Structure of 2086301-13-3
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| CAS No. : | 2086301-13-3 |
| Formula : | C17H22ClN3O2S |
| M.W : | 367.89 |
| SMILES Code : | O=C([C@H]1NC[C@H](O)C1)N[C@H](C2=CC=C(C3=C(C)N=CS3)C=C2)C.[H]Cl |
| English Name : | (2S,4R)-4-Hydroxy-N-((S)-1-(4-(4-methylthiazol-5-yl)phenyl)ethyl)pyrrolidine-2-carboxamide hydrochloride |
| MDL No. : | MFCD34603380 |
| InChI Key : | GVZLAESMLXHEDD-LGSZIGQDSA-N |
| Pubchem ID : | 139530909 |
* All experimental methods are cited from the reference, please refer to the original source for details. We do not guarantee the accuracy of the content in the reference.

| Yield | Reaction Conditions | Operation in experiment |
|---|---|---|
| 100% | With hydrogenchloride; methanol In ethyl acetate at 20℃; | 1.1.3 (2S,4R)-4-hydroxy-2-((S)-1-(4-methylthiazole-5-yl)phenyl)ethyl)carbamoyl)pyrrolidine-1-carboxylic acid tert-butyl ester (compound C-8) (16.60g, 39mmol) methanol (30mL) solution, stirred to dissolve. At room temperature, the solution of ethyl acetate (2M, 77mL, 154mmol) of hydrogen chloride was added to the system dropwise, and the stirring reaction was overnight. Concentrate the reaction liquid to give a yellow solid product (2S, 4R)-4-hydroxy-2- (((S)-1-(4-(4-methylthiazole-5-yl) phenyl) carbamoyl) pyrrolidine hydrochloride (compound C-9) (14.15g, yield 100%). |
| 96% | With hydrogenchloride In 1,4-dioxane at 25℃; for 16h; Inert atmosphere; | 4 Step 4: (2S, 4R)-4-hydroxy-N-[(1S)-1-[4-(4-methyl-1, 3-thiazol-5- yl)phenyl]ethyl]pyrrolidine-2-carboxamide hydrochloride. To a stirred solution of tert-butyl (2S,4R)-4-hydroxy-2-[[(1S)-1-[4-(4-methyl-1,3-thiazol-5-yl)phenyl]ethyl]carbamoyl]pyrrolidine- 1-carboxylate (46.0 g, 107 mmol) in dioxane (300 mL) was added a solution of HCl (gas) in 1,4- dioxane (4 M, 150 mL) dropwise at 25 °C under nitrogen atmosphere. The resulting mixture was stirred for 16 h at 25 °C under nitrogen atmosphere. The resulting mixture was filtered. The filter cake was washed with CH2Cl2 (3 x 50.0 mL) and dried to give the title compound as a light yellow solid (42.0 g, 96%): 1H NMR (400 MHz, CD3OD) d 10.11 (s, 1H), 7.66-7.60 (d, J = 8.3 Hz, 2H), 7.57 (d, J = 8.3 Hz, 2H), 5.13 (q, J = 7.0 Hz, 1H), 4.67-4.54 (m, 2H), 3.41 (dd, J = 12.1, 3.5 Hz, 1H), 3.37-3.29 (m, 1H), 2.65 (s, 3H), 2.56 (ddt, J = 13.5, 7.5, 1.7 Hz, 1H), 1.98 (ddd, J = 13.4, 10.6, 4.0 Hz, 1H), 1.56 (d, J = 7.0 Hz, 3H); LC/MS (ESI, m/z): [(M + H)]+ = 332.1. |
| 83% | With hydrogenchloride In methanol at 20℃; for 2h; | 5 Step 5: Preparation of (2S,4R)-4-hydroxy-N-((S)-l-(4-(4-methylthiazol-5- yl)phenyl)ethyl)pyrrolidine-2-carboxamide hydrochloride Into a 500-mL round-bottom flask, was placed a solution of tert-butyl (2S,4R)-4- hydroxy-2- [ [( 1 S )- 1 - [4-(4-methyl- 1 ,3 -thiazol-5-yl)phenyl] ethyl] carbamoyl] pyrrolidine- 1 - carboxylate (5.0 g, 11.59 mmol, 1.00 equiv) in methanol (200 mL), then hydrogen chloride (gas) was bubbled into the reaction mixture for 2 hours at room temperature. The resulting mixture was concentrated under vacuum. This resulted in 3.2 g (83%) of (2S,4R)-4-hydroxy-N-[(lS)-l- [4-(4-methyl-l,3-thiazol-5-yl)phenyl]ethyl]pyrrolidine-2-carboxamide as a red solid. |
| 83% | With hydrogenchloride In methanol at 20℃; for 2h; | 5 Step 5: Preparation of (2S,4R)-4-hydroxy-N-((S)-1-(4-(4-methylthiazol-5- yl)phenyl)ethyl)pyrrolidine-2-carboxamide hydrochloride Into a 500-mL round-bottom flask, was placed a solution of tert-butyl (2S,4R)-4- hydroxy-2-[[(1S)-1-[4-(4-methyl-1,3-thiazol-5-yl)phenyl]ethyl]carbamoyl]pyrrolidine-1- carboxylate (5.0 g, 11.59 mmol, 1.00 equiv) in methanol (200 mL), then hydrogen chloride (gas) was bubbled into the reaction mixture for 2 h at room temperature. The resulting mixture was concentrated under vacuum. This resulted in 3.2 g (83%) of (2S,4R)-4-hydroxy-N-[(1S)-1-[4-(4- methyl-1,3-thiazol-5-yl)phenyl]ethyl]pyrrolidine-2-carboxamide as a red solid. |
| 83% | With hydrogenchloride In methanol at 20℃; for 2h; | 5 Step 5: Preparation of (2S,4R)-4-hydroxy-N-((S)-1-(4-(4-methylthiazol-5-yl)phenyl)ethyl)pyrrolidine-2-carboxamide hydrochloride Into a 500-mL round-bottom flask, was placed a solution of tert-butyl (2S,4R)-4-hydroxy-2-[[(1S)-1-[4-(4-methyl-1,3-thiazol-5-yl)phenyl]ethyl]carbamoyl]pyrrolidine-1-carboxylate (5.0 g, 11.59 mmol, 1.00 equiv) in methanol (200 mL), then hydrogen chloride (gas) was bubbled into the reaction mixture for 2 hours at room temperature. The resulting mixture was concentrated under vacuum. This resulted in 3.2 g (83%) of (2S,4R)-4-hydroxy-N-[(1S)-1-[4-(4-methyl-1,3-thiazol-5-yl)phenyl]ethyl]pyrrolidine-2-carboxamide as a red solid. |
| With hydrogenchloride In 1,4-dioxane at 0 - 20℃; for 16h; | Step-e: Synthesis of (2S,4R)-4-hydroxy-N-((S)-1-(4-(4-methylthiazol-5-yl)phenyl)ethyl)pyrrolidine-2-carboxamide hydrochloride (17e) To a stirred solution of tert-butyl (2S,4R)-4-hydroxy-2-(((S)-1-(4-(4-methylthiazol-5-yl)phenyl)ethyl)carbamoyl)pyrrolidine-1-carboxylate (10.0 g, 23.20 mmol) in l,4-dioxane (50 mL) at 0 °C was added 4M HC1 in l,4-dioxane (50 mL). The reaction mixture was stirred at room temperature for 16 h. The solvents were evaporated under reduced pressure and the residue was washed with diethyl ether to afford the title compound (8.5 g, 100 %) which was used in the next step without further purification. 1H NMR (400 MHz, DMSO-d6): d 10.20 (bs, 1H), 9.22- 9.30 (m, 1H), 9.15 (s, 1H), 8.67 (bs, 1H), 7.46-7.42 (m, 3H), 5.01-4.95 (m, 1H) ,4.50-4.40 (m, 2H), 3.30-3.20 (m, 1H), 3.20-3.00 (m, 1H), 2.50 (s, 3H), 2.40-2.35 (m, 2H), 1.80-1.70 (m, 1H), 1.43 (d, / = 8.4 Hz, 3H), LC-MS: m/z 332.1 (M+l)+. | |
| 1.33 g | With hydrogenchloride In 1,4-dioxane; methanol; dichloromethane; water at 20℃; for 6h; | 210.210-2 (210-2) (2S, 4R) -4-hydroxy-N-{(1S) -1- [4- (4-methyl-1,3-thiazole-5-yl) phenyl] ethyl} pyrrolidine-2- Carboxamide hydrochloride (Example compound 210-2) Example Compound 210-1 (2.02 g) was added to a mixed solution of dichloromethane (4.0 mL) and methanol (6.0 mL), and a 4M hydrogen chloride / dioxane solution (4.3 mL) was added, and the mixture was stirred at room temperature for 6 hours. The reaction mixture was concentrated under reduced pressure and then azeotroped with toluene. Hexane was added to the residue and concentrated under reduced pressure, and the obtained solid was suspended and washed with diethyl ether to give the title compound (1.33 g) as a light brown powder. |
| With hydrogenchloride In 1,4-dioxane; methanol; water at 0 - 20℃; for 2h; | 1.10 Step 10: Preparation of (2S,4R)-4-hydroxy-N-((S)-1-(4-(4-methylthiazol-5-yl)phenyl)ethyl)pyrrolidine-2- carboxamide hydrochloride To a stirred solution of tert-butyl (2S,4R)-4-hydroxy-2-(((S)-l-(4-(4-methylthiazol-5- yl)phenyl)ethyl)carbamoyl)pyrrolidine-l-carboxylate (8.33 g, 19.3 mmol) at 0 °C was added a solution of HCI in 1 ,4-dioxane (4 N, 50 mL, 200 mmol) resulting in a sticky yellow gum. 15 mL of MeOH was added to the mixture and the mixture was stirred at room temperature for 2 h. The solvents were removed under reduced pressure and the residue was washed with diethyl ether to afford (2S,4R)-4-hydroxy-N-((S)-1-(4- (4-methylthiazol-5-yl)phenyl)ethyl)pyrrolidine-2-carboxamide hydrochloride which was used in the next step without further purification. | |
