ISOQUINOLINONE DERIVATIVES SERVING AS ROCK PROTEIN KINASE INHIBITORS AND USE THEREOF
20230024516 · 2023-01-26
Assignee
Inventors
- Yizhi Liu (Guangzhou, CN)
- Yandong Wang (Guangzhou, CN)
- Lingyun Wu (Guangzhou, CN)
- Xu You (Guangzhou, CN)
- Zheming Xiao (Guangzhou, CN)
- Shuhui Chen (Guangzhou, CN)
Cpc classification
International classification
C07D401/12
CHEMISTRY; METALLURGY
Abstract
Disclosed are a series of isoquinolone derivatives as ROCK protein kinase inhibitors and uses thereof in preparing medicaments for ROCK protein kinase inhibitor-related glaucoma or ocular hypertension diseases. Specially, disclosed are a compound of formula (I), an isomer thereof or a pharmaceutically acceptable salt thereof.
##STR00001##
Claims
1. A compound of formula (I), an isomer thereof or a pharmaceutically acceptable salt thereof, ##STR00166## wherein, T.sub.1 is —(CH.sub.2).sub.n—; T.sub.2 is selected from a group consisting of —(CH.sub.2).sub.m— and —C(R.sub.7)(R.sub.8)—; R.sub.1 is selected from a group consisting of C.sub.1-16 alkyl, phenyl, C.sub.3-7 cycloalkyl, 3-8 membered heterocycloalkyl and 5-10 membered heteroaryl, each of C.sub.1-16 alkyl, phenyl, C.sub.3-7 cycloalkyl, 3-8 membered heterocycloalkyl and 5-10 membered heteroaryl is optionally substituted by 1, 2 or 3 R.sub.a; each of R.sub.2 and R.sub.3 is independently selected from a group consisting of H, F, Cl, Br, I, OH, NH.sub.2, CN and C.sub.1-3 alkyl; R.sub.4 is selected from a group consisting of H, F, Cl, Br, I, OH, NH.sub.2, CN, and C.sub.1-3 alkyl which is optionally substituted by 1, 2 or 3 R.sub.b; R.sub.5 is NR.sub.9R.sub.10; R.sub.6 is selected from a group consisting of H, F, Cl, Br, I, OH, NH.sub.2, CN and C.sub.1-3 alkyl; each of R.sub.7 and R.sub.8 is independently selected from a group consisting of H, F, Cl, Br, I, OH, NH.sub.2, CN, and C.sub.1-3 alkyl which is optionally substituted by 1, 2 or 3 R.sub.c; or, R.sub.7 and R.sub.8 together with the atom that they attached to form a C.sub.3-5 cycloalkyl which is optionally substituted by 1, 2 or 3 R.sub.d; each of R.sub.9 and R.sub.10 is independently selected from a group consisting of H, and C.sub.1-3 alkyl which is optionally substituted by 1, 2 or 3 R.sub.e; L is selected from a group consisting of a single bond, —O— and —NR.sub.11—; R.sub.11 is selected from a group consisting of H and C.sub.1-3 alkyl; n is selected from a group consisting of 0, 1 and 2; m is selected from a group consisting of 0, 1, 2 and 3; R.sub.a is selected from a group consisting of F, Cl, Br, I, OH, NH.sub.2, CN, C.sub.1-3 alkyl and C.sub.1-3 alkoxy, wherein each of C.sub.1-3 alkyl and C.sub.1-3 alkoxy is optionally substituted by 1, 2 or 3 R; each of R.sub.b, R.sub.c, R.sub.d and R.sub.e is independently selected from a group consisting of H, F, Cl, Br, I, OH, NH.sub.2, CN and C.sub.1-3 alkyl; R is selected from a group consisting of F, Cl, Br, I, OH, NH.sub.2, CN and CH.sub.3; wherein, each of the 3-8 membered heterocycloalkyl and 5-10 membered heteroaryl is independently include 1, 2, 3 or 4 heteroatom(s) or heteroatom group(s) which is/are independently selected from a group consisting of —NH—, —O—, —S— and N.
2. The compound, the isomer thereof or the pharmaceutically acceptable salt thereof according to claim 1, wherein, R.sub.a is selected from a group consisting of F, Cl, Br, I, OH, NH.sub.2, CN, CH.sub.3, CF.sub.3, CH.sub.2F, CHF.sub.2, CH.sub.2CH.sub.3 and OCH.sub.3.
3. The compound, the isomer thereof or the pharmaceutically acceptable salt thereof according to claim 1, wherein, R.sub.1 is selected from a group consisting of C.sub.1-12 alkyl, phenyl, cyclobutyl, cyclopentyl, cyclohexyl, tetrahydrofuranyl, tetrahydropyranyl, piperidinyl, thienyl, furyl, pyrrolyl and benzofuryl, wherein, each of C.sub.1-12 alkyl, phenyl, cyclobutyl, cyclopentyl, cyclohexyl, tetrahydrofuranyl, tetrahydropyranyl, piperidinyl, thienyl, furyl, pyrrolyl and benzofuryl is optionally substituted by 1, 2 or 3 R.sub.a.
