DEY FABIAN (CH)
XU HONGTAO (CN)
ZHANG WEIXING (CN)
ZHU WEI (CN)
HOFFMANN LA ROCHE (US)
WO2015057655A1 | 2015-04-23 | |||
WO2015057659A1 | 2015-04-23 | |||
WO2017106607A1 | 2017-06-22 | |||
WO2018031434A1 | 2018-02-15 | |||
WO2020207991A1 | 2020-10-15 | |||
WO2020020800A1 | 2020-01-30 |
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CLAIMS 1. A compound of formula (I), , wherein R1 is , , , , , or ; wherein R4 is C1-6alkyl, C1-6alkoxy, haloC1-6alkyl, halogen, nitro or cyano; R4a is C1-6alkyl or C3-7cycloalkyl; R5, R5a and R5b are independently selected from H and deuterium; R6 is H or halogen; R2 is C1-6alkyl; R3 is ((amino(C1-6alkoxy)pyrrolidinyl)phenyl)azetidinyl, (amino(C1- 6alkoxy)pyrrolidinyl)pyridinyl, (amino(C1-6alkoxy)pyrrolidinyl)pyridinyloxy, (amino-1,4-oxazepanyl)pyridinyl, (aminoazetidinyl)pyridinyl, (morpholinylC1- 6alkyl)phenyl, (morpholinylC1-6alkyl)phenylamino, (piperazinylphenyl)azetidinyl, (piperazinylphenyl)C1-6alkylamino, aminohalopyrrolidinyl, morpholinylphenyl, morpholinylphenylamino, piperazinylphenyl, piperazinylpyridinyl, piperazinylpyridinyloxy, piperazinylpyrimidinyloxy or pyridinylpiperazinyl; or a pharmaceutically acceptable salt thereof. 2. A compound according to claim 1, wherein R1 is ; wherein R4 is cy 5 ano; R is H or deuterium. 3. A compound according to claim 2, wherein R3 is ((3-amino-4-methoxy-pyrrolidin-1- yl)phenyl)azetidin-1-yl; (3-amino-4-methoxy-pyrrolidin-1-yl)-3-pyridinyl; (3-amino-4-methoxy- pyrrolidin-1-yl)-3-pyridinyloxy; (3-aminoazetidin-1-yl)-3-pyridinyl; (4-morpholin-2- ylmethyl)phenyl; (4-morpholin-2-ylmethyl)phenylamino; (4-piperazin-1-ylphenyl)azetidin-1-yl; (4-piperazin-1-ylphenyl)methylamino; (6-amino-1,4-oxazepan-4-yl)-3-pyridinyl; 2-piperazin-1- ylpyrimidin-5-yloxy; 3-amino-4-fluoro-pyrrolidin-1-yl; 4-morpholin-2-ylphenyl; 4-morpholin-2- ylphenylamino; 4-piperazin-1-ylphenyl; 4-pyridinylpiperazin-1-yl; 5-piperazin-1-yl-2- pyridinyloxy; 5-piperazin-1-yl-3-pyridinyloxy; 6-piperazin-1-yl-3-pyridinyl or 6-piperazin-1-yl- 3-pyridinyloxy. 4. A compound according to any one of claims 1 to 3, wherein R2 is methyl. 5. A compound according to claim 2, wherein R3 is (morpholinylC1-6alkyl)phenylamino, (piperazinylphenyl)azetidinyl, morpholinylphenyl, morpholinylphenylamino, piperazinylpyridinyloxy or piperazinylpyrimidinyloxy. 6. A compound according to claim 2, wherein R3 is (4-morpholin-2-ylmethyl)phenylamino, (4- piperazin-1-ylphenyl)azetidin-1-yl, 4-morpholin-2-ylphenyl, 4-morpholin-2-ylphenylamino, 6- piperazin-1-yl-3-pyridinyloxy or 2-piperazin-1-ylpyrimidin-5-yloxy. 7. A compound according to claim 5, wherein R1 is ; wherein R4 is cyano; R5 is H or deuterium; R2 is C1-6alkyl; R3 is (morpholinylC1-6alkyl)phenylamino, (piperazinylphenyl)azetidinyl, morpholinylphenyl, morpholinylphenylamino, piperazinylpyridinyloxy or piperazinylpyrimidinyloxy; or a pharmaceutically acceptable salt thereof. 8. A compound according to claim 7, wherein R1 is ; wherein R4 is cyano; R5 is H or deuterium; R2 is methyl; R3 is (4-morpholin-2-ylmethyl)phenylamino, (4-piperazin-1-ylphenyl)azetidin-1-yl, 4- morpholin-2-ylphenyl, 4-morpholin-2-ylphenylamino, 6-piperazin-1-yl-3- pyridinyloxy or 2-piperazin-1-ylpyrimidin-5-yloxy; or a pharmaceutically acceptable salt thereof. 9. A compound selected from: 5-[(4R,8R,9aS)-4-methyl-8-[(4-piperazin-1-ylphenyl)methylamino]-1,3,4,6,7,8,9,9a- octahydropyrido[1,2-a]pyrazin-2-yl]quinoline-8-carbonitrile; 5-[(4R,8S,9aS)-4-methyl-8-[(4-piperazin-1-ylphenyl)methylamino]-1,3,4,6,7,8,9,9a- octahydropyrido[1,2-a]pyrazin-2-yl]quinoline-8-carbonitrile; 5-[(4R,8R)-4-methyl-8-(6-piperazin-1-yl-3-pyridyl)-1,3,4,6,7,8,9,9a-octahydropyrido[1,2- a]pyrazin-2-yl]quinoline-8-carbonitrile; 5-[(4R,8R)-4-methyl-8-(4-piperazin-1-ylphenyl)-1,3,4,6,7,8,9,9a-octahydropyrido[1,2- a]pyrazin-2-yl]quinoline-8-carbonitrile; 5-[(4R,8R,9aS)-4-methyl-8-(6-piperazin-1-yl-3-pyridyl)-1,3,4,6,7,8,9,9a- octahydropyrido[1,2-a]pyrazin-2-yl]quinoline-8-carbonitrile; 5-[(4R,8R,9aS)-8-[6-[(3S,4S)-3-amino-4-methoxy-pyrrolidin-1-yl]-3-pyridyl]-4-methyl- 1,3,4,6,7,8,9,9a-octahydropyrido[1,2-a]pyrazin-2-yl]quinoline-8-carbonitrile; 5-[(4R,8R,9aS)-8-[6-(3-aminoazetidin-1-yl)-3-pyridyl]-4-methyl-1,3,4,6,7,8,9,9a- octahydropyrido[1,2-a]pyrazin-2-yl]quinoline-8-carbonitrile; 5-[(4R,8R,9aS)-4-methyl-8-[3-(4-piperazin-1-ylphenyl)azetidin-1-yl]-1,3,4,6,7,8,9,9a- octahydropyrido[1,2-a]pyrazin-2-yl]quinoline-8-carbonitrile; 5-[(4R,8S,9aS)-4-methyl-8-[(5-piperazin-1-yl-2-pyridyl)oxy]-1,3,4,6,7,8,9,9a- octahydropyrido[1,2-a]pyrazin-2-yl]quinoline-8-carbonitrile; 5-[(4R,8R,9aS)-8-[6-[(3R,4S)-3-amino-4-methoxy-pyrrolidin-1-yl]-3-pyridyl]-4-methyl- 1,3,4,6,7,8,9,9a-octahydropyrido[1,2-a]pyrazin-2-yl]quinoline-8-carbonitrile; 5-[(4R,8R,9aS)-8-[6-[(3S,4R)-3-amino-4-methoxy-pyrrolidin-1-yl]-3-pyridyl]-4-methyl- 1,3,4,6,7,8,9,9a-octahydropyrido[1,2-a]pyrazin-2-yl]quinoline-8-carbonitrile; 5-[(4R,8R,9aS)-8-[6-[(3R,4R)-3-amino-4-methoxy-pyrrolidin-1-yl]-3-pyridyl]-4-methyl- 1,3,4,6,7,8,9,9a-octahydropyrido[1,2-a]pyrazin-2-yl]quinoline-8-carbonitrile; 5-[(4R,8R,9aR)-4-methyl-8-(6-piperazin-1-yl-3-pyridyl)-1,3,4,6,7,8,9,9a- octahydropyrido[1,2-a]pyrazin-2-yl]quinoline-8-carbonitrile; 5-[(4R,8S,9aS)-4-methyl-8-[(6-piperazin-1-yl-3-pyridyl)oxy]-1,3,4,6,7,8,9,9a- octahydropyrido[1,2-a]pyrazin-2-yl]quinoline-8-carbonitrile; 5-[(4R,8S,9aS)-4-methyl-8-(2-piperazin-1-ylpyrimidin-5-yl)oxy-1,3,4,6,7,8,9,9a- octahydropyrido[1,2-a]pyrazin-2-yl]quinoline-8-carbonitrile; 5-[(4R,8R,9aS)-8-[6-[(3R,4S)-3-amino-4-fluoro-pyrrolidin-1-yl]-3-pyridyl]-4-methyl- 1,3,4,6,7,8,9,9a-octahydropyrido[1,2-a]pyrazin-2-yl]quinoline-8-carbonitrile; 5-[(4R,8R,9aS)-4-methyl-8-[4-[(2S)-morpholin-2-yl]anilino]-1,3,4,6,7,8,9,9a- octahydropyrido[1,2-a]pyrazin-2-yl]quinoline-8-carbonitrile; 5-[(4R,8R,9aS)-4-methyl-8-[4-[[(2R)-morpholin-2-yl]methyl]anilino]-1,3,4,6,7,8,9,9a- octahydropyrido[1,2-a]pyrazin-2-yl]quinoline-8-carbonitrile; 5-[(4R,8R,9aS)-4-methyl-8-[4-[[(2R)-morpholin-2-yl]methyl]anilino]-1,3,4,6,7,8,9,9a- octahydropyrido[1,2-a]pyrazin-2-yl]-2-deuterio-quinoline-8-carbonitrile5-[(4R,8R,9aS)-4- methyl-8-[4-[[(2R)-morpholin-2-yl]methyl]anilino]-1,3,4,6,7,8,9,9a-octahydropyrido[1,2- a]pyrazin-2-yl]-2-deuterio-quinoline-8-carbonitrile; 5-[(4R,8R,9aS)-8-[6-[(6S)-6-amino-1,4-oxazepan-4-yl]-3-pyridyl]-4-methyl- 1,3,4,6,7,8,9,9a-octahydropyrido[1,2-a]pyrazin-2-yl]quinoline-8-carbonitrile; 5-[(4R,8S,9aS)-4-methyl-8-(4-morpholin-2-ylphenyl)-1,3,4,6,7,8,9,9a- octahydropyrido[1,2-a]pyrazin-2-yl]quinoline-8-carbonitrile; 5-[(4R,8R,9aS)-4-methyl-8-[4-[(2R)-morpholin-2-yl]phenyl]-1,3,4,6,7,8,9,9a- octahydropyrido[1,2-a]pyrazin-2-yl]quinoline-8-carbonitrile; 5-[(4R,8R,9aS)-4-methyl-8-[4-[[(2R)-morpholin-2-yl]methyl]phenyl]-1,3,4,6,7,8,9,9a- octahydropyrido[1,2-a]pyrazin-2-yl]quinoline-8-carbonitrile; 5-[(4R,8S,9aS)-4-methyl-8-[4-(morpholin-2-ylmethyl)phenyl]-1,3,4,6,7,8,9,9a- octahydropyrido[1,2-a]pyrazin-2-yl]quinoline-8-carbonitrile; 5-[(4R,8S,9aS)-8-[[6-[(3R,4S)-3-amino-4-methoxy-pyrrolidin-1-yl]-3-pyridyl]oxy]-4- methyl-1,3,4,6,7,8,9,9a-octahydropyrido[1,2-a]pyrazin-2-yl]quinoline-8-carbonitrile; 5-[(4R,8S,9aS)-4-methyl-8-[(5-piperazin-1-yl-3-pyridyl)oxy]-1,3,4,6,7,8,9,9a- octahydropyrido[1,2-a]pyrazin-2-yl]quinoline-8-carbonitrile; 5-[(4R,8R,9aS)-8-[3-[4-[(3R,4R)-3-amino-4-methoxy-pyrrolidin-1-yl]phenyl]azetidin-1- yl]-4-methyl-1,3,4,6,7,8,9,9a-octahydropyrido[1,2-a]pyrazin-2-yl]quinoline-8-carbonitrile; 5-[(4R,8R,9aS)-8-[3-[4-[(3R,4S)-3-amino-4-methoxy-pyrrolidin-1-yl]phenyl]azetidin-1- yl]-4-methyl-1,3,4,6,7,8,9,9a-octahydropyrido[1,2-a]pyrazin-2-yl]quinoline-8-carbonitrile; 5-[(4R,8R,9aS)-4-methyl-8-[4-[[(2S)-morpholin-2-yl]methyl]anilino]-1,3,4,6,7,8,9,9a- octahydropyrido[1,2-a]pyrazin-2-yl]-2-deuterio-quinoline-8-carbonitrile; 5-[(4R,8R,9aS)-8-[3-[4-[(3R,4S)-3-amino-4-methoxy-pyrrolidin-1-yl]phenyl]azetidin-1- yl]-4-methyl-1,3,4,6,7,8,9,9a-octahydropyrido[1,2-a]pyrazin-2-yl] ]-2-deuterio-quinoline-8- carbonitrile; and 5-[(4R,8R,9aS)-4-methyl-8-[4-(4-pyridyl)piperazin-1-yl]-1,3,4,6,7,8,9,9a- octahydropyrido[1,2-a]pyrazin-2-yl]-2-deuterio-quinoline-8-carbonitrile; or a pharmaceutically acceptable salt thereof. 