| With hydrogenchloride In 1,4-dioxane; methanol; water at 0 - 20℃; for 2h; | 1.10 Step 10: Preparation of (2S,4R)-4-hydroxy-N-((S)-1-(4-(4-methylthiazol-5-yl)phenyl)ethyl)pyrrolidine-2- carboxamide hydrochloride To a stirred solution of tert-butyl (2S,4R)-4-hydroxy-2-(((S)-l-(4-(4-methylthiazol-5- yl)phenyl)ethyl)carbamoyl)pyrrolidine-l-carboxylate (8.33 g, 19.3 mmol) at 0 °C was added a solution of HCI in 1 ,4-dioxane (4 N, 50 mL, 200 mmol) resulting in a sticky yellow gum. 15 mL of MeOH was added to the mixture and the mixture was stirred at room temperature for 2 h. The solvents were removed under reduced pressure and the residue was washed with diethyl ether to afford (2S,4R)-4-hydroxy-N-((S)-1-(4- (4-methylthiazol-5-yl)phenyl)ethyl)pyrrolidine-2-carboxamide hydrochloride which was used in the next step without further purification. | |
| With hydrogenchloride In 1,4-dioxane; dichloromethane at 20℃; | 3.2 Step 2: (25,4A)-4-Hydroxy-A-((S)-l-(4-(4-methylthiazol-5- yl)phenyl)ethyl)pyrrolidine-2-carboxamide (hydrochloride salt) A solution of tert-butyl (2k,4/?)-4-hydroxy-2-((fS')- l -(4-(4-methylthiazol-5- yl)phenyl)ethyl)carbamoyl)pyrrolidine-l -carboxylate (6.89 g, 13.5 mmol) in 4 M HCl/dioxane (50 mL) and dichloromethane (100 mL) was stirred at room temperature for 1 hour. The solvent was concentrated under vacuum to yield 5.8 g (crude) of the title compound as a white solid. LC-MS: (ESI, m/z)'. [M+H]+= 332. | |
| With hydrogenchloride In 1,4-dioxane at 20℃; | 1.10 Step 10: Preparation of (2S,4R)-4-hydroxy-N-((S)-1-(4-(4-methylthiazol-5- yl)phenyl)ethyl)pyrrolidine-2-carboxamide hydrochloride. To tert-butyl (2S,4R)-4-hydroxy-2-(((S)-1-(4-(4-methylthiazol-5- yl)phenyl)ethyl)carbamoyl)pyrrolidine-1 -carboxylate (8.33 g, 19.3 mmol) at 0 °C was added a solution of HCI in 1 ,4-dioxane (4 N, 50 mL, 200 mmol) resulting in a sticky yellow gum. 15 mL of MeOH were added to the mixture and the mixture was stirred at room temperature for 2 h. The solvents were removed under reduced pressure and the residue was washed with diethyl ether to afford (2S,4R)-4-hydroxy-N-((S)-1-(4-(4-methylthiazol-5-yl)phenyl)ethyl)pyrrolidine-2-carboxamide hydrochloride which was used in the next step without further purification. | |
| 100 % | With hydrogenchloride In 1,4-dioxane; dichloromethane at 20℃; | General procedure: (9H-fluoren-9-yl)methyl((R)-1-((2R,4R)-4-hydroxy-2-(((S)-1-(4-(4-methylthiazol-5-yl)phenyl)ethyl)carbamoyl)pyrrolidin-1-yl)-3-methyl-1-oxo-3-(tritylthio)butan-2-yl)carbamatewas prepared following general procedure 4. 190 mg(9H-fluoren-9-yl)methyl ((R)-1-((2R,4R)-4-hydroxy-2-(((S)-1-(4-(4-methylthiazol-5-yl)phenyl)ethyl)carbamoyl)pyrrolidin-1-yl)-3-methyl-1-oxo-3-(tritylthio)butan-2-yl)carbamatewas obtained from 118 mgtert-butyl (2S,4R)-4-hydroxy-2-(((S)-1-(4-(4-methylthiazol-5-yl)phenyl)ethyl)carbamoyl)pyrrolidine-1-carboxylate,75% yield.1H NMR (400 MHz, CDCl3) δ 8.67 (s, 1H), 7.78 - 7.72 (m, 2H), 7.62 - 7.52 (m, 8H), 7.41 - 7.17 (m, 18H), 5.73 (d, J = 5.3 Hz, 1H), 5.00 (p, J = 7.0 Hz, 1H), 4.65 (t, J = 8.0 Hz, 1H), 4.42 - 4.15 (m, 4H), 3.56 (d, J = 5.3 Hz, 1H), 3.45 (d, J = 11.6 Hz, 1H), 3.17 (dd,J= 11.6, 3.5 Hz, 1H), 2.83 (br, s, 1H), 2.51 (s, 3H), 2.44 - 2.32 (m, 1H), 2.08 - 1.96 (m, 1H), 1.39 - 1.31 (m, 6H), 1.17 (s, 3H).13C NMR (101 MHz, CDCl3) δ 170.91, 169.57, 156.61, 150.35, 148.68, 144.60, 144.02, 143.58, 143.26, 141.41, 130.99, 130.04, 129.68, 128.03, 127.94, 127.28, 127.08, 126.58, 125.29, 125.13, 120.18, 70.25, 67.79, 58.61, 58.38, 56.62, 48.92, 47.14, 35.86, 26.12, 26.05, 22.34, 16.23. LC-MS,ESI+,m/z927 [M+H]+. |
| With hydrogenchloride In 1,4-dioxane at 20℃; | 1.10 Step 10: Preparation of (2S,4R)-4-hydroxy-N-((S)-1-(4-(4-methylthiazol-5- yl)phenyl)ethyl)pyrrolidine-2-carboxamide hydrochloride. To tert-butyl (2S,4R)-4-hydroxy-2-(((S)-1-(4-(4-methylthiazol-5- yl)phenyl)ethyl)carbamoyl)pyrrolidine-1 -carboxylate (8.33 g, 19.3 mmol) at 0 °C was added a solution of HCI in 1 ,4-dioxane (4 N, 50 mL, 200 mmol) resulting in a sticky yellow gum. 15 mL of MeOH were added to the mixture and the mixture was stirred at room temperature for 2 h. The solvents were removed under reduced pressure and the residue was washed with diethyl ether to afford (2S,4R)-4-hydroxy-N-((S)-1-(4-(4-methylthiazol-5-yl)phenyl)ethyl)pyrrolidine-2-carboxamide hydrochloride which was used in the next step without further purification. | |
| 100 % | With hydrogenchloride In 1,4-dioxane at 0 - 20℃; | 4.6.5. (2S,4R)-4-hydroxy-N-((S)-1-(4-(4-methylthiazol-5-yl)phenyl)ethyl)pyrrolidine-2-carboxamide hydrochloride (52) To the solution of 50 (1.0 g, 2.317 mmol) in dioxane (3.2 mL) and EA(0.8 mL), 4.0 N HCl in dioxane (4.4 mL) was added dropwise at 0 C. Thereaction was stirred at room temperature for 18 h. Then, the solvent wasevaporated to dryness and the residue was tritured with DEE (10 mL),collected by filtration, and dried in vacuo to afford the titled compoundas a yellow solid (0.852 g, two-step yield 100 %). 1H NMR (400 MHz,DMSO-d6) δ 10.16 (bs, 1H), 9.28 (d, J = 7.5 Hz, 1H), 9.09 (s, 1H), 8.58(bs, 1H), 7.44 (dd, J = 25.3, 8.2 Hz, 4H), 5.06-4.93 (m, 1H), 4.45-4.36(m, 2H), 3.35-3.24 (m, 1H), 3.12-3.03 (m, 1H), 2.68 (s, 3H), 2.46 (s,3H), 2.40-2.31 (m, 1H), 1.84-1.75 (m, 1H), 1.42 (d, J = 6.9 Hz, 3H). |
| 100 % | With hydrogenchloride In 1,4-dioxane at 0 - 20℃; | 4.6.5. (2S,4R)-4-hydroxy-N-((S)-1-(4-(4-methylthiazol-5-yl)phenyl)ethyl)pyrrolidine-2-carboxamide hydrochloride (52) To the solution of 50 (1.0 g, 2.317 mmol) in dioxane (3.2 mL) and EA(0.8 mL), 4.0 N HCl in dioxane (4.4 mL) was added dropwise at 0 C. Thereaction was stirred at room temperature for 18 h. Then, the solvent wasevaporated to dryness and the residue was tritured with DEE (10 mL),collected by filtration, and dried in vacuo to afford the titled compoundas a yellow solid (0.852 g, two-step yield 100 %). 1H NMR (400 MHz,DMSO-d6) δ 10.16 (bs, 1H), 9.28 (d, J = 7.5 Hz, 1H), 9.09 (s, 1H), 8.58(bs, 1H), 7.44 (dd, J = 25.3, 8.2 Hz, 4H), 5.06-4.93 (m, 1H), 4.45-4.36(m, 2H), 3.35-3.24 (m, 1H), 3.12-3.03 (m, 1H), 2.68 (s, 3H), 2.46 (s,3H), 2.40-2.31 (m, 1H), 1.84-1.75 (m, 1H), 1.42 (d, J = 6.9 Hz, 3H). |
| 100 % | With hydrogenchloride In 1,4-dioxane; dichloromethane at 20℃; | General procedure: (9H-fluoren-9-yl)methyl((R)-1-((2R,4R)-4-hydroxy-2-(((S)-1-(4-(4-methylthiazol-5-yl)phenyl)ethyl)carbamoyl)pyrrolidin-1-yl)-3-methyl-1-oxo-3-(tritylthio)butan-2-yl)carbamatewas prepared following general procedure 4. 190 mg(9H-fluoren-9-yl)methyl ((R)-1-((2R,4R)-4-hydroxy-2-(((S)-1-(4-(4-methylthiazol-5-yl)phenyl)ethyl)carbamoyl)pyrrolidin-1-yl)-3-methyl-1-oxo-3-(tritylthio)butan-2-yl)carbamatewas obtained from 118 mgtert-butyl (2S,4R)-4-hydroxy-2-(((S)-1-(4-(4-methylthiazol-5-yl)phenyl)ethyl)carbamoyl)pyrrolidine-1-carboxylate,75% yield.1H NMR (400 MHz, CDCl3) δ 8.67 (s, 1H), 7.78 - 7.72 (m, 2H), 7.62 - 7.52 (m, 8H), 7.41 - 7.17 (m, 18H), 5.73 (d, J = 5.3 Hz, 1H), 5.00 (p, J = 7.0 Hz, 1H), 4.65 (t, J = 8.0 Hz, 1H), 4.42 - 4.15 (m, 4H), 3.56 (d, J = 5.3 Hz, 1H), 3.45 (d, J = 11.6 Hz, 1H), 3.17 (dd,J= 11.6, 3.5 Hz, 1H), 2.83 (br, s, 1H), 2.51 (s, 3H), 2.44 - 2.32 (m, 1H), 2.08 - 1.96 (m, 1H), 1.39 - 1.31 (m, 6H), 1.17 (s, 3H).13C NMR (101 MHz, CDCl3) δ 170.91, 169.57, 156.61, 150.35, 148.68, 144.60, 144.02, 143.58, 143.26, 141.41, 130.99, 130.04, 129.68, 128.03, 127.94, 127.28, 127.08, 126.58, 125.29, 125.13, 120.18, 70.25, 67.79, 58.61, 58.38, 56.62, 48.92, 47.14, 35.86, 26.12, 26.05, 22.34, 16.23. LC-MS,ESI+,m/z927 [M+H]+. |
| 7.29 g | With hydrogenchloride In methanol at 4℃; | |
| With hydrogenchloride In 1,4-dioxane; methanol at 0 - 20℃; | Step 4: Dissolve 2-c (3.2g, 7.4mmol) in MeOH (30mL), add HCl/dioxane solution (30mL, 4M) at 0°C, raise the reaction solution to room temperature and stir for 2 hours. The reaction is concentrated to obtain intermediate 2 (3.1g, brown solid, crude product). | |
| 5.4 g | With hydrogenchloride In 1,4-dioxane; dichloromethane; water at 20℃; for 1h; Inert atmosphere; | Step 2. (2S,4R)-4-hydroxy-N-[(1S)-1-[4-(4-methyl-1 ,3-thiazol-5-yl)phenyl]ethyl]pyrrolidine-2- carboxamide A solution of tert-butyl (2S,4R)-4-hydroxy-2-[(1 S)-1 -[4-(4-methyl-1 ,3-thiazol-5- yl)phenyl]ethyl]carbamoyl}pyrrolidine-1-carboxylate (11 .7 g, 27.11 mmol) in 4N HCI in 1 ,4-dioxane (70 mL) and DCM (70 mL) was stirred for 1 h at room temperature. The resulting mixture was concentrated under reduced pressure, The mixture was purified by reversed-phase flash chromatography with the following conditions: column, C18 silica gel; mobile phase, MeCN in Water (0.05% TFA), 0% to 100% gradient in 15 min; detector, UV 254 nm, to afford the title compound (5.4 g) as a brown solid. LCMS (ESI) m/z [M+H]+= 332.3. 1H NMR (300 MHz, DMSO-d6) 5 = 9.01 (s, 1 H), 8.22 - 8.06 (m, 1 H), 7.48 (d, J = 8.3 Hz, 2H), 7.40 (d, J = 8.3 Hz, 2H), 5.10 - 4.93 (m, 1 H), 4.47 - 4.25 (m, 3H), 3.11 (s, 1 H), 2.46 (s, 3H), 2.36 (dd, J = 13.2, 7.2 Hz, 3H), 1 .92 - 1 .75 (m, 1 H), 1 .43 (d, J = 7.0 Hz, 3H). LCMS (ESI) m/z [M+H]+= 332.2. |
| With hydrogenchloride In 1,4-dioxane; dichloromethane at 20℃; for 1h; | Step 2: (2S,4R)-4-Hydroxy-N-((S)-1-(4-(4-methylthiazol-5-yl)phenyl)ethyl) pyrrolidine-2 carboxamide To a solution of tert-butyl (2S,4R)-4-hydroxy-2-[[(1S)-1-[4-(4-methylthiazol-5-yl) phenyl]ethyl]carbamoy1]pyrrolidine-1-carboxylate (3.77 g, 8.74 mmol) in DCM (20 mL) was added 4M HCl-dioxane (20 mL, 80 mmol) and the reaction mixture was stirred at room temperature for 1 h. The solvent was evaporated under reduced pressure to afford the title compound. | |
| With hydrogenchloride In dichloromethane; water at 0 - 20℃; for 2h; |

| Yield | Reaction Conditions | Operation in experiment |
|---|---|---|
| 85.6% | With N-ethyl-N,N-diisopropylamine; N-[(dimethylamino)-3-oxo-1H-1,2,3-triazolo[4,5-b]pyridin-1-yl-methylene]-N-methylmethanaminium hexafluorophosphate In N,N-dimethyl-formamide at 20℃; for 3h; | 1.1.3 (2S,4R)-4-hydroxy-2-(((S)-1-(4-(4-methylthiazole-5-yl)phenyl)ethyl)carbamoyl)pyrrolidine hydrochloride (Compound C-9) (14.15g, 39mmol), (S)-2-((tert-butoxycarbonyl)amino)-3,3-dimethylbutyric acid (compound C-10) (9.79g, 43mmol), diisopropylethylamine (30.83g, 240mmol) was added to N, N-Dimethylformamide (70mL), stir well. At room temperature, HATU (17.55g, 47mmol) was added to the system and stirred for 3 hours. Add 200mL of water to the reaction solution to quench, there is a white solid product precipitation, filtration, filtrate with ethyl acetate extraction, organic phase and then washed with saturated sodium chloride aqueous solution 2-3 times, organic phase with anhydrous sodium sulfate drying, filtering, concentrated filtrate, to obtain oily crude products. Column chromatography purification (petroleum ether: ethyl acetate = 1:1~0:1) was combined with a white solid product to give a product ((S)-1-((2S,4R)-4-hydroxy-2-((S)-1-(4-(4-methylthiazole-5-yl)phenyl)ethyl) carbamoyl)pyrrolidine-1-yl)-3,3-dimethyl-1-oxobutane-2-yl) carbamic acid tert-butyl ester (compound C-11) (17.94g, yield 85.6%). |
| 84% | With N-ethyl-N,N-diisopropylamine; HATU In N,N-dimethyl-formamide at 20℃; for 12h; | 6 Step 6: Preparation of tert- butyl ((S)-l-((2S,4R)-4-hydroxy-2-(((S)-l-(4-(4- methylthiazol-5-yl)phenyl)ethyl)carbamoyl)pyrrolidin-l-yl)-3, 3-dimethyl- l-oxobutan-2- yl)carbamate Into a 25-mL round-bottom flask, was placed (2S)-2-[(tert-butoxy)carbonyl] amino- 3, 3 -dimethylbutanoic acid (2.0 g, 8.65 mmol, 0.99 equiv) in N,N-dimethylformamide (30 mL). N-ethyl-N-isopropylpropan-2-amine (3.4 g, 3.00 equiv), o-(7- Azabenzotriazol- 1 -yl)-N,N,N',N’- te-tramethyluronmm hexafluorophosphate (5.0 g, 1.50 equiv), (2S,4R)-4-hydroxy-N-[(lS)-l-[4- (4-methyl-l,3-thiazol-5-yl)phenyl]ethyl]pyrrolidine-2-carboxamide hydrochloride (3.2 g, 8.70 mmol, 1.00 equiv). The resulting solution was stirred for 12 hours at room temperature. The resulting solution was extracted with ethyl acetate (60 mL x 3) and washed with water (100 mL x 2). The organic layers combined and dried, concentrated under vacuum. The residue was applied onto a silica gel column with ethyl acetate/petroleum ether (1:3). This resulted in 4.0 g (84%) of tert-butyl N - [(2S )- 1 - [(2S ,4R)-4-hydroxy-2- [ [( 1 S )- 1 - [4-(4-methyl- 1 ,3 -thiazol-5- yl)phenyl]ethyl]carbamoyl]pyrrolidin-l-yl]-3, 3-dimethyl- l-oxobutan-2-yl]carbamate as a yellow solid. LC/MS (ESI) m/z: 545.30 [M+l] +. |
| 84% | With N-ethyl-N,N-diisopropylamine; HATU In N,N-dimethyl-formamide at 20℃; for 12h; | 6 Step 6: Preparation of tert- butyl ((S)-1-((2S,4R)-4-hydroxy-2-(((S)-1-(4-(4- methylthiazol-5-yl)phenyl)ethyl)carbamoyl)pyrrolidin-1-yl)-3,3-dimethyl-1-oxobutan-2- yl)carbamate Into a 25-mL round-bottom flask, was placed (2S)-2-[(tert-butoxy)carbonyl]amino- 3,3-dimethylbutanoic acid (2.0 g, 8.65 mmol, 0.99 equiv) in N,N-dimethylformamide (30 mL). N-ethyl-N-isopropylpropan-2-amine (3.4 g, 3.00 equiv), o-(7-Azabenzotriazol-1-yl)-N,N,N',N'- te-tramethyluronium hexafluorophosphate (5.0 g, 1.50 equiv), (2S,4R)-4-hydroxy-N-[(1S)-1-[4- (4-methyl-1,3-thiazol-5-yl)phenyl]ethyl]pyrrolidine-2-carboxamide hydrochloride (3.2 g, 8.70 mmol, 1.00 equiv). The resulting solution was stirred for 12 h at room temperature. The resulting solution was extracted with ethyl acetate (60 mL x 3) and washed with water (100 mL x 2). The organic layers combined and dried, concentrated under vacuum. The residue was applied onto a silica gel column with ethyl acetate/petroleum ether (1:3). This resulted in 4.0 g (84%) of tert- butyl N-[(2S)-1-[(2S,4R)-4-hydroxy-2-[[(1S)-1-[4-(4-methyl-1,3-thiazol-5- yl)phenyl]ethyl]carbamoyl]pyrrolidin-1-yl]-3,3-dimethyl-1-oxobutan-2-yl]carbamate as a yellow solid. LC/MS (ESI) m/z: 545.30 [M+1] +. |
| 84% | With N-ethyl-N,N-diisopropylamine; N-[(dimethylamino)-3-oxo-1H-1,2,3-triazolo[4,5-b]pyridin-1-yl-methylene]-N-methylmethanaminium hexafluorophosphate In N,N-dimethyl-formamide at 20℃; for 12h; | 6 Step 6: Preparation of tert-butyl ((S)-1-((2S,4R)-4-hydroxy-2-(((S)-1-(4-(4-methylthiazol-5-yl)phenyl)ethyl)carbamoyl)pyrrolidin-1-yl)-3,3-dimethyl-1-oxobutan-2-yl)carbamate Into a 25-mL round-bottom flask, was placed (2S)-2-[(tert-butoxy)carbonyl]amino-3,3-dimethylbutanoic acid (2.0 g, 8.65 mmol, 0.99 equiv) in N,N-dimethylformamide (30 mL). N-ethyl-N-isopropylpropan-2-amine (3.4 g, 3.00 equiv), o-(7-Azabenzotriazol-1-yl)-N,N,N',N'-te-tramethyluronium hexafluorophosphate (5.0 g, 1.50 equiv), (2S,4R)-4-hydroxy-N-[(1S)-1-[4-(4-methyl-1,3-thiazol-5-yl)phenyl]ethyl]pyrrolidine-2-carboxamide hydrochloride (3.2 g, 8.70 mmol, 1.00 equiv). The resulting solution was stirred for 12 hours at room temperature. The resulting solution was extracted with ethyl acetate (60 mL x 3) and washed with water (100 mL x 2). The organic layers combined and dried, concentrated under vacuum. The residue was applied onto a silica gel column with ethyl acetate/petroleum ether (1:3). This resulted in 4.0 g (84%) of tert-butyl N-[(2S)-1-[(2S,4R)-4-hydroxy-2-[[(1S)-1-[4-(4-methyl-1,3-thiazol-5-yl)phenyl]ethyl]carbamoyl]pyrrolidin-1-yl]-3,3-dimethyl-1-oxobutan-2-yl]carbamate as a yellow solid. LC/MS (ESI) m/z: 545.30 [M+1] + |
| 76% | With triethylamine; HATU In N,N-dimethyl-formamide at 0 - 25℃; Inert atmosphere; | 5 Step 5: Tert-butyl N-[(2S)-1-[(2S,4R)-4-hydroxy-2-[[(1S)-1-[4-(4-methyl-1,3-thiazol- 5-yl)phenyl]ethyl]carbamoyl]pyrrolidin-1-yl]-3,3-dimethyl-1-oxobutan-2-yl]carbamate. To a stirred mixture of (2S)-2-[(tert-butoxycarbonyl)amino]-3,3-dimethylbutanoic acid (27.7 g, 120 mmol) and TEA (45.3 mL, 448 mmol) in DMF (400 mL) were added (2S,4R)-4-hydroxy-N-[(1S)- 1-[4-(4-methyl-1,3-thiazol-5-yl)phenyl]ethyl]pyrrolidine-2-carboxamide hydrochloride (40.0 g, 109 mmol) and HATU (53.7 g, 142 mmol) in portions at 0 °C under nitrogen atmosphere. The resulting mixture was stirred for 3 h at 25 °C under nitrogen atmosphere. The reaction was quenched by the addition of water (300 mL) at 25 °C. The resulting mixture was extracted with EtOAc (3 x 300 mL). The combined organic layers were washed with brine (3 x 300 mL), dried over anhydrous Na2SO4. After filtration, the filtrate was concentrated under reduced pressure. The residue was purified by trituration with EtOAc (300 mL). The precipitated solids were collected by filtration and washed with EtOAc (2 x 40.0 mL). This resulted in the title compound as a light yellow solid (50.0 g, 76%): 1H NMR (400 MHz, CD3OD) d 8.90 (s, 1H), 7.50-7.36 (m, 4H), 6.40 (d, J = 9.3 Hz, 1H), 5.02 (p, J = 7.0 Hz, 1H), 4.62 (t, J = 8.3 Hz, 1H), 4.46 (s, 1H), 4.34-4.27 (m, 1H), 3.87 (d, J = 11.1 Hz, 1H), 3.76 (dd, J = 10.9, 4.0 Hz, 1H), 2.50 (s, 3H), 2.24 (dd, J = 13.2, 7.8 Hz, 1H), 2.06-1.93 (m, 1H), 1.56-1.48 (m, 3H), 1.46 (s, 9H), 1.03 (s, 9H); LC/MS (ESI, m/z): [(M + H)]+ = 545.40. |
| 66.6% | With N-[(dimethylamino)-3-oxo-1H-1,2,3-triazolo[4,5-b]pyridin-1-yl-methylene]-N-methylmethanaminium hexafluorophosphate In N,N-dimethyl-formamide at 25 - 30℃; for 16h; Inert atmosphere; Alkaline conditions; | |
| 9.8 g | With N-ethyl-N,N-diisopropylamine; N-[(dimethylamino)-3-oxo-1H-1,2,3-triazolo[4,5-b]pyridin-1-yl-methylene]-N-methylmethanaminium hexafluorophosphate In dichloromethane at 20℃; for 16h; | Step-f: Synthesis of tert-butyl ((S)-1-((2S,4R)-4-hydroxy-2-(((S)-1-(4-(4-methylthiazol-5-yl)phenyl)ethyl)carbamoyl)pyrrolidin-1-yl)-3,3-dimethyl-1-oxobutan-2-yl)carbamate (17f) To a solution of (2S,4R)-4-hydroxy-N-((S)-1-(4-(4-methylthiazol-5-yl)phenyl)ethyl)pyrrolidine-2-carboxamide hydrochloride (10.0 g, 27.24 mmol) and (S)-2-((tert-butoxycarbonyl)amino)-3,3-dimethylbutanoic acid (6.29 g, 27.24 mmol) in DCM (150 mL) at 0 °C was added HATU (12.4 g, 32,69 mmol) followed by dropwise addition of DIPEA (25.0 mL, 136.2 mmol) and stirred for 16 h at RT. Then the reaction mixture was poured into ice cold water and the resulting mixture was extracted with DCM (2 X 500 mL). The combined organic layer was washed with water (200 mL), saturated sodium bicarbonate solution (100 mL), brine (200 mL), dried over anhydrous sodium sulphate and concentrated under vacuum to give the crude product which was washed with chilled acetone and filtered to afford the title compound (9.8 g, 67.5 %) which was used in the next step without further purification. 1H NMR (400 MHz, DMSO-d6): d 8.97 (s, 1H), 8.38 (d, J = 6.4 Hz, 1H), 7.44-7.31 (m, 4H), 6.38 (d, / = 8.4 Hz, 1H), 5.10 (s, 1H), 4.91-4.88 (m, 1H), 4.28 (bs, 1H), 4.14 (d, / = 8.8 Hz, 1H), 3.62-3.58 (m, 2H), 2.45 (s, 3H), 2.08-2.00 (m, 2H), 1.78-1.76 (m, 1H), 1.48 (s, 9H), 1.44 (d, J = 6.9 Hz, 3H), 0.93 (s, 9H); LC-MS: m/z 545.3 (M+l)+. |
| 1.873 g | Stage #1: N-tert-butyloxycarbonyl-L-tert-leucine With N-ethyl-N,N-diisopropylamine; N-[(dimethylamino)-3-oxo-1H-1,2,3-triazolo[4,5-b]pyridin-1-yl-methylene]-N-methylmethanaminium hexafluorophosphate In dichloromethane at 0℃; Inert atmosphere; Stage #2: (2S,4R)-4-hydroxy-N-((S)-1-(4-(4-methylthiazol-5-yl)phenyl)ethyl)pyrrolidine-2-carboxamide hydrochloride In dichloromethane at 20℃; Inert atmosphere; | 4.6.6. Tert-butyl ((S)-1-((2S,4R)-4-hydroxy-2-(((S)-1-(4-(4-methylthiazol-5-yl)phenyl)ethyl)carbamoyl)pyrrolidin-1-yl)-3,3-dimethyl-1-oxobutan-2-yl)carbamate (54) Under nitrogen atmosphere, to a stirred solution of (S)-2-((tertbutoxycarbonyl)amino)-3,3-dimethylbutanoic acid (0.953 g, 4.123mmol) in dry DCM (10.5 mL), DIPEA (1.8 mL, 10.307 mmol) and HATU(1.834 g, 4.824 mmol) were added at 0 C. After 1 h, 52 (0.910 g, 2.473mmol) was added slowly. The mixture was stirred for 18 h at roomtemperature. Then, DCM (50 mL) was added to the reaction mixture andwas washed with 10 % citric acid (12 mL) twice, saturated NaHCO3solution (12 mL) twice, water (12 mL) twice, and brine (20 mL) once.The organic phase was dried over anhydrous Na2SO4 and concentratedunder reduced pressure to afford the desired compound as a yellow oil(1.873 g) which was used directly for the next step. |
| 1.873 g | Stage #1: N-tert-butyloxycarbonyl-L-tert-leucine With N-ethyl-N,N-diisopropylamine; N-[(dimethylamino)-3-oxo-1H-1,2,3-triazolo[4,5-b]pyridin-1-yl-methylene]-N-methylmethanaminium hexafluorophosphate In dichloromethane at 0℃; Inert atmosphere; Stage #2: (2S,4R)-4-hydroxy-N-((S)-1-(4-(4-methylthiazol-5-yl)phenyl)ethyl)pyrrolidine-2-carboxamide hydrochloride In dichloromethane at 20℃; Inert atmosphere; | 4.6.6. Tert-butyl ((S)-1-((2S,4R)-4-hydroxy-2-(((S)-1-(4-(4-methylthiazol-5-yl)phenyl)ethyl)carbamoyl)pyrrolidin-1-yl)-3,3-dimethyl-1-oxobutan-2-yl)carbamate (54) Under nitrogen atmosphere, to a stirred solution of (S)-2-((tertbutoxycarbonyl)amino)-3,3-dimethylbutanoic acid (0.953 g, 4.123mmol) in dry DCM (10.5 mL), DIPEA (1.8 mL, 10.307 mmol) and HATU(1.834 g, 4.824 mmol) were added at 0 C. After 1 h, 52 (0.910 g, 2.473mmol) was added slowly. The mixture was stirred for 18 h at roomtemperature. Then, DCM (50 mL) was added to the reaction mixture andwas washed with 10 % citric acid (12 mL) twice, saturated NaHCO3solution (12 mL) twice, water (12 mL) twice, and brine (20 mL) once.The organic phase was dried over anhydrous Na2SO4 and concentratedunder reduced pressure to afford the desired compound as a yellow oil(1.873 g) which was used directly for the next step. |

| Yield | Reaction Conditions | Operation in experiment |
|---|---|---|
| Multi-step reaction with 4 steps 1: potassium acetate; palladium diacetate / N,N-dimethyl acetamide / 2 h / 120 °C 2: hydrogenchloride / methanol / 2 h / 20 °C 3: N-ethyl-N,N-diisopropylamine; HATU / N,N-dimethyl-formamide / 12 h / 20 °C 4: hydrogenchloride / methanol / 2 h / 20 °C | ||
| Multi-step reaction with 4 steps 1: potassium acetate; palladium diacetate / N,N-dimethyl-formamide / 16 h / 80 °C / Inert atmosphere 2: hydrogenchloride / 1,4-dioxane / 25 °C / Inert atmosphere 3: HATU; N-ethyl-N,N-diisopropylamine / N,N-dimethyl-formamide / 0 - 25 °C / Inert atmosphere 4: hydrogenchloride / 1,4-dioxane / 16 h / 25 °C / Inert atmosphere | ||
| Multi-step reaction with 4 steps 1: potassium acetate; palladium diacetate / N,N-dimethyl acetamide / 7 h / 90 °C / Inert atmosphere 2: hydrogenchloride / water; 1,4-dioxane / 5 h / 20 °C 3: N-ethyl-N,N-diisopropylamine; N-[(dimethylamino)-3-oxo-1H-1,2,3-triazolo[4,5-b]pyridin-1-yl-methylene]-N-methylmethanaminium hexafluorophosphate / N,N-dimethyl-formamide / 1.5 h / 20 °C 4: hydrogenchloride / dichloromethane; water; 1,4-dioxane; methanol / 6 h / 20 °C |
| Multi-step reaction with 4 steps 1: palladium diacetate; potassium acetate / N,N-dimethyl acetamide / 2 h / 120 °C / Inert atmosphere 2: hydrogenchloride / methanol / 2 h / 20 °C 3: N-ethyl-N,N-diisopropylamine; N-[(dimethylamino)-3-oxo-1H-1,2,3-triazolo[4,5-b]pyridin-1-yl-methylene]-N-methylmethanaminium hexafluorophosphate / N,N-dimethyl-formamide / 12 h / 20 °C 4: hydrogenchloride / methanol / 2 h / 20 °C | ||
| Multi-step reaction with 4 steps 1: palladium diacetate; potassium acetate / N,N-dimethyl-formamide / 4 h / 100 °C / Inert atmosphere 2: hydrogenchloride / ethyl acetate / 12 h / 20 °C 3: N-ethyl-N,N-diisopropylamine; N-[(dimethylamino)-3-oxo-1H-1,2,3-triazolo[4,5-b]pyridin-1-yl-methylene]-N-methylmethanaminium hexafluorophosphate / N,N-dimethyl-formamide / 3 h / 20 °C 4: methanol; hydrogenchloride / ethyl acetate / 20 °C | ||
| Multi-step reaction with 4 steps 1: palladium diacetate; potassium acetate / N,N-dimethyl acetamide / 110 °C / Inert atmosphere 2: hydrogenchloride / 1,4-dioxane; dichloromethane / 1 h / 20 °C 3: triethylamine; N-[(dimethylamino)-3-oxo-1H-1,2,3-triazolo[4,5-b]pyridin-1-yl-methylene]-N-methylmethanaminium hexafluorophosphate / dichloromethane / 2 h / 20 °C 4: hydrogenchloride / 1,4-dioxane; dichloromethane / 1 h / 20 °C | ||
| Multi-step reaction with 4 steps 1: potassium acetate; palladium diacetate / N,N-dimethyl acetamide / 4 h / 130 °C / Inert atmosphere 2: hydrogenchloride / 1,4-dioxane; dichloromethane / 18 h / 0 - 20 °C 3: N-[(dimethylamino)-3-oxo-1H-1,2,3-triazolo[4,5-b]pyridin-1-yl-methylene]-N-methylmethanaminium hexafluorophosphate; triethylamine / N,N-dimethyl-formamide; dichloromethane / 18 h / 0 - 20 °C / Inert atmosphere 4: hydrogenchloride / 1,4-dioxane / 18 h / 0 - 20 °C | ||
| Multi-step reaction with 4 steps 1.1: potassium acetate; palladium diacetate / 12 h / 95 °C / Inert atmosphere 2.1: hydrogenchloride / methanol; 1,4-dioxane / 2 h / 0 - 20 °C 3.1: N-[(dimethylamino)-3-oxo-1H-1,2,3-triazolo[4,5-b]pyridin-1-yl-methylene]-N-methylmethanaminium hexafluorophosphate / N,N-dimethyl-formamide / 20 min / Cooling with ice 3.2: 20 °C 4.1: hydrogenchloride / methanol; 1,4-dioxane / 2 h / 0 - 20 °C |

| Yield | Reaction Conditions | Operation in experiment |
|---|---|---|
| Multi-step reaction with 3 steps 1: hydrogenchloride / methanol / 2 h / 20 °C 2: N-ethyl-N,N-diisopropylamine; HATU / N,N-dimethyl-formamide / 12 h / 20 °C 3: hydrogenchloride / methanol / 2 h / 20 °C | ||
| Multi-step reaction with 3 steps 1: hydrogenchloride / 1,4-dioxane / 25 °C / Inert atmosphere 2: HATU; N-ethyl-N,N-diisopropylamine / N,N-dimethyl-formamide / 0 - 25 °C / Inert atmosphere 3: hydrogenchloride / 1,4-dioxane / 16 h / 25 °C / Inert atmosphere | ||
| Multi-step reaction with 3 steps 1: hydrogenchloride / water; 1,4-dioxane / 5 h / 20 °C 2: N-ethyl-N,N-diisopropylamine; N-[(dimethylamino)-3-oxo-1H-1,2,3-triazolo[4,5-b]pyridin-1-yl-methylene]-N-methylmethanaminium hexafluorophosphate / N,N-dimethyl-formamide / 1.5 h / 20 °C 3: hydrogenchloride / dichloromethane; water; 1,4-dioxane; methanol / 6 h / 20 °C |
| Multi-step reaction with 3 steps 1: hydrogenchloride / methanol / 2 h / 20 °C 2: N-ethyl-N,N-diisopropylamine; N-[(dimethylamino)-3-oxo-1H-1,2,3-triazolo[4,5-b]pyridin-1-yl-methylene]-N-methylmethanaminium hexafluorophosphate / N,N-dimethyl-formamide / 12 h / 20 °C 3: hydrogenchloride / methanol / 2 h / 20 °C | ||
| Multi-step reaction with 3 steps 1: hydrogenchloride / ethyl acetate / 12 h / 20 °C 2: N-ethyl-N,N-diisopropylamine; N-[(dimethylamino)-3-oxo-1H-1,2,3-triazolo[4,5-b]pyridin-1-yl-methylene]-N-methylmethanaminium hexafluorophosphate / N,N-dimethyl-formamide / 3 h / 20 °C 3: methanol; hydrogenchloride / ethyl acetate / 20 °C | ||
| Multi-step reaction with 3 steps 1: hydrogenchloride / 1,4-dioxane; dichloromethane / 1 h / 20 °C 2: triethylamine; N-[(dimethylamino)-3-oxo-1H-1,2,3-triazolo[4,5-b]pyridin-1-yl-methylene]-N-methylmethanaminium hexafluorophosphate / dichloromethane / 2 h / 20 °C 3: hydrogenchloride / 1,4-dioxane; dichloromethane / 1 h / 20 °C | ||
| Multi-step reaction with 3 steps 1: hydrogenchloride / 1,4-dioxane; dichloromethane / 18 h / 0 - 20 °C 2: N-[(dimethylamino)-3-oxo-1H-1,2,3-triazolo[4,5-b]pyridin-1-yl-methylene]-N-methylmethanaminium hexafluorophosphate; triethylamine / N,N-dimethyl-formamide; dichloromethane / 18 h / 0 - 20 °C / Inert atmosphere 3: hydrogenchloride / 1,4-dioxane / 18 h / 0 - 20 °C | ||
| Multi-step reaction with 3 steps 1.1: hydrogenchloride / methanol; 1,4-dioxane / 2 h / 0 - 20 °C 2.1: N-[(dimethylamino)-3-oxo-1H-1,2,3-triazolo[4,5-b]pyridin-1-yl-methylene]-N-methylmethanaminium hexafluorophosphate / N,N-dimethyl-formamide / 20 min / Cooling with ice 2.2: 20 °C 3.1: hydrogenchloride / methanol; 1,4-dioxane / 2 h / 0 - 20 °C | ||
| Multi-step reaction with 3 steps 1.1: hydrogenchloride / dichloromethane; water / 2 h / 0 - 20 °C 2.1: N-ethyl-N,N-diisopropylamine / N,N-dimethyl-formamide / 0.08 h / 20 °C 2.2: 12 h / 0 - 20 °C 3.1: hydrogenchloride / dichloromethane; water / 2 h / 0 - 20 °C |

| Yield | Reaction Conditions | Operation in experiment |
|---|---|---|
| Multi-step reaction with 2 steps 1: N-ethyl-N,N-diisopropylamine; HATU / N,N-dimethyl-formamide / 12 h / 20 °C 2: hydrogenchloride / methanol / 2 h / 20 °C | ||
| Multi-step reaction with 2 steps 1: HATU; N-ethyl-N,N-diisopropylamine / N,N-dimethyl-formamide / 0 - 25 °C / Inert atmosphere 2: hydrogenchloride / 1,4-dioxane / 16 h / 25 °C / Inert atmosphere | ||
| Multi-step reaction with 2 steps 1: N-ethyl-N,N-diisopropylamine; N-[(dimethylamino)-3-oxo-1H-1,2,3-triazolo[4,5-b]pyridin-1-yl-methylene]-N-methylmethanaminium hexafluorophosphate / N,N-dimethyl-formamide / 1.5 h / 20 °C 2: hydrogenchloride / dichloromethane; water; 1,4-dioxane; methanol / 6 h / 20 °C |
| Multi-step reaction with 2 steps 1: N-ethyl-N,N-diisopropylamine; N-[(dimethylamino)-3-oxo-1H-1,2,3-triazolo[4,5-b]pyridin-1-yl-methylene]-N-methylmethanaminium hexafluorophosphate / N,N-dimethyl-formamide / 12 h / 20 °C 2: hydrogenchloride / methanol / 2 h / 20 °C | ||
| Multi-step reaction with 2 steps 1: N-[(dimethylamino)-3-oxo-1H-1,2,3-triazolo[4,5-b]pyridin-1-yl-methylene]-N-methylmethanaminium hexafluorophosphate; N-ethyl-N,N-diisopropylamine / dichloromethane / 16 h / 0 - 20 °C 2: hydrogenchloride / water; 1,4-dioxane; methanol / 2 h / 0 - 20 °C | ||
| Multi-step reaction with 2 steps 1: N-ethyl-N,N-diisopropylamine; N-[(dimethylamino)-3-oxo-1H-1,2,3-triazolo[4,5-b]pyridin-1-yl-methylene]-N-methylmethanaminium hexafluorophosphate / N,N-dimethyl-formamide / 3 h / 20 °C 2: methanol; hydrogenchloride / ethyl acetate / 20 °C | ||
| Multi-step reaction with 2 steps 1: N-[(dimethylamino)-3-oxo-1H-1,2,3-triazolo[4,5-b]pyridin-1-yl-methylene]-N-methylmethanaminium hexafluorophosphate; N-ethyl-N,N-diisopropylamine / dichloromethane / 16 h / 0 - 20 °C 2: hydrogenchloride / 1,4-dioxane / 2 h / 20 °C | ||
| Multi-step reaction with 2 steps 1: triethylamine; N-[(dimethylamino)-3-oxo-1H-1,2,3-triazolo[4,5-b]pyridin-1-yl-methylene]-N-methylmethanaminium hexafluorophosphate / dichloromethane / 2 h / 20 °C 2: hydrogenchloride / 1,4-dioxane; dichloromethane / 1 h / 20 °C | ||
| Multi-step reaction with 2 steps 1: N-[(dimethylamino)-3-oxo-1H-1,2,3-triazolo[4,5-b]pyridin-1-yl-methylene]-N-methylmethanaminium hexafluorophosphate; triethylamine / N,N-dimethyl-formamide; dichloromethane / 18 h / 0 - 20 °C / Inert atmosphere 2: hydrogenchloride / 1,4-dioxane / 18 h / 0 - 20 °C |

| Yield | Reaction Conditions | Operation in experiment |
|---|---|---|
| Multi-step reaction with 2 steps 1: N-ethyl-N,N-diisopropylamine; HATU / N,N-dimethyl-formamide / 12 h / 20 °C 2: hydrogenchloride / methanol / 2 h / 20 °C | ||
| Multi-step reaction with 2 steps 1: N-ethyl-N,N-diisopropylamine; N-[(dimethylamino)-3-oxo-1H-1,2,3-triazolo[4,5-b]pyridin-1-yl-methylene]-N-methylmethanaminium hexafluorophosphate / N,N-dimethyl-formamide / 1.5 h / 20 °C 2: hydrogenchloride / dichloromethane; water; 1,4-dioxane; methanol / 6 h / 20 °C | ||
| Multi-step reaction with 2 steps 1: N-ethyl-N,N-diisopropylamine; N-[(dimethylamino)-3-oxo-1H-1,2,3-triazolo[4,5-b]pyridin-1-yl-methylene]-N-methylmethanaminium hexafluorophosphate / N,N-dimethyl-formamide / 12 h / 20 °C 2: hydrogenchloride / methanol / 2 h / 20 °C |
| Multi-step reaction with 2 steps 1: N-[(dimethylamino)-3-oxo-1H-1,2,3-triazolo[4,5-b]pyridin-1-yl-methylene]-N-methylmethanaminium hexafluorophosphate; N-ethyl-N,N-diisopropylamine / dichloromethane / 16 h / 0 - 20 °C 2: hydrogenchloride / water; 1,4-dioxane; methanol / 2 h / 0 - 20 °C | ||
| Multi-step reaction with 2 steps 1: N-ethyl-N,N-diisopropylamine; N-[(dimethylamino)-3-oxo-1H-1,2,3-triazolo[4,5-b]pyridin-1-yl-methylene]-N-methylmethanaminium hexafluorophosphate / N,N-dimethyl-formamide / 3 h / 20 °C 2: methanol; hydrogenchloride / ethyl acetate / 20 °C | ||
| Multi-step reaction with 2 steps 1: N-[(dimethylamino)-3-oxo-1H-1,2,3-triazolo[4,5-b]pyridin-1-yl-methylene]-N-methylmethanaminium hexafluorophosphate; N-ethyl-N,N-diisopropylamine / dichloromethane / 16 h / 0 - 20 °C 2: hydrogenchloride / 1,4-dioxane / 2 h / 20 °C | ||
| Multi-step reaction with 2 steps 1: triethylamine; N-[(dimethylamino)-3-oxo-1H-1,2,3-triazolo[4,5-b]pyridin-1-yl-methylene]-N-methylmethanaminium hexafluorophosphate / dichloromethane / 2 h / 20 °C 2: hydrogenchloride / 1,4-dioxane; dichloromethane / 1 h / 20 °C | ||
| Multi-step reaction with 2 steps 1: N-[(dimethylamino)-3-oxo-1H-1,2,3-triazolo[4,5-b]pyridin-1-yl-methylene]-N-methylmethanaminium hexafluorophosphate; triethylamine / N,N-dimethyl-formamide; dichloromethane / 18 h / 0 - 20 °C / Inert atmosphere 2: hydrogenchloride / 1,4-dioxane / 18 h / 0 - 20 °C | ||
| Multi-step reaction with 3 steps 1: 1-ethyl-(3-(3-dimethylamino)propyl)-carbodiimide hydrochloride; benzotriazol-1-ol; N-ethyl-N,N-diisopropylamine / dichloromethane; N,N-dimethyl-formamide / 18 h / -10 - 20 °C 2: potassium carbonate; Trimethylacetic acid; [1,3-bis(2,6-diisopropylphenyl)imidazol-2-ylidene](3-chloropyridyl)palladium(ll) dichloride / N,N-dimethyl acetamide / 125 °C 3: hydrogenchloride / methanol / 2 h / 4 °C | ||
| Multi-step reaction with 2 steps 1: N-[(dimethylamino)-3-oxo-1H-1,2,3-triazolo[4,5-b]pyridin-1-yl-methylene]-N-methylmethanaminium hexafluorophosphate; triethylamine / dichloromethane / 2 h / 25 °C 2: hydrogenchloride / dichloromethane; 1,4-dioxane / 1 h / 20 °C | ||
| Multi-step reaction with 2 steps 1.1: N-ethyl-N,N-diisopropylamine / N,N-dimethyl-formamide / 0.08 h / 20 °C 1.2: 12 h / 0 - 20 °C 2.1: hydrogenchloride / dichloromethane; water / 2 h / 0 - 20 °C |

| Yield | Reaction Conditions | Operation in experiment |
|---|---|---|
| Multi-step reaction with 2 steps 1: N-ethyl-N,N-diisopropylamine; HATU / N,N-dimethyl-formamide / 12 h / 20 °C 2: hydrogenchloride / methanol / 2 h / 20 °C | ||
| Multi-step reaction with 2 steps 1: triethylamine; HATU / N,N-dimethyl-formamide / 0 - 25 °C / Inert atmosphere 2: hydrogenchloride / 1,4-dioxane / 16 h / 25 °C / Inert atmosphere | ||
| Multi-step reaction with 2 steps 1: N-ethyl-N,N-diisopropylamine; N-[(dimethylamino)-3-oxo-1H-1,2,3-triazolo[4,5-b]pyridin-1-yl-methylene]-N-methylmethanaminium hexafluorophosphate / N,N-dimethyl-formamide / 12 h / 20 °C 2: hydrogenchloride / methanol / 2 h / 20 °C |
| Multi-step reaction with 2 steps 1: N-ethyl-N,N-diisopropylamine; N-[(dimethylamino)-3-oxo-1H-1,2,3-triazolo[4,5-b]pyridin-1-yl-methylene]-N-methylmethanaminium hexafluorophosphate / N,N-dimethyl-formamide / 3 h / 20 °C 2: methanol; hydrogenchloride / ethyl acetate | ||
| Multi-step reaction with 2 steps 1.1: N-[(dimethylamino)-3-oxo-1H-1,2,3-triazolo[4,5-b]pyridin-1-yl-methylene]-N-methylmethanaminium hexafluorophosphate; N-ethyl-N,N-diisopropylamine / dichloromethane / 1 h / 0 °C / Inert atmosphere 1.2: 18 h / 20 °C / Inert atmosphere 2.1: hydrogenchloride / methanol; 1,4-dioxane; dichloromethane / 0 - 20 °C | ||
| Multi-step reaction with 3 steps 1: sodium hydroxide / methanol; dichloromethane; water / 4 °C / pH 12.5 - 13 2: 1-ethyl-(3-(3-dimethylamino)propyl)-carbodiimide hydrochloride; benzotriazol-1-ol; N-ethyl-N,N-diisopropylamine / dichloromethane; N,N-dimethyl-formamide / -10 - 20 °C 3: hydrogenchloride / methanol / 2 h / 4 °C |

| Yield | Reaction Conditions | Operation in experiment |
|---|---|---|
| Multi-step reaction with 5 steps 1: triethylamine / dichloromethane / 2 h / 20 °C 2: potassium acetate; palladium diacetate / N,N-dimethyl acetamide / 2 h / 120 °C 3: hydrogenchloride / methanol / 2 h / 20 °C 4: N-ethyl-N,N-diisopropylamine; HATU / N,N-dimethyl-formamide / 12 h / 20 °C 5: hydrogenchloride / methanol / 2 h / 20 °C | ||
| Multi-step reaction with 5 steps 1: sodium hydrogencarbonate / ethyl acetate; water / 20 °C / Cooling with ice 2: potassium acetate; palladium diacetate / N,N-dimethyl acetamide / 7 h / 90 °C / Inert atmosphere 3: hydrogenchloride / water; 1,4-dioxane / 5 h / 20 °C 4: N-ethyl-N,N-diisopropylamine; N-[(dimethylamino)-3-oxo-1H-1,2,3-triazolo[4,5-b]pyridin-1-yl-methylene]-N-methylmethanaminium hexafluorophosphate / N,N-dimethyl-formamide / 1.5 h / 20 °C 5: hydrogenchloride / dichloromethane; water; 1,4-dioxane; methanol / 6 h / 20 °C | ||
| Multi-step reaction with 5 steps 1: triethylamine / dichloromethane / 2 h / 20 °C 2: palladium diacetate; potassium acetate / N,N-dimethyl acetamide / 2 h / 120 °C / Inert atmosphere 3: hydrogenchloride / methanol / 2 h / 20 °C 4: N-ethyl-N,N-diisopropylamine; N-[(dimethylamino)-3-oxo-1H-1,2,3-triazolo[4,5-b]pyridin-1-yl-methylene]-N-methylmethanaminium hexafluorophosphate / N,N-dimethyl-formamide / 12 h / 20 °C 5: hydrogenchloride / methanol / 2 h / 20 °C |
| Multi-step reaction with 5 steps 1: triethylamine / dichloromethane / 12 h / 20 °C 2: palladium diacetate; potassium acetate / N,N-dimethyl-formamide / 4 h / 100 °C / Inert atmosphere 3: hydrogenchloride / ethyl acetate / 12 h / 20 °C 4: N-ethyl-N,N-diisopropylamine; N-[(dimethylamino)-3-oxo-1H-1,2,3-triazolo[4,5-b]pyridin-1-yl-methylene]-N-methylmethanaminium hexafluorophosphate / N,N-dimethyl-formamide / 3 h / 20 °C 5: methanol; hydrogenchloride / ethyl acetate / 20 °C | ||
| Multi-step reaction with 5 steps 1: triethylamine / dichloromethane / 0 - 20 °C 2: palladium diacetate; potassium acetate / N,N-dimethyl acetamide / 110 °C / Inert atmosphere 3: hydrogenchloride / 1,4-dioxane; dichloromethane / 1 h / 20 °C 4: triethylamine; N-[(dimethylamino)-3-oxo-1H-1,2,3-triazolo[4,5-b]pyridin-1-yl-methylene]-N-methylmethanaminium hexafluorophosphate / dichloromethane / 2 h / 20 °C 5: hydrogenchloride / 1,4-dioxane; dichloromethane / 1 h / 20 °C | ||
| Multi-step reaction with 5 steps 1: sodium hydrogencarbonate / water / 18 h / 20 °C 2: potassium acetate; palladium diacetate / N,N-dimethyl acetamide / 4 h / 130 °C / Inert atmosphere 3: hydrogenchloride / 1,4-dioxane; dichloromethane / 18 h / 0 - 20 °C 4: N-[(dimethylamino)-3-oxo-1H-1,2,3-triazolo[4,5-b]pyridin-1-yl-methylene]-N-methylmethanaminium hexafluorophosphate; triethylamine / N,N-dimethyl-formamide; dichloromethane / 18 h / 0 - 20 °C / Inert atmosphere 5: hydrogenchloride / 1,4-dioxane / 18 h / 0 - 20 °C | ||
| Multi-step reaction with 3 steps 1: 1-ethyl-(3-(3-dimethylamino)propyl)-carbodiimide hydrochloride; benzotriazol-1-ol; N-ethyl-N,N-diisopropylamine / dichloromethane; N,N-dimethyl-formamide / 18 h / -10 - 20 °C 2: potassium carbonate; Trimethylacetic acid; [1,3-bis(2,6-diisopropylphenyl)imidazol-2-ylidene](3-chloropyridyl)palladium(ll) dichloride / N,N-dimethyl acetamide / 125 °C 3: hydrogenchloride / methanol / 2 h / 4 °C | ||
| Multi-step reaction with 4 steps 1.1: sodium hydrogencarbonate / water / 2 h / 0 °C / Inert atmosphere 1.2: 12 h / 90 °C / Inert atmosphere 2.1: hydrogenchloride / dichloromethane; water / 2 h / 0 - 20 °C 3.1: N-ethyl-N,N-diisopropylamine / N,N-dimethyl-formamide / 0.08 h / 20 °C 3.2: 12 h / 0 - 20 °C 4.1: hydrogenchloride / dichloromethane; water / 2 h / 0 - 20 °C |

| Yield | Reaction Conditions | Operation in experiment |
|---|---|---|
| 84% | With N-ethyl-N,N-diisopropylamine; HATU In N,N-dimethyl-formamide at 15℃; for 1h; | 5; 6 Step 5: Preparation of (2S,4R)-1-(2-(5-(2,2-dimethoxyethoxy)-1-methyl-1H-pyrazol- 3-yl)-3-methylbutanoyl)-4-hydroxy-N-((S)-1-(4-(4-methylthiazol-5-yl)phenyl)ethyl)pyrrolidine- 2-carboxamide To a solution of (2S,4R)-4-hydroxy-N-[(1S)-1-[4-(4-methylthiazol-5-yl)phenyl]ethyl] pyrrolidine-2-carboxamide (334 mg, 0.90 mmol, 1 eq, Hydrochloride) 2-[5-(2,2- dimethoxyethoxy)-1-methyl-pyrazol-3-yl]-3-methyl-butanoic acid (260 mg, 0.90 mmol, 1 eq) in dimethylformamide (5 mL) was added O-(7-Azabenzotriazol-1-yl)-N,N,N’,N’- tetramethyluronium hexafluorophosphate (517 mg, 1.36 mmol, 1.5 eq) and N,N- diisopropylethylamine (352 mg, 2.72 mmol, 3 eq) .The mixture was stirred at 15 °C for 1 hr. The reaction mixture was diluted with water 30 mL and extracted with ethyl acetate (50 mL x 3). The combined organic layers were washed with brine (50 mL x 3), dried over anhydrous sodium sulfate, filtered and concentrated under reduced pressure. The residue was purified by flash silica gel chromatography (Eluent of 0~10% dichloromethane/methanol). Compound (2S,4R)-1-[2-[5- (2,2-dimethoxyethoxy) -1-methyl-pyrazol-3-yl]-3-methyl-butanoyl]-4-hydroxy-N-[(1S)-1-[4-(4- methylthiazol-5-yl)phenyl]ethyl]pyrrolidine-2-carboxamide (300 mg, 0.47 mmol, 52.33% yield, 95% purity) was obtained as a white gum. LC/MS (ESI) m/z: 600.3 [M+1] +; 1H-NMR (400MHz, CDCl3) d 8.68 (s, 1H), 7.44 - 7.30 (m, 5H), 5.57 - 5.53 (m, 1H), 4.82 - 4.95 (m, 1H), 4.74 - 4.53 (m, 3H), 4.11 - 3.95 (m, 2H), 3.93 - 3.82 (m, 1H), 3.58 (d, J = 3.2 Hz, 4H), 3.45 - 3.42 (m, 3H), 3.41 (s, 2H), 3.38 (s, 2H), 2.53 (d, J = 1.6 Hz, 3H), 2.39 (s, 1H), 1.48 (d, J = 7.2 Hz, 1H), 1.32 (d, J = 7.2 Hz, 3H), 1.03 (d, J = 6.6 Hz, 3H), 0.94 - 0.86 (m, 2H), 0.86 - 0.77 (m, 3H). |
| Yield | Reaction Conditions | Operation in experiment |
|---|---|---|
| With benzotriazol-1-ol; 1-ethyl-(3-(3-dimethylamino)propyl)-carbodiimide hydrochloride; N-ethyl-N,N-diisopropylamine In N,N-dimethyl-formamide at 25℃; for 12h; | 10; 11 Step 10: Preparation of tert-butyl 4-(3-(1-((2S,4R)-4-hydroxy-2-(((S)-1-(4-(4- methylthiazol-5-yl)phenyl)ethyl)carbamoyl)pyrrolidin-1-yl)-3-methyl-1-oxobutan-2-yl)-1- methyl-1H-pyrazol-5-yl)piperazine-1-carboxylate To a mixture of 2-[5-(4-tert-butoxycarbonylpiperazin-1-yl)-1-methyl-pyrazol-3-yl]-3- methyl- butanoic acid (330 mg, 0.79 mmol, 1 eq) in dimethylformamide (5 mL) was added diisopropylethylamine (306 mg, 2.37 mmol, 3 eq) , 1-hydroxybenzotriazole (128 mg, 0.95 mmol, 1.2 eq) and 1-(3-dimethylaminopropyl)-3-ethylcarbodiimide (227 mg, 1.19 mmol, 1.5 eq) in one portion at 25 °C. The mixture was stirred at 25 °C for 10 min, then (2S,4R)-4-hydroxy-N-[(1S)- 1-[4-(4-methylthiazol-5-yl)phenyl]ethyl]pyrrolidine-2-carboxamide (291 mg, 0.79 mmol, 1 eq, hydrochloride) was added, the mixture was stirred at 25 °C for 11 hour 50 min. Water (40 mL) was poured into the mixture and stirred for 1 min. The aqueous phase was extracted with ethyl acetate (20 mL x 3). The combined organic phase was washed with brine (20 mL x 3), dried with anhydrous sodium sulfate, filtered and concentrated in vacuum. The residue was purified by silica gel column chromatography (dichloromethane/ methanol=200/1, 40/1). The product tert- butyl 4-[5-[1-[(2S,4R)-4- hydroxy-2-[[(1S)-1-[4-(4-methylthiazol-5- yl)phenyl]ethyl]carbamoyl]pyrrolidine-1-carbonyl]-2-methyl-propyl]-2-methyl-pyrazol-3- yl]piperazine-1-carboxylate (360 mg, 0.44 mmol, 56% yield, 84% purity) was obtained as a colorless oil. LC/MS (ESI) m/z: 680.3 [M+1] +. |
| Yield | Reaction Conditions | Operation in experiment |
|---|---|---|
| 76% | Stage #1: (2S,4R)-4-hydroxy-N-((S)-1-(4-(4-methylthiazol-5-yl)phenyl)ethyl)pyrrolidine-2-carboxamide hydrochloride; 2-(3-(4-(4-(4-(3-(2,6-difluoro-3-(((R)-3-fluoropyrrolidine)-1-sulfonamido)benzoyl)-1H-pyrrolo[2,3-b]pyridin-5-yl)phenyl)piperazin-1-yl)butoxy)isoxazol-5-yl)-3-methylbutanoic acid With benzotriazol-1-ol; 1-ethyl-(3-(3-dimethylamino)propyl)-carbodiimide hydrochloride; triethylamine In N,N-dimethyl-formamide at 30℃; for 12h; Stage #2: formic acid | 5; 6 Step 5: Preparation of (2S,4R)-1-(2-(3-(4-(4-(4-(3-(2,6-difluoro-3-(((R)-3- fluoropyrrolidine)-1-sulfonamido)benzoyl)-1H-pyrrolo[2,3-b]pyridin-5-yl)phenyl)piperazin-1- yl)butoxy)isoxazol-5-yl)-3-methylbutanoyl)-4-hydroxy-N-((S)-1-(4-(4-methylthiazol-5- yl)phenyl)ethyl)pyrrolidine-2-carboxamide To a solution of 2-[3-[4-[4-[4-[3-[2,6-difluoro-3-[[(3R)-3-fluoropyrrolidin-1- yl]sulfonylamino] benzoyl]-1H-pyrrolo[2,3-b]pyridin-5-yl]phenyl]piperazin-1- yl]butoxy]isoxazol-5-yl]-3-methyl-butanoic acid (200 mg, 0.24 mmol, 1 eq) and (2S,4R)-4- hydroxy-N-[(1S)-1-[4-(4- methylthiazol-5-yl)phenyl]ethyl]pyrrolidine-2-carboxamide (89. mg, 0.24 mmol, 1.00 eq, HCl) in N,N-dimethylformamide (5 mL) was added triethylamine (36 mg, 0.36 mmol, 1.5 eq), 1-(3-dimethylaminopropyl)-3-ethylcarbodiimide hydrochloride (55 mg, 0.29 mmol, 1.2 eq) and hydroxybenzotriazole (39 mg, 0.29 mmol, 1.2 eq). The mixture was stirred at 30 °C for 12 hours. Water (50 mL) was poured into the mixture and stirred for 1 minute. The aqueous phase was extracted with dichloromethane (20 mL x 3). The combined organic phase was washed with brine (20 mL x 2), dried with anhydrous sodium sulfate, filtered and concentrated in vacuum. The residue was purified by prep-HPLC. Compound (2S,4R)-1-[2-[3- [4-[4-[4-[3-[2,6-difluoro-3-[[(3R)-3-fluoropyrrolidin-1-yl]sulfonylamino]benzoyl]-1H- pyrrolo[2,3-b]pyridin-5-yl]phenyl]piperazin-1-yl]butoxy]isoxazol-5-yl]-3-methyl-butanoyl]-4- hydroxy-N-[(1S)-1-[4-(4-methylthiazol-5-yl)phenyl]ethyl]pyrrolidine-2-carboxamide (130 mg, 0.10 mmol, 43% yield, 96% purity, formate) was obtained as a white solid. LC/MS (ESI) m/z: 569.4 [M/2+1] +. |
| Yield | Reaction Conditions | Operation in experiment |
|---|---|---|
| 1: 68% 2: 76% | With N-ethyl-N,N-diisopropylamine; HATU In N,N-dimethyl-formamide at 15℃; for 0.5h; | 21; 22 Step 21: Preparation of tert-butyl 4-(5-(1-((2S,4R)-4-hydroxy-2-(((S)-1-(4-(4- methylthiazol-5-yl)phenyl)ethyl)carbamoyl)pyrrolidin-1-yl)-3-methyl-1-oxobutan-2-yl)isoxazol- 3-yl)piperazine-1-carboxylate To a solution of 2-[3-(4-tert-butoxycarbonylpiperazin-1-yl)isoxazol-5-yl]-3-methyl- butanoic acid (100 mg, 0.28 mmol, 1 eq) and o-(7-azabenzotriazol-1-yl)-n,n,n',n'- tetramethyluronium hexafluorophosphate (129 mg, 0.33 mmol, 1.2 eq) in dimethyl formamide (5 mL) were added (2S,4R)-4-hydroxy-N-[(1S)-1-[4-(4-methylthiazol-5- yl)phenyl]ethyl]pyrrolidine-2-carboxamide (104 mg, 0.28 mmol, 1 eq, Hydrochloride) and diisopropyl ethyl amine (109 mg, 0.84 mmol, 3 eq). The reaction mixture was stirred at 15°C for 0.5 h. The reaction mixture was poured into water (20 mL), extracted with ethyl acetate (30 mL x 3). The combined organic layers were washed with brine (50 mL x 3), dried over anhydrous sodium sulfate, filtered and concentrated under reduced pressure. The residue was purified by prep-TLC (dichloromethane: methanol = 10:1). tert-butyl 4-[5-[1-[(2S,4R)-4-hydroxy-2-[[(1S)- 1-[4-(4-methylthiazol-5-yl)phenyl]ethyl]carbamoyl]pyrrolidine-1-carbonyl]-2-methyl- propyl]isoxazol-3-yl]piperazine-1-carboxylate (180 mg, 0.26 mmol, 92% yield, 97% purity) was obtained as a colorless oil. LC/MS (ESI) m/z: 667.4 [M+1] +; 1H-NMR (400MHz, CDCl3) d 8.70 (d, J=1.2 Hz, 1H), 8.03 (s, 1H), 7.45 - 7.39 (m, 3H), 7.38 - 7.31 (m, 2H), 5.94 (d, J=10.0 Hz, 1H), 5.13 - 4.95 (m, 1H), 4.79 (dd, J=4.4, 8.4 Hz, 1H), 4.70 - 4.59 (m, 2H), 3.84 - 3.71 (m, 1H), 3.67 - 3.55 (m, 4H), 3.54 - 3.47 (m, 5H), 3.21 (td, J=5.2, 13.2 Hz, 4H), 2.98 (s, 4H), 2.90 (s, 4H), 2.82 (s, 6H), 2.55 (d, J=1.6 Hz, 4H), 2.45 (ddd, J=6.4, 9.6, 16.0 Hz, 1H), 2.08 - 1.92 (m, 1H), 1.53 - 1.37 (m, 13H), 1.06 (dd, J=2.4, 6.4 Hz, 3H), 0.94 (d, J=6.8 Hz, 3H). |
| Yield | Reaction Conditions | Operation in experiment |
|---|---|---|
| 1: 12% 2: 8% | Stage #1: (2S,4R)-4-hydroxy-N-((S)-1-(4-(4-methylthiazol-5-yl)phenyl)ethyl)pyrrolidine-2-carboxamide hydrochloride; 5-(4-(3-(2,6-difluoro-3-(((R)-3-fluoropyrrolidine)-1-sulfonamido)benzoyl)-1H-pyrrolo[2,3-b]pyridin-5-yl)phenoxy)-2-(3-methylisoxazol-5-yl)pentanoic acid With benzotriazol-1-ol; N-ethyl-N,N-diisopropylamine In N,N-dimethyl-formamide at 15℃; for 0.5h; Stage #2: With 1-ethyl-(3-(3-dimethylamino)propyl)-carbodiimide hydrochloride In N,N-dimethyl-formamide at 40℃; for 2h; | 9 To a solution of 5-[4-[3-[2,6-difluoro-3-[[(3R)-3-fluoropyrrolidin-1- yl]sulfonylamino] benzoyl]-1H-pyrrolo[2,3-b]pyridin-5-yl]phenoxy]-2-(3-methylisoxazol-5- yl)pentanoic acid (130 mg, 0.19 mmol, 1 eq) and (2S,4R)-4-hydroxy-N-[(1S)-1-[4-(4- methylthiazol-5-yl)phenyl]ethyl] pyrrolidine-2-carboxamide (69 mg, 0.19 mmol, 1 eq, HCl) in N,N-dimethylformamide (5 mL) was added 1-hydroxybenzotriazole (31 mg, 0.23 mmol, 1.2 eq) and N,N-diisopropylethylamine (73 mg, 0.56 mmol, 3 eq), and the mixture was stirred at 15°C for 30 min, and N-(3-dimethylaminopropyl)-N-ethylcarbodiimide hydrochloride (43 mg, 0.23 mmol, 1.2 eq) was added into the mixture, then stirred at 40°C for 2 h. The mixture was concentrated in the vacuum. The residue was purified by pre-HPLC. The first product, (2S,4R)- 1-[(2S)-5-[4-[3-[2,6-difluoro-3-[[(3R)-3-fluoropyrrolidin-1-yl] sulfonylamino]benzoyl]-1H- pyrrolo[2,3-b]pyridin-5-yl]phenoxy]-2-(3-methylisoxazol-5-yl)pentanoyl]-4-hydroxy-N-[(1S)-1- [4-(4-methylthiazol-5-yl)phenyl]ethyl]pyrrolidine-2-carboxamide (23.7 mg, 0.02 mmol, 12 % yield, 99 % purity), was obtained as white solid. LC/MS (ESI) m/z: 1011.3 [M+1] +; 1H-NMR (300MHz, DMSO-d6) d 9.04 - 8.91 (m, 1H), 8.65 (s, 1H), 8.56 (s, 1H), 8.41 (d, J=7.5 Hz, 1H), 8.07 (s, 1H), 7.72 - 7.55 (m, 3H), 7.49 - 7.31 (m, 5H), 7.21 (s, 1H), 7.13 - 7.00 (m, 2H), 6.36 - 6.03 (m, 1H), 5.40 - 5.19 (m, 1H), 5.13 (s, 1H), 5.00 - 4.84 (m, 1H), 4.48 - 4.37 (m, 1H), 4.31 (s, 1H), 4.18 (t, J=7.1 Hz, 1H), 4.11 - 3.95 (m, 2H), 3.78 (dd, J=4.5, 10.5 Hz, 1H), 3.58 (d, J=11.4 Hz, 1H), 3.45 (s, 1H), 2.47 - 2.39 (m, 4H), 2.27 - 2.16 (m, 4H), 2.15 - 2.00 (m, 4H), 2.00 - 1.88 (m, 2H), 1.88 - 1.65 (m, 3H), 1.25 - 0.98 (m, 1H), 1.38 (d, J=6.8 Hz, 3H). The second product, (2S,4R)-1-[(2R)-5-[4-[3-[2,6-difluoro-3-[[(3R)-3-fluoropyrrolidin-1-yl] sulfonylamino]benzoyl]- 1H-pyrrolo[2,3-b]pyridin-5-yl]phenoxy]-2-(3-methylisoxazol-5-yl)pentanoyl]-4-hydroxy-N- [(1S)-1-[4-(4-methylthiazol-5-yl)phenyl]ethyl]pyrrolidine-2-carboxamide (15.7 mg, 0.02 mmol, 8 % yield, 98 % purity), was obtained as white solid. LC/MS (ESI) m/z: 1011.3 [M+1] +; 1H- NMR (300MHz, DMSO-d6) d 9.04 - 8.94 (m, 1H), 8.65 (d, J=2.1 Hz, 1H), 8.56 (s, 1H), 8.40 (d, J=7.7 Hz, 1H), 8.05 (s, 1H), 7.67 (d, J=8.7 Hz, 2H), 7.62 - 7.53 (m, 1H), 7.48 - 7.41 (m, 2H), 7.40 - 7.30 (m, 3H), 7.21 - 7.01 (m, 1H), 7.11 - 7.01 (m, 2H), 6.28 - 6.22 (m, 1H), 6.33 (s, 1H), 5.43 - 5.19 (m, 1H), 5.17 (s, 1H), 5.03 (s, 1H), 4.92 (t, J=7.3 Hz, 1H), 4.65 - 4.56 (m, 1H), 4.54 - 4.37 (m, 1H), 4.29 (s, 1H), 4.21 (t, J=7.2 Hz, 1H), 4.06 (t, J=6.1 Hz, 2H), 4.02 - 3.90 (m, 1H), 3.73 (t, J=7.7 Hz, 1H), 3.60 (d, J=10.0 Hz, 1H), 2.47 - 2.43 (m, 4H), 2.33 (d, J=1.8 Hz, 1H), 2.25 - 2.16 (m, 4H), 2.13 - 1.91 (m, 5H), 1.78 (dd, J=7.5, 12.5 Hz, 3H), 1.26 - 1.13 (m, 1H), 1.44 - 1.10 (m, 5H). |
| Yield | Reaction Conditions | Operation in experiment |
|---|---|---|
| 1: 28% 2: 11% | Stage #1: (2S,4R)-4-hydroxy-N-((S)-1-(4-(4-methylthiazol-5-yl)phenyl)ethyl)pyrrolidine-2-carboxamide hydrochloride; 6-(4-(3-(2,6-difluoro-3-(((R)-3-fluoropyrrolidine)-1-sulfonamido)benzoyl)-1H-pyrrolo[2,3-b]pyridin-5-yl)phenoxy)-2-(3-methylisoxazol-5-yl)hexanoic acid With benzotriazol-1-ol; N-ethyl-N,N-diisopropylamine In N,N-dimethyl-formamide at 15℃; for 0.5h; Stage #2: With 1-ethyl-(3-(3-dimethylamino)propyl)-carbodiimide hydrochloride In N,N-dimethyl-formamide at 40℃; for 2h; | 6 To a solution of 6-[4-[3-[2,6-difluoro-3-[[(3R)-3-fluoropyrrolidin-1- yl]sulfonylamino]benzoyl] -1H-pyrrolo[2,3-b]pyridin-5-yl]phenoxy]-2-(3-methylisoxazol-5- yl)hexanoic acid (110 mg, 0.16 mmol, 1 eq) and (2S,4R)-4-hydroxy-N-[(1S)-1-[4-(4- methylthiazol-5-yl)phenyl]ethyl] pyrrolidine-2-carboxamide (57 mg, 0.16 mmol, 1 eq, HCl) in N,N-dimethylformamide (5 mL) was added 1-hydroxybenzotriazole (25 mg, 0.19 mmol, 1.2 eq) and N,N-diisopropylethylamine (60 mg, 0.47 mmol, 3 eq), and the mixture was stirred at 15 °C for 30 min, and N-(3-dimethylaminopropyl)-N-ethylcarbodiimide hydrochloride (36 mg, 0.19 mmol, 1.2 eq) was added into the mixture, then stirred at 40 °C for 2 h. The mixture was concentrated in the vacuum. The residue was purified by pre-HPLC. The product 1 (2S,4R)-1- [(2S)-6-[4-[3-[2,6-difluoro-3-[[(3R)-3-fluoropyrrolidin-1-yl] sulfonylamino]benzoyl]-1H- pyrrolo[2,3-b]pyridin-5-yl]phenoxy]-2-(3-methylisoxazol-5-yl)hexanoyl]-4-hydroxy-N-[(1S)-1- [4-(4-methylthiazol-5-yl)phenyl]ethyl]pyrrolidine-2-carboxamide (45.2 mg, 0.04 mmol, 28% yield, 98% purity) was obtained as white solid. LC/MS (ESI) m/z: 1025.3 [M+1] +; 1H-NMR (400MHz, DMSO-d6) d 9.04 - 8.92 (m, 1H), 8.64 (d, J=2.0 Hz, 1H), 8.55 (s, 1H), 8.38 (d, J=8.0 Hz, 1H) , 8.06 (s, 1H), 7.75 - 7.54 (m, 3H), 7.52 - 7.27 (m, 4H), 7.19 (t, J=8.8 Hz, 1H), 7.11 - 6.99 (m, 2H), 6.34 - 6.02 (m, 1H), 5.38 - 5.04 (m, 2H), 5.01 - 4.80 (m, 1H), 4.76 - 4.58 (m, 1H), 4.51 - 4.35 (m, 1H), 4.30 (s, 1H), 4.10 (t, J=7.6 Hz, 1H), 4.05 - 3.86 (m, 3H), 3.76 (dd, J=4.4, 10.0 Hz, 1H), 3.56 - 3.49 (m, 2H), 3.51 - 3.43 (m, 1H), 2.47 - 2.35 (m, 5H), 2.26 - 2.14 (m, 3H), 2.13 - 1.89 (m, 4H), 1.88 - 1.64 (m, 4H), 1.59 - 1.09 (m, 5H). The product 2 (2S,4R)-1-[(2R)-6- [4-[3-[2,6-difluoro-3-[[(3R)-3-fluoropyrrolidin-1-yl] sulfonylamino]benzoyl]-1H-pyrrolo[2,3- b]pyridin-5-yl]phenoxy]-2-(3-methylisoxazol-5-yl)hexanoyl]-4-hydroxy-N-[(1S)-1-[4-(4- methylthiazol-5-yl)phenyl]ethyl]pyrrolidine-2-carboxamide (18.6 mg, 0.02 mmol, 11% yield, 97% purity) was obtained as white solid. LC/MS (ESI) m/z: 1026.2 [M+1] +; 1H-NMR (400MHz, DMSO-d6) d 9.02 - 8.94 (m, 1H), 8.65 (d, J=1.6 Hz, 1H), 8.56 (s, 1H), 8.38 (d, J=7.6 Hz, 1H), 8.20 (s, 1H), 8.07 (s, 1H), 7.72 - 7.56 (m, 3H), 7.48 - 7.37 (m, 3H), 7.36 - 7.27 (m, 2H), 7.22 (t, J=8.8 Hz, 1H), 7.07 (d, J=8.8 Hz, 2H), 6.34 - 6.17 (m, 1H), 5.35 (s, 1H), 5.22 (s, 1H), 5.13 (s, 1H), 5.04 - 4.86 (m, 1H), 4.51 - 4.38 (m, 1H), 4.28 (s, 1H), 4.16 - 3.90 (m, 2H), 3.73 (d, J=7.6 Hz, 1H), 3.62 - 3.51 (m, 1H), 3.46 (s, 2H), 2.42 (d, J=8.8 Hz, 5H), 2.27 - 2.14 (m, 3H), 2.07 (d, J=15.2 Hz, 2H), 2.12 - 2.01 (m, 1H), 1.99 - 1.88 (m, 2H), 1.87 - 1.69 (m, 3H), 1.52 - 1.35 (m, 5H). |