Patent ID: 12240849

DETAILED DESCRIPTION

The present disclosure will be described in detail through examples below, but it is not intended to adversely limit the present disclosure. The present disclosure has been described in detail herein, in which the specific embodiments thereof have also been disclosed. For those skilled in the art, it will be obvious to make various variations and improvements to the specific embodiments of the present disclosure without departing from the spirits and scope of the present disclosure.

Synthetic Route:

Step 1: Synthesis of Compound 001-02

001-02B (473.30 mg, 5.50 mmol, 1.1 eq), 001-02A (1.0 g, 5.00 mmol, 4.99 mL, 1.0 eq), potassium carbonate (1.73 g, 12.50 mmol, 2.5 eq) and N, N-dimethylformamide (2 mL) were added into a flask and stirred at 90° C. for 16 h. The reaction solution was diluted with 5 mL of dichloromethane and filtered, the solid was soaked in 5 mL of dichloromethane, and the obtained solution was stirred at 50° C. for 15 min and filtered hot, and two filtrates were combined and concentrated, diluted with 20 mL of ethyl acetate, and washed with 20 mL of saturated sodium sulfate for three times. The organic phase was dried with anhydrous sodium sulfate for 30 min, and dried by a spinning method to obtain compound 001-02.

Step 2: Synthesis of Compound 001-3

001-01 (150 mg, 731.04 μmol, 1 eq), 001-02 (278.93 mg, 1.10 mmol, 1.5 eq), titanium tetraisopropoxide (2 mL), sodium cyanoborohydride (229.70 mg, 3.66 mmol, 5.0 eq) and ethylene glycol dimethyl ether (10 mL) were added into a 10 mL flask, and stirred at 70° C. for 10 h. 20 mL of water was added, insoluble substances were removed by filtration, the filtrate was separated to obtain an organic phase, the organic phase was dried by a spinning method to obtain a crude product, and the crude product was purified by column chromatography (ethyl acetate:petroleum ether=0-33%) to obtain compound 001-03. LCMS: MS (ESI) m/z: 444.2 [M+1]+.

Step 3: Synthesis of Compound 001-04

001-03 (160 mg, 338.18 μmol, 1 eq) was added into a 50 mL flask, hydrogen chloride/dioxane (1 mL) was added, and the reaction solution was stirred at 25° C. for 6 h. The reaction solution was dried by a spinning method to obtain a crude product 001-04. LCMS: MS (ESI) m/z: 344.1 [M+1]+.

Step 4: Synthesis of Compound 001 and Compound 002

001-05 (50 mg, 223.55 μmol, 1.0 eq), 001-04 (115.17 mg, 335.33 μmol, 1.5 eq), diisopropylethylamine (130.01 mg, 1.01 mmol, 175.22 μL, 4.5 eq) and isopropanol (2 mL) were added into a 50 mL flask, and subjected to microwave reaction for 1 h at 120° C. The reaction solution was dried by a spinning method to obtain a crude product. The crude product was initially purified by column chromatography (methanol:dichloromethane=0-5%) to obtain 001-h, and 001-h was separated by a chiral preparative column (chromatographic column: DAICEL CHIRALPAK AS (250 mm×30 mm, 10 μm); mobile phase: [A: CO2B: 0.1% ammonia water ethanol]; B %: 50%-50%) to obtain target compounds namely compound 001 and compound 002.

Compound 001 (peak position: 0.949 min)

1H NMR (400 MHz, deuterated methanol) δ ppm 1.16-1.46 (m, 5H) 1.55 (br s, 3H) 1.67 (br s, 3H) 2.19 (s, 3H) 2.31 (s, 2H) 2.36 (brs, 2H) 2.64 (br s, 1H) 3.44-3.68 (m, 3H) 3.71-3.86 (m, 2H) 4.64 (s, 2H) 6.06 (br s, 1H) 6.22 (br s, 1H) 7.68 (d, J=4.27 Hz, 1H) 7.89-7.98 (m, 2H) 8.40 (s, 1H) 8.50 (d, J=4.27 Hz, 1H)

LCMS: MS (ESI) m/z: 531.1 [M+1]+.

Compound 002 (peak position: 2.405 min)

1H NMR (400 MHz, deuterated methanol) δ ppm 1.16-1.46 (m, 5H) 1.55 (br s, 3H) 1.67 (br s, 3H) 2.19 (s, 3H) 2.31 (s, 2H) 2.36 (br s, 2H) 2.64 (br s, 1H) 3.44-3.68 (m, 31H) 3.71-3.86 (m, 2H) 4.64 (s, 2H) 6.06 (br s, 1H) 6.22 (br s, 1H) 7.68 (d, J=4.27 Hz, 1H) 7.89-7.98 (m, 2H) 8.40 (s, 1H) 8.50 (d, J=4.27 Hz, 1H) LCMS: MS (ESI) m/z: 531.1 [M+1]+.

Example 2

Synthetic Route:

Step 1: Synthesis of Compound 003-02

Compound 001-02 (200 mg, 832.15 μmol, 1.5 eq), 003-01 (113.83 mg, 554.77 μmol, 1 eq) and sodium cyanoborohydride (104.59 mg, 1.66 mmol, 3 eq) were added into a mixture of tetraisopropyl titanate (5 mL) and dichloroethane (5 mL), air in the reaction system was evacuated and nitrogen was filled for protection, and the reaction solution was stirred at 70° C. for 3.5 h. 20 mL of water was added, insoluble substances were removed by filtration, the filtrate was separated to obtain an organic phase, and the organic phase was dried by a spinning method to obtain a crude product. The crude product was purified by a chromatography column (dichloromethane/methanol=1/0 to 20/1) to obtain compound 003-02. LCMS (ESI) m/z: 430.2[M+1]+.

Step 2: Synthesis of Compound 003-03

Compound 003-02 (220 mg, 512.19 μmol, 1 eq) was added to dichloromethane (2.5 mL), trifluoroacetic acid (1.36 g, 11.89 mmol, 880.00 μL, 23.21 eq) was added, and the reaction solution was stirred at 25° C. for 1.5 h. The reaction solution was directly dried by a spinning method, 10 mL of dichloromethane was added, and the reaction solution was dried by a spinning method, and the above operations were repeated for three times. Compound 003-03 was obtained without further purification. LCMS (ESI) m/z: 330.1 [M+1]+.

Step 2: Synthesis of Compound 003 and Compound 004

Compound 003-03 (120 mg, 364.28 μmol, 2 eq) and compound 001-05 (40.74 mg, 182.14 μmol, 1 eq) were added into n-butanol (3 mL), N,N-diisopropylethylamine (1.11 g, 8.61 mmol, 1.50 mL, 47.28 eq) was added, and the reaction solution was heated at 130° C. and stirred for 16 h. The reaction solution was directly dried by a spinning method to obtain a crude product. The crude product was purified by a chromatographic column (dichloromethane/methanol=20/1) to obtain compound 003-h. Compound 003-h was subjected to chiral resolution (column: DAICEL CHIRALCEL OJ-H (250 mm×30 mm, 5 m); mobile phase: [A: CO2B: 0.1% ammonia water EtOH]; B %: 35%-35%), to obtain target compounds namely compound 003 and compound 004.

Compound 003 (peak position: 4.488 min)

LCMS (ESI) m/z: 517.1 [M+1]+

1H NMR (400 MHz, deuterated methanol) δ ppm 1.45 (d, J=6.53 Hz, 3H) 1.60 (q, J=6.02 Hz, 4H) 1.70-1.82 (m, 2H) 2.19 (s, 3H) 2.28 (s, 3H) 2.40 (d, J=9.29 Hz, 1H) 2.47-2.62 (m, 2H) 2.72-2.83 (m, 1H) 3.57-3.71 (m, 2H) 3.73-3.93 (m, 2H) 6.04 (s, 1H) 5.92-6.08 (m, 1H) 6.15 (s, 1H) 7.69 (d, J=4.02 Hz, 1H) 7.86-8.02 (m, 2H) 8.39 (d, J=1.25 Hz, 1H) 8.51 (d, J=4.27 Hz, 1H)

Compound 004 (peak position: 4.755 min)

LCMS (ESI) m/z: 517.1 [M+1]+

1H NMR (400 MHz, deuterated methanol) δ ppm 1.45 (d, J=6.53 Hz, 3H) 1.60 (q, J=6.19 Hz, 4H) 1.69-1.81 (m, 2H) 2.19 (s, 3H) 2.28 (s, 3H) 2.39 (d, J=9.79 Hz, 1H) 2.45-2.60 (m, 2H) 2.73-2.83 (m, 1H) 3.60-3.70 (m, 2H) 3.73-3.93 (m, 2H) 6.04 (s, 1H) 5.92-6.08 (m, 1H) 6.14 (s, 1H) 7.67-7.72 (m, 1H) 7.89-8.00 (m, 2H) 8.39 (d, J=1.51 Hz, 1H) 8.51 (dd, J=4.52, 0.75 Hz, 1H)

Example 3

Synthetic Route:

Step 1: Synthesis of Compound 005-01

Compound 001-02 was dissolved into a mixture of dichloromethane (5 mL) and methanol (10 mL), sodium borohydride (276.57 mg, 7.31 mmol, 3 eq) was added at 20° C., and the reaction solution was reacted for 0.5 h. The reaction solution was concentrated under reduced pressure at 43° C. The concentrate was dissolved into 30 mL of water and 30 mL of dichloromethane, and extracted, and meanwhile a water phase was extracted with dichloromethane (30 mL×3). Organic phases were combined and dried by a spinning method to obtain a crude product. The crude product was purified by a chromatographic column (petroleum ether/ethyl acetate=3:1) to obtain compound 005-01, for using in the next step.

Step 2: Synthesis of Compound 005-02

Compound 005-01 was dissolved into dichloromethane (5 mL), dibromosulfoxide (5.02 g, 24.13 mmol, 1.87 mL, 10 eq) was added dropwise at 0° C., and the mixture was stirred at 50° C. for 3 h. The reaction solution was concentrated under reduced pressure at 43° C. The concentrate was dissolved into 10 mL of dichloromethane, and concentrated under reduced pressure. The above operations were repeated for three times. The resulting crude product was purified by a chromatographic column (petroleum ether/ethyl acetate=5/1) to obtain compound 005-02. LCMS (ESI) m/z: 269.8 [M+1]+.

Step 3: Synthesis of Compound 005-04

Compound 005-03 (155.36 mg, 610.88 μmol, 1.1 eq), sodium hydride (26.65 mg, 666.42 μmol, 60% purity, 1.2 eq) were added into N,N-dimethylformamide (3 mL) at 0° C., air in the reaction system was evacuated and nitrogen was filled for protection, compound 005-02 (150 mg, 555.35 μmol, 1 eq) was added after stirring the reaction solution for 10 min, and the reaction solution was stirred for 2.5 h at 20° C. Water was added into 10 mL of the reaction solution, and a water phase was extracted for three times with 10 mL of ethyl acetate. The organic phase was collected, dried with anhydrous sodium sulfate, and dried by a spinning method to obtain a crude product. The crude product was purified by a chromatographic column (petroleum ether/ethyl acetate=3/1) to obtain compound 005-04. LCMS (ESI) m/z: 344.3 [M-Boc+1]+.

Step 4: Synthesis of Compound 005-05

Compound 005-04 (100 mg, 225.47 μmol, 1 eq) was added into dichloromethane (5 mL), trifluoroacetic acid (2.57 g, 22.51 mmol, 1.67 mL, 99.84 eq) was added, and reaction solution was stirred at 25° C. for 1.5 h. The reaction solution was directly dried by a spinning method, 10 mL of dichloromethane was added for drying by a spinning method, and the above operations were repeated for 3 times. Compound 005-05 was obtained without further purification. LCMS (ESI) m/z: 344.4 [M+1]+.

Step 5: Synthesis of Compounds 005 and 006

Compound 005-05 (50 mg, 145.60 μmol, 1 eq) and compound 001-05 (32.57 mg, 145.60 μmol, 1 eq) were added into n-butanol (3 mL), N,N-diisopropylethylamine (742.00 mg, 5.74 mmol, 1 mL, 39.43 eq) was added, and the reaction solution was heated and stirred at 130° C. for 16 h. The reaction solution was directly dried by a spinning method to obtain a crude product. The crude product is purified by a high performance liquid chromatographic column (column: Waters Xbridge BEH C18 150×25 mm, 5 μm; mobile phase: [water (10 mM ammonium bicarbonate)-acetonitrile]; acetonitrile %: 37%-67%, 9.5 min) to obtain compound 005-h. Compound 005-h was separated by a chiral preparative column: (DAICEL CHIRALPAK AS-H (250 mm×30 mm, 5 μm); mobile phase: [A: CO2, B: 0.1% ammonia water-ethanol]; B %: 50%-50%) to obtain target compounds namely compound 005 and compound 006.

005-h

1H NMR (400 MHz, deuterated methanol) δ ppm 1.42 (d, J=13.80 Hz, 1H) 1.48-1.57 (m, 1H) 1.62 (d, J=7.28 Hz, 3H) 1.85 (qd, J=12.51, 4.39 Hz, 2H) 2.01-2.16 (m, 2H) 2.20 (s, 3H) 2.26 (s, 3H) 3.04-3.21 (m, 3H) 3.43-3.60 (m, 1H) 4.51-4.69 (m, 2H) 5.42 (q, J=6.94 Hz, 1H) 5.87-6.45 (m, 2H) 7.69 (d, J=4.02 Hz, 1H) 7.86-7.91 (m, 1H) 7.91-7.97 (m, 1H) 8.37 (d, J=1.76 Hz, 1H) 8.51 (d, J=4.27 Hz, 1H)

Target compound 005 (peak position: 4.323)

LCMS (ESI) m/z: 531.5 [M+1]+

1H NMR (400 MHz, deuterated methanol) δ ppm 1.42 (d, J=13.80 Hz, 1H) 1.48-1.57 (m, 1H) 1.62 (d, J=7.28 Hz, 3H) 1.85 (qd, J=12.51, 4.39 Hz, 2H) 2.01-2.16 (m, 2H) 2.20 (s, 3H) 2.26 (s, 3H) 3.04-3.21 (m, 3H) 3.43-3.60 (m, 1H) 4.51-4.69 (m, 2H) 5.42 (q, J=6.94 Hz, 1H) 5.87-6.45 (m, 2H) 7.69 (d, J=4.02 Hz, 1H) 7.86-7.91 (m, 1H) 7.91-7.97 (m, 1H) 8.37 (d, J=1.76 Hz, 1H) 8.51 (d, J=4.27 Hz, 1H)

Target compound 006 (peak position: 5.496)

LCMS (ESI) m/z: 531.5 [M+1]+

1H NMR (400 MHz, deuterated methanol) δ ppm 1.42 (d, J=13.80 Hz, 1H) 1.48-1.57 (m, 1H) 1.62 (d, J=7.28 Hz, 3H) 1.85 (qd, J=12.51, 4.39 Hz, 2H) 2.01-2.16 (m, 2H) 2.20 (s, 3H) 2.26 (s, 3H) 3.04-3.21 (m, 3H) 3.43-3.60 (m, 1H) 4.51-4.69 (m, 2H) 5.42 (q, J=6.94 Hz, 1H) 5.87-6.45 (m, 2H) 7.69 (d, J=4.02 Hz, 1H) 7.86-7.91 (m, 1H) 7.91-7.97 (m, 1H) 8.37 (d, J=1.76 Hz, 1H) 8.51 (d, J=4.27 Hz, 1H)

Example 4

Synthetic Route:

Step 1: Synthesis of Compound 007-03

007-01 (323.96 mg, 2.50 mmol, 1.2 eq) and N,N-dimethylformamide (0.2 mL) were added into a 10 mL flask, nitrogen was pumped and replaced for three times, sodium hydride (125.03 mg, 3.13 mmol, 60% purity, 1.5 eq) was added at 0° C., the reaction solution was stirred at 25° C. for 30 min, and 005-02 (500 mg, 2.08 mmol, 1 eq) was added and stirred for 6 h. The reaction solution was washed with 10 mL of water and extracted with ethyl acetate (10 mL×3). The organic phases were combined, dried with anhydrous sodium sulfate, and dried by a spinning method to obtain a crude product. The crude product was purified by column chromatography to obtain compound 007-03. LCMS: MS (ESI) m/z: 358.4 [M-Boc+1]+.

Step 2: Synthesis of Compound 007-04

007-03 (140 mg, 305.98 μmol, 1 eq), a hydrochloric acid/dioxane solution (4 M, 764.96 μL, 10 eq) and dioxane (0.5 mL) were added into a 10 mL flask, and stirred at 25° C. for 6 h. The reaction solution was dried by a spinning method to obtain a crude product 007-04.

LCMS: MS (ESI) m/z: 358.3 [M+1]+.

Step 3: Synthesis of Compound 007-06

001-05 (62.58 mg, 279.78 μmol, 1.0 eq), 007-04 (100 mg, 279.78 μmol, 1.0 eq), diisopropylethylamine (162.71 mg, 1.26 mmol, 219.29 L, 4.5 eq) and isopropanol (2 mL) were added into a 50 mL flask, and stirred at 120° C. for 30 min under microwave. The reaction solution was dried by a spinning method, and purified by preparative chromatography (chromatographic column: Agela ASB 150×25 mm×5 μm; mobile phase: [water (0.05% HCl)-ACN]; acetonitrile %: 25%-45%, 10 min) to obtain 007-06. 007-06 was separated by a chiral preparative column (separation conditions: chromatographic column: DAICEL CHIRALPAK AS (250 mm×30 mm, 10 μm); mobile phase: [A: CO2B: 0.1% ammonia EtOH]; B %: 55%-55%) to obtain target compound 007 and compound 008.

Target compound 007 (peak position: 4.002 min)1H NMR (400 MHz, deuterated methanol) δ ppm 1.57 (br s, 1H) 1.60 (d, J=7.28 Hz, 3H) 1.65 (br s, 1H) 1.76-1.94 (m, 3H) 1.99-2.09 (m, 2H) 2.09-2.20 (m, 2H) 2.22 (s, 3H) 2.28 (s, 3H) 2.92-3.01 (m, 1H) 3.21-3.30 (m, 2H) 3.34-3.39 (m, 1H) 4.41-4.54 (m, 2H) 5.98 (q, J=7.28 Hz, 1H) 6.06-6.23 (m, 1H) 7.71 (d, J=4.27 Hz, 1H) 7.82-7.89 (m, 1H) 7.91-7.97 (m, 1H) 8.35 (d, J=2.26 Hz, 1H) 8.52 (d, J=4.52 Hz, 11H).

LCMS: MS (ESI) m/z: 545.0 [M+1]+.

Target compound 008 (peak position: 6.072 min)

1H NMR (400 MHz, deuterated methanol) δ ppm 1.57 (br s, 1H) 1.60 (d, J=7.28 Hz, 3H) 1.65 (br s, 1H) 1.76-1.94 (m, 31H) 1.99-2.09 (m, 2H) 2.09-2.20 (m, 2H) 2.22 (s, 3H) 2.28 (s, 3H) 2.92-3.01 (m, 1H) 3.21-3.30 (m, 2H) 3.34-3.39 (m, 1H) 4.41-4.54 (m, 2H) 5.98 (q, J=7.28 Hz, 1H) 6.06-6.23 (m, 1H) 7.71 (d, J=4.27 Hz, 1H) 7.82-7.89 (m, 1H) 7.91-7.97 (m, 1H) 8.35 (d, J=2.26 Hz, 1H) 8.52 (d, J=4.52 Hz, 1H).

LCMS: MS (ESI) m/z: 545.4 [M+1]+.

Example 5

Synthetic Route:

Step 1: Synthesis of Compound 009-03

009-1 (20 g, 93.35 mmol, 1 eq) was dissolved into tetrahydrofuran (100 mL) under nitrogen atmosphere at −78° C., lithium diisopropylamide (2 M, 51.34 mL, 1.1 eq) was slowly dropwise added, the reaction solution was stirred for 1 h at 0° C. and cooled to −78° C., bromopropionitrile (18.76 g, 140.02 mmol, 11.51 mL, 1.5 eq) was added, and the reaction solution was stirred at 25° C. for 16 h. 50 mL of a saturated ammonium chloride solution was added into the reaction solution. The reaction solution was extracted with 100 mL of ethyl acetate for three times, and organic phases were combined and dried by a spinning method to obtain a crude product. The crude product was purified by column chromatography to obtain 009-03. LCMS: MS (ESI) m/z: [M+1]+268.2

Step 2: Synthesis of Compound 009-04

009-03 (12 g, 44.89 mmol, 1 eq), methanol (100 mL), raney nickel (384.57 mg, 4.49 mmol, 0.1 eq) and potassium carbonate (18.61 g, 134.67 mmol, 3.0 eq) were added into a IL hydrogenation flask, and reacted for 16 h at 45° C. under a hydrogen pressure of 45 psi. The reaction solution was filtered to remove insoluble substances. The filtrate was dried by a spinning method to obtain a crude product, and the crude product was purified by column chromatography to obtain 009-04. LCMS: MS (ESI) m/z: 226.1 [M+1]+

Step 3: Synthesis of Compound 009-06

009-04 (2.2 g, 9.77 mmol, 1 eq), sodium hydride (468.70 mg, 11.72 mmol, 60% purity, 1.2 eq) and N,N-dimethylformamide (10 mL) were added at 0° C. and stirred for 30 min, and 005-02 (2.95 g, 10.94 mmol, 1.12 eq) was added. The reaction solution was stirred at 25° C. for 16 h. The reaction solution was spin-dried by an oil pump, 50 mL of water was added, and the reaction solution was extracted with 50 mL of ethyl acetate for three times, and organic phases were combined and dried by a spinning method to obtain a crude product. The crude product was purified by column chromatography to obtain 009-06. LCMS: MS (ESI) m/z: 415.1 [M+1]+.

Step 4: Synthesis of Compound 009-07

009-06 (3.5 g, 8.44 mmol, 1 eq) was dissolved into tetrahydrofuran (20 mL), hydrochloric acid (12 M, 7.04 mL, 10 eq) was added, and the reaction solution was stirred at 45° C. for 4 h. 100 mL of a saturated sodium carbonate solution was added in the reaction solution under the pH of 9-10, and the reaction solution was extracted with 100 mL of dichloromethane for three times. The organic phases were combined and dried by a spinning method to obtain a crude product. The crude product was purified by column chromatography to obtain 009-07. LCMS: MS (ESI) m/z: 371.0 [M+1]+.

Step 5: Synthesis of Compound 009-09

009-07 was added under nitrogen atmosphere at −78° C., tetrahydrofuran (40 mL) was added, lithium diisopropylamide (1 M, 5.67 mL, 1.4 eq) was slowly added dropwise, 009-08 was added after stirring the reaction solution for 30 min, and then the reaction solution was continued to stir for 16 h. 20 mL of saturated amine chloride was added into the reaction solution for quenching, and the reaction solution was extracted with 50 mL of dichloromethane for three times, and the organic phases were combined. The obtained crude product was purified by column chromatography to obtain 009-09. LCMS: MS (ESI) m/z: 503.1 [M+1]+.

Step 6: Synthesis of Compound 009-10

009-09 (1.63 g, 3.24 mmol, 1 eq), bis(pinacolato)diboron (823.75 mg, 3.24 mmol, 1 eq), [1,1′-bis(diphenylphosphine)ferrocene]palladium dichloride dichloromethane complex (397.36 mg, 486.59 μmol, 0.15 eq), potassium acetate (318.36 mg, 3.24 mmol, 1 eq) and dioxane (20 mL) were added into a 100 mL flask, nitrogen was pumped for three times, and the reaction solution was reacted for 16 h at 110° C. The reaction solution was filtered to remove insoluble substances, the filtrate was dried by a spinning method to obtain a crude product. The crude product was purified by column chromatography to obtain 009-10. LCMS: MS (ESI) m/z: 480.9 [M+1]+.

Step 7: Synthesis Compound 009-12

009-10 (500 mg, 1.04 mmol, 1 eq), 001-05 (246.76 mg, 1.10 mmol, 1.06 eq), [1,1′-bis(diphenylphosphine)ferrocene]palladium dichloride dichloromethane complex (85.00 mg, 104.00 μmol, 0.1 eq), potassium carbonate (431.20 mg, 3.12 mmol, 3.0 eq), dioxane (5 mL) and water (1.25 mL) were added into a 100 mL flask, nitrogen was pumped for three times, and the reaction solution was stirred for 2 h at 80° C. The reaction solution was filtered to remove insoluble substances, the filtrate was dried by a spinning method, 5 mL of methanol was added for dissolution, and the rustled solution was separated by preparative high performance liquid chromatography to obtain 009-12. Separation conditions were as below: column: Boston Green ODS 150×30 mm×5 μm; mobile phase: [water (0.075% trifluoroacetic acid)-acetonitrile]: acetonitrile %: 25%-55%, 12 min. LCMS: MS (ESI) m/z: 542.2 [M+1]+.

Step 8: Synthesis of Compounds 009-13 and 14

009-12 (130 mg, 240.02 μmol, 1 eq) was separated by a chiral column to obtain 009-13 and 14. Separation conditions were as below: column: DAICEL CHIRALPAK AS-H (250 mm×30 mm, 5 μm); mobile phase: [A: CO2, B: 0.1% ammonia water-methanol]; B %: 45%-45%.

013 LCMS: MS (ESI) m/z: 542.2 [M+1]+

014 LCMS: MS (ESI) m/z: 542.2 [M+1]+

Step 9: Synthesis of Compounds 009-15 and 009-16

009-13 (51 mg, 94.16 μmol, 1 eq), palladium on carbon (0.5 mg, 10% purity, 1.00 eq) and methanol (2 mL) were added into a 100 mL hydrogenation flask, and reacted for 16 h at 45° C. under the hydrogen pressure of 45 psi. The reaction solution was filtered to remove insoluble substances. The filtrate was dried by a spinning method to obtain a crude product, and the crude product was purified by column chromatography to obtain compound 009-15 or compound 009-16.

009-14 (51 mg, 94.16 μmol, 1 eq), palladium on carbon (0.5 mg, 10% purity, 1.00 eq) and methanol (2 mL) were added into a 100 mL hydrogenation flask, and reacted for 16 h at 45° C. under the hydrogen pressure of 45 psi. The reaction solution was filtered to remove insoluble substances. The filtrate was dried by a spinning method to obtain a crude product, and the crude product was purified by column chromatography to obtain compound 009-16 or compound 009-15.

009-15 LCMS: MS (ESI) m/z: 544.3 [M+1]+

009-16 LCMS: MS (ESI) m/z: 544.3 [M+1]+

Step 10: Synthesis of Compounds 009, 010, 011 and 012

009-15 (51 mg, 94.16 μmol, 1 eq) was separated in a chiral column to obtain compound 009 and compound 010. Separation condition were as below: column: DAICEL CHIRALPAK IC (250 mm×30 mm, 10 μm); mobile phase: [A: CO2, B: 0.1% ammonia water-EtOH]: B %: 50%-50%,

009 LCMS: MS (ESI) m/z: 544.5 [M+1]+

010 LCMS: MS (ESI) m/z: 544.5 [M+1]+

009-16 (71.0 mg, 130.60 μmol, 1 eq) was separated in a chiral column o obtain compound 011 and compound 012. Separation condition were as below: column: DAICEL CHIRALPAK AS (250 mm×30 mm, 10 μm); mobile phase: [A: CO2, B: 0.1% ammonia water-methanol]: B %:40%-40%.

011 LCMS: MS (ESI) m/z: 544.5 [M+1]+

012 LCMS: MS (ESI) m/z: 544.5[M+1]+

Target compound 009 (SFC peak position: 2.230)

1H NMR (400 MHz, deuterated methanol) δ ppm 1.22-1.34 (m, 2H) 1.47 (d, J=7.03 Hz, 3H) 1.64 (brd, J=4.77 Hz, 2H) 1.67-1.79 (m, 3H) 2.05-2.12 (m, 2H) 2.15 (s, 3H) 2.23 (s, 3H) 2.35 (br dd, J=19.95, 10.16 Hz, 2H) 2.65 (br s, 1H) 2.73-2.91 (m, 1H) 3.11-3.31 (m, 3H) 5.93 (br d, J=6.02 Hz, 1H) 6.48-6.76 (m, 1H) 7.57 (d, J=4.27 Hz, 1H) 7.69-7.89 (m, 2H) 8.23 (s, 1H) 8.39 (d, J=4.27 Hz, 1H)

Target compound 010 (SFC peak position: 3.943)

1H NMR (400 MHz, deuterated methanol) δ ppm 1.30-1.37 (m, 6H) 1.62 (d, J=7.28 Hz, 3H) 1.73-1.95 (m, 7H) 2.01-2.24 (m, 4H) 2.30 (s, 2H) 2.35 (s, 2H) 2.78 (brs, 1H) 2.87-3.01 (m, 1H) 6.01-6.36 (m, 1H) 6.74 (brs, 1H) 7.71 (d, J=3.76 Hz, 1H) 7.80-8.03 (m, 2H) 8.37 (s, 1H) 8.53 (d, J=4.52 Hz, 1H)

Target compound 011 (SFC peak position: 1.763)

1H NMR (400 MHz, deuterated methanol) δ ppm 1.22-1.34 (m, 2H) 1.47 (d, J=7.03 Hz, 3H) 1.64 (brd, J=4.77 Hz, 2H) 1.67-1.79 (m, 3H) 2.05-2.12 (m, 2H) 2.15 (s, 3H) 2.23 (s, 3H) 2.35 (br dd, J=19.95, 10.16 Hz, 2H) 2.65 (br s, 1H) 2.73-2.91 (m, 1H) 3.11-3.31 (m, 3H) 5.93 (brd, J=6.02 Hz, 1H) 6.48-6.76 (m, 1H) 7.57 (d, J=4.27 Hz, 1H) 7.69-7.89 (m, 2H) 8.23 (s, 1H) 8.39 (d, J=4.27 Hz, 1H)

Target compound 012 (SFC peak position: 4.660)

1H NMR (400 MHz, deuterated methanol) δ ppm 1.30-1.37 (m, 6H) 1.62 (d, J=7.28 Hz, 3H) 1.73-1.95 (m, 7H) 2.01-2.24 (m, 4H) 2.30 (s, 2H) 2.35 (s, 2H) 2.78 (br s, 1H) 2.87-3.01 (m, 1H) 6.01-6.36 (m, 1H) 6.74 (br s, 1H) 7.71 (d, J=3.76 Hz, 1H) 7.80-8.03 (m, 2H) 8.37 (s, 1H) 8.53 (d, J=4.52 Hz, 1H)

Example 6

Synthetic Route:

Step 1: Synthesis of Compound 013-02

Under nitrogen protection, compound 013-01 (53 g, 247.37 mmol, 1 eq) was dissolved into tetrahydrofuran (250 mL) at 0° C., lithium hexamethyldisilazide (1 M, 371.05 mL, 1.5 eq) was added, the reaction solution was stirred for 1 h, oxygen was introduced at 0° C., and then the reaction solution was stirred for 2 h. 250 mL of sodium sulfite was added into the reaction solution for quenching, the water phase was extracted for 4 times with 250 mL of ethyl acetate, and the organic phase was collected, dried with anhydrous sodium sulfate, and concentrated under reduced pressure to obtain a crude product. The obtained crude product was purified by a chromatographic column (petroleum ether/ethyl acetate=1/0 to 4/1) to obtain Compound 013-02.

Step 2: Synthesis of Compound 013-03

Compound 013-02 (9.2 g, 39.96 mmol, 1 eq) was dissolved into N,N-dimethylformamide (70 mL) at 0° C., sodium hydrogen (3.20 g, 79.91 mmol, 60% purity, 2 eq) was added, the reaction solution was stirred for 0.5 h, 3-bromopropene (14.50 g, 119.87 mmol, 3 eq) was slowly added dropwise, and then the reaction solution was heated to 20° C., and continued to stir for 1 h. The reaction solution was quenched with 50 mL of saturated ammonium chloride, and extracted with ethyl acetate (50 mL×3). The organic phases were combined, dried with anhydrous sodium sulfate, filtered, and concentrated under reduced pressure to obtain a crude product. The obtained crude product was separated and purified by column chromatography (silica gel, ethyl acetate:petroleum ether=1:20 to 1:5) to obtain compound 013-03.

Step 3: Synthesis of Compound 013-04

Compound 013-03 (20 g, 73.99 mmol, 1 eq) was dissolved into a mixture of dichloromethane (85 mL) and methanol (15 mL), sodium bicarbonate (9.32 g, 110.98 mmol, 4.32 mL, 1.5 eq) was added, the reaction solution was cooled to −78° C., ozone (3.55 g, 73.99 mmol, 1 eq) was introduced. The reaction solution was stirred for 0.5 h under a pressure of 15 psi, and then it turned blue, oxygen was continuously introduced until blue disappeared, triphenylphosphine (23.29 g, 88.78 mmol, 1.2 eq) was added, and the reaction solution was heated to 20° C. and continued to stir for 1 h. The reaction solution was directly concentrated under reduced pressure to obtain a crude product. The obtained crude product was separated and purified by column chromatography (silica gel, ethyl acetate:petroleum ether=1:10 to 1:1) to obtain Compound 013-04.

Step 4: Synthesis of Compound 013-05

N,N-dibenzylamine (4.97 g, 25.21 mmol, 4.83 mL, 0.8 eq) was dissolved into 1,2-dichloroethane (80 mL) at 0° C., sodium acetate borohydride (10.02 g, 47.27 mmol, 1.5 eq) and compound 013-04 (13 g, 31.51 mmol, 1 eq) were added, and the reaction solution was heated to 20° C. and reacted for 1 h under stirring. The reaction solution was quenched with 50 mL of saturated sodium bicarbonate, and extracted with ethyl acetate (50 mL×3). The organic phases were combined, dried with anhydrous sodium sulfate, filtered, and concentrated under reduced pressure to obtain a crude product. The obtained crude product was separated and purified by column chromatography (ethyl acetate:petroleum ether=1:20 to 1:1) to obtain compound 013-05.1H NMR (400 MHz, deuterated methanol) δ=7.37 (d, J=7.3 Hz, 4H), 7.32-7.26 (m, 4H), 7.24-7.18 (m, 2H), 4.11 (q, J=7.3 Hz, 2H), 3.92 (s, 4H), 3.65 (s, 4H), 3.40 (t, J=6.0 Hz, 2H), 2.67 (t, J=6.0 Hz, 2H), 1.99-1.86 (m, 4H), 1.76 (dt, J=5.8, 12.2 Hz, 2H), 1.61-1.52 (m, 2H), 1.19 (t, J=7.2 Hz, 3H).

Step 5: Synthesis of Compound 013-06

Compound 013-05 (10.6 g, 23.37 mmol, 1 eq) was dissolved into methanol (100 mL), palladium/carbon (1 g, 23.37 mmol, 10% purity, 1.00 eq) and potassium carbonate (6.46 g, 46.74 mmol, 2 eq) were added, hydrogen (94.42 mg, 46.74 mmol, 2 eq) was introduced, and the reaction solution was stirred for 10 h at 45° C. under the pressure of 45 psi. The reaction solution was filtered through a layer of diatomite and washed with dichloromethane, and the filtrate was concentrated under reduced pressure. The obtained residue was dissolved into a mixture of methanol (90 mL) and water (30 mL), potassium carbonate (6.46 g, 46.76 mmol, 2 eq) was added, and the reaction solution was stirred at 20° C. for 2 h. A thin-layer silica gel plate (silica gel plate, ethyl acetate:petroleum ether=1:0, Rf=0.30) showed that raw materials completely reacted to generate a target compound with reduced polarity. The reaction solution was concentrated under reduced pressure to remove methanol, the water phase was extracted with ethyl acetate, and organic phases were combined, dried, filtered, and concentrated under reduced pressure to obtain compound 013-06 which may be directly used in the next step without purification.1H NMR (400 MHz, deuterated chloroform) 6=7.03 (br s, 1H), 3.94 (s, 4H), 3.83 (t, J=5.0 Hz, 2H), 3.39 (brt, J=6.0 Hz, 2H), 2.24-2.11 (m, 2H), 1.94 (brd, J=13.3 Hz, 2H), 1.83 (dt, J=4.1, 13.5 Hz, 2H), 1.62 (brd, J=13.1 Hz, 2H).

Step 6: Synthesis of Compound 013-07

Compound 013-06 (1.7 g, 7.48 mmol, 1 eq) was dissolved into N,N-dimethylformamide (17 mL) at 0° C., sodium hydrogen (448.79 mg, 11.22 mmol, 60% purity, 1.5 eq) was added, the reaction solution was stirred for 1 h, compound 005-02 (2.22 g, 8.23 mmol, 1.1 eq) was added, and the reaction solution was heated to 20° C. and reacted for 1 h under stirring. After being quenched with 20 mL of saturated ammonium chloride, the reaction solution was extracted with ethyl acetate (30 mL×3). The organic phases were combined, washed with saturated salt solution (20 mL×2), dried with anhydrous sodium sulfate, filtered and concentrated under reduced pressure to obtain compound 013-07.

Step 7: Synthesis of Compound 013-08

Compound 013-07 (3.12 g, 7.49 mmol, 1 eq) was dissolved into tetrahydrofuran (60 mL), hydrochloric acid aqueous solution (1 M, 74.92 mL, 10 eq) was added, the reaction solution was heated to 45° C. and reacted for 1 h under stirring. After being quenched with saturated sodium bicarbonate, the reaction solution was extracted with ethyl acetate. The organic phases were combined, dried, filtered, and concentrated under reduced pressure to obtain a crude product. The obtained crude product was separated and purified by column chromatography (silica gel column, ethyl acetate:petroleum ether=1:1 to 2:1) To obtain compound 013-08.

Step 8: Synthesis of Compound 013-09

Compound 013-08 (2 g, 5.37 mmol, 1 eq) was dissolved into tetrahydrofuran (20 mL) at −78° C., potassium hexamethyldisilazide (1 M, 7.52 mL, 1.4 eq) was slowly added, the reaction solution was stirred for 0.5 h, compound 009-08 (2.53 g, 6.44 mmol, 1.2 eq) was added, and the reaction solution was continuously stirred for 1 h. The reaction solution was quenched with 20 mL of saturated ammonium chloride at −78° C. and heated to 20° C. The water phase was extracted with ethyl acetate (50 mL×3). The organic phases were combined, washed with a saturated salt solution (20 mL×2), dried with anhydrous sodium sulfate, filtered, and concentrated under reduced pressure to obtain a crude product. The obtained crude product was separated and purified by column chromatography (silica gel, ethyl acetate:petroleum ether=1:10 to 1:3) to obtain compound 013-09.

Step 9: Synthesis of Compound 013-10

Compound 013-09 (2.48 g, 4.92 mmol, 1 eq) and bis(pinacolato)diboron (1.25 g, 4.92 mmol, 1 eq) were dissolved into dioxane (25 mL), [1,]′-bis(diphenylphosphino)ferrocene]dichloropalladium(II)dichloromethane (401.48 mg, 491.62 μmol, 0.1 eq) and potassium acetate (1.45 g, 14.75 mmol, 3 eq) were added, nitrogen was replaced for three times, and the reaction solution was heated to 110° C. and reacted for 1 h under stirring. After being cooled, the reaction solution was directly concentrated under reduced pressure to obtain a crude product. The obtained crude product was separated and purified by column chromatography (silica gel, ethyl acetate:petroleum ethe=1:10 to 1:1, Rf=0.80) to obtain compound 013-10.

Step 10: Synthesis of Compound 013-11

Compound 013-10 (1 g, 2.07 mmol, 1 eq) and compound 001-05 (510.06 mg, 2.28 mmol, 1.1 eq) were dissolved into dioxane (12 mL) and water (3 mL), [1,1′-bis(diphenylphosphino)ferrocene]dichloropalladium (II) (151.69 mg, 207.32 μmol, 0.1 eq) and potassium phosphate (1.32 g, 6.22 mmol, 3 eq) were added, nitrogen was replaced for three times, the reaction solution was heated to 90° C. and reacted for 1 h under stirring. The reaction solution was diluted with 200 mL of dichloromethane and washed with a saturated salt solution (30 mL×3), and the organic phase was dried with anhydrous sodium sulfate, filtered, and concentrated under reduced pressure to obtain a crude product. The obtained crude product was separated and purified by column chromatography (silica gel, dichloromethane:methanol=100:0 to 20:1) to obtain compound 013-11. LCMS: MS (ESI) m/z: 544.4 [M+1]+.

Step 11: Synthesis of Compound 013-12

Compound 013-11 (500 mg, 919.80 μmol, 1 eq), palladium on carbon (100 mg, 91.98 μmol, 10% purity, 0.1 eq) and methanol (5 mL) were added into a hydrogenation flask, and heated and stirred for 16 h at 45° C. under the hydrogen (919.80 μmol) pressure of 45 psi. The reaction solution was filtered to remove palladium on carbon, and the filtrate was dried by a spinning method to obtain compound 013-12. LCMS: MS (ESI) m/z: 546.4 [M+1]+

Step 12: Synthesis of Compounds 013, 014, 015 and 016

Compound 013-12 (420 mg, 769.78 μmol, 1 eq) was separated by a chiral preparative column to obtain target compounds 013, 014, 015 and 016. Separation conditions were as below: column: DAICEL CHIRALPAK AD (250 mm×30 mm, 10 μm); mobile phase: [A: CO2, B: 0.1% ammonia water-isopropanol]: B %: 45%-45%.

Target compound: 013 (SFC peak position: 0.746)

1H NMR (400 MHz, deuterated methanol) δ ppm 1.48 (brd, J=7.13 Hz, 3H) 1.67 (br s, 2H) 1.81-2.02 (m, 6H) 2.16 (s, 3H) 2.20 (s, 3H) 2.64 (br s, 1H) 2.76-2.97 (m, 1H) 3.23-3.41 (m, 1H) 3.54-3.95 (m, 2H) 5.81 (q, J=6.92 Hz, 1H) 6.09 (br s, 1H) 6.56 (br s, 1H) 7.57 (d, J=4.13 Hz, 1H) 7.66-7.91 (m, 2H) 8.23 (s, 11H) 8.37 (d, J=4.25 Hz, 11H). LCMS: MS (ESI) m/z: 546.4 [M+1]+

Target compound: 014 (SFC peak position: 0.997)

1H NMR (400 MHz, deuterated methanol) 8 ppm 1.48 (brd, J=7.13 Hz, 3H) 1.67 (brs, 2H) 1.81-2.02 (m, 6H) 2.16 (s, 3H) 2.20 (s, 3H) 2.64 (brs, 1H) 2.76-2.97 (m, 1H) 3.23-3.41 (m, 1H) 3.54-3.95 (m, 2H) 5.81 (q, J=6.92 Hz, 1H) 6.09 (brs, 1H) 6.56 (brs, 1H) 7.57 (d, J=4.13 Hz, 1H) 7.66-7.91 (m, 2H) 8.23 (s, 1H) 8.37 (d, J=4.25 Hz, 1H).

LCMS: MS (ESI) m/z: 546.4 [M+1]+

Target compound: 015 (SFC peak position: 0.595)

1H NMR (400 MHz, deuterated methanol) δ ppm 1.61 (d, J=7.03 Hz, 3H) 1.65-1.81 (m, 2H) 1.91-2.12 (m, 2H) 2.23-22.37 (m, 9H) 2.91-3.07 (m, 2H) 3.37 (s, 1H) 3.42-3.52 (m, 1H) 3.74-3.98 (m, 2H) 5.93 (q, J=7.03 Hz, 1H) 6.16-6.90 (m, 2H) 7.70 (d, J=4.02 Hz, 1H) 7.83-7.98 (m, 2H) 8.38 (s, 1H) 8.51 (d, J=4.27 Hz, 11H). LCMS: MS (ESI) m/z: 546.4 [M+1]+

Target compound: 016 (SFC peak position: 0.610)

1H NMR (400 MHz, deuterated methanol) δ ppm 1.61 (d, J=7.03 Hz, 3H) 1.65-1.81 (m, 2H) 1.91-2.12 (m, 2H) 2.23-22.37 (m, 9H) 2.91-3.07 (m, 2H) 3.37 (s, 1H) 3.42-3.52 (m, 1H) 3.74-3.98 (m, 2H) 5.93 (q, J=7.03 Hz, 1H) 6.16-6.90 (m, 2H) 7.70 (d, J=4.02 Hz, 11H) 7.83-7.98 (m, 2H) 8.38 (s, 11H) 8.51 (d, J=4.27 Hz, 1H). LCMS: MS (ESI) m/z: 546.4 [M+1]+

Example 7

Synthetic Route:

Step 1: Synthesis of Compound 017-02

Compound 017-01 (10 g, 38.57 mmol, 1 eq) and sodium hydrogen (3.08 g, 77.13 mmol, 60% purity, 2 eq) were added into N,N-dimethylformamide (50 mL) at 0° C. and stirred for 0.25 h, 3-bromopropene (23.33 g, 192.83 mmol, 5 eq) was added, and the reaction solution was stirred at 20° C. for 3 h. 20 mL of a saturated ammonium chloride solution was added into the reaction solution, the water phase was extracted for three times with 30 mL of ethyl acetate, the organic phase was collected and most of ethyl acetate was spun. The remaining part was washed for 6 times with 40 mL of a saturated salt solution and the organic phase was dried by a spinning method to obtain a crude product. The crude product was purified by a chromatographic column (petroleum ether/ethyl acetate=3/1) to obtain compound 017-02. LCMS: MS (ESI) m/z: 300 [M+1]+

Step 2: Synthesis of Compound 017-03

Compound 017-02 (19 g, 63.47 mmol, 1 eq) and sodium bicarbonate (8.00 g, 95.20 mmol, 3.70 mL, 1.5 eq) were added into a mixture of dichloromethane (85 mL) and methanol (15 mL), ozone (3.34 mmol) was introduced at −78° C. for 30 min, the solution turned blue, and nitrogen was introduced until blue disappeared. Triphenylphosphine (18.31 g, 69.82 mmol, 1.1 eq) was added, and the reaction solution was stirred at 25° C. for 0.5 h. After ozone was introduced, oxygen was introduced until the reaction solution faded from light blue, triphenylphosphorus was added, reaction was conducted for 0.5 h, and the reaction solution was directly dried by a spinning method to obtain a crude product. The crude product was purified by a chromatographic column (petroleum ether/ethyl acetate=1-0-6/1-5/1) to obtain compound 017-03. LCMS: MS (ESI) m/z: 302 [M+1]+

Step 3: Synthesis of Compound 017-04

Compound 017-03 (14.5 g, 48.12 mmol, 1 eq) and N,N-dibenzylamine (7.59 g, 38.50 mmol, 7.37 mL, 0.8 eq) were dissolved into dichloroethane (70 mL) at 20° C., sodium acetate borohydride (12.24 g, 57.74 mmol, 1.2 eq) was added, and the reaction solution was stirred for 5 min. 20 mL of saturated ammonium chloride was added into the reaction solution for quenching, the resulting reaction solution was extracted with dichloromethane (40 mL×2). The organic phases were combined, dried with anhydrous sodium sulfate, filtered, and concentrated under reduced pressure to obtain a crude product. The obtained crude product was purified by a chromatographic column (petroleum ether/ethyl acetate=1/0 to 5/1) to obtain compound 017-04. LCMS: MS (ESI) m/z: 483 [M+1]+

Step 4: Synthesis of Compound 017-05

Compound 017-04 (6.1 g, 12.64 mmol, 1 eq) was added into a hydrogenation flask containing methanol (60 mL), palladium/carbon (0.6 g, 10%) was added, and air in the hydrogenation flask was replaced with argon for 4 times. Hydrogen (45 psi) was continuously introduced, and the reaction solution was stirred at 45° C. for 12 h. After the reaction solution was filtrated with diatomite, the diatomite was washed with methanol for 3 times, and the organic phase was collected and dried by a spinning method to obtain a crude product. The crude product was added into methanol (30 mL) and water (10 mL), potassium carbonate (6.37 g, 46.10 mmol, 3 eq) and palladium/carbon (1.2 g, 10% purity) were added, the air was exchanged, and the reaction was conducted at 25° C. for 3 h under hydrogen (45 psi). The reaction solution was filtered with diatomite, and the diatomite was washed for 3 times with 20 mL of methanol. The organic phase was collected and dried by a spinning method to obtain a crude product. The obtained crude product was added into a hydrogenation flask containing methanol (50 mL), the air in the hydrogenation flask was replaced with argon, and reaction solution was stirred for 24 h at 60° C. under hydrogen (50 psi). The reaction solution was filtered with diatomite, the diatomite was washed 3 times with 20 mL of methanol, and the organic phase was collected and dried by a spinning method to obtain compound 017-05.

Step 5: Synthesis of Compound 017-06

Compound 017-05 (1.5 g, 5.20 mmol, 1 eq) was added into N,N-dimethylformamide (10 mL), diisopropylethylamine (2.02 g, 15.61 mmol, 2.72 mL, 3 eq) and propyl phosphoric anhydride (8.28 g, 13.01 mmol, 7.73 mL, 50% purity, 2.5 eq) were added, and the reaction solution was stirred at 25° C. for 0.5 h. 2 mL of 1 M hydrochloric acid was added into the reaction solution, 5 mL of water was added, and the water phase was extracted with 10 mL of ethyl acetate for 3 times. The organic phase was collected and washed with 20 mL of a saturated salt solution for 4 times, dried with anhydrous sodium sulfate, and dried by a spinning method to obtain compound 017-06.

Step 6: Synthesis of Compound 017-07

Compound 017-06 (1.2 g, 4.44 mmol, 1 eq) was dissolved into N,N-dimethylfomamide (10 mL) at 0° C., sodium hydrogen (213.06 mg, 5.33 mmol, 60% purity, 1.2 eq) was added, the reaction solution was stirred for 0.2 h, compound 005-02 (1.26 g, 4.66 mmol, 1.05 eq) was added, and the reaction solution was heated to 20° C. and continued to stir for 0.5 h. The reaction solution was poured into 10 mL of water, and filtered to obtain filter cake. The filter cake was washed with 10 mL of water, and then washed with 5 mL of petroleum ether. The filter cake was collected, and dried by a spinning method under reduced pressure to obtain compound 017-07.

Step 7: Synthesis of Compound 017-08

Compound 017-07 (1.9 g, 4.13 mmol, 1 eq) was added into dichloromethane (20 mL), trifluoroacetic acid (6.16 g, 54.03 mmol, 4 mL, 13.07 eq) was added, and the reaction solution was stirred at 25° C. for 0.5 h. The reaction solution was directly dried by a spinning method, 20 mL of dichloromethane was added, and then continued to dry by a spinning method, and the operations were repeated for 3 times. Trifluoroacetate of compound 017-08 was obtained.

Step 8: Synthesis of Compound 017-h

The trifluoroacetate of compound 017-08 (787.08 mg, 3.52 mmol, 0.85 eq) and compound 001-05 (1.96 g, 4.14 mmol, 1 eq, TFA) were added into n-butanol (10 mL), diisopropylethylamine (1.61 g, 12.42 mmol, 2.16 mL, 3 eq) was added, and the reaction solution was heated and stirred at 130° C. for 16 h. The reaction solution was directly dried by a spinning method to obtain a crude product. The crude product was purified by a chromatographic column (dichloromethane/methanol=1/0-4% methanol), and purified by high performance chromatography (column: VenusilASB Phenyl 150×30 mm×5 μm; mobile phase: [water (0.05% hydrochloric acid)-acetonitrile]; acetonitrile %: 30%-60%, 9 min) to obtain compound 017-h. LCMS: MS (ESI) m/z: 547 [M+1]+

Step 9: Synthesis of Compound 017 or 018

The compound 017-h (0.75 g, 1.37 mmol, 1 eq) was subjected to chiral resolution (column: DAICEL CHIRALPAK AS (250 mm×50 mm, 10 μm); mobile phase: [A: CO2, B: 0.1% ammonia water-ethanol]; B %: 50%-50%) to obtain compound 017 and compound 018.

Target compound: 017 (SFC peak position: 3.645)

1H NMR (400 MHz, deuterated methanol) δ ppm 1.62 (d, J=6.8 Hz, 3H) 1.86-1.93 (m, 2H) 2.09-2.13 (m, 2H) 2.22 (s, 3H) 2.28 (s, 3H) 3.02-3.20 (m, 3H) 3.46-3.48 (m, 1H) 3.87-3.97 (m, 2H) 4.58-4.61 (m, 2H) 4.90 (m, 1H) 5.89-5.94 (m, 1H) 6.11-6.22 (m, 1H) 7.70 (d, J=4 Hz, 1H) 7.87-7.96 (m, 2H) 8.38 (s, 1H) 8.52 (d, J=4.4 Hz, 1H). LCMS: MS (ESI) m/z: 547 [M+1]+

Target compound: 018 (SFC peak position: 4.906)

1H NMR (400 MHz, deuterated methanol) δ ppm 1.62 (d, J=6.8 Hz, 3H) 1.86-1.93 (m, 2H) 2.09-2.13 (m, 2H) 2.22 (s, 3H) 2.28 (s, 3H) 3.02-3.20 (m, 3H) 3.46-3.48 (m, 1H) 3.87-3.97 (m, 2H) 4.58-4.61 (m, 2H) 4.90 (m, 1H) 5.89-5.94 (m, 1H) 6.11-6.22 (m, 1H) 7.70 (d, J=4 Hz, 1H) 7.87-7.96 (m, 2H) 8.38 (s, 1H) 8.52 (d, J=4.4 Hz, 1H). LCMS: MS (ESI) m/z: 547 [M+1]+

Biological Test Data:

Experimental Example 1: Evaluation on Inhibitory Activity In-Vitro of Wild-Type and V804M Mutant Kinases

33P isotope-labeled kinase activity test (Reaction Biology Corp) was used to determine IC50value so as to evaluate an inhibitory ability of a test compound on human wild-type, V804M and V804L mutant RET.

Buffer solution condition: 20 mM 4-(2-hydroxyethyl)piperazine-1-ethanesulfonic acid (Hepes) (pH 7.5), 10 mM MgCl2, 1 mM 1,2-bis[2-[bis(carboxymethyl)amino]ethoxy]ethane (EGTA), 0.02% polyoxyethylene lauryl ether (Brij35), 0.02 mg/mL bovine serum albumin (BSA), 0.1 mM of Na3VO4, 2 mM dithiothreitol (DTT) and 1% DMSO.

Compound treatment: the compound to be tested was dissolved in 100% DMSO, and then the obtained solution was continuously diluted with DMSO by using Integra Viaflo Assist until a specific concentration was obtained.

Test procedures: a substrate was dissolved in the newly prepared buffer solution, a kinase to be tested was added therein, and the obtained solution was gently and evenly mixed. DMSO solution in which a compound to be test was dissolved was added in the above evenly mixed reaction solution by using an acoustic technology (Echo 550), and then incubated for 20 minutes at room temperature. The concentrations of the compounds in the reaction solutions were 3 μM, 1 μM, 0.333 μM, 0.111 μM, 0.0370 μM, 0.0123 μM, 4.12 nM, 1.37 nM, 0.457 nM and 0.152 nM, respectively. After incubation for 15 min,33P-ATP (activity of 0.01 μCi/μL, Kmconcentration) was added and the reaction was started. After the reaction was conducted at room temperature for 120 min, radioactivity was detected by a filter binding method. Kinase activity data was expressed by comparing the kinase activity of the compound to be tested with the kinase activity of a blank group (containing only DMSO). The IC50value was obtained by curve fitting with Prism4 software (GraphPad). The experimental results were shown in Table 1.

TABLE 1In-vitro screening test results of the compounds of the present disclosureIC50(nM)CompoundWild-typeV804MV804L001-h92.99128.70ND003-h73.87133.60ND005-h3.0013.02ND00768.88116.30ND0080.360.71ND00991.03NDND010119.9NDND011233.60NDND0120.901.060.4701346.7NDND0141.124.11.2301548.30NDND01675.23NDND01770.3274.30ND0181.291.971.23Note:“ND” refers to that no detection was performed.
Conclusion: The compounds of the present disclosure showed relatively good inhibitory activity on the wild-type, V804M and V804L mutant RET.

Experimental Example 2: Pharmacokinetic Evaluation on Compounds

Experiment objective: to test the pharmacokinetic of compound in mice

Experiment material: CD-1 mice (male)

Experiment operations: the pharmacokinetic characteristics of rodents after intravenous injection and oral administration of the compounds were obtained by using a standard scheme. In the experiment, candidate compounds were formulated into clear solutions, and mice were administrated by a single intravenous injection and oral administration. The solvent of the intravenous injection and oral administration was 10% PEG400 (polyethylene glycol 400)+90%(10% hydroxypropyl-β-cyclodextrin). The whole blood samples were collected within 24 hours. All the blood samples were added respectively into labeled plastic centrifuge tubes in which 0.5 M K2-EDTA anticoagulants were pre-added. The blood samples were collected, and centrifuged at 3,000 g for 10 min at 4° C. The supernatant plasma was collected by absorbing and quickly placed in dry ice to keep at −20° C. or lower. Blood concentrations were quantitatively analyzed by using an LC-MS/MS analysis method, and pharmacokinetic parameters, such as peak concentration, peak time, clearance rate, half-life, area under drug concentration-time curve, bioavailability, etc. were calculated.

The experiment results were shown in Table 2.

TABLE 2Pharmacokinetic test results of the compounds of the present disclosureInjection Administration (2 mpk)SamplesHalf-Oral Administration (10 mpk)to beClearancelifeIntegratedIntegratedTestRateT1/2ConcentrationConcentrationBioavailability(Compounds)(mL/min/kg)(h)AUC (nM · hr)AUC (nM · hr)F (%)Compound1.652.53164144961953.7014Compound6.552.5693012772359.7018

Conclusion: The pharmacokinetic of the compounds of the present disclosure in mice have good pharmacokinetic indexes.

Experimental Example 3: Evaluation on Efficacy of Compounds in Ba/F3 KIF5B-RET-V804L Tumor Cell Xenograft Model

1. Experiment Objective:

An engineered cell line Ba/F3 KIF5B-RET-V804L tumor cell xenograft model was established in NPSG mice, and efficacy of drugs to be tested alone on the xenograft model of NPSG mice was verified.

2. Experiment Design:

1) Ba/F3 KIF5B-RET-V804L cells were resuscitated and cultured in vitro to obtain 5×107cells.

2) Forty-five 6-8 week old female mice were adaptively fed for 1 week and weighed.

3) Ba/F3 KIF5B-RET-V804L tumor cells were subcutaneously inoculated into the right scapula position of the mice so as to establish the engineered cell line Ba/F3 KIF5B3-RET-V804L tumor cell xenograft model. Inoculation conditions were as follows: (see Table 3)

TABLE 3Inoculation informationVolume ofTotalInoculationInoculationAmountNumberCellCellofAnimalofType of InoculationInoculationAmountSuspensionNeededStrainAnimalsCellsPosition(Cell)(mL)CellsNPSG45Ba/F3 KIF5B-RET-Subcutaneous1 × 1060.1 mL5 × 107V804Ltissue

4) After inoculation, the volume and weight of tumors were measured once a week. When the average tumor volume reached 124.5 mm3, mice with the tumor volume between 82.9 mm3and 145.4 mm3were selected and randomly grouped according to the volume and weight of tumors with 6 mice in each group. The administration was started immediately after the grouping. The start date of the administration was recorded as day 0. Administration and grouping information were seen in Table 4.

TABLE 4Grouping and administration informationNumberAdministrationAdministrationofSample forDosageAdministrationAmountAdministrationGroupAnimalsTest(m/kg)RouteμL/gPeriod16BlankNAp.o.10Day 0-Day 9ControlBID26Compound10p.o.10Day 0-Day 9014BID36Compound10p.o.10Day 0-Day 9018BIDNote:NA represents no administration; Day 0-Day 9 represents from day 0 to day 9.

5) After the administration was started, the mice were continuously administrated for 9 days, and the body weight and tumor volume of the mice were measured on day 3, day 6, and day 9.

6) Data statistics were analyzed by one-way ANOVA. Variance homogeneity difference of data was firstly detected. If there was no difference in the variance homogeneity, an LSD method was used for analysis; and if there was a difference in the variance homogeneity, Dunnett's T3 was selected for data analysis. All data were analyzed by using SPSS 17.0. A p value less than 0.05 was considered a significant difference.

3. Experimental Results

3.1 Body Weight

The average body weights of each group at different time points were shown in Table 5.

TABLE 5Body weight changes of mice in each group in experimentNumberCompound 014Compound 018of DayBlank Control10 mg/kg10 mg/kg018.7 ± 0.519.5 ± 0.5318.6 ± 0.49319.6 ± 0.5220.3 ± 0.7218.8 ± 0.48620 ± 0.6320.5 ± 0.7218.6 ± 0.87920.4 ± 0.4520.3 ± 0.8419 ± 0.64
3.2 Inhibition of Tumor Growth

Inhibition of tumor growth in each group was shown in Table 6.

TABLE 6Tumor-inhibition effect of mice in each groupTumorTumorVolume onVolume onTGIΔVACPTreatmentDay 0 (mm3)aDay 9 (mm3)a(%)(%)valueBlank Control126.3 ± 4.66827.8 ± 44.98///Compound014123.8 ± 7.284.8 ± 6.25106%−6%<0.00110 mg/kgCompound018125 ± 3.9675.5 ± 7.11107%−7%<0.00110 mg/kgNote:ameans an average nunber ± standard error;b means that the data was analyzed by using One-way ANOVA, and a Dunnetts T3 method was used for post-mortem analysis due to uneven variance;TGI represents tumor growth inhibition rate and its value is 1-ΔT/ΔC;ΔT represents increased tumor volume in the experimental group;ΔC represents increased tumor volume in the control group.

Conclusion: in the engineered cell line Ba/F3 KIF5B-RET-V804L xenograft model of mice, the compounds of the present disclosure showed relatively strong efficacy for 9 consecutive administration days.

Experimental Example 4: Evaluation on Efficacy of Compounds in Engineered Cell Line Ba/F3 KIF5-RET Xenograft Model of Female NPSG Mice

1. Experiment Objective:

To verify efficacy of drugs to be tested alone on the engineered cell line Ba/F3 KIF5B-RET xenograft model of NPSG mice.

2. Experiment Design:

1) Ba/F3 KIF5B-RET cells were resuscitated and cultured in vitro to obtain 4×107cells.

2) Sixty 6-8 week old female NPSG mice were adaptively fed for one week and weighed.

3) The Ba/F3 KIF5B-RET cells were subcutaneously inoculated on right scapula of mice according to the inoculation conditions obtained in the pre-test (see Table 7) to establish the Ba/F3 KIF5B-RET tumor cell xenograft model of NPSG mice.

TABLE 7Inoculation informationVolume ofTotalInoculationInoculationAmountNumberCellCellofAnimalofType of InoculationInoculationAmountSuspensionNeededStrainAnimalsCellsPosition(Cell)(mL)CellsNPSG60Ba/F3 KIF5B-RETSubcutaneous1 × 1060.1 mL6 × 107tissue

4) After inoculation, the volume and weight of tumors were measured once a week. When the average tumor volume reached 104 mm3, mice were randomly grouped according to the volume and weight of tumors with 6 mice in each group. The administration was started immediately after the grouping. The start date of the administration was recorded as day 0. Administration and grouping information were shown in Table 8.

TABLE 8Grouping and administration informationNumberAdministrationAdministrationofSample forDosageAdministrationAmountAdministrationGroupAnimalsTest(m/kg)RouteμL/gPeriod16BlankNAp.o.10Day 0-Day 10ControlBID26Compound10p.o.10Day 0-Day 10014BID36Compound10p.o.10Day 0-Day 10018BID46Compound20p.o.10Day 0-Day 10018BIDNote:NA represents no administration; Day 0-Day 10 represents from day 0 to day 10.

5) After administration was started, the body weight and tumor volume of the mice were measured respectively on the day 0, day 3, day 6, and day 10.

6) For the experiment of two groups, a T-Test analysis method was used. For a comparison of three or more groups, One-Way ANOVA was used for analysis. For comparing potential synergistic effects, Two-Way ANOVA was used for analysis. All data were analyzed by using SPSS 17.0. A p value less than 0.05 was considered to be a significant difference, and a p value less than 0.01 was considered to be a very significant difference.

3. Experimental Results

3.1 Body Weight

The average body weights of each group at different time points were shown in Table 9.

TABLE 9Body weight changes in each groupNumberBlankCompound 014Compound 018Compound 018of DaysControl10 mg/kg10 mg/kg20 mg/kg017.2 ± 0.2618 ± 0.5717.7 ± 0.4717.6 ± 0.58318 ± 0.3618.5 ± 0.4718.2 ± 0.5118.1 ± 0.56618.4 ± 0.2418.5 ± 0.7518.5 ± 0.5718.3 ± 0.531019.6 ± 0.3118.6 ± 0.6217.7 ± 0.8618.5 ± 0.63
3.2 Inhibition of Tumor Growth

Inhibition of tumor growth in each group was shown in Table 10.

TABLE 10Tumor-inhibition effect of drugs in each groupTumorTumorVolume onVolume onTGIΔVACPTreatmentDay 0 (mm3)aDay 9 (mm3)a(%)(%)valuebBlank Control105.8 ± 7.751063.5 ± 65.21///014102.3 ± 7.72116.6 ± 10.7799%1%<0.00110 mg/kgCompound018106.3 ± 7.14205.1 ± 29.8790%10%<0.00110 mg/kgCompound018102.2 ± 7.0932.8 ± 6.81107%−7%<0.00120 mg/kgNote:ameans an average number ± standard error;bmeans that the data was analyzed by using One-way ANOVA and a Dunnett's T3 method was used for post-mortem analysis due to uneven variance;TGI represents tumor growth inhibition rate and its value is 1-ΔT/ΔC;ΔT represents increased tumor volume in the experimental group;ΔC represents increased tumor volume in the control group.

Conclusion: in the engineered cell line Ba/F3 KIF5B-RET xenograft model of mice, the compounds of the present disclosure showed relatively strong efficacy for 10 consecutive administration days.