10. A process for the preparation of a compound according to any one of claims 1 to 19 comprising any one of the following steps: a) Buchwald-Hartwig amination reaction between compound of formula (VIII), , and amine (V), b) reductive amination of compound of formula (IV), , with amine (V), ; c) Buchwald-Hartwig amination reaction between compound of formula (XIV), , and amine (V), ; d) Buchwald-Hartwig amination reaction between compound of formula (XVIII), , and amine (V), ; wherein X is halogen; R7 is H; R8 is heterocyclylheteroaryl, heterocyclylC1- 6alkylheteroaryl, heterocyclylheteroarylC1-6alkyl, heterocyclylC1-6alkylaryl, heterocyclylarylC1- 6alkyl; or R7 and R8 together with the nitrogen they are attached to form a heterocyclyl; W is heteroaryl or aryl; R1 and R2 are defined as in any one of claims 1 to 8. 11. A compound or pharmaceutically acceptable salt according to any one of claims 1 to 9 for use as therapeutically active substance. 12. A pharmaceutical composition comprising a compound in accordance with any one of claims 1 to 9 and a therapeutically inert carrier. 13. The use of a compound according to any one of claims 1 to 9 for the treatment or prophylaxis of systemic lupus erythematosus or lupus nephritis. 14. The use of a compound according to any one of claims 1 to 9 for the preparation of a medicament for the treatment or prophylaxis of systemic lupus erythematosus or lupus nephritis. 15. The use of a compound according to any one of claims 1 to 9 as the TLR7 or TLR8 or TLR9 antagonist. 16. The use of a compound according to any one of claims 1 to 9 for the preparation of a medicament for TLR7 and TLR8 and TLR9 antagonist. 17. A compound or pharmaceutically acceptable salt according to any one of claims 1 to 9 for the treatment or prophylaxis of systemic lupus erythematosus or lupus nephritis. 18. A compound or pharmaceutically acceptable salt according to any one of claims 1 to 9, when manufactured according to a process of claim 10. 19. A method for the treatment or prophylaxis of systemic lupus erythematosus or lupus nephritis, which method comprises administering a therapeutically effective amount of a compound as defined in any one of claims 1 to 9. |
Step 1 : tert-butyl 4-(4-((((4R,9aS)-2-(8-cyanoquinolin-5-yl)-4-methyloctahydro- 2H- pyrido[1,2-a]pyrazin-8-yl)amino)methyl)phenyl)piperazine-1-c arboxylate (compound 1b) A solution of 5-((4R,9aS)-4-methyl-8-oxooctahydro-2H-pyrido[1,2-a]pyrazin- 2- yl)quinoline-8-carbonitrile (Intermediate C1, 32 mg, 99.9 µmol), tert-butyl 4-(4- (aminomethyl)phenyl)piperazine-1-carboxylate (Compound 1a, CAS: 852180-47-3, Vendor: Accela ChemBio Inc, 29 mg, 99.9 µmol) in MeOH (4 mL) was stirred at room temperature for 1 hour. Then the reaction mixture was cooled at 0°C and sodium cyanotrihydroborate (12 mg, 200 µmol) was added. The reaction mixture was stirred at room temperature for 12 hrs. The mixture was quenched with water (10 mL), extracted with EA twice. The combined organic layer was washed with brine, dried over Na 2 SO 4 , filtered and concentrated to afford crude tert-butyl 4-(4- ((((4R,9aS)-2-(8-cyanoquinolin-5-yl)-4-methyloctahydro-2H-py rido[1,2-a]pyrazin-8- yl)amino)methyl)phenyl)piperazine-1-carboxylate (Compound 1b, 50 mg) which was used for next step without purification. MS: calc’d 596 (MH + ), measured 596 (MH + ). Step 2 : 5-[(4R,8R,9aS)-4-methyl-8-[(4-piperazin-1-ylphenyl)methylami no]- 1,3,4,6,7,8,9,9a-octahydropyrido[1,2-a]pyrazin-2-yl]quinolin e-8-carbonitrile (Example 1) 5-[(4R,8S,9aS)-4-methyl-8-[(4-piperazin-1-ylphenyl)methylami no]-1,3,4,6,7,8,9,9a- octahydropyrido[1,2-a]pyrazin-2-yl]quinoline-8-carbonitrile (Example 17) To a solution of tert-butyl 4-(4-((((4R,9aS)-2-(8-cyanoquinolin-5-yl)-4-methyloctahydro- 2H-pyrido[1,2-a]pyrazin-8-yl)amino)methyl)phenyl)piperazine- 1-carboxylate (Compound 1b, 50 mg, 83.9 µmol) in DCM (4 mL) was added TFA (0.5 mL) at 0 o C. The reaction mixture was stirred at rt for 2 hrs. Then the mixture was concentrated to crude product, which was purified by prep-HPLC to afford Example 1 (3 mg) as a light yellow solid and Example 17 (4 mg) as a light yellow solid. Example 1: The stereochemistry was confirmed by NOESY. MS calc’d 496 (MH + ), measured 496 (MH + ). 1 H NMR (400 MHz, METHANOL-d 4 ) δ 8.99 (dd, J = 1.7, 4.2 Hz, 1H), 8.62 (dd, J = 1.7, 8.6 Hz, 1H), 8.16 (d, J = 8.1 Hz, 1H), 7.65 (dd, J = 4.2, 8.6 Hz, 1H), 7.26 (dd, J = 8.4, 11.9 Hz, 3H), 6.98 (d, J = 8.7 Hz, 2H), 3.78 (s, 2H), 3.46 - 3.37 (m, 2H), 3.19 - 3.11 (m, 4H), 3.03 - 2.97 (m, 4H), 2.89 - 2.51 (m, 6H), 2.15 - 1.92 (m, 3H), 1.58 - 1.45 (m, 1H), 1.33 - 1.23 (m, 1H), 1.19 (d, J = 6.2 Hz, 3H). Example 17: The stereochemistry was confirmed by NOESY. MS calc’d 496 (MH + ), measured 496 (MH + ). 1 H NMR (400 MHz, METHANOL-d 4 ) δ 8.99 (dd, J = 1.7, 4.2 Hz, 1H), 8.62 (dd, J = 1.7, 8.6 Hz, 1H), 8.15 (d, J = 8.1 Hz, 1H), 7.65 (dd, J = 4.2, 8.6 Hz, 1H), 7.29 (d, J = 8.6 Hz, 2H), 7.22 (d, J = 8.1 Hz, 1H), 6.97 (d, J = 8.7 Hz, 2H), 3.81 - 3.70 (m, 2H), 3.44 - 3.38 (m, 1H), 3.29 - 3.23 (m, 1H), 3.18 - 3.09 (m, 5H), 3.04 - 2.94 (m, 6H), 2.89 - 2.71 (m, 3H), 2.50 - 2.41 (m, 1H), 1.96 - 1.69 (m, 3H), 1.59 - 1.50 (m, 1H), 1.19 (d, J = 5.9 Hz, 3H). Example 2 5-[(4R,8R)-4-methyl-8-(6-piperazin-1-yl-3-pyridyl)-1,3,4,6,7 ,8,9,9a-octahydropyrido[1,2- a]pyrazin-2-yl]quinoline-8-carbonitrile
The title compound was prepared according to the following scheme: Step 1 : N'-((4R,E)-2-(8-cyanoquinolin-5-yl)-4-methyloctahydro-8H-pyr ido[1,2- a]pyrazin-8-ylidene)-4-methylbenzenesulfonohydrazide (compound 2c) To a solution of 5-((4R)-4-methyl-8-oxooctahydro-2H-pyrido[1,2-a]pyrazin-2-yl )quinoline-8- carbonitrile (Intermediate C1, 387 mg, 1.21 mmol) in ethanol (8 mL) was added 4- methylbenzenesulfonohydrazide (Compound 2b, 225 mg, 1.21 mmol). The reaction mixture was stirred at 25 °C for 3 hrs. LCMS showed the starting material consumed, the reaction mixture was concentrated to afford crude N'-((4R,E)-2-(8-cyanoquinolin-5-yl)-4- methyloctahydro-8H-pyrido[1,2-a]pyrazin-8-ylidene)-4-methylb enzenesulfonohydrazide (compound 2c, 400 mg, 48.8 % yield) which can be used in next step without purification as a yellow solid. MS calc’d 489 (MH + ); measured 489 (MH + ). Step 2 : 5-((4R)-8-(6-bromopyridin-3-yl)-4-methyloctahydro-2H-pyrido[ 1,2- a]pyrazin-2-yl)quinoline-8-carbonitrile (compound 2e) To a solution of N'-((4R,E)-2-(8-cyanoquinolin-5-yl)-4-methyloctahydro-8H-pyr ido[1,2- a]pyrazin-8-ylidene)-4-methylbenzenesulfonohydrazide (compound 2c, 300 mg, 614 µmol) in 1,4-dioxane (2 mL) was added (6-bromopyridin-3-yl)boronic acid (compound 2d, 198 mg, 982 µmol), and Cs 2 CO 3 (320 mg, 982 µmol). After being stirred at 120 °C for 16 hrs under N2 atmosphere, then the mixture was concentrated to afford crude product, which was purified by silica gel column (PE/EA=3/1) to afford 5-((4R)-8-(6-bromopyridin-3-yl)-4-methyloctahydro- 2H-pyrido[1,2-a]pyrazin-2-yl)quinoline-8-carbonitrile (compound 2e, 50 mg, 17.6 % yield) as yellow oil. MS calc’d 463 (MH + ), measured 463 (MH + ). Step 3 : tert-butyl 4-(5-((4R)-2-(8-cyanoquinolin-5-yl)-4-methyloctahydro-2H- pyrido[1,2-a]pyrazin-8-yl)pyridin-2-yl)piperazine-1-carboxyl ate (compound 2g) To a microwave tube was added 5-((4R)-8-(6-bromopyridin-3-yl)-4-methyloctahydro-2H- pyrido[1,2-a]pyrazin-2-yl)quinoline-8-carbonitrile (compound 2e, 50 mg, 108 µmol), tert-butyl piperazine-1-carboxylate (compound 2f, 26.2 mg, 141 µmol), sodium tert-butoxide (162 µl, 324 µmol) and 1,4-dioxane (5 mL), the suspension was bubbled with N2 for 5 mins and tBuXPhos PD G3 (8.59 mg, 10.8 µmol) was added. After being stirred at 100 °C for 12 hrs, the mixture was cooled to room temperature, quenched with saturated NaHCO 3 (5 mL) solution and extracted with EtOAc (10 mL) three times. The combined organic layer was washed with brine, dried over Na 2 SO 4 , filtered and concentrated. The residue was purified by flash chromatography (silica gel, 0% to 15% DCM in MeOH) to afford tert-butyl 4-(5-((4R)-2-(8-cyanoquinolin-5-yl)-4- methyloctahydro-2H-pyrido[1,2-a]pyrazin-8-yl)pyridin-2-yl)pi perazine-1-carboxylate (compound 2g, 30 mg, 52.8 µmol)as an orange solid. MS: calc’d 568 (MH + ), measured 568 (MH + ). Step 4 : 5-[(4R,8R)-4-methyl-8-(6-piperazin-1-yl-3-pyridyl)-1,3,4,6,7 ,8,9,9a- octahydropyrido[1,2-a]pyrazin-2-yl]quinoline-8-carbonitrile (Example 2) To a solution of tert-butyl 4-(5-((4R)-2-(8-cyanoquinolin-5-yl)-4-methyloctahydro-2H- pyrido[1,2-a]pyrazin-8-yl)pyridin-2-yl)piperazine-1-carboxyl ate (compound 2g, 30 mg, 52.8 µmol) in DCM (4 mL) was added TFA (0.5 mL) at 0 o C. Then the mixture was concentrated to crude product, which was purified by prep-HPLC to afford Example 2 (2 mg) as a light yellow solid. MS calc’d 468 (MH + ), measured 468 (MH + ). 1 H NMR (400 MHz, METHANOL-d 4 ) δ 8.90 (dd, J = 1.7, 4.3 Hz, 1H), 8.56 (dd, J = 1.6, 8.6 Hz, 1H), 8.06 (d, J = 7.9 Hz, 1H), 7.96 (d, J = 2.2 Hz, 1H), 7.56 (dd, J = 4.3, 8.7 Hz, 1H), 7.47 (dd, J = 2.5, 8.7 Hz, 1H), 7.16 (d, J = 8.1 Hz, 1H), 6.76 (d, J = 8.7 Hz, 1H), 3.50 - 3.43 (m, 4H), 3.37 - 3.29 (m, 2H), 3.00 - 2.93 (m, 4H), 2.82 - 2.74 (m, 2H), 2.70 - 2.64 (m, 2H), 2.18 - 2.08 (m, 1H), 1.91 - 1.84 (m, 1H), 1.78 - 1.67 (m, 2H), 1.52 - 1.40 (m, 1H), 1.23 - 1.21 (m, 1H), 1.20 - 1.17 (m, 1H), 1.14 (d, J = 6.0 Hz, 3H). Example 3 5-[(4R,8R)-4-methyl-8-(4-piperazin-1-ylphenyl)-1,3,4,6,7,8,9 ,9a-octahydropyrido[1,2- a]pyrazin-2-yl]quinoline-8-carbonitrile The title compound was prepared in analogy to the preparation of Example 2 by using (4- bromophenyl)boronic acid instead of (6-bromopyridin-3-yl)boronic acid (compound 2d). Example 3 (17 mg) was obtained as a light grey solid. The stereochemistry was confirmed by NOESY. MS: calc’d 467 (MH + ), measured 467 (MH + ). 1 H NMR (400 MHz, METHANOL-d 4 ) δ 9.07 - 9.00 (m, 1H), 8.71 (dd, J = 1.7, 8.6 Hz, 1H), 8.26 - 8.18 (m, 1H), 7.76 - 7.66 (m, 1H), 7.38 (d, J = 8.1 Hz, 1H), 7.29 - 7.22 (m, 2H), 7.08 - 7.00 (m, 2H), 4.26 - 4.05 (m, 1H), 4.03 - 3.80 (m, 2H), 3.79 - 3.64 (m, 2H), 3.63 - 3.49 (m, 1H), 3.48 - 3.34 (m, 8H), 3.24 (br d, J = 11.7 Hz, 4H), 2.31 - 1.98 (m, 3H), 1.93 - 1.84 (m, 1H), 1.84 - 1.78 (m, 1H), 1.54 (d, J = 6.5 Hz, 1H). Example 4 5-[(4R,8R,9aS)-4-methyl-8-(6-piperazin-1-yl-3-pyridyl)-1,3,4 ,6,7,8,9,9a- octahydropyrido[1,2-a]pyrazin-2-yl]quinoline-8-carbonitrile The title compound was prepared in analogy to the preparation of Example 2 by using (6- bromopyridin-3-yl)boronic acid instead of (6-bromopyridin-3-yl)boronic acid (compound 2d). Example 4 (8 mg) was obtained as a brown solid. The stereochemistry was confirmed by NOESY. MS: calc’d 468 (MH + ), measured 468 (MH + ). 1 H NMR (400 MHz, METHANOL-d 4 ) δ 8.91 (dd, J = 1.6, 4.3 Hz, 1H), 8.59 (dd, J = 1.7, 8.6 Hz, 1H), 8.09 (d, J = 7.9 Hz, 1H), 8.00 (d, J = 2.3 Hz, 1H), 7.67 (dd, J = 2.3, 9.0 Hz, 1H), 7.59 (dd, J = 4.3, 8.7 Hz, 1H), 7.27 (d, J = 8.1 Hz, 1H), 7.01 (d, J = 8.9 Hz, 1H), 4.03 - 3.96 (m, 1H), 3.86 - 3.78 (m, 1H), 3.77 - 3.69 (m, 5H), 3.67 - 3.54 (m, 2H), 3.30 - 3.24 (m, 4H), 3.18 - 3.08 (m, 3H), 3.08 - 2.97 (m, 1H), 2.19 - 2.07 (m, 2H), 2.03 - 1.91 (m, 1H), 1.86 - 1.75 (m, 1H), 1.43 (d, J = 6.4 Hz, 3H). Example 6 5-[(4R,8R,9aS)-8-[6-[(3S,4S)-3-amino-4-methoxy-pyrrolidin-1- yl]-3-pyridyl]-4-methyl- 1,3,4,6,7,8,9,9a-octahydropyrido[1,2-a]pyrazin-2-yl]quinolin e-8-carbonitrile
The title compound was prepared in analogy to the preparation of Example 2 by using tert- butyl N-[(3S,4S)-4-methoxypyrrolidin-3-yl]carbamate (CAS: 1627185-88-9, PharmaBlock, Catalog# PBZ4724) instead of tert-butyl piperazine-1-carboxylate (compound 2f). Example 6 (21 mg) was obtained as a light yellow solid. The stereochemistry was confirmed by NOESY. MS: calc’d 498 (MH + ), measured 498 (MH + ). 1 H NMR (400 MHz, METHANOL-d 4 ) δ 9.00 (dd, J = 1.5, 4.3 Hz, 1H), 8.68 (dd, J = 1.6, 8.7 Hz, 1H), 8.18 (d, J = 7.9 Hz, 1H), 8.03 (dd, J = 2.0, 9.3 Hz, 1H), 7.93 (d, J = 2.0 Hz, 1H), 7.68 (dd, J = 4.3, 8.6 Hz, 1H), 7.36 (d, J = 8.1 Hz, 1H), 7.12 (d, J = 9.3 Hz, 1H), 4.30 - 4.23 (m, 1H), 4.14 - 3.99 (m, 4H), 3.92 (br s, 1H), 3.88 - 3.79 (m, 2H), 3.79 - 3.62 (m, 3H), 3.49 (s, 3H), 3.29 - 3.12 (m, 4H), 2.31 - 2.19 (m, 2H), 2.16 - 2.04 (m, 1H), 2.01 - 1.89 (m, 1H), 1.53 (d, J = 6.5 Hz, 3H). Example 7 5-[(4R,8R,9aS)-8-[6-(3-aminoazetidin-1-yl)-3-pyridyl]-4-meth yl-1,3,4,6,7,8,9,9a- octahydropyrido[1,2-a]pyrazin-2-yl]quinoline-8-carbonitrile
The title compound was prepared in analogy to the preparation of Example 2 by using tert- butyl N-[(3S,4S)-4-methoxypyrrolidin-3-yl]carbamate (CAS: 1627185-88-9, PharmaBlock, Catalog# PBZ4724) instead of tert-butyl piperazine-1-carboxylate (compound 2f). Example 7 (14 mg) was obtained as a light yellow solid. The stereochemistry was confirmed by NOESY. MS: calc’d 454 (MH + ), measured 454 (MH + ). 1 H NMR 1 H NMR (400 MHz, METHANOL-d 4 ) δ 9.05 (dd, J = 4.3, 1.6 Hz, 1H), 8.70 (dd, J = 8.6, 1.6 Hz, 1H), 8.23 (d, J = 7.9 Hz, 1H), 7.98 (d, J = 2.0 Hz, 1H), 7.90 (dd, J = 9.0, 2.2 Hz, 1H), 7.72 (dd, J = 8.6, 4.3 Hz, 1H), 7.39 (d, J = 8.1 Hz, 1H), 6.84 (d, J = 9.0 Hz, 1H), 4.58 (dd, J = 9.8, 7.2 Hz, 2H), 4.37 – 4.22 (m, 3H), 4.16 – 4.08 (m, 1H), 3.98 – 3.80 (m, 2H), 3.80 – 3.64 (m, 2H), 3.27 – 3.07 (m, 4H), 2.21 (br s, 2H), 2.13 – 1.99 (m, 1H), 1.90 (br d, J = 14.1 Hz, 1H), 1.54 (d, J = 6.5 Hz, 3H). Example 9 5-[(4R,8R,9aS)-4-methyl-8-[3-(4-piperazin-1-ylphenyl)azetidi n-1-yl]-1,3,4,6,7,8,9,9a- octahydropyrido[1,2-a]pyrazin-2-yl]quinoline-8-carbonitrile
The title compound was prepared according to the following scheme: Step 1 : 5-[(4R,8R,9aS)-8-[3-(4-bromophenyl)azetidin-1-yl]-4-methyl-1 ,3,4,6,7,8,9,9a- octahydropyrido[1,2-a]pyrazin-2-yl]quinoline-8-carbonitrile compound 9b) A solution of 5-((4R,9aS)-4-methyl-8-oxooctahydro-2H-pyrido[1,2-a]pyrazin- 2- yl)quinoline-8-carbonitrile (Intermediate C1, 30 mg, 93 µmol), 3-(4-bromophenyl)azetidine (Compound 9a, 20 mg, 93 µmol) in MeOH (10 mL) was stirred at room temperature for 1 hour. Then the reaction mixture was cooled at 0°C and sodium cyanotrihydroborate (17 mg, 281 µmol) was added. After being stirred at room temperature for 12 hrs, the mixture was quenched with water (10 mL), extracted with EA twice. The combined organic layer was washed with brine, dried over Na 2 SO 4 , filtered and concentrated to afford crude product which was purified by prep- HPLC to afford 5-[(4R,8R,9aS)-8-[3-(4-bromophenyl)azetidin-1-yl]-4-methyl-1 ,3,4,6,7,8,9,9a- octahydropyrido[1,2-a]pyrazin-2-yl]quinoline-8-carbonitrile (compound 9b, 20 mg). MS: calc’d 517 (MH + ), measured 517 (MH+). The stereochemistry was confirmed by NOESY. Step 2 : tert-butyl 4-[4-[1-[(4R,8R,9aS)-2-(8-cyano-5-quinolyl)-4-methyl- 1,3,4,6,7,8,9,9a-octahydropyrido[1,2-a]pyrazin-8-yl]azetidin -3-yl]phenyl]piperazine-1- carboxylate(compound 9d) A mixture of 5-[(4R,8R,9aS)-8-[3-(4-bromophenyl)azetidin-1-yl]-4-methyl- 1,3,4,6,7,8,9,9a-octahydropyrido[1,2-a]pyrazin-2-yl]quinolin e-8-carbonitrile.