4. The compound, the isomer thereof or the pharmaceutically acceptable salt thereof according to claim 3, wherein, R.sub.1 is selected from a group consisting of CH.sub.3, CH.sub.2CH.sub.3, (CH.sub.2).sub.2CH.sub.3, (CH.sub.2).sub.3CH.sub.3, (CH.sub.2).sub.4CH.sub.3, (CH.sub.2).sub.5CH.sub.3, (CH.sub.2).sub.6CH.sub.3, (CH.sub.2).sub.10CH.sub.3, CH(CH.sub.3).sub.2, C(CH.sub.3).sub.3, ##STR00167##
5. The compound, the isomer thereof or the pharmaceutically acceptable salt thereof according to claim 4, wherein, R.sub.1 is selected from a group consisting of CH.sub.3, CH.sub.2CH.sub.3, (CH.sub.2).sub.2CH.sub.3, (CH.sub.2).sub.3CH.sub.3, (CH.sub.2).sub.4CH.sub.3, (CH.sub.2).sub.5CH.sub.3, (CH.sub.2).sub.6CH.sub.3, (CH.sub.2).sub.10CH.sub.3, CH(CH.sub.3).sub.2, C(CH.sub.3).sub.3, ##STR00168##
6. The compound, the isomer thereof or the pharmaceutically acceptable salt thereof according to claim 1, wherein, each of R.sub.2 and R.sub.3 is independently selected from a group consisting of H, F, Cl, Br, I, OH and NH.sub.2.
7. The compound, the isomer thereof or the pharmaceutically acceptable salt thereof according to claim 1, wherein, R.sub.4 is selected from a group consisting of H, F, Cl, Br, I, OH, NH.sub.2, CN, CH.sub.3 and CH.sub.2CH.sub.3.
8. The compound, the isomer thereof or the pharmaceutically acceptable salt thereof according to claim 1, wherein, each of R.sub.9 and Rim is independently selected from a group consisting of H, CH.sub.3 and CH.sub.2CH.sub.3.
9. The compound, the isomer thereof or the pharmaceutically acceptable salt thereof according to claim 1, wherein, R.sub.5 is selected from a group consisting of NH.sub.2, NH(CH.sub.3) and N(CH.sub.3).sub.2.
10. The compound, the isomer thereof or the pharmaceutically acceptable salt thereof according to claim 1, wherein, R.sub.6 is selected from a group consisting of H, F, Cl, Br, I, OH, NH.sub.2, CN and CH.sub.3.
11. The compound, the isomer thereof or the pharmaceutically acceptable salt thereof according to claim 1, wherein, each of R.sub.7 and R.sub.8 is independently selected from a group consisting of H, F, Cl, Br, I, OH, NH.sub.2, CN and CH.sub.3.
12. The compound, the isomer thereof or the pharmaceutically acceptable salt thereof according to claim 1, wherein, R.sub.7 and R.sub.8 together with the atom that they attached to form a cyclopropyl which is optionally substituted by 1, 2 or 3 R.sub.d.
13. The compound, the isomer thereof or the pharmaceutically acceptable salt thereof according to claim 12, wherein, R.sub.7 and R.sub.8 together with the atom that they attached to form a cyclopropyl.
14. The compound, the isomer thereof or the pharmaceutically acceptable salt thereof according to claim 1, wherein, L is selected from a group consisting of a single bond, —O—, —NH— and —N(CH.sub.3)—.
15. The compound, the isomer thereof or the pharmaceutically acceptable salt thereof according to claim 1, wherein, T2 is selected from a group consisting of —CH.sub.2—, —(CH.sub.2).sub.2— and ##STR00169##
16. The compound, the isomer thereof or the pharmaceutically acceptable salt thereof according to claim 1, wherein, the structure unit ##STR00170## is selected from a group consisting of ##STR00171##
17. The compound, the isomer thereof or the pharmaceutically acceptable salt thereof according to claim 16, wherein, the structure unit ##STR00172## is selected from a group consisting of ##STR00173##
18. The compound, the isomer thereof or the pharmaceutically acceptable salt thereof according to claim 1, wherein, the structure unit ##STR00174## is selected from a group consisting of CH.sub.3, CH.sub.2CH.sub.3, (CH.sub.2).sub.2CH.sub.3, (CH.sub.2).sub.3CH.sub.3, (CH.sub.2).sub.4CH.sub.3, (CH.sub.2).sub.5CH.sub.3, (CH.sub.2).sub.6CH.sub.3, (CH.sub.2).sub.10CH.sub.3, CH(CH.sub.3).sub.2, C(CH.sub.3).sub.3, OCH.sub.3, OCH.sub.2CH.sub.3, O(CH.sub.2).sub.2CH.sub.3, O(CH.sub.2).sub.3CH.sub.3, O(CH.sub.2).sub.4CH.sub.3, O(CH.sub.2).sub.5CH.sub.3, O(CH.sub.2).sub.6CH.sub.3, OCH(CH.sub.3).sub.2, OC(CH.sub.3).sub.3, N(CH.sub.3).sub.2, ##STR00175##
19. The compound, the isomer thereof or the pharmaceutically acceptable salt thereof according to claