1-[6-[(3R)-3-methylmorpholin-4-yl]-2-(1H-pyrrolo[2,3-b]pyridin-4-yl)pyrimidin-4-yl]cyclopropyl]-oxo-λ6-sulfane (1) as a whit solid (41.4 kg, 92% yield): Assay (HPLC) 99.9%; Assay (NMR) 99% wt/wt.

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//////AZD6738, AZD-6738, AZD 6738, AstraZeneca, University of Pennsylvania, Phase II, Breast cancer, Gastric cancer, Non-small cell lung cancer, Ovarian cancer, Ceralasertib

C[C@@H]1COCCN1c2cc(nc(n2)c3cncc4[nH]ccc34)C5(CC5)[S@](=N)(=O)C

SEVITERONEL, севитеронел , سيفيتيرونيل , 赛维罗奈 ,

June 19, 2019 1:08 pm / Leave a comment

SEVITERONEL

CAS Registry Number 1610537-15-9

Molecular formulaC18 H17 F4 N3 O3, MW 399.34

1H-1,2,3-Triazole-5-methanol, α-[6,7-bis(difluoromethoxy)-2-naphthalenyl]-α-(1-methylethyl)-, (αS)-

(αS)-α-[6,7-Bis(difluoromethoxy)-2-naphthalenyl]-α-(1-methylethyl)-1H-1,2,3-triazole-5-methanol

8S5OIN36X4

севитеронел [Russian] [INN]

سيفيتيرونيل [Arabic] [INN]

赛维罗奈 [Chinese] [INN]

Mechanism of ActionAndrogen receptor antagonists; Estrogen receptor antagonists; Steroid 17-alpha-hydroxylase inhibitors; Steroid 17-alpha-hydroxylase modulators
WHO ATC codeL01 (Antineoplastic Agents)L01X-X (Other antineoplastic agents)
EPhMRA codeL1 (Antineoplastics)L1X9 (All other antineoplastics)

1H-1,2,3-Triazole-5-methanol, alpha-(6,7-bis(difluoromethoxy)-2-naphthalenyl)-alpha-(1-methylethyl)-, (alphaS)-

Seviteronel (developmental codes VT-464 and, formerly, INO-464) is an experimental cancer medication which is under development by Viamet Pharmaceuticals and Innocrin Pharmaceuticals for the treatment of prostate cancer and breast cancer.^[1] It is a nonsteroidal CYP17A1 inhibitor and works by inhibiting the production of androgens and estrogens in the body.^[1] As of July 2017, seviteronel is in phase II clinical trials for both prostate cancer and breast cancer.^[1] In January 2016, it was designated fast-track status by the United States Food and Drug Administration for prostate cancer.^[1]^[2] In April 2017, seviteronel received fast-track designation for breast cancer as well.^[1]

Originator Viamet Pharmaceuticals
Developer Innocrin Pharmaceuticals
Clas sAntiandrogens; Antineoplastics; Fluorine compounds; Naphthalenes; Propanols; Small molecules; Triazoles
Mechanism of Action Androgen receptor antagonists; Estrogen receptor antagonists; Steroid 17-alpha-hydroxylase inhibitors; Steroid 17-alpha-hydroxylase modulators

Phase II Breast cancer; Prostate cancer; Solid tumours

31 Jan 2019 Innocrin Pharmaceutical completes a phase II trial in Prostate Cancer (Second-line therapy or greater, Hormone refractory) in the US (NCT02445976)
31 Jan 2019 Innocrin Pharmaceutical completes a phase II trial for Prostate Cancer (Hormone refractory) in the US, UK, Switzerland and Greece (NCT02012920)
31 Jan 2019 Innocrin Pharmaceuticals completes the phase I/II CLARITY-01 trial for Breast cancer (Late stage disease) in USA (NCT02580448)
CYP-17 useful for treating fungal infections, prostate cancer, and polycystic ovary syndrome, assigned to Viamet Pharmaceuticals Inc , naming Hoekstra and Rafferty. Innocrin Pharmaceuticals , a spin-out of Viamet is developing oral seviteronel, the lead dual selective inhibitors of the 17,20-lyase activity of P450c17 (CYP17) and androgen receptor antagonist, which also includes VT-478 and VT-489, developed using the company’s Metallophile technology, for treating castration-resistant prostate cancer (CRPC) in men, breast cancer and androgen (AR) related cancers.

Pharmacology

Pharmacodynamics

Seviteronel is a nonsteroidal antiandrogen, acting specifically as an androgen synthesis inhibitor via inhibition of the enzyme CYP17A1, for the treatment of castration-resistant prostate cancer.^[3]^[4]^[5]^[6]^[7]^[8] It has approximately 10-fold selectivity for the inhibition of 17,20-lyase (IC₅₀ = 69 nM) over 17α-hydroxylase (IC₅₀ = 670 nM), which results in less interference with corticosteroid production relative to the approved CYP17A1 inhibitor abiraterone acetate (which must be administered in combination with prednisone to avoid glucocorticoid deficiency and mineralocorticoid excess due to 17α-hydroxylase inhibition) and hence may be administerable without a concomitant exogenous glucocorticoid.^[4]^[5]^[6]^[7]^[8] Seviteronel is 58-fold more selective for inhibition of 17,20-lyase than abiraterone (the active metabolite of abiraterone acetate), which has IC₅₀ values for inhibition of 17,20-lyase and 17α-hydroxylase of 15 nM and 2.5 nM, respectively.^[7] In addition, in in vitro models, seviteronel appears to possess greater efficacy as an antiandrogen relative to abiraterone.^[6] Similarly to abiraterone acetate, seviteronel has also been found to act to some extent as an antagonist of the androgen receptor.^[6]

Society and culture

Generic names

Seviteronel is the generic name of the drug and its INN.^[9]

PATENT

WO2012064943

PATENT

WO-2019113312

https://patentscope.wipo.int/search/en/detail.jsf?docId=WO2019113312&redirectedID=true

The present invention relates to a process for preparing compound 1 that is useful as an anticancer agent. In particular, the invention seeks to provide a new methodology for preparing compound 1 and substituted derivatives thereof.

Living organisms have developed tightly regulated processes that specifically import metals, transport them to intracellular storage sites and ultimately transport them to sites of use. One of the most important functions of metals such as zinc and iron in biological systems is to enable the activity of metalloenzymes. Metalloenzymes are enzymes that incorporate metal ions into the enzyme active site and utilize the metal as a part of the catalytic process. More than one-third of all characterized enzymes are metalloenzymes.

The function of metalloenzymes is highly dependent on the presence of the metal ion in the active site of the enzyme. It is well recognized that agents which bind to and inactivate the active site metal ion dramatically decrease the activity of the enzyme. Nature employs this same strategy to decrease the activity of certain metalloenzymes during periods in which the enzymatic activity is undesirable. For example, the protein TIMP (tissue inhibitor of metalloproteases) binds to the zinc ion in the active site of various matrix metalloprotease enzymes and thereby arrests the enzymatic activity. The pharmaceutical industry has used the same strategy in the design of therapeutic agents. For example, the azole antifungal agents fluconazole and voriconazole contain a l-( 1,2, 4-triazole) group that binds to the heme iron present in the active site of the target enzyme lanosterol demethylase and thereby inactivates the enzyme.

In the design of clinically safe and effective metalloenzyme inhibitors, use of the most appropriate metal-binding group for the particular target and clinical indication is critical. If a weakly binding metal-binding group is utilized, potency may be suboptimal. On the other hand, if a very tightly binding metal-binding group is utilized, selectivity for the target enzyme versus related metalloenzymes may be suboptimal. The lack of optimal selectivity can be a cause for clinical toxicity due to unintended inhibition of these off-target metalloenzymes.

One example of such clinical toxicity is the unintended inhibition of human drug metabolizing enzymes such as CYP2C9, CYP2C19 and CYP3A4 by the currently-available azole antifungal agents such as fluconazole and voriconazole. It is believed that this off-target inhibition is caused primarily by the indiscriminate binding of the currently utilized l-(l,2,4-triazole) to iron in the active site of CYP2C9, CYP2C19 and CYP3A4. Another example of this is the joint pain that has been observed in many clinical trials of matrix metalloproteinase inhibitors. This toxicity is considered to be related to inhibition of off-target metalloenzymes due to indiscriminate binding of the hydroxamic acid group to zinc in the off-target active sites.

Therefore, the search for metal-binding groups that can achieve a better balance of potency and selectivity remains an important goal and would be significant in the realization of therapeutic agents and methods to address currently unmet needs in treating and preventing diseases, disorders and symptoms thereof. Similarly, methods of synthesizing such therapeutic agents on the laboratory and, ultimately, commercial scale is needed. Addition of metal-based nucleophiles (Zn, Zr, Ce, Ti, Mg, Mn, Li) to azole-methyl substituted ketones have been effected in the synthesis of voriconazole (M. Butters, Org. Process Res. Dev. 2001, 5, 28-36). The nucleophile in these examples was an ethyl-pyrimidine substrate. Similarly, optically active azole-methyl epoxide has been prepared as precursor electrophile toward the synthesis of ravuconazole (A. Tsuruoka, Chem. Pharm. Bull. 1998, 46, 623-630). Despite this, the development of methodology with improved efficiency and selectivity is desirable

Preparation of Compound 4:

Acetone (850 L), 2,3-dihydroxynaphthalene (85.00 kg, 530.7 moles), and potassium carbonate (219.3 kg, 1,586.7 moles) were charged to a clean, fixed reactor with stirring and with the temperature maintained at 20 – 35 °C. Dimethyl sulfate (200.6 kg, 2131.09) was added to the stirred reaction at a rate that maintains the internal temperature of the exothermic reaction below 60 °C. This addition typically requires about 3 hours. At the end of the dimethyl sulfate addition, the reaction is continued to allow to stir while maintaining the internal temperature at 50 – 60 °C. After about 3 hours, the reaction was analyzed by HPLC. The reaction was concentrated by atmospheric pressure distillation of acetone. The distillation was continued until 340 – 425 L of distillate was collected. This represents 40 – 50 % of the initial charge of acetone. At the end of the distillation, the reaction mass is present as a thick suspension. While maintaining the internal temperature below 60 °C, the reactor contents were slowly diluted with water (850 L). When the addition is complete, the reaction was cooled to an internal temperature of 25 – 35 °C and stirring was continued for 1 – 2 hours after the designated internal temperature was reached. Compound 2 was isolated by filtration and the cake was washed with water (at least 3 X 85 L). Compound 2 was dried at 40 – 45 °C and full vacuum until the water content by Karl Fisher titration is found to be NMT 2.0 %. Typically, greater than 90 kg of dry product is obtained with an assay of >99.5% AUC by HPLC.

Dichloromethane (with a water content by Karl Fisher Titration of NMT 0.50%) (928 L) and 2,3-dimethoxynaphthalene (2, 116.00 kg, 616.3 moles) were charged to a clean, fixed reactor with stirring and with the temperature maintained at 20 – 35 °C. The reactor contents were cooled to an internal temperature of -5 to 0 °C. Aluminum chloride (164.72 kg, 1235.3 moles, 2.00 molar equivalents) was carefully added in portions to the reaction, while maintaining the internal temperature at -5 to +5 °C. This addition typically requires 5 – 6 hours. At the end of the addition, the reactor contents were cooled to an internal temperature of -15 to -5 °C. Isobutyryl chloride (102.08 kg, 958.05 moles, 1.55 molar equivalents) was slowly added to the reaction while maintaining the internal temperature at -15 to -5 °C. The addition typically requires about 3 hours. At the end of the isobutyryl chloride addition, the reaction was warmed to an internal temperature of 20 – 35 °C. When the temperature was reached, these conditions were maintained for 2 – 3 hours until the IPC indicated a level of residual starting material of NMT 2.0 % AUC by HPLC. The reactor contents were then cooled to 0 – 5 °C. The reaction was quenched by adding the reaction to a precooled (0 – 5 °C) 3M aqueous solution of hydrochloric hcid (Water, 754 L: cone. HC1, 406 L). The mixture was vigorously stirred for 15 – 20 minutes then the layers were allowed to settle. The lower, dichloromethane, product-containing layer was washed sequentially with 10 % aqueous sodium bicarbonate (1044 L), water (1160 L), then 10 % aqueous sodium chloride (1044 L). The reaction was concentrated by distillation under full vacuum and at an internal temperature of NMT 40 °C. The reaction concentrate was cooled to 20 – 35 °C and diluted with hexanes (812 L). The resultant slurry was warmed to 45 – 50 °C and these conditions were maintained for 1 – 2 hours. The reactor contents were cooled to 20 – 35 °C for 1 – 2 hours. Compound 3 was isolated by filtration. The cake was washed with fresh hexanes (232 L) twice, the filter was cooled, and the cake was washed an additional two times with hexanes. Compound 3 was dried under full vacuum at a jacket temperature of 45 °C. Typically, about 95 kg of dry product was isolated with a product purity of >90% by HPLC.

Acetic acid (212.5 L L) and l-(6,7-dimethoxynaphthalene-2-yl)-2-methylpropane-l- one (42.5 kg, 164.5 moles) were charged to a clean, fixed reactor with stirring and with the temperature maintained at 25 – 45 °C. Concentrated hydrochloric acid (425.0 L) was added carefully to the stirring reactor contents while maintaining reactor contents at an internal temperature of 25 – 45 °C. When the addition was complete, the internal temperature of the reaction was raised to 100 – 105 °C. Note that the reaction is a heterogeneous mixture. The reaction was stirred under these conditions for 6 – 8 hours. The reaction was cooled to 85 – 90 °C to which was carefully added a fresh portion of hydrochloric acid (127.5 L). The reaction was warmed to 100 – 105 °C and stirred for another 6 – 8 hours. The reaction was cooled to 85 – 90 °C. The reaction was cooled further to 70 – 80 °C. Water (212.5 L) was added to the well stirred reaction and the reactor contents were cooled to an internal temperature of 35 – 45 °C and stirred for 3 – 4 hours. Compound 4 was collected by filtration. The wet cake was washed with water (212.5 L). The wet cake was added to a clean reactor with a 5% aqueous sodium bicarbonate solution and stirred at an internal temperature of 35 – 45 °C for 1 – 2 hours.

Compound 4 was collected by filtration and washed with water (212.5 L). Compound 4 was dried under full vacuum and a temperature of < 50 °C until the water content of the dried material was found to be NMT 5.0% by Karl Fisher Titration. The yield is typically >31 kg with a purity >99.5 %.

Preparation of Compound 5:

The following difluoromethylation conditions listed in Table 1 were investigated:

Preparation 1:

The reaction flask was dried under an argon flow at 120 °C. (lS,2R)-l-Phenyl-2-(l- pyrrolidinyl)propan-l-ol (ligand 45) (196.6 g, 0.96 mol, 2.2 eq.) was added into the flask and then toluene (195 mL) was added. The solution was cooled to <12 °C. A solution of diethyl zinc (716.4 g, 0.87 mol, 15 wt%, 2 eq.) in toluene was added through a septum over 30 min at 0-10 °C. Further, a solution of ((Trimethylsilyl)ethynyl)-magnesium bromide in THF (1.81 kg; 0.87 mol, 9.7 wt%, 2 eq.) was added over 30 min at 0-10 °C. Finally, trifluoroethanol (87.0 g; 0.87 mol; 2 eq.) was added over 10 min at 0-10 °C. The reaction solution was stirred at 10-12 °C for 3 h. Compound 5 (143.4 g; 0.434 mol; 1 eq.) was added (as a solid) at room

temperature. The reaction mixture was stirred at room temperature for 1 h and at 55 °C for 17 h. The reaction solution was cooled to room temperature and dosed with aqueous HC1 (3600 mL; 7.5 wt%) within 20 min. The temperature of the mixture was kept below 25 °C. Toluene (1250 mL) was added and the mixture was stirred at room temperature for 5 min. The aqueous phase was separated and stored for the recycling of ligand 45. The organic phases were washed with water (638 mL) and concentrated via distillation under reduced pressure (50 mbar). The residue (approx. 184 g) was treated with heptane (200 mL), which was removed

via distillation. The residue was dissolved in heptane (2050 mL) at 50 °C. The mixture was cooled to room temperature and subsequently to -8 °C within 2 hours. The obtained suspension was stirred at -8 °C for 1 h. Crystallized compound 5 (20.0 g; 14%) was isolated via filtration, washed twice with cold (0 °C) heptane (2×20 mL) and dried under vacuum at 50 °C for 12 hours. The combined heptane phases were concentrated under reduced pressure to obtain a 48 wt% solution of compound 18b in heptane (yield: 83.0%). The solution was directly used for the next step.

1H-NMR (600.6 MHz, DMSO-D₆) d: 0.23 (s, 9H), 0.77 (d, J = 6.7 Hz, 3H), 0.93 (d, 7 = 6.7 Hz, 3H), 2.04 (sept., 7 = 6.7 Hz, 1H), 6.11 (s, 1H), 7.32 (t, ²7H,F = 73.4 Hz, 1H), 7.35 (t, ²7H,F = 73.4 Hz, 1H), 7.68 (dd, 7 = 8.6, 1.5 Hz, 1H), 7.84 (s, 1H), 7.87 (s, 1H), 7.93 (d, 7 = 8.6 Hz, 1H), 8.03 (s (broad), 1H);

HPLC (purity): 94%;

chiral HPLC: e.r. = 18:82.

Preparation 2:

(7S,2R)-l-Phenyl-2-(l-pyrrolidinyl)propan-l-ol (ligand 45) (13.0 kg, 63.3 mol, 2.2 eq.) was charged into the reactor and toluene (60 L) was added. The solution was cooled to < 12 °C. A solution of diethyl zinc (35.6 kg, 57.3 mol, 20 wt%, 2 eq.) in toluene was added via mass flow controller at 8-16 °C. Further, a solution of ((trimethylsilyl)ethynyl)-magnesium bromide in THF (11.5 kg; 57.3 mol, 9.7 wt%, 2 eq.) was added at 8-16 °C. Finally, trifluoroethanol (5.7 kg; 57.3 mol; 2 eq.) was added over 10 min at 8-16 °C.The reaction solution was stirred at 22-25 °C for 3 h. A solution of compound 5 (9.5 kg; 28.7 mol; 1 eq.) in toluene (20 L) was added at room temperature. The reaction mixture was stirred at 25 °C for 1 h and at 55 °C for 17 h. The reaction solution was cooled to room temperature and dosed in aqueous HC1 (225L; 7.5 wt%) within 20 min. The temperature of the mixture should be kept below 25 °C. Toluene (80 L) was added and the mixture was stirred at room temperature for 5 min. The organic phases was washed with water (50 L) and concentrated via distillation under reduced pressure (50 mbar). The residue was treated with heptane (100 L), which was removed via distillation. The residue was dissolved in heptane (100 L) at 50°C, which was removed via distillation. The residue was dissolved in heptane (25 L). Heptane (110 L) was added, the mixture was cooled to room temperature and subsequently to 0-5 °C and seeded with compound 5 (0.15 kg). The obtained suspension was cooled to -8 °C within 1 h and stirred at this temperature for 2 h. Crystallized compound 5 was removed via filtration. The filtrate was concentrated under reduced pressure to obtain a 48 wt% solution of compound 18b in heptane (calculated 8.8 kg, 71.6%). This solution was directly used for the next step.

HPLC (purity): 94%;

chiral HPLC: e.r. = 18:82.

Recovery of the chiral ligand ( lS,2R)-l-Phenvl-2-
-l-ol from the

Preparation 1:

The above acidic aqueous phase was diluted with toluene (1000 mL) and the mixture was treated with sodium hydroxide (50 wt% solution) to adjust the pH to 12. The mixture was warmed to 50 °C and sodium chloride (100 g) was added. The aqueous phase was separated and washed with toluene (1000 mL). The combined organic phases were washed with water (200 mL). The combined toluene phases were treated with water (1000 mL) and the pH was adjusted to 2 by the addition of a cone. HC1 solution. The aqueous phase was separated and the mixture was treated with sodium hydroxide (50 wt% solution) at 5 °C to adjust the pH to 12. After seeding, the suspension was stirred at 5 °C for 30 min. The solids were isolated, washed with cold (0 °C) water (4×100 mL) and dried under vacuum at 30 °C for 24 hours. Ligand 45 (178.9g; 91%) was obtained as slightly yellow crystalline solid.

HPLC (purity): 99%.

Preparation 2:

The acidic aqueous phase containing ligand 45 (500 L) was diluted with toluene (125 L) and treated with“Kieselgur” (20 L). The mixture was treated with sodium hydroxide (40 L; 50 wt% solution) to adjust the pH to 12 whereas the temperature was kept <55 °C. The suspension was stirred for 15-20 min and filtered to remove all solids. Toluene (80 L) was added and the aqueous phase was separated. The organic phase was treated with water (150 mL) and the pH was adjusted to 1.5-2 by the addition of an aqueous HC1 solution (10 L; 32 wt%). The aqueous phase was separated, toluene (150 L) was added, and the mixture was treated with sodium hydroxide (5 L; 50 wt% solution) at 5 °C to adjust the pH to 12-12.5. The organic phase was separated, washed with water (30 L), and concentrated under reduced

pressure at 50 °C. Approx. 100L of distillate was removed. A sample of the solution of ligand 45 in toluene was analyzed:

The NMR results indicated a 21.6 wt% solution of ligand 45 in toluene which corresponds to a calculated amount of 118.4 kg (83.6%) of ligand 45.

Preparation of Compound 18a

Preparation 1:

A solution of tertiary alcohol 18b (320 g; 48 wt%; 0.36 mol; 1 eq.) in heptane was dissolved in methanol (800 mL). Potassium carbonate (219 g; 1.58 mol; 4.4 eq.) was added (temperature was kept < 30 °C) and the suspension was stirred at room temperature for 3 h. Water (1250 mL) was added and the mixture was treated with a cone. HC1 solution (approx. 130 mL) to adjust the pH to 7.8. The reaction mixture was extracted twice with methyl- /-butyl ether (MTBE; 2×465 mL). The combined MTBE phases were washed with water (155 mL). Water (190 mL) was added to the MTBE phase and the organic solvent was distilled off under reduced pressure (50 mbar). The obtained emulsion of compound 18a (yield: 99%) was directly used for the next step.

1H-NMR (600.6 MHz, CDC13) d: 0.87 (d, J = 6.8 Hz, 3H), 1.09 (d, / = 6.8 Hz, 3H), 2.20 (sept. / = 6.8 Hz, 1H), 2.47 (s, 1H), 2.77 (s, 1H), 6.63 (t, ²7H,F = 73.5 Hz, 1H), 6.63 (t, ²/H,F = 73.5 Hz, 1H), 7.65 (s, 1H), 7.69 (s, 1H), 7.74 (dd, 7 = 8.6, 1.7 Hz, 1H), 7.79 (d, / =

8.6 Hz, 1H), 8.06 (s (broad), 1H);

HPLC (purity): 95%.

Preparation 2:

The solution of tertiary alcohol 18b (48 wt%; 57.5 mol; 1 eq.) in heptane was dissolved in methanol (128 L). Potassium carbonate (35.0 kg; 253 mol; 4.4 eq.) was added (temperature was kept < 30 °C) and the suspension was stirred at 20-30 °C for 3 h. Water (200 L) was added and the mixture was treated with an aqueous HC1 solution (approx. 25 L; 32 wt%) to adjust the pH to 7.5 – 7.8. The reaction mixture was extracted twice with MTBE

(2×66.6 L). The combined MTBE phases were washed with water (25 L). Water (30 L) was added to the MTBE phase and the organic solvent was distilled off under reduced pressure (<80 mbar; 55°C). The residue was dissolved in tert-butanol (25 L). The resulting 18a was cooled to <30°C and used directly in the next step.

^-NMR (600.6 MHz, CDC13) d: 0.87 (d, / = 6.8 Hz, 3H), 1.09 (d, / = 6.8 Hz, 3H), 2.20 (sept. / = 6.8 Hz, 1H), 2.47 (s, 1H), 2.77 (s, 1H), 6.63 (t, ²7H,F = 73.5 Hz, 1H), 6.63 (t, ²/H,F = 73.5 Hz, 1H), 7.65 (s, 1H), 7.69 (s, 1H), 7.74 (dd, 7 = 8.6, 1.7 Hz, 1H), 7.79 (d, / = 8.6 Hz, 1H), 8.06 (s (broad), 1H);

HPLC (purity): 95%.

Preparation of Compound 31

Preparation 1:

Benzyl bromide (39.4 g; 0.23 mol; 1 eq.) was dissolved in water (177 mL) and t-BuOH (200 mL). Diisopropylethylamine (DIPEA; 59.4 g; 0.46 mol; 2 eq.) and sodium azide (15.0 g; 0.23 mol; 1 eq.) were added. The suspension was stirred for 5 min at room temperature. A suspension of compound 18a (82 g; 0.23 mol; 1 eq.) in water (123 mL) was treated with t-BuOH (100 mL) and copper (I) iodide (8.8 g; 46 mmol; 0.2 eq.) was added and the temperature was kept below 30 °C. The yellow-brown suspension was stirred for 5 h at room temperature. Zinc powder (5.0 g; 76 mmol) and ammonium chloride (7.4 g; 0.14 mol) were added and the reaction mixture was stirred at room temperature for 3 hours. The mixture was diluted with MTBE (800 mL), water (280 mL), and an aqueous ammonia solution (120 g; 25 wt%). Solids were removed by filtration and additional MTBE (200 mL) and brine (200 mL) were added. The aqueous phase was separated and extracted with MTBE (400 mL). The combined organic phases were treated with water (150 mL) and MTBE was distilled off under reduced pressure (100 mbar). The obtained suspension of compound 31 (113 g; 50 wt%) in water (approx. 113 mL) was directly used for the next step.

Ή-NMEI (600.6 MHz, DMSO-D6) d: 0.66 (d, / = 6.8 Hz, 3H), 0.83 (d, / = 6.7 Hz, 3H), 2.78 (sept. / = 6.8 Hz, 1H), 5.55 (s, 2H), 5.68 (s, 1H), 7.29 (t, ²7H,F = 73.4 Hz, 1H), 7.32 (t, ²7H,F = 73.4 Hz, 1H), 7.36 – 7.26 (m, 5H), 7.79 (s, 1H), 7.82 (s, 1H), 7.82 (dd, 7 = 8.8, 1.7 Hz, 1H), 7.86 (d, / = 8.8 Hz, 1H), 7.94 (s, 1H), 8.10 (s (broad), 1H);

HPLC (purity): 87%.

Preparation 2:

Benzyl bromide (11.0 kg g; 64.4 mol; 1,12 eq.) was dissolved in water (40 L) and t-BuOH (60 L). DIPEA (16.4 kg; 126.5 mol; 2,2 eq.) and sodium azide (4.12 kg; 63.3 mol; 1 eq.) were added. The suspension was stirred 5 min at room temperature. A mixture of compound 18a (20.5 kg; 57.5 mol; 1 eq.) in ieri-butanol (see previous step) was added together with water (5 L) and copper (I) iodide (2.2 kg; 11.5 mol; 0.2 eq.) at a temperature < 30 °C. The yellow-brown suspension was stirred for 5 h at room temperature. Zinc powder (1.25 kg; 19 mol, 0.33 eq.) and an aqueous solution of ammonium chloride (2.14 kg; 20 wt%; 40 mol; 0.7 eq.) were added and the reaction mixture was stirred at 20-30 °C for 2 hours. The reaction mixture was concentrated under vacuum (<200 mbar, 55 °C). The residue was diluted with MTBE (200 L), water (30 L), and an aqueous ammonia solution (30 kg; 25 wt%). Solids were removed by filtration over a pad of“Kieselgur NF” (2 kg). Brine (50 L) was added for a better phase separation. The aqueous phase was separated and washed with MTBE (200 L). The combined organic phases were washed with an aqueous HC1 solution (1 N, 52 L) and water (50 L). MTBE was distilled off under reduced pressure (<400 mbar, 55°C; distillate min. 230L). The oily residue was dissolved in ethanol (150 L), which was distilled off under reduced pressure (<300 mbar; 55°C; distillate min. 150-155L) and the residue was dissolved in additional ethanol (60 L). To the resulting solution of compound 31 was added water (24 L) and the mixture was warmed to 50-55 °C. The mixture was cooled to 30 °C and crystallization started. The suspension was stirred at 30 °C for 1 h, cooled to <0 °C within 2 hours, and stirred at -5-0 °C for an additional 2 hours. The solids were isolated and washed with ethanol/water (1/1; v/v) (2 x 12 L). The wet product was dissolved in ethanol (115L) at 60 °C and water (24 L) was added. The mixture was cooled to 40 °C and the crystallization started. The suspension was stirred at 30 °C for 1 h, cooled to <0 °C within 2 hours, and stirred at -5-0 °C for additional 2 hours. The solids were isolated and washed (without stirring) with ethanol/water (1/1; v/v) (3 x 8 L). Pure, wet compound 31 was isolated as a white solid, which was used for the next step without drying. 14.0 kg of wet 31 were obtained with a 31 content of 81.6 wt%. Based on the determined content, the calculated amount of pure 31 was 11.4 kg with a yield of 41% over two steps (from 18b).

1H-NMR (600.6 MHz, DMSO-D6) d: 0.66 (d, J = 6.8 Hz, 3H), 0.83 (d, / = 6.7 Hz, 3H), 2.78 (sept. / = 6.8 Hz, 1H), 5.55 (s, 2H), 5.68 (s, 1H), 7.29 (t, ²7H,F = 73.4 Hz, 1H), 7.32 (t, ²7H,F = 73.4 HZ, 1H), 7.36 – 7.26 (m, 5H), 7.79 (s, 1H), 7.82 (s, 1H), 7.82 (dd, 7 = 8.8, 1.7 Hz, 1H), 7.86 (d, / = 8.8 Hz, 1H), 7.94 (s, 1H), 8.10 (s (broad), 1H);

HPLC (purity): 87%.

Preparation 3: Synthesis of compound 31 directly from compound 18b

Benzyl bromide (1.64 g, 9.59 mmol, 1.12 eq) was dissolved in water (2.4 mL) and

MeOH (2.4 mL). K2CO3 (2.38 g, 17.2 mmol, 2.00 eq), sodium ascorbate (0.34 g, 1.72 mmol, 0.20 eq) and finally sodium azide (0.62 g, 9.40 mmol, 1.10 eq.) were added. The suspension was stirred for 5 min at room temperature. A suspension of 18b (3.08 g; 8.64 mmol, 1.00 eq) in water (2.5 mL) and MeOH (2.5 mL) and the resulting mixture was stirred for 10 min.

CuS0₄ (0.21 g, 1.30 mmol, 0.15 eq) were added (slightly exothermic reaction). The reaction mixture was stirred for 19 h and the conversion was determined by HPLC (conv. 100%, purity of compound 31 by HPLC: 83 area%). To the yellow-green suspension was added zinc powder (0.24 g, 4.13 mmol, 0.43 eq) and ammonium chloride (0.34 g, 6.36 mmol, 0.74 eq) were added and the reaction mixture was stirred at room temperature for 2 hours. The reaction mixture was concentrated under reduced pressure (150 mbar, 50 °C). The mixture was diluted with MTBE (40 mL), water (15 mL), and an aqueous ammonia solution (6.5 mL). Solids were removed by filtration and brine (5.5 mL) was added. The aqueous phase was separated and extracted with MTBE (20 mL). The combined organic phases were treated with water (10 mL) and the pH was adjusted to a pH of 1 by addition of cone. HC1. After phase separation, the organic layer was washed with water (10 mL). MTBE was distilled off under reduced pressure (100 mbar, 50°C) to give the crude compound 31 as an oil. Water (2.5 mL) and EtOH (30 mL) were added and the mixture was warmed to 50 °C. After cooling to 30 °C, the mixture was seeded with compound 31 and compound 31 started to precipitate. The mixture was kept for 1 h at 30 °C, then cooled to 0 °C over 2 h and kept at 0 °C for 2 h. The resulting product, 31, was collected by filtration and the filter cake was washed with small portions of EtOH/water (1:1). After drying, the product (2.97 g) was obtained as a pale yellow, crystalline solid with an HPLC purity of 79 area% and a NMR content of ca. 70 wt%.

Recrystallization of
31

Preparation 1:

To a suspension of compound 31 (96 g; 0.196 mol; 50 wt%) in water (96 mL) was added ethanol (480 mL) and the mixture was warmed to 50 °C. The mixture was cooled to 30 °C and crystallization started. The suspension was stirred at 30 °C for 1 h, cooled to 0 °C within 2 hours and stirred at 0 °C for additional 2 hours. The solids were isolated and washed with ethanol/water (1/1; v/v) (3 x 40 mL). The wet product was dissolved in ethanol (280 mL) at 60 °C and water (56 mL) was added. The mixture was cooled to 40 °C and crystallization started. The suspension was stirred at 30 °C for 1 h, cooled to 0 °C within 2 hours, and stirred at 0 °C for an additional 2 hours. The solids were isolated and washed with ethanol/water (1/1; v/v) (3 x 28 mL). Pure, wet compound 31 (46.8 g on dried basis; 49 % over 2 steps) was isolated as a white solid, which was used for the next step without drying.

HPLC (purity): 99.5%;

chiral HPLC: e.r.: 0.2:99.8%.

mp of dried product: 110 °C.

Preparation 2:

14 kg of ethanol-wet 31 (content 81.6 wt%, calculated 11.4 kg, 23.7 mol) were suspended in ethanol (46 L) and the mixture was warmed to 50-55 °C, forming a homogenous solution at this temperature. Water (9 L) was added at 50-55 °C and the mixture was cooled to 40-45 °C. After the crystallization had started, the suspension was stirred at 40-45 °C for 1 h, cooled to 0 °C within 2 hours, and stirred at 0 °C for additional 2 hours. The solids were isolated and washed with ethanol/water (1/1; v/v) (3 x 8 L). Pure, wet compound 31 (14.5 kg) was isolated as a white solid, which was used for the next step without drying.

1H-NMR (600.6 MHz, DMSO-D6) d: 0.66 (d, / = 6.8 Hz, 3H), 0.83 (d, / = 6.7 Hz, 3H), 2.78 (sept. / = 6.8 Hz, 1H), 5.55 (s, 2H), 5.68 (s, 1H), 7.29 (t, ²7H,F = 73.4 Hz, 1H), 7.32 (t, ²7H,F = 73.4 Hz, 1H), 7.36 – 7.26 (m, 5H), 7.79 (s, 1H), 7.82 (s, 1H), 7.82 (dd, 7 = 8.8, 1.7 Hz, 1H), 7.86 (d, / = 8.8 Hz, 1H), 7.94 (s, 1H), 8.10 (s (broad), 1H);

HPLC (purity): 99.8%;

chiral HPLC: e.r.: 0.2:99.8%.

mp of dried product: 110 °C.

Preparation of Azidomethyl Pivalate Protected Triazole (6) from Compound 18a

Azidomethyl pivalate (1.42 g, 9.00 mmol, 1.05 eq) was suspended in water (6.0 mL) and t-BuOH (7.2 mL) and the suspension was stirred for 5 min. Compound 18a (theor. 3.08 g, 8.64 mmol, 1.00 eq), sodium ascorbate (0.48 g, 2.4 mmol, 0.30 eq), and CuS0₄ (0.08 g, 0.40 mmol, 0.05 eq.) were added. The reaction mixture was stirred for 19 h and conversion was determined by HPLC (conv. 98%, purity of the product by HPLC: 81 area%). To the green suspension was added MTBE (20 mL), water (10 mL), and an aqueous ammonia solution (2 g). A biphasic turbid mixture was formed. To improve phase separation, additional MTBE (20 mL) and water (10 mL) were added. The aqueous phase was separated and extracted with MTBE (20 mL). The combined organic phases were concentrated under reduced pressure (100 mbar, 50 °C) to give the crude product as a brown oil that solidified upon standing. HPLC purity: ca. 65 area%; NMR content of ca. 73 wt%.

1H-NMR (600.6 MHz, CDCL) d: 0.79 (d, 3H), 0.93 (d, 3H), 1.15 (s. 9H), 2.86 (sept, 1H), 3.12 (s, 1H), 6.20 (s, 2H), 6.59 (t/t, ²7H,F = 73.5 Hz, 2H), 7.61 (1, 1H), 7.64 (s, 1H), 7.70 – 7.82 (m, 3H), 8.04 (s, 1H).

Preparation of Azidomethyl Pivalate Protected Triazole (6) from 18b

In a reaction flask, sodium ascorbate (277 mg, 1.4 mmol, 1.20 eq) and CuS0₄ (37 mg, 0.23 mmol, 0.20 eq.) were suspended in MeOH (11 mL). Azidomethyl pivalate (183 mg, 1.16 mmol, 1.00 eq) and 18b (183 mg, 1.16 mmol, 1.00 eq) were added and the mixture was warmed to 60 °C. The reaction mixture was stirred for 19 h and worked up. To the green suspension was added an aq NH₄Cl solution (2 mL) and zinc powder, and the mixture was stirred for 2 h. MTBE (2 mL) was added and the aqueous phase was separated and extracted with MTBE (2 mL). The combined organic phases were concentrated under reduced pressure (100 mbar, 50 °C) to give 6 as a brown oil that solidified upon standing. HPLC purity: ca. 81 area%; NMR content of ca. 57 wt%.

Preparation of Compound 1

Preparation 1:

Compound 31 (26 g; 53 mmol; 1 eq.) was dissolved in ethanol (260 mL) and Noblyst Pl 155 (2.2 g; 10 % Pd; 54 wt% water) was added. The autoclave was flushed with nitrogen and hydrogen (5 bar) was added. The reaction mixture was stirred at room temperature for 32 hours. The reaction mixture was treated with charcoal (2 g), stirred for 15 min, and the charcoal was filtered off. The filtrate was concentrated via distillation and the residue (approximately 42 g) was diluted with heptane (200 mL). The mixture was heated to reflux to

obtain a clear solution. The solution was cooled to room temperature within 1 h and the resulting suspension was cooled to 0 °C and stirred for 2 hours at 0 °C. The solids were isolated via filtration and washed with heptane/ethanol (10:1; v/v; 3×10 mL). Compound 1 (18.0 g; 85 %) was dried under vacuum at 60 °C for 24 hours and obtained as a white, crystalline solid.

1H-NMR (600 MHz) d: 0.80 (d, J = 6.8 Hz, 3H), 0.97 (d, / = 6.7 Hz, 3H), 2.83 (sept. / = 6.8 Hz, 1H), 6.60 (t, ²7H,F = 73.5 Hz, 1H), 6.61 (t, ²7H,F = 73.5 Hz, 1H), 7.61 (s, 1H), 7.65 (s, 1H), 7.68 (dd, / = 8.7, 1.6 Hz, 1H), 7.74 (s, 1H), 7.75 (d, / = 8.7 Hz, 1H), 8.02 (s (broad), 1H); HPLC (purity): 100%.

Preparation 2:

Compound 31 (26.5 kg; 53.5 mol; 1 eq.) was dissolved in ethanol (265 L) and Pd/C (2.0 kg; 10 % Pd; 54 wt% water) was added. The reactor was flushed with nitrogen, and hydrogen (4.5 bar) was added. The reaction mixture was stirred at 28-32 °C until the reaction was complete. The reaction mixture was treated with charcoal (1.3 kg) at a temperature of <

33 °C, stirred for 10 min, and the charcoal was filtered off, and the filter was washed with ethanol (10 L).The filtrates from two reactions were combined and concentrated via distillation under reduced pressure (max. 65 °C; distillate: min 480 L). The residue (approx. 50-60 L) was diluted with isopropylacetate (250 L). The mixture was again concentrated via distillation under reduced pressure (max. 65 °C; distillate: min 240-245 L). The residue (approx. 60-70 L) was cooled to 35-40 °C and isopropylacetate (125 L) and heptane (540 L) were added. The suspension was heated to reflux (approx. 88 °C) and stirred under reflux for 15-20 min. Subsequently, the mixture was cooled to 0-5 °C within 2 h and stirred at 0-5 °C for 2 hours. The solids were isolated via filtration and washed with heptane/isopropylacetate (5:1; v/v; 2×30 L; 0-5 °C). Wet 1 was dried under vacuum at 60 °C and was obtained as a white, crystalline solid (35.4 kg, 81.9%).

1H-NMR (600 MHz) d: 0.80 (d, / = 6.8 Hz, 3H), 0.97 (d, / = 6.7 Hz, 3H), 2.83 (sept. / = 6.8 Hz, 1H), 6.60 (t, ²7H,F = 73.5 Hz, 1H), 6.61 (t, ²7H,F = 73.5 Hz, 1H), 7.61 (s, 1H), 7.65 (s, 1H), 7.68 (dd, / = 8.7, 1.6 Hz, 1H), 7.74 (s, 1H), 7.75 (d, / = 8.7 Hz, 1H), 8.02 (s (broad), 1H); HPLC (purity): 100%.

Preparation 3: Preparation of Compound 1 from Compound 6

At room temperature, 6 (3.00 g, 5.84 mmol) was dissolved in MeOH (19.8 mL). NaOH (1.0 M, 19.8 mL) was added in one portion and the reaction mixture was stirred for 1 h at room temperature. The reaction progress was monitored by HPLC, which showed 98% conversion after 1 h. Aq. HC1 (19.8 mL) was added and the mixture was diluted with water (120 mL) and MTBE (60 mL), resulting in a clear biphasic solution. After phase separation, the organic phase was washed with aq NaHC0₃ (20 mL). The organic layer was concentrated under high vacuum (25 mbar, 45 °C) to yield 2.77 g of 1 as a greenish oil. The identity was confirmed by comparison of HPLC retention time with an authentic sample of 1 as well as by 1H NMR.

Recrystallization of Compound 1

Wet 1 (40 kg; isopropylacetate/heptane wet) was treated with isopropylacetate (110 L) and heptane (440 L). The suspension was heated to reflux (approx. 88 °C) and stirred under reflux for 15-20 min. Subsequently, the mixture was cooled to 0-5 °C within 2 h and stirred at 0-5 °C for 2 hours. The solids were isolated via filtration and washed with

heptane/isopropylacetate (5:1; v/v; 2×30 L; 0-5 °C). A sample was taken for analysis

(criterion: a) purity; NLT 99.0 A% by HPLC; b) single impurities, NMT 0.15 A% by HPLC; c) enantiomer VT-463, NMT 1.0 A% by HPLC). Wet 1 was dried under vacuum at 60 °C for not less than 12 h. A sample was taken for analysis: criterion: a) LOD; NMT 0.5 wt% by gravimetry; b) residual toluene, NMT 890 ppm by HS-GC. 1 was obtained as a white, crystalline solid (28.5 kg, 66.7% from 31).

PAPER

Bioorganic & Medicinal Chemistry Letters (2014), 24(11), 2444-2447.

https://www.sciencedirect.com/science/article/pii/S0960894X14003606

PATENT

WO 2016040896

https://patents.google.com/patent/WO2016040896A1/en

References

^ Jump up to:^a ^b ^c ^d ^e http://adisinsight.springer.com/drugs/800035241
^ http://www.pharmaceutical-technology.com/news/newsfda-grants-fast-track-status-innocrins-seviteronel-treat-metastatic-crpc-4770025
^ Yin L, Hu Q, Hartmann RW (2013). “Recent progress in pharmaceutical therapies for castration-resistant prostate cancer”. Int J Mol Sci. 14 (7): 13958–78. doi:10.3390/ijms140713958. PMC 3742227. PMID 23880851.
^ Jump up to:^a ^b Stein MN, Patel N, Bershadskiy A, Sokoloff A, Singer EA (2014). “Androgen synthesis inhibitors in the treatment of castration-resistant prostate cancer”. Asian J. Androl. 16 (3): 387–400. doi:10.4103/1008-682X.129133. PMC 4023364. PMID 24759590.
^ Jump up to:^a ^b Rafferty SW, Eisner JR, Moore WR, Schotzinger RJ, Hoekstra WJ (2014). “Highly-selective 4-(1,2,3-triazole)-based P450c17a 17,20-lyase inhibitors”. Bioorg. Med. Chem. Lett. 24 (11): 2444–7. doi:10.1016/j.bmcl.2014.04.024. PMID 24775307.
^ Jump up to:^a ^b ^c ^d Toren PJ, Kim S, Pham S, Mangalji A, Adomat H, Guns ES, Zoubeidi A, Moore W, Gleave ME (2015). “Anticancer activity of a novel selective CYP17A1 inhibitor in preclinical models of castrate-resistant prostate cancer”. Mol. Cancer Ther. 14 (1): 59–69. doi:10.1158/1535-7163.MCT-14-0521. PMID 25351916.
^ Jump up to:^a ^b ^c Stephen Neidle (30 September 2013). Cancer Drug Design and Discovery. Academic Press. pp. 341–342. ISBN 978-0-12-397228-6.
^ Jump up to:^a ^b Wm Kevin Kelly; Edouard J. Trabulsi, MD; Nicholas G. Zaorsky, MD (17 December 2014). Prostate Cancer: A Multidisciplinary Approach to Diagnosis and Management. Demos Medical Publishing. pp. 342–. ISBN 978-1-936287-59-8.
^ http://www.who.int/medicines/publications/druginformation/innlists/RL76.pdf

External links[

Seviteronel – AdisInsight

Seviteronel
Clinical data

Synonyms	VT-464; INO-464
Routes of administration	By mouth
Drug class	Androgen biosynthesis inhibitor; Nonsteroidal antiandrogen
ATC code	None
Identifiers
IUPAC name [show]
CAS Number	1610537-15-9
PubChem CID	78357816
ChemSpider	32738723
UNII	8S5OIN36X4
Chemical and physical data
Formula	C₁₈H₁₇F₄N₃O₃
Molar mass	399.339 g/mol g·mol⁻¹
3D model (JSmol)	Interactive image
SMILES [hide] CC(C)C(C1=CC2=CC(=C(C=C2C=C1)OC(F)F)OC(F)F)(C3=NNN=C3)O
InChI [hide] InChI=1S/C18H17F4N3O3/c1-9(2)18(26,15-8-23-25-24-15)12-4-3-10-6-13(27-16(19)20)14(28-17(21)22)7-11(10)5-12/h3-9,16-17,26H,1-2H3,(H,23,24,25)/t18-/m0/s1 Key:ZBRAJOQFSNYJMF-SFHVURJKSA-N

References

Innocrin Pharmaceuticals Created as a Spin-out of the Prostate Cancer Program from Viamet Pharmaceuticals.