(compound 9b, 20 mg, 38.7 µmol), tert-butyl piperazine-1-carboxylate (compound 9c, 9.38 mg, 50.3 µmol), tBuXPhos Pd G3 (1.54 mg, 1.94 µmol) and sodium tert-butoxide (18.6 mg, 194 µmol) in 1,4- dioxane (10 mL) was charged with N2. After being stirred at 90 o C overnight, the mixture was cooled to room temperature, the solid was filtered off and washed with EA (10 mL) twice. The filtrate was concentrated to crude product tert-butyl 4-[4-[1-[(4R,8R,9aS)-2-(8-cyano-5- quinolyl)-4-methyl-1,3,4,6,7,8,9,9a-octahydropyrido[1,2-a]py razin-8-yl]azetidin-3- yl]phenyl]piperazine-1-carboxylate(compound 9d, 25 mg)which can be used in next step without purification. MS: calc’d 622 (MH + ), measured 622 (MH + ). Step 3 : 5-[(4R,8R,9aS)-4-methyl-8-[3-(4-piperazin-1-ylphenyl)azetidi n-1-yl]- 1,3,4,6,7,8,9,9a-octahydropyrido[1,2-a]pyrazin-2-yl]quinolin e-8-carbonitrile (Example 9) To a solution of tert-butyl 4-[4-[1-[(4R,8R,9aS)-2-(8-cyano-5-quinolyl)-4-methyl- 1,3,4,6,7,8,9,9a-octahydropyrido[1,2-a]pyrazin-8-yl]azetidin -3-yl]phenyl]piperazine-1- carboxylate (compound 9d, 25 mg) in DCM (4 mL) was added TFA (0.5 mL) at 0 o C. The reaction mixture was stirred at rt for 2 hrs. Then the mixture was concentrated to crude product, which was purified by prep-HPLC to afford Example 9 (11 mg) as a light yellow solid. MS: calc’d 522 (MH + ), measured 522 (MH + ). 1 H NMR (400 MHz, METHANOL-d 4 ) δ 8.92 (dd, J = 4.3, 1.6 Hz, 1H), 8.55 (dd, J = 8.6, 1.7 Hz, 1H), 8.09 (d, J = 7.9 Hz, 1H), 7.58 (dd, J = 8.6, 4.3 Hz, 1H), 7.25 (t, J = 8.4 Hz, 3H), 6.92-7.04 (m, 2H), 4.35-4.51 (m, 2H), 4.11-4.30 (m, 2H), 3.80- 4.07 (m, 2H), 3.38-3.71 (m, 5H), 3.26-3.37 (m, 6H), 2.90-3.12 (m, 2H), 2.69-2.85 (m, 1H), 2.18- 2.42 (m, 2H), 1.47-1.81 (m, 2H), 1.32 ppm (d, J = 6.4 Hz, 3H). Example 11 5-[(4R,8S,9aS)-4-methyl-8-[(5-piperazin-1-yl-2-pyridyl)oxy]- 1,3,4,6,7,8,9,9a- octahydropyrido[1,2-a]pyrazin-2-yl]quinoline-8-carbonitrile The title compound was prepared according to the following scheme:
Step 1 : 5-((4R,8R)-8-hydroxy-4-methyloctahydro-2H-pyrido[1,2-a]pyraz in-2- yl)quinoline-8-carbonitrile (compound 11a) To a solution of 5-((4R)-4-methyl-8-oxooctahydro-2H-pyrido[1,2-a]pyrazin-2- yl)quinoline-8-carbonitrile (Intermediate C1, 500 mg, 1.56 mmol) in MeOH (10 mL) was added sodium tetrahydroborate (70.8 mg, 1.87 mmol). After being stirred at room temperature for 2 hours, the reaction was quenched carefully with 10% HCl solution. The resulting mixture was neutralized with K 2 CO 3 and extracted with EtOAc (30 mL) twice. The combined organic layer was washed with brine and dried over Na 2 SO 4 , filtered and concentrated to afford 5-((4R,8R)-8- hydroxy-4-methyloctahydro-2H-pyrido[1,2-a]pyrazin-2-yl)quino line-8-carbonitrile (compound 11a, 500 mg) which was used directly for the next step without further purification. The stereochemistry was confirmed by NOESY. MS: calc’d 323 (MH + ), measured 323 (MH + ).1H NMR (DMSO-d6, 400 MHz) δ 9.03 (dd, 1H, J=1.6, 4.2 Hz), 8.51 (dd, 1H, J=1.5, 8.6 Hz), 8.22 (d, 1H, J=8.1 Hz), 7.67 (dd, 1H, J=4.3, 8.6 Hz), 7.18 (d, 1H, J=8.2 Hz), 4.69 (d, 1H, J=4.6 Hz), 3.48 (dt, 1H, J=5.6, 10.4 Hz), 3.2-3.3 (m, 1H), 3.20 (br d, 1H, J=11.4 Hz), 2.6-2.8 (m, 2H), 2.3- 2.4 (m, 1H), 1.7-1.9 (m, 3H), 1.3-1.5 (m, 1H), 1.0-1.1 (m, 1H), 1.0-1.1 (m, 3H). Step 2 : 5-((4R,8S,9aS)-8-((5-chloropyridin-2-yl)oxy)-4-methyloctahyd ro-2H- pyrido[1,2-a]pyrazin-2-yl)quinoline-8-carbonitrile (compound 11c) A solution of 5-((4R,9aS)-8-hydroxy-4-methyloctahydro-2H-pyrido[1,2-a]pyra zin-2- yl)quinoline-8-carbonitrile (compound 11a, 60 mg, 186 µmol), 5-chloropyridin-2-ol (compound 11b, 24 mg, 186 µmol) and PPh3 (98 mg, 372 µmol) in THF (4 mL) at 0 °C was treated with DIAD (72 µL, 372 µmol), stirred at room temperature for 1 hour. After being stirred at 70 °C for 1 hour, the mixture was cooled to room temperature, diluted with EtOAc, washed with water and brine, dried over Na2SO4 and concentrated. The residue was purified by silica gel column chromatography (20% to 100% EtOAc/PE) to afford 5-((4R,8S,9aS)-8-((5- chloropyridin-2-yl)oxy)-4-methyloctahydro-2H-pyrido[1,2-a]py razin-2-yl)quinoline-8- carbonitrile ((compound 11c, 40 mg) as a pale yellow solid. MS: calc’d 434 (MH + ), measured 434 (MH + ). The stereochemistry was confirmed by NOESY. Step 3 : tert-butyl 4-(6-(((4R,8S,9aS)-2-(8-cyanoquinolin-5-yl)-4-methyloctahydr o-2H- pyrido[1,2-a]pyrazin-8-yl)oxy)pyridin-3-yl)piperazine-1-carb oxylate (compound 11e) To a microwave tube was added 5-((4R,8S,9aS)-8-((5-chloropyridin-2-yl)oxy)-4- methyloctahydro-2H-pyrido[1,2-a]pyrazin-2-yl)quinoline-8-car bonitrile (compound 11c, 40 mg, 92.2 µmol), tert-butyl piperazine-1-carboxylate (compound 11d, 22 mg, 120 µmol), sodium tert-butoxide (138 µL, 277 µmol, 2M in THF) and 1,4-dioxane (5 mL), the suspension was bubbled with N2 for 5 mins and tBuXPhos PD G3 (7 mg, 9.22 µmol) was added. After being stirred at 100 °C for 12 hrs, the mixture was cooled to room temperature, diluted with saturated NaHCO 3 (5 mL) solution and extracted with EtOAC (10 mL) three times. The combined organic layer was washed with brine, dried over Na2SO4, and concentrated to afford tert-butyl 4-(6- (((4R,8S,9aS)-2-(8-cyanoquinolin-5-yl)-4-methyloctahydro-2H- pyrido[1,2-a]pyrazin-8- yl)oxy)pyridin-3-yl)piperazine-1-carboxylate (compound 11e, 20 mg) as an orange solid which can be used in next step without purification . MS: calc’d 584(MH + ), measured 584 (MH + ). Step 4 : 5-[(4R,8S,9aS)-4-methyl-8-[(5-piperazin-1-yl-2-pyridyl)oxy]- 1,3,4,6,7,8,9,9a- octahydropyrido[1,2-a]pyrazin-2-yl]quinoline-8-carbonitrile (Example 11) To a solution of tert-butyl 4-(6-(((4R,8S,9aS)-2-(8-cyanoquinolin-5-yl)-4- methyloctahydro-2H-pyrido[1,2-a]pyrazin-8-yl)oxy)pyridin-3-y l)piperazine-1-carboxylate (compound 11e, 20 mg) in DCM (2 mL) was added TFA (0.5 mL) at 0 o C. The reaction mixture was stirred at rt for 2 hrs. Then the mixture was concentrated to crude product, which was purified by prep-HPLC to afford Example 11 (16 mg) as a light brown solid. MS calc’d 484 (MH + ), measured 484 (MH + ). 1 H NMR (400 MHz, METHANOL-d 4 ) δ 9.02 (dd, J = 4.2, 1.7 Hz, 1H), 8.70 (dd, J = 8.6, 1.7 Hz, 1H), 8.20 (d, J = 8.1 Hz, 1H), 7.89 (d, J = 2.9 Hz, 1H), 7.69 (dd, J = 8.7, 4.3 Hz, 1H), 7.57 (dd, J = 9.0, 3.1 Hz, 1H), 7.38 (d, J = 8.1 Hz, 1H), 6.93 (d, J = 8.9 Hz, 1H), 5.45 (br s, 1H), 4.17 – 4.09 (m, 1H), 4.00 – 3.87 (m, 2H), 3.78 – 3.71 (m, 1H), 3.66 – 3.59 (m, 1H), 3.45 – 3.34 (m, 9H), 3.21 (td, J = 14.0, 11.1 Hz, 2H), 2.48 – 2.36 (m, 2H), 2.27 – 2.16 (m, 1H), 2.12 – 2.01 (m, 1H), 1.52 (d, J = 6.5 Hz, 3H). Example 12 5-[(4R,8R,9aS)-8-[6-[(3R,4S)-3-amino-4-methoxy-pyrrolidin-1- yl]-3-pyridyl]-4-methyl- 1,3,4,6,7,8,9,9a-octahydropyrido[1,2-a]pyrazin-2-yl]quinolin e-8-carbonitrile The title compound was prepared in analogy to the preparation of Example 2 by using tert- butyl ((3R,4S)-4-methoxypyrrolidin-3-yl)carbamate (CAS: 1932508-77-4, PharmaBlock) instead of tert-butyl piperazine-1-carboxylate (compound 2f). Example 12 (9 mg) was obtained as a light yellow solid. The stereochemistry was confirmed by NOESY. MS: calc’d 498 (MH + ), measured 498 (MH + ). 1 H NMR (400 MHz, METHANOL-d 4 ) δ 9.02 (dd, J = 4.3, 1.6 Hz, 1H), 8.70 (dd, J = 8.6, 1.6 Hz, 1H), 8.20 (d, J = 8.1 Hz, 1H), 8.06 (dd, J = 9.4, 2.0 Hz, 1H), 7.99 – 7.94 (m, 1H), 7.70 (dd, J = 8.6, 4.3 Hz, 1H), 7.38 (d, J = 8.1 Hz, 1H), 7.14 (d, J = 9.4 Hz, 1H), 4.36 (dt, J = 4.5, 2.3 Hz, 1H), 4.22 – 4.04 (m, 3H), 3.99 – 3.65 (m, 7H), 3.53 (s, 3H), 3.31 – 3.14 (m, 4H), 2.34 – 2.07 (m, 3H), 2.05 – 1.92 (m, 1H), 1.55 (d, J = 6.5 Hz, 3H). Example 13 5-[(4R,8R,9aS)-8-[6-[(3S,4R)-3-amino-4-methoxy-pyrrolidin-1- yl]-3-pyridyl]-4-methyl- 1,3,4,6,7,8,9,9a-octahydropyrido[1,2-a]pyrazin-2-yl]quinolin e-8-carbonitrile The title compound was prepared in analogy to the preparation of Example 2 by using tert- butyl ((3S,4R)-4-methoxypyrrolidin-3-yl)carbamate (CAS: 1931911-57-7, PharmaBlock, Catalog# PBZ4730) instead of tert-butyl piperazine-1-carboxylate (compound 2f). Example 13 (15 mg) was obtained as a light yellow solid. The stereochemistry was confirmed by NOESY. MS: calc’d 498 (MH + ), measured 498 (MH + ). 1 H NMR (400 MHz, METHANOL-d 4 ) δ 9.02 (dd, J = 4.2, 1.5 Hz, 1H), 8.70 (dd, J = 8.6, 1.6 Hz, 1H), 8.20 (d, J = 7.9 Hz, 1H), 8.06 (dd, J = 9.4, 2.0 Hz, 1H), 7.99 – 7.93 (m, 1H), 7.70 (dd, J = 8.6, 4.3 Hz, 1H), 7.38 (d, J = 7.9 Hz, 1H), 7.14 (d, J = 9.3 Hz, 1H), 4.36 (td, J = 4.5, 2.4 Hz, 1H), 4.21 – 4.05 (m, 3H), 3.99 – 3.90 (m, 2H), 3.89 – 3.79 (m, 2H), 3.78 – 3.65 (m, 3H), 3.53 (s, 3H), 3.31 – 3.14 (m, 4H), 2.34 – 2.07 (m, 3H), 2.04 – 1.92 (m, 1H), 1.55 (d, J = 6.4 Hz, 3H). Example 14 5-[(4R,8R,9aS)-8-[6-[(3R,4R)-3-amino-4-methoxy-pyrrolidin-1- yl]-3-pyridyl]-4-methyl- 1,3,4,6,7,8,9,9a-octahydropyrido[1,2-a]pyrazin-2-yl]quinolin e-8-carbonitrile
The title compound was prepared in analogy to the preparation of Example 2 by using tert- butyl ((3R,4R)-4-methoxypyrrolidin-3-yl)carbamate (CAS: 1932066-52-8, PharmaBlock, Catalog# PBZ4728) instead of tert-butyl piperazine-1-carboxylate (compound 2f). Example 13 (10 mg) was obtained as a light yellow solid. The stereochemistry was confirmed by NOESY. MS: calc’d 498 (MH + ), measured 498 (MH + ). 1 H NMR (400 MHz, METHANOL-d 4 ) δ 9.04 (dd, J = 4.2, 1.5 Hz, 1H), 8.71 (dd, J = 8.6, 1.7 Hz, 1H), 8.22 (d, J = 7.9 Hz, 1H), 8.01 – 7.95 (m, 2H), 7.71 (dd, J = 8.6, 4.2 Hz, 1H), 7.39 (d, J = 7.9 Hz, 1H), 7.06 (d, J = 10.0 Hz, 1H), 4.26 (dt, J = 5.5, 2.8 Hz, 1H), 4.16 – 4.09 (m, 1H), 4.09 – 3.99 (m, 3H), 3.98 – 3.82 (m, 2H), 3.82 – 3.66 (m, 4H), 3.51 (s, 3H), 3.30 – 3.13 (m, 4H), 2.34 – 2.20 (m, 2H), 2.16 – 2.03 (m, 1H), 2.00 – 1.88 (m, 1H), 1.55 (d, J = 6.4 Hz, 3H). Example 15 5-[(4R,8R,9aR)-4-methyl-8-(6-piperazin-1-yl-3-pyridyl)-1,3,4 ,6,7,8,9,9a- octahydropyrido[1,2-a]pyrazin-2-yl]quinoline-8-carbonitrile
The title compound was prepared in analogy to the preparation of Example 2 by using 5- ((4R,8R,9aR)-8-(6-bromopyridin-3-yl)-4-methyloctahydro-2H-py rido[1,2-a]pyrazin-2- yl)quinoline-8-carbonitrile instead of 5-((4R)-8-(6-bromopyridin-3-yl)-4-methyloctahydro-2H- pyrido[1,2-a]pyrazin-2-yl)quinoline-8-carbonitrile (compound 2e). Example 15 (4 mg) was obtained as a light yellow solid. The stereochemistry was confirmed by NOESY. MS: calc’d 468 (MH + ), measured 468 (MH + ). 1 H NMR (400 MHz, METHANOL-d 4 ) δ 8.99 (dd, J = 4.2, 1.7 Hz, 1H), 8.73 (dd, J = 8.6, 1.7 Hz, 1H), 8.16 (d, J = 8.1 Hz, 1H), 8.03 (d, J = 2.3 Hz, 1H), 7.66 (dd, J = 8.6, 4.3 Hz, 1H), 7.54 (dd, J = 8.7, 2.4 Hz, 1H), 7.27 (d, J = 8.1 Hz, 1H), 6.83 (d, J = 8.7 Hz, 1H), 3.52 – 3.35 (m, 8H), 3.31 – 3.26 (m, 1H), 3.25 – 3.14 (m, 1H), 2.98 – 2.82 (m, 6H), 2.80 – 2.65 (m, 2H), 1.93 – 1.77 (m, 2H), 1.65 – 1.49 (m, 1H), 1.48 – 1.43 (m, 3H). Example 16 5-[(4R,8S,9aS)-4-methyl-8-[(6-piperazin-1-yl-3-pyridyl)oxy]- 1,3,4,6,7,8,9,9a- octahydropyrido[1,2-a]pyrazin-2-yl]quinoline-8-carbonitrile The title compound was prepared in analogy to the preparation of Example 11 by using 5- bromo-2-hydroxypyridine (PharmaBlock, CAS: 13466-38-1) instead of 5-chloropyridin-2-ol (compound 11b). Example 16 (50 mg) was obtained as a white solid. The stereochemistry was confirmed by NOESY. MS: calc’d 484 (MH + ), measured 484 (MH + ). 1 H NMR (400 MHz, METHANOL-d 4 ) δ 8.94 (dd, J = 4.2, 1.6 Hz, 1H), 8.57 (dd, J = 8.6, 1.6 Hz, 1H), 8.11 (d, J = 7.9 Hz, 1H), 7.76 (d, J = 2.8 Hz, 1H), 7.61 (dd, J = 8.6, 4.2 Hz, 1H), 7.45 (dd, J = 8.9, 3.1 Hz, 1H), 7.20 (d, J = 7.9 Hz, 1H), 6.79 (d, J = 8.9 Hz, 1H), 5.21 (t, J = 2.4 Hz, 1H), 3.40 (br d, J = 11.2 Hz, 1H), 3.26 (br d, J = 11.7 Hz, 1H), 3.21 – 3.15 (m, 1H), 3.07 – 3.01 (m, 4H), 3.01 – 2.94 (m, 5H), 2.85 – 2.72 (m, 3H), 2.49 – 2.36 (m, 1H), 2.14 – 2.06 (m, 1H), 2.01 – 1.87 (m, 2H), 1.70 – 1.61 (m, 1H), 1.18 (d, J = 5.9 Hz, 3H). Example 18 5-[(4R,8S,9aS)-4-methyl-8-(2-piperazin-1-ylpyrimidin-5-yl)ox y-1,3,4,6,7,8,9,9a- octahydropyrido[1,2-a]pyrazin-2-yl]quinoline-8-carbonitrile The title compound was prepared in analogy to the preparation of Example 11 by using 2- Chloro-5-hydroxypyrimidine (Accela ChemBio Inc, CAS: 4983-28-2) instead of 5- chloropyridin-2-ol (compound 11b). Example 18 (27 mg) was obtained as a light yellow solid. The stereochemistry was confirmed by NOESY. MS: calc’d 485 (MH + ), measured 485 (MH + ). 1 H NMR (400 MHz, METHANOL-d 4 ) δ 9.04 (dd, J = 4.2, 1.7 Hz, 1H), 8.71 (dd, J = 8.6, 1.7 Hz, 1H), 8.37 (s, 2H), 8.22 (d, J = 7.9 Hz, 1H), 7.71 (dd, J = 8.6, 4.3 Hz, 1H), 7.38 (d, J = 7.9 Hz, 1H), 4.79 (br s, 1H), 4.21 – 4.12 (m, 1H), 4.06 – 4.00 (m, 4H), 3.99 – 3.87 (m, 2H), 3.79 – 3.72 (m, 1H), 3.70 – 3.62 (m, 1H), 3.50 – 3.40 (m, 1H), 3.32 – 3.28 (m, 4H), 3.21 (td, J = 13.7, 11.2 Hz, 2H), 2.42 – 2.35 (m, 2H), 2.25 – 2.13 (m, 1H), 2.10 – 2.00 (m, 1H), 1.52 (d, J = 6.5 Hz, 3H). Example 19 5-[(4R,8R,9aS)-8-[6-[(3R,4S)-3-amino-4-fluoro-pyrrolidin-1-y l]-3-pyridyl]-4-methyl- 1,3,4,6,7,8,9,9a-octahydropyrido[1,2-a]pyrazin-2-yl]quinolin e-8-carbonitrile The title compound was prepared in analogy to the preparation of Example 2 by using tert- butyl N-[(3R,4S)-4-fluoropyrrolidin-3-yl]carbamate (CAS: 1033718-91-0, PharmaBlock, Catalog# PB09204) instead of tert-butyl piperazine-1-carboxylate (compound 2f). Example 19 (5 mg) was obtained as a light yellow solid. The stereochemistry was confirmed by NOESY. MS: calc’d 486 (MH + ), measured 486 (MH + ). 1 H NMR (400 MHz, METHANOL-d 4 ) δ 9.04 (dd, J = 4.3, 1.6 Hz, 1H), 8.71 (dd, J = 8.6, 1.6 Hz, 1H), 8.22 (d, J = 7.9 Hz, 1H), 8.05 (dd, J = 9.2, 2.3 Hz, 1H), 8.00 (d, J = 1.8 Hz, 1H), 7.71 (dd, J = 8.6, 4.3 Hz, 1H), 7.39 (d, J = 7.9 Hz, 1H), 7.12 (d, J = 9.3 Hz, 1H), 5.69 – 5.51 (m, 1H), 4.34 – 4.20 (m, 2H), 4.17 – 4.03 (m, 3H), 4.00 – 3.80 (m, 3H), 3.79 – 3.65 (m, 3H), 3.31 – 3.14 (m, 4H), 2.34 – 2.21 (m, 2H), 2.19 – 2.06 (m, 1H), 2.03 – 1.91 (m, 1H), 1.55 (d, J = 6.4 Hz, 3H). Example 20 5-[(4R,8R,9aS)-4-methyl-8-[4-[(2S)-morpholin-2-yl]anilino]-1 ,3,4,6,7,8,9,9a- octahydropyrido[1,2-a]pyrazin-2-yl]quinoline-8-carbonitrile
The title compound was prepared in analogy to the preparation of Example 1 by using tert- butyl (2S)-2-(4-aminophenyl)morpholine-4-carboxylate instead of tert-butyl 4-(4- (aminomethyl)phenyl)piperazine-1-carboxylate (Compound 1a) in step 1. Example 20 (7 mg) was obtained as a light yellow solid. The stereochemistry was confirmed by NOESY. MS: calc’d 483 (MH + ), measured 483 (MH + ). 1 H NMR (400 MHz, METHANOL-d 4 ) δ 9.04 (dd, J = 4.3, 1.6 Hz, 1H), 8.71 (dd, J = 8.6, 1.6 Hz, 1H), 8.22 (d, J = 7.9 Hz, 1H), 7.72 (dd, J = 8.6, 4.3 Hz, 1H), 7.38 (d, J = 8.1 Hz, 1H), 7.20 (d, J = 8.6 Hz, 2H), 6.75 (d, J = 8.6 Hz, 2H), 4.60 (dd, J = 11.2, 2.1 Hz, 1H), 4.21 (dd, J = 12.9, 3.1 Hz, 1H), 3.60-4.12 (m, 7H), 3.05-3.25 (m, 8H), 2.44 (br t, J = 14.7 Hz, 2H), 1.68-1.90 (m, 1H), 1.45 (s, 3H). Example 21 5-[(4R,8R,9aS)-4-methyl-8-[4-[[(2R)-morpholin-2-yl]methyl]an ilino]-1,3,4,6,7,8,9,9a- octahydropyrido[1,2-a]pyrazin-2-yl]quinoline-8-carbonitrile The title compound was prepared in analogy to the preparation of Example 1 by using tert- butyl (2R)-2-[(4-aminophenyl)methyl]morpholine-4-carboxylate instead of tert-butyl 4-(4- (aminomethyl)phenyl)piperazine-1-carboxylate (Compound 1a) in step 1. Example 21 (8 mg) was obtained as a light yellow solid. The stereochemistry was confirmed by NOESY. MS: calc’d 497 (MH + ), measured 497 (MH + ). 1 H NMR (400 MHz, METHANOL-d 4 ) δ 9.04 (dd, J = 4.2, 1.7 Hz, 1H), 8.71 (dd, J = 8.6, 1.7 Hz, 1H), 8.22 (d, J = 7.9 Hz, 1H), 7.72 (dd, J = 8.6, 4.3 Hz, 1H), 7.38 (d, J = 8.1 Hz, 1H), 7.06 (d, J = 8.6 Hz, 2H), 6.73 (d, J = 8.4 Hz, 2H), 4.07 (br dd, J = 13.0, 3.5 Hz, 2H), 3.63-3.99 (m, 7H), 3.03-3.27 (m, 6H), 2.61-2.92 (m, 3H), 2.32-2.54 (m, 2H), 1.68- 1.88 (m, 1H), 1.56-1.60 (m, 1H), 1.40 (s, 3H). Example 22 5-[(4R,8R,9aS)-4-methyl-8-[4-[[(2R)-morpholin-2-yl]methyl]an ilino]-1,3,4,6,7,8,9,9a- octahydropyrido[1,2-a]pyrazin-2-yl]-2-deuterio-quinoline-8-c arbonitrile The title compound was prepared in analogy to the preparation of Example 1 by using tert- butyl (R)-2-(4-aminobenzyl)morpholine-4-carboxylate and 2-deuterio-5-[(4R)-4-methyl-8-oxo- 3,4,6,7,9,9a-hexahydro-1H-pyrido[1,2-a]pyrazin-2-yl]quinolin e-8-carbonitrile (Intermediate D1) instead of tert-butyl 4-(4(aminomethyl)phenyl)piperazine-1-carboxylate (Compound 1a) and 5- ((4R,9aS)-4-methyl-8-oxooctahydro-2H-pyrido[1,2-a]pyrazin-2- yl)quinoline-8-carbonitrile (Intermediate C1). Example 22 (39 mg) was obtained as a light yellow solid. The stereochemistry was confirmed by NOESY. MS: calc’d 498 (MH + ), measured 498 (MH + ). 