Preparation of [4-amino-2-[(1-methanesulfonylpiperidin-4-yl)amino]pyrimidin-5-yl](2,3-difluoro-6-methoxyphenyl)methanone_ Kain Industrial Additives

Background and overview^[1]

R547 ([4-Amino-2-[(1-methanesulfonylpiperidin-4-yl)amino]pyrimidin-5-yl](2,3-difluoro-6-methoxyphenyl) Methone) is a potent inhibitor of CDKs 1, 2, 4, 7, and 9 and has shown broad antitumor activity in preclinical models. The Phase Ia study evaluated safety, pharmacokinetic (PK) and pharmacodynamic (PD) endpoints. The method is: R547 ([4-amino-2-[(1-methanesulfonylpiperidin-4-yl)amino]pyrimidin-5-yl](2,3-difluoro-6-methoxyphenyl) ketone) given as a 90 or 180 minute infusion on D1, D8 (21 day cycle). The results showed that 41 patients tolerated R547 ([4-amino-2-[(1-methanesulfonylpiperidin-4-yl)amino]pyrimidin-5-yl](2, 3-Difluoro-6-methoxyphenyl)methanone).

Adverse reactions^[1]

The main associated events were nausea (54%), fatigue (34%), vomiting (34%), headache (34%) and hypotension (32%).

Pharmacokinetics^[1]

PK: Mean AUC, PK of 20 patients receiving ≥155 mg/m 2 R547 ([4-amino-2-[(1-methanesulfonylpiperidin-4-yl)amino]pyrimidin-5-yl] (2,3-difluoro-6-methoxyphenyl)methanone) exceeds effective exposure in xenograft studies. At equivalent doses, a period of 180 minutes produced equivalent AUC but a 30% reduction in Cmax.

PD: For patients, at R547 ([4-amino-2-[(1-methanesulfonylpiperidin-4-yl)amino]pyrimidin-5-yl](2,3-difluoro-6- An exposure-dependent decrease in the pRB/total RB ratio was observed from 1.5 to 24 hours after methoxyphenyl)methanone).

Preparation^[2]

Step 1: Preparation of 4-amino-2-ethylsulfanylpyrimidine-5-carboxylic acid methoxymethylamide (8)

To 4-amino-2-ethylthiopyrimidine-5-carboxylic acid (7) (1.00 g, 5.02 mmol) and diisopropylethylamine (2.79 g, 21.58 mmol, 4 equiv.) in DMF (20 mL ) solution, add 1-hydroxybenzotriazole (at 0°C, add 1.09g, 8.07mmol, 1.5 equivalents) and O-benzotriazole-1-yl-N, N, N, N-tetramethyl Urea hexafluorophosphate (3.06 g, 8.07 mmol, 1.5 equivalents). After stirring for 15 minutes, N,O-dimethylhydroxylamine hydrochloride (790 mg, 8.10 mmol) was added, and the reaction was stirred at 0-20°C for 2 hours. The resulting reaction mixture was treated with H2O and extracted with EtOAc. The combined organic extracts were washed with brine, dried over NaSO and concentrated. The crude product was purified on silica gel with Hexane/EtOAc (1:1) to give 8 as a white solid (1.19g): 4-amino-2-ethylsulfanylpyrimidine-5-carboxylic acid methoxymethyl amide (8), yield 98%.

Step 2 Preparation of (4-amino-2-ethylthiopyrimidin-5-yl) (2-methoxyphenyl)-methanone (10c)

Add a solution of 2-iodoanisole (20.0g, 85.47mmol) in anhydrous THF (130mL) to a 1.6M n-butyllithium hexane solution (51.0mL, 81.6mmol) within 30 minutes, and stir The mixture was heated at -78°C for an additional 40 minutes, resulting in a clear solution. A portion of freshly prepared 2-methoxyphenyllithium reagent (70.0 mL, ~30 mmol, ~3.5 equiv) was slowly added to a solution of amide 8 (2.07 g, 8.54 mmol) in anhydrous THF (30 mL) and the mixture was stirred at The mixture was kept at -78°C for 1-2 hours until complete consumption of amide 8, and the resulting mixture was quenched with aqueous NH4Cl and extracted with EtOAc (3x). The combined organic extracts were washed with brine (2x), dried over Na2SO4 and evaporated. The residue was purified on silica gel with Hexane/EtOAc (80/20 (60/40)) to give 10c as a light yellow solid (1.85g)

Step 3: Preparation of (4-amino-2-ethylsulfonylpyrimidin-5-yl)(2-methoxyphenyl)-methanone (11c)

To a cooled solution of 10c (2.66 g, 9.19 mmol) in chloroform (120 mL) add 3-chloroperoxybenzoic acid (~77% purity, 5.20 g, ~23 mmol, 2.5 equiv) in three portions and mix the mixture Stir at 0°C for 1 hour. The resulting mixture was diluted with CHCl (80 mL), washed with 10% aqueous sodium thiosulfate (2 x 30 mL) and brine (2 x 20 mL), dried over NaSO, and evaporated. The residue was purified on silica gel with hexanes/. EtOAc (60/40), obtained 11c as a white solid (2.30, yield 78%):

Step 4: [4-amino-2-[(1-methanesulfonylpiperidin-4-yl)amino]pyrimidin-5-yl](2,3-difluoro-6-methoxybenzene Preparation of methyl ketone

A suspension of 11c (43.1 mg, 0.134 mmol) and ethyl 4-amino-1-piperidinecarboxylate (30.0 mg, 0.174 mmol, 1.3 equiv) in i-PrOH (4 mL) was diluted at 100-110 Heated under microwave radiation at ℃. The resulting mixture was concentrated for 1 h and the residue was purified on silica gel with CH2Cl2/MeOH (95/5) to give 26 as a pale yellow solid (47.1 mg): [4-amino-2-[(1-methanesulfonyl Piperidin-4-yl)amino]pyrimidin-5-yl](2,3-difluoro-6-methoxyphenyl)methanone: white solid; melting point 218-219°C.

Main reference materials

[1] Diab S, Eckhardt S, Tan A, et al. A phase I study of R547, a novel, selective inhibitor of cell cycle and transcriptional cyclin dependent kinases (CDKs)[J]. Journal of Clinical Oncology, 2007, 25(18_suppl): 3528-3528.

[2] Chu X J, DePinto W, Bartkovitz D, et al. Discovery of [4-amino-2-(1-methanesulfonylpiperidin-4-ylamino) pyrimidin-5-yl](2, 3-difluoro-6 -methoxyphenyl) methanone (R547), a potent and selective cyclin-dependent kinase inhibitor with significant in vivo antitumor activity[J]. Journal of medicinal chemistry, 2006, 49(22): 6549-6560.