Media Release
Viamet Pharmaceuticals and the Novartis Option Fund Enter Agreement for Development of Novel Metalloenzyme Inhibitors.

Media Release
Innocrin Pharmaceuticals, Inc. Granted SME Status Designation by the European Medicines Agency.

Media Release
A Single arm, open label, signal seeking, Phase II a trial of the activity of seviteronel in patients with androgen receptor (AR) positive solid tumours

ctiprofile
Innocrin Pharmaceuticals and the Prostate Cancer Foundation (PCF) Join Forces for Innovative Phase 2 Clinical Study.

Media Release
A Phase 2 Open-label Study to Evaluate the Efficacy and Safety of Seviteronel in Subjects With Castration-Resistant Prostate Cancer Progressing on Enzalutamide or Abiraterone

ctiprofile
Innocrin Pharmaceuticals, Inc. Granted Fast Track Designation by FDA for VT-464 Treatment of Patients with Metastatic Castrate-resistant Prostate Cancer.

Media Release
Innocrin Pharmaceuticals, Inc. Begins Phase 2 Study of Seviteronel in Women with Estrogen Receptor-positive or Triple-negative Breast Cancer and Expands Two Phase 2 Studies of Seviteronel in Men with Metastatic Castrate-resistant Prostate Cancer.

Media Release
A Phase 2 Open-Label Study to Evaluate the Efficacy and Safety of VT-464 in Patients With Metastatic Castration Resistant Prostate Cancer Who Have Previously Been Treated With Enzalutamide, Androgen Receptor Positive Triple-Negative Breast Cancer Patients, and Men With ER Positive Breast Cancer

ctiprofile
Innocrin Pharmaceuticals Inc. to Present Interim Results from Its Phase 1/2 Prostate Cancer Clinical Study and Preclinical Results That Demonstrate VT-464 Efficacy in a Clinically-Relevant Enzalutamide-Resistant Mouse Model.

Media Release
A Phase 1/2 Open-Label Study to Evaluate the Safety, Pharmacokinetics, and Pharmacodynamics of Seviteronel in Subjects With Castration-Resistant Prostate Cancer

ctiprofile
A Phase 1/2 Open-Label, Multiple-Dose Study to Evaluate the Safety, Tolerability, Pharmacokinetics, and Pharmacodynamics of Once-Daily VT-464 in Patients With Castration-Resistant Prostate Cancer

ctiprofile
Viamet Pharmaceuticals Appoints Former Novartis Executive Marc Rudoltz, M.D. as Chief Medical Officer.

Media Release
VIAMET PHARMACEUTICALS AND THE NATIONAL INSTITUTES OF HEALTH TO JOINTLY DEVELOP NOVEL VIAMET COMPOUND.

Media Release
Viamet Pharmaceuticals Initiates Phase 1/2 Clinical Trial of Novel Prostate Cancer Therapy, VT-464.

Media Release
Viamet Pharmaceuticals to Present at the 32nd Annual J.P. Morgan Healthcare Conference.

Media Release
VIAMET PHARMACEUTICALS TO PRESENT AT THE 31st Annual J.P. MORGAN HEALTHCARE CONFERENCE.

Media Release
Innocrin Pharmaceuticals, Inc. Initiates Phase 2 Castration-Resistant Prostate Cancer (CRPC) Study in Men Who Have Failed Enzalutmaide or Abiraterone.

Media Release
Innocrin Pharmaceuticals Appoints Fred Eshelman, PharmD as CEO and is Granted Fast Track Designation by FDA for Seviteronel Treatment of Women with Triple-negative Breast Cancer and Women or Men with Estrogen Receptor-positive Breast Cancer.

Media Release
Gucalp A, Bardia A, Gabrail N, DaCosta N, Danso M, Elias AD, et al. Phase 1/2 study of oral seviteronel (VT-464), a dual CYP17-lyase inhibitor and androgen receptor (AR) antagonist, in patients with advanced AR positive triple negative (TNBC) or estrogen receptor (ER) positive breast cancer (BC). SABCS-2016 2016; abstr. P2-08-04.

Available from: URL:http://www.abstracts2view.com/sabcs/view.php?nu=SABCS16L_1479
Innocrin Pharmaceuticals Presents Data from the Ongoing Phase 2 Trial of Seviteronel in Estrogen Receptor-positive or Triple-negative Breast Cancer (CLARITY-01) at the San Antonio Breast Cancer Symposium.

Media Release
Innocrin Pharmaceuticals, Inc. Appoints Edwina Baskin-Bey, MD as Chief Medical Officer and Expands the Ongoing Phase 2 Study of Seviteronel in Women with Estrogen Receptor-positive or Triple-negative Breast Cancer (TNBC).

Media Release
Innocrin Pharmaceuticals, Inc. Raises $28 Million in Series D Financing.

Media Release
A Phase 1/2 Open-Label Study to Evaluate the Safety, Pharmacokinetics, Pharmacodynamics and Efficacy of Seviteronel in Subjects With Advanced Breast Cancer

ctiprofile
Speers CW, Chandler B, Zhao S, Liu M, Wilder-Romans K, Olsen E, et al. Radiosensitization of androgen receptor (AR)-positive triple-negative breast cancer (TNBC) cells using seviteronel (SEVI), a selective CYP17 lyase and AR inhibitor. ASCO-2017 2017; abstr. e12102.

Available from: URL: http://abstracts.asco.org/199/AbstView_199_193240.html
Innocrin Pharmaceuticals, Inc. Appoints Charles F. Osborne Jr. as its Chief Financial Officer.

Media Release
Viamet Pharmaceuticals Secures $18 Million Financing.

Media Release
Viamet Pharmaceuticals Raises $4 Million Round of Financing.

Media Release

///////////SEVITERONEL, VT-464, INO-464, VT 464, INO 464, Phase II, Breast cancer, Prostate cancer, Solid tumours, viamet, CANCER, севитеронел , سيفيتيرونيل , 赛维罗奈 ,

C1(=CN=NN1)C(C1=CC2=C(C=C1)C=C(C(=C2)OC(F)F)OC(F)F)(C(C)C)O

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