1 H NMR (400 MHz, METHANOL-d 4 ) δ 8.67 (d, J = 8.7 Hz, 1H), 8.17 (d, J = 8.1 Hz, 1H), 7.68 (d, J = 8.6 Hz, 1H), 7.31 (d, J = 7.9 Hz, 1H), 7.04 (d, J = 8.4 Hz, 2H), 6.69 (d, J = 8.6 Hz, 2H), 4.06 (dd, J = 12.8, 3.5 Hz, 1H), 3.88 – 3.38 (m, 9H), 3.28 – 3.19 (m, 2H), 3.16 – 3.01 (m, 3H), 2.87 (dd, J = 12.7, 11.1 Hz, 1H), 2.82 – 2.75 (m, 1H), 2.71 – 2.63 (m, 1H), 2.39 – 2.24 (m, 2H), 1.75 – 1.62 (m, 1H), 1.51 – 1.43 (m, 1H), 1.40 (d, J = 6.4 Hz, 3H). Example 23 5-[(4R,8R,9aS)-8-[6-[(6S)-6-amino-1,4-oxazepan-4-yl]-3-pyrid yl]-4-methyl-1,3,4,6,7,8,9,9a- octahydropyrido[1,2-a]pyrazin-2-yl]quinoline-8-carbonitrile The title compound was prepared in analogy to the preparation of Example 2 by using tert- butyl N-[(6S)-1,4-oxazepan-6-yl]carbamate (PharmaBlock, Catalog# PB97931) instead of tert- butyl piperazine-1-carboxylate (compound 2f). Example 23 (1 mg) was obtained as a light yellow solid. The stereochemistry was confirmed by NOESY. MS: calc’d 498 (MH + ), measured 498 (MH + ). 1 H NMR (400 MHz, METHANOL-d 4 ) δ 8.92 (dd, J = 4.2, 1.5 Hz, 1H), 8.59 (dd, J = 8.6, 1.6 Hz, 1H), 8.10 (d, J = 7.9 Hz, 1H), 7.98 (d, J = 2.3 Hz, 1H), 7.60 (dd, J = 8.6, 4.3 Hz, 1H), 7.51 (dd, J = 8.9, 2.5 Hz, 1H), 7.27 (d, J = 7.9 Hz, 1H), 6.79 (d, J = 8.8 Hz, 1H), 4.17 – 4.08 (m, 1H), 4.02 – 3.88 (m, 3H), 3.85 – 3.52 (m, 10H), 3.17 – 3.07 (m, 2H), 3.07 – 2.90 (m, 2H), 2.18 – 2.05 (m, 2H), 2.02 – 1.91 (m, 1H), 1.83 – 1.71 (m, 1H), 1.42 (d, J = 6.4 Hz, 3H). Example 24 & 27 5-[(4R,8S,9aS)-4-methyl-8-(4-morpholin-2-ylphenyl)-1,3,4,6,7 ,8,9,9a-octahydropyrido[1,2- a]pyrazin-2-yl]quinoline-8-carbonitrile (Example 24)
5-[(4R,8R,9aS)-4-methyl-8-[4-[(2R)-morpholin-2-yl]phenyl]-1, 3,4,6,7,8,9,9a- octahydropyrido[1,2-a]pyrazin-2-yl]quinoline-8-carbonitrile (Example 27) The title compound was prepared in analogy to the preparation of Example 2 by using tert- butyl (R)-2-(4-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)phenyl )morpholine-4-carboxylate instead of 6-bromopyridin-3-yl)boronic acid (compound 2d). Example 24 (11 mg) was obtained as a light yellow solid and Example 27 (17 mg) was obtained as a light yellow solid. Example 24 The stereochemistry was confirmed by NOESY. MS: calc’d 468 (MH + ), measured 468 (MH + ). 1 H NMR (400 MHz, METHANOL-d 4 ) δ 9.03 – 8.98 (m, 1H), 8.70 (dd, J = 8.6, 1.5 Hz, 1H), 8.59 (dd, J = 8.6, 1.4 Hz, 1H), 8.73 – 8.56 (m, 1H), 8.23 – 8.20 (m, 1H), 8.18 (br d, J = 7.9 Hz, 1H), 8.24 – 8.16 (m, 1H), 7.71 – 7.63 (m, 1H), 7.58 – 7.46 (m, 2H), 7.45 – 7.33 (m, 2H), 4.79 (ddd, J = 16.6, 11.3, 2.0 Hz, 1H), 4.26 (dt, J = 12.6, 4.2 Hz, 1H), 4.44 – 4.20 (m, 1H), 4.09 – 3.63 (m, 5H), 3.60 – 3.41 (m, 3H), 3.18 – 3.01 (m, 2H), 2.74 – 2.60 (m, 1H), 2.51 – 2.01 (m, 3H), 1.52 – 1.43 (m, 3H). Example 27 The stereochemistry was confirmed by NOESY. MS: calc’d 468 (MH + ), measured 468 (MH + ). 1 H NMR (400 MHz, METHANOL-d 4 ) δ 9.05 – 9.01 (m, 1H), 8.71 (dd, J = 8.6, 1.5 Hz, 1H), 8.21 (d, J = 7.9 Hz, 1H), 7.71 (dd, J = 8.6, 4.3 Hz, 1H), 7.45 – 7.36 (m, 5H), 4.77 (dd, J = 11.2, 2.3 Hz, 1H), 4.26 (dd, J = 13.0, 3.3 Hz, 1H), 4.15 – 4.08 (m, 1H), 4.05 – 3.81 (m, 3H), 3.80 – 3.65 (m, 2H), 3.46 (br d, J = 13.0 Hz, 1H), 3.40 – 3.35 (m, 1H), 3.31 – 3.05 (m, 6H), 2.32 – 2.19 (m, 2H), 2.16 – 2.02 (m, 1H), 1.93 (q, J = 13.1 Hz, 1H), 1.54 (d, J = 6.4 Hz, 3H). Example 25 & 26 5-[(4R,8R,9aS)-4-methyl-8-[4-[[(2R)-morpholin-2-yl]methyl]ph enyl]-1,3,4,6,7,8,9,9a- octahydropyrido[1,2-a]pyrazin-2-yl]quinoline-8-carbonitrile (Example 25) 5-[(4R,8S,9aS)-4-methyl-8-[4-(morpholin-2-ylmethyl)phenyl]-1 ,3,4,6,7,8,9,9a- octahydropyrido[1,2-a]pyrazin-2-yl]quinoline-8-carbonitrile (Example 26) The title compound was prepared in analogy to the preparation of Example 2 by using tert- butyl (R)-2-(4-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)phenyl )morpholine-4-carboxylate instead of 6-bromopyridin-3-yl)boronic acid (compound 2d). Example 25 (15 mg) was obtained as a light yellow solid and Example 26 (8 mg) was obtained as a light yellow solid. Example 25: The stereochemistry was confirmed by NOESY. MS: calc’d 482 (MH + ), measured 482 (MH + ). 1 H NMR (400 MHz, METHANOL-d 4 ) δ 9.02 (dd, J = 4.3, 1.6 Hz, 1H), 8.71 (dd, J = 8.6, 1.7 Hz, 1H), 8.20 (d, J = 8.1 Hz, 1H), 7.70 (dd, J = 8.6, 4.3 Hz, 1H), 7.38 (d, J = 8.1 Hz, 1H), 7.27 (s, 4H), 4.14 – 4.02 (m, 2H), 3.98 – 3.83 (m, 3H), 3.83 – 3.64 (m, 3H), 3.29 – 3.08 (m, 7H), 2.97 – 2.77 (m, 3H), 2.31 – 2.17 (m, 2H), 2.15 – 2.01 (m, 1H), 1.98 – 1.86 (m, 1H), 1.54 (d, J = 6.5 Hz, 3H). Example 26: The stereochemistry was confirmed by NOESY. MS: calc’d 482 (MH + ), measured 482 (MH + ). 1 H NMR (400 MHz, METHANOL-d 4 ) δ 9.04 – 8.99 (m, 1H), 8.73 – 8.58 (m, 1H), 8.21 (dd, J = 10.1, 8.0 Hz, 1H), 7.68 (ddd, J = 13.0, 8.6, 4.3 Hz, 1H), 7.52 – 7.31 (m, 3H), 7.25 (s, 2H), 4.42 – 4.20 (m, 1H), 4.10 – 3.65 (m, 7H), 3.60 – 3.41 (m, 2H), 3.30 – 3.22 (m, 2H), 3.18 – 3.04 (m, 3H), 2.98 – 2.81 (m, 3H), 2.74 – 2.59 (m, 1H), 2.48 – 1.98 (m, 3H), 1.47 (dd, J = 14.2, 6.7 Hz, 3H). Example 28 5-[(4R,8S,9aS)-8-[[6-[(3R,4S)-3-amino-4-methoxy-pyrrolidin-1 -yl]-3-pyridyl]oxy]-4-methyl- 1,3,4,6,7,8,9,9a-octahydropyrido[1,2-a]pyrazin-2-yl]quinolin e-8-carbonitrile The title compound was prepared in analogy to the preparation of Example 11 by using 2- Chloro-5-hydroxypyridine (CAS: 41288-96-4) and tert-butyl ((3R,4S)-4-methoxypyrrolidin-3- yl)carbamate (CAS: 1932508-77-4, PharmaBlock) instead of 5-chloropyridin-2-ol (compound 11b) and tert-butyl piperazine-1-carboxylate (compound 11d). Example 28 (6 mg) was obtained as a light yellow solid. The stereochemistry was confirmed by NOESY. MS: calc’d 514 (MH + ), measured 514 (MH + ). 1 H NMR (400 MHz, METHANOL-d 4 ) δ 8.88 (dd, J = 4.2, 1.3 Hz, 1H), 8.57 (dd, J = 8.5, 1.3 Hz, 1H), 8.06 (d, J = 7.9 Hz, 1H), 7.84 (dd, J = 9.7, 2.7 Hz, 1H), 7.79 (d, J = 2.7 Hz, 1H), 7.57 (dd, J = 8.6, 4.2 Hz, 1H), 7.24 (d, J = 7.9 Hz, 1H), 6.99 (d, J = 9.7 Hz, 1H), 4.76 – 4.72 (m, 1H), 4.27 – 4.20 (m, 1H), 4.09 – 3.98 (m, 2H), 3.96 – 3.89 (m, 1H), 3.88 – 3.74 (m, 3H), 3.71 – 3.56 (m, 3H), 3.55 – 3.49 (m, 1H), 3.40 (s, 3H), 3.36 – 3.26 (m, 1H), 3.19 – 3.05 (m, 2H), 2.31 – 2.22 (m, 2H), 2.20 – 2.06 (m, 1H), 2.05 – 1.94 (m, 1H), 1.41 (d, J = 6.5 Hz, 3H). Example 29 5-[(4R,8S,9aS)-4-methyl-8-[(5-piperazin-1-yl-3-pyridyl)oxy]- 1,3,4,6,7,8,9,9a- octahydropyrido[1,2-a]pyrazin-2-yl]quinoline-8-carbonitrile The title compound was prepared in analogy to the preparation of Example 11 by using 3- Bromo-5-hydroxypyridine (CAS: 74115-13-2) instead of 5-chloropyridin-2-ol (compound 11b). Example 29 (3 mg) was obtained as a light yellow solid. The stereochemistry was confirmed by NOESY. MS: calc’d 484 (MH + ), measured 484 (MH + ). 1 H NMR (400 MHz, METHANOL-d 4 ) δ 9.02 (dd, J = 1.6, 4.3 Hz, 1H), 8.70 (dd, J = 1.6, 8.6 Hz, 1H), 8.21 (d, J = 7.9 Hz, 1H), 8.13 (br d, J = 19.7 Hz, 2H), 7.69 (dd, 1H, J = 4.3, 8.6 Hz), 7.61 (s, 1H), 7.38 (d, J = 7.9 Hz, 1H), 5.14 (br s, 1H), 4.16 (br t, J = 11.6 Hz, 1H), 3.9-4.0 (m, 2H), 3.7-3.8 (m, 1H), 3.6-3.7 (m, 5H), 3.4-3.5 (m, 5H), 3.2-3.3 (m, 2H), 2.4-2.5 (m, 2H), 2.2-2.4 (m, 1H), 2.1-2.2 (m, 1H), 1.53 (d, J = 6.5 Hz, 3H). Example 30 5-[(4R,8R,9aS)-8-[3-[4-[(3R,4R)-3-amino-4-methoxy-pyrrolidin -1-yl]phenyl]azetidin-1-yl]- 4-methyl-1,3,4,6,7,8,9,9a-octahydropyrido[1,2-a]pyrazin-2-yl ]quinoline-8-carbonitrile
The title compound was prepared in analogy to the preparation of Example 9 by using tert- butyl ((3R,4R)-4-methoxypyrrolidin-3-yl)carbamate (CAS: 1932066-52-8, PharmaBlock) instead of tert-butyl piperazine-1-carboxylate (compound 9c). Example 30 (4 mg) was obtained as a light yellow solid. The stereochemistry was confirmed by NOESY. MS: calc’d 552 (MH + ), measured 552 (MH + ). 1 H NMR (400 MHz, METHANOL-d 4 ) δ 8.87 (dd, J = 4.3, 1.6 Hz, 1H), 8.51 (dd, J = 8.6, 1.6 Hz, 1H), 8.04 (d, J = 8.1 Hz, 1H), 7.53 (dd, J = 8.6, 4.2 Hz, 1H), 7.01-7.18 (m, 3H), 6.49 (m, 2H), 3.70-3.98 (m, 3H), 3.41-3.66 (m, 4H), 2.99-3.16 (m, 11H), 2.37-2.81 (m, 5H), 1.71-2.04 (m, 3H), 0.90-1.38 ppm (m, 4H). Example 32 5-[(4R,8R,9aS)-8-[3-[4-[(3R,4S)-3-amino-4-methoxy-pyrrolidin -1-yl]phenyl]azetidin-1-yl]-4- methyl-1,3,4,6,7,8,9,9a-octahydropyrido[1,2-a]pyrazin-2-yl]q uinoline-8-carbonitrile
The title compound was prepared in analogy to the preparation of Example 9 by using tert- butyl ((3R,4S)-4-methoxypyrrolidin-3-yl)carbamate instead of tert-butyl piperazine-1- carboxylate (compound 9c). The stereochemistry was confirmed by NOESY. Example 32 (2 mg) was obtained as a light yellow solid. MS: calc’d 552 (MH + ), measured 552 (MH + ). 1 H NMR (400 MHz, METHANOL-d 4 ) δ 9.04 (dd, J = 4.2, 1.5 Hz, 1H), 8.67 (dd, J = 8.6, 1.7 Hz, 1H), 8.22 (d, J = 7.9 Hz, 1H), 7.70 (dd, J = 8.6, 4.3 Hz, 1H), 7.34 (dd, J = 19.0, 8.4 Hz, 3H), 6.68 (d, J = 8.7 Hz, 2H), 4.46-4.66 (m, 2H), 4.20-4.39 (m, 3H), 3.94-4.17 (m, 3H), 3.40-3.83 (m, 10H), 3.05-3.25 (m, 3H), 2.78-3.01 (m, 2H), 2.33-2.53 (m, 2H), 1.59-1.99 (m, 2H), 1.46 (s, 3H). Example 34 5-[(4R,8R,9aS)-4-methyl-8-[4-[[(2S)-morpholin-2-yl]methyl]an ilino]-1,3,4,6,7,8,9,9a- octahydropyrido[1,2-a]pyrazin-2-yl]-2-deuterio-quinoline-8-c arbonitrile The title compound was prepared in analogy to the preparation of Example 1 by using tert- butyl (S)-2-(4-aminobenzyl)morpholine-4-carboxylate and 2-deuterio-5-[(4R)-4-methyl-8-oxo- 3,4,6,7,9,9a-hexahydro-1H-pyrido[1,2-a]pyrazin-2-yl]quinolin e-8-carbonitrile (Intermediate D1) instead of tert-butyl 4-(4(aminomethyl)phenyl)piperazine-1-carboxylate (Compound 1a) and 5- ((4R,9aS)-4-methyl-8-oxooctahydro-2H-pyrido[1,2-a]pyrazin-2- yl)quinoline-8-carbonitrile (Intermediate C1). Example 34 (34 mg) was obtained as a light yellow solid. The stereochemistry was confirmed by NOESY. MS: calc’d 498 (MH + ), measured 498 (MH + ). 1 H NMR (400 MHz, METHANOL-d 4 ) δ 8.58 (d, J = 8.6 Hz, 1H), 8.08 (d, J = 7.9 Hz, 1H), 7.59 (d, J = 8.6 Hz, 1H), 7.25 (d, J = 7.9 Hz, 1H), 6.92 (d, J = 8.6 Hz, 2H), 6.58 (d, J = 8.6 Hz, 2H), 3.98 – 3.89 (m, 2H), 3.82 – 3.50 (m, 7H), 3.15 – 2.95 (m, 6H), 2.78 – 2.64 (m, 2H), 2.59 – 2.51 (m, 1H), 2.36 – 2.23 (m, 2H), 1.72 – 1.59 (m, 1H), 1.50 – 1.41 (m, 1H), 1.39 (d, J = 6.4 Hz, 3H). Example 36 5-[(4R,8R,9aS)-8-[3-[4-[(3R,4S)-3-amino-4-methoxy-pyrrolidin -1-yl]phenyl]azetidin-1-yl]-4- methyl-1,3,4,6,7,8,9,9a-octahydropyrido[1,2-a]pyrazin-2-yl] ]-2-deuterio-quinoline-8- carbonitrile The title compound was prepared in analogy to the preparation of Example 9 by using 2- deuterio-5-[(4R)-4-methyl-8-oxo-3,4,6,7,9,9a-hexahydro-1H-py rido[1,2-a]pyrazin-2- yl]quinoline-8-carbonitrile (Intermediate D1) and tert-butyl N-[(3R,4S)-4-methoxypyrrolidin-3- yl]carbamate (CAS: 1932508-77-4, PharmaBlock) instead of 5-((4R,9aS)-4-methyl-8- oxooctahydro-2H-pyrido[1,2-a]pyrazin-2-yl)quinoline-8-carbon itrile (Intermediate C1) and tert-butyl piperazine-1-carboxylate (compound 9c ). Example 36 (7 mg) was obtained as a light yellow solid. MS: calc’d 553 (MH + ), measured 553 (MH + ). 1 H NMR (400 MHz, METHANOL- d4) δ 8.56 (d, J = 8.7 Hz, 1H), 8.10 (d, J = 7.9 Hz, 1H), 7.59 (d, J = 8.6 Hz, 1H), 7.23 (dd, J = 18.5, 8.3 Hz, 3H), 6.61 (d, J = 8.8 Hz, 2H), 4.48 – 4.38 (m, 2H), 4.25 – 4.14 (m, 2H), 4.04 – 3.94 (m, 3H), 3.82 – 3.73 (m, 2H), 3.70 – 3.63 (m, 1H), 3.62 – 3.52 (m, 5H), 3.38 – 3.36 (m, 3H), 3.35 – 3.31 (m, 1H), 3.18 – 3.01 (m, 3H), 2.95 – 2.84 (m, 1H), 2.40 – 2.27 (m, 2H), 1.84 – 1.72 (m, 1H), 1.67 – 1.56 (m, 1H), 1.36 (d, J = 6.2 Hz, 3H). Example 37 5-[(4R,8R,9aS)-4-methyl-8-[4-(4-pyridyl)piperazin-1-yl]-1,3, 4,6,7,8,9,9a- octahydropyrido[1,2-a]pyrazin-2-yl]-2-deuterio-quinoline-8-c arbonitrile The title compound was prepared in analogy to the preparation of Example 1 by using 1- (pyridin-4-yl)piperazine and 2-deuterio-5-[(4R)-4-methyl-8-oxo-3,4,6,7,9,9a-hexahydro-1H- pyrido[1,2-a]pyrazin-2-yl]quinoline-8-carbonitrile (Intermediate D1) instead of tert-butyl 4-(4- (aminomethyl)phenyl)piperazine-1-carboxylate (Compound 1a) and 5-((4R,9aS)-4-methyl-8- oxooctahydro-2H-pyrido[1,2-a]pyrazin-2-yl)quinoline-8-carbon itrile (Intermediate C1) in step 1. Example 37 (3 mg) was obtained as a light yellow solid. MS: calc’d 469 (MH + ), measured 469 (MH + ). 1 H NMR (400 MHz, METHANOL-d 4 ) δ 8.52 (d, J = 8.6 Hz, 1H), 7.96-8.11 (m, 3H), 7.53 (d, J = 8.7 Hz, 1H), 7.13 (d, J = 8.1 Hz, 1H), 6.66-6.83 (m, 2H), 3.26-3.45 (m, 9H), 2.51- 2.80 (m, 7H), 1.74-2.04 (m, 3H), 1.43-1.62 (m, 1H), 1.18-1.35 (m, 1H), 1.09 (d, J = 6.2 Hz, 3H). Example 38 The following tests were carried out in order to determine the activity of the compounds of formula (I) and (Ia) in HEK293-Blue-hTLR-7/8/9 cells assay. HEK293-Blue-hTLR-7 cells assay: A stable HEK293-Blue-hTLR-7 cell line was purchased from InvivoGen (Cat.#: hkb-htlr7, San Diego, California, USA). These cells were originally designed for studying the stimulation of human TLR7 by monitoring the activation of NF-κB. A SEAP (secreted embryonic alkaline phosphatase) reporter gene was placed under the control of the IFN-β minimal promoter fused to five NF-κB and AP-1-binding sites. The SEAP was induced by activating NF-κB and AP-1 via stimulating HEK-Blue hTLR7 cells with TLR7 ligands. Therefore the reporter expression was declined by TLR7 antagonist under the stimulation of a ligand, such as R848 (Resiquimod), for incubation of 20 hrs. The cell culture supernatant SEAP reporter activity was determined using QUANTI-Blue™ kit (Cat.#: rep-qb1, Invivogen, San Diego, Ca, USA) at a wavelength of 640 nm, a detection medium that turns purple or blue in the presence of alkaline phosphatase. HEK293-Blue-hTLR7 cells were incubated at a density of 250,000~450,000 cells/mL in a volume of 170 µL in a 96-well plate in Dulbecco's Modified Eagle's medium (DMEM) containing 4.5 g/L glucose, 50 U/mL penicillin, 50 mg/mL streptomycin, 100 mg/mL Normocin, 2 mM L-glutamine, 10% (v/v) heat-inactivated fetal bovine serum with addition of 20 µL test compound in a serial dilution in the presence of final DMSO at 1% and 10 µL of 20uM R848 in above DMEM, perform incubation under 37 ºC in a CO 2 incubator for 20 hrs. Then 20 µL of the supernatant from each well was incubated with 180 µL Quanti-blue substrate solution at 37 o C for 2 hrs and the absorbance was read at 620~655 nm using a spectrophotometer. The signaling pathway that TLR7 activation leads to downstream NF-κB activation has been widely accepted, and therefore similar reporter assay was modified for evaluating TLR7 antagonist. HEK293-Blue-hTLR-8 cells assay: A stable HEK293-Blue-hTLR-8 cell line was purchased from InvivoGen (Cat.#: hkb-htlr8, San Diego, California, USA). These cells were originally designed for studying the stimulation of human TLR8 by monitoring the activation of NF-κB. A SEAP (secreted embryonic alkaline phosphatase) reporter gene was placed under the control of the IFN-β minimal promoter fused to five NF-κB and AP-1-binding sites. The SEAP was induced by activating NF-κB and AP- 1 via stimulating HEK-Blue hTLR8 cells with TLR8 ligands. Therefore the reporter expression was declined by TLR8 antagonist under the stimulation of a ligand, such as R848, for incubation of 20 hrs. The cell culture supernatant SEAP reporter activity was determined using QUANTI- Blue™ kit (Cat.#: rep-qb1, Invivogen, San Diego, Ca, USA) at a wavelength of 640 nm, a detection medium that turns purple or blue in the presence of alkaline phosphatase. HEK293-Blue-hTLR8 cells were incubated at a density of 250,000~450,000 cells/mL in a volume of 170 µL in a 96-well plate in Dulbecco's Modified Eagle's medium (DMEM) containing 4.5 g/L glucose, 50 U/mL penicillin, 50 mg/mL streptomycin, 100 mg/mL Normocin, 2 mM L-glutamine, 10% (v/v) heat-inactivated fetal bovine serum with addition of 20 µL test compound in a serial dilution in the presence of final DMSO at 1% and 10 µL of 60uM R848 in above DMEM, perform incubation under 37 ºC in a CO 2 incubator for 20 hrs. Then 20 µL of the supernatant from each well was incubated with 180 µL Quanti-blue substrate solution at 37 o C for 2 hrs and the absorbance was read at 620~655 nm using a spectrophotometer. The signaling pathway that TLR8 activation leads to downstream NF-κB activation has been widely accepted, and therefore similar reporter assay was modified for evaluating TLR8 antagonist. HEK293-Blue-hTLR-9 cells assay: A stable HEK293-Blue-hTLR-9 cell line was purchased from InvivoGen (Cat.#: hkb-htlr9, San Diego, California, USA). These cells were originally designed for studying the stimulation of human TLR9 by monitoring the activation of NF-κB. A SEAP (secreted embryonic alkaline phosphatase) reporter gene was placed under the control of the IFN-β minimal promoter fused to five NF-κB and AP-1-binding sites. The SEAP was induced by activating NF-κB and AP- 1 via stimulating HEK-Blue hTLR9 cells with TLR9 ligands. Therefore the reporter expression was declined by TLR9 antagonist under the stimulation of a ligand, such as ODN2006 (Cat.#: tlrl-2006-1, Invivogen, San Diego, California, USA), for incubation of 20 hrs. The cell culture supernatant SEAP reporter activity was determined using QUANTI-Blue™ kit (Cat.#: rep-qb1, Invivogen, San Diego, California, USA) at a wavelength of 640 nm, a detection medium that turns purple or blue in the presence of alkaline phosphatase. HEK293-Blue-hTLR9 cells were incubated at a density of 250,000~450,000 cells/mL in a volume of 170 µL in a 96-well plate in Dulbecco's Modified Eagle's medium (DMEM) containing 4.5 g/L glucose, 50 U/mL penicillin, 50 mg/mL streptomycin, 100 mg/mL Normocin, 2 mM L-glutamine, 10% (v/v) heat-inactivated fetal bovine serum with addition of 20 µL test compound in a serial dilution in the presence of final DMSO at 1% and 10 µL of 20uM ODN2006 in above DMEM, perform incubation under 37 ºC in a CO 2 incubator for 20 hrs. Then 20 µL of the supernatant from each well was incubated with 180 µL Quanti-blue substrate solution at 37 o C for 2 h and the absorbance was read at 620~655 nm using a spectrophotometer. The signaling pathway that TLR9 activation leads to downstream NF-κB activation has been widely accepted, and therefore similar reporter assay was modified for evaluating TLR9 antagonist. The compounds of formula (I) have human TLR7 and/or TLR8 inhibitory activities (IC50 value) <0.5 µM. Moreover, some compounds also have human TLR9 inhibitory activity <0.5µM. Activity data of the compounds of the present invention were shown in Table 2. Table 2. The activity of the compounds of present invention in HEK293-Blue-hTLR-7/8/9 cells assays Example 39 hERG channel inhibition assay: The hERG channel inhibition assay is a highly sensitive measurement that identifies compounds exhibiting hERG inhibition related to cardiotoxicity in vivo. The hERG K + channels were cloned in humans and stably expressed in a CHO (Chinese hamster ovary) cell line. CHO hERG cells were used for patch-clamp (voltage-clamp, whole-cell) experiments. Cells were stimulated by a voltage pattern to activate hERG channels and conduct I KhERG currents (rapid delayed outward rectifier potassium current of the hERG channel). After the cells were stabilized for a few minutes, the amplitude and kinetics of I KhERG were recorded at a stimulation frequency of 0.1 Hz, (6 bpm). Thereafter, the test compound was added to the preparation at increasing concentrations. For each concentration, an attempt was made to reach a steady-state effect, usually, this was achieved within 3-10 min at which time the next highest concentration was applied. The amplitude and kinetics of I KhERG are recorded in each concentration of the drug which were compared to the control values (taken as 100%). (references: Redfern WS, Carlsson L, Davis AS, Lynch WG, MacKenzie I, Palethorpe S, Siegl PK, Strang I, Sullivan AT, Wallis R, Camm AJ, Hammond TG.2003; Relationships between preclinical cardiac electrophysiology, clinical QT interval prolongation and torsade de pointes for a broad range of drugs: evidence for a provisional safety margin in drug development. Cardiovasc. Res.58:32-45, Sanguinetti MC, Tristani-Firouzi M.2006; hERG potassium channels and cardiac arrhythmia. Nature 440:463- 469, Webster R, Leishman D, Walker D.2002; Towards a drug concentration effect relationship for QT prolongation and torsades de pointes. Curr. Opin. Drug Discov. Devel.5:116-26). Results of hERG are given in Table 3. A safety ratio (hERG IC 20 /EC 50 ) > 30 suggests a sufficient window to differentiate the pharmacology by inhibiting TLR7/8/9 pathways from the potential hERG related cardiotoxicity. According to the calculation of hERG IC 20 / TLR7/8/9 IC 50 below which serves as early selectivity index to assess hERG liability, obviously reference compounds ER-887258, ER-888285, ER-888286, R1 and R2 have much narrower safety window compared to the compounds of this invention. Table 3. hERG and safety ratio results Example 40 The compounds would be desirable to have minimal DDI liabilities. Therefore, the effects of compounds of formula (I) of (Ia) on major CYP isoforms, e.g. CYP2C9, CYP2D6 and CYP3A4, are determined. CYP inhibition assay This is a high throughput screening assay used for assessment of reversible inhibition of CYP2C9, CYP2D6, and CYP3A4 activity of test compounds in human liver microsome (HLM) in early discovery stage. Table 4. Chemicals and materials used in the CYP inhibition assay Procedure 10 mM DMSO stock solutions of test compounds were diluted in DMSO to generate 2 mM intermediate stock solution.250 nL of intermediate stock solution were transferred in duplicate into 3 separate 384 well microtitre plates (assay-ready plates). A mixture of HLM and each substrate was made up.45 µL of HLM substrate mix was then transferred to each well of an assay ready plate and mixed. The negative (solvent) and positive controls (standard inhibitor for each CYP) were included in each assay ready plate. The assay ready plate was warmed to 37 °C in an incubator over 10 minutes.5 µL pre-warmed NADPH regenerating system was added to each incubation well to start the reaction. Final incubation volume was 50 µL. The assay plate then was placed back in the 37 °C incubator. After incubation (10 minutes for CYP2D6) for 5 minutes, incubates were quenched by addition of 50 µL 100% acetonitrile containing internal standards (400 ng/mL 13C6-4’-OH-Diclofenac, 20 ng/mL D3-Dextrorphan and 20 ng/mL D4- 1’OH-Midazolam). The supernatants were collected for RapidFire/MS/MS analysis. RapidFire online solid phase extraction/sample injection system (Agilent) coupled with API4000 triple quadrupole mass spectrometer (AB Sciex) were used for sample analysis. The mobile phase composed of acetonitrile and water supplemented with 0.1% formic acid. A C4 solid phase extraction cartridge is used for sample separation. MS detection is achieved in positive ion MRM mode. Data analysis Peak areas for substrate, metabolite and internal standard are determined using the RapidFire integrator software (version 3.6.12009.12296). Peak area ratios (PAR) of metabolite and internal standard (stable-labelled metabolite) are then calculated. The measurement window for each experiment is then defined: PAR (0% activity) = average PAR for all incubations containing concentrated inhibitor; Par (100% activity) = average PAR for all incubations containing no inhibitor (DMSO controls); % Activity (test inhibitor) = [PAR(test inhibitor)-PAR(0% activity)]/[PAR(100% activity)-PAR(0% activity)]; % Inhibition (test inhibitor) = 100-% Activity (test inhibitor). The compounds of present invention were found to have low CYP inhibition for CYP2D6 determined in the assays described above. Table 5. CYP inhibition of the compounds of this invention for CYP2D6
percentage inhibition <0: not or weak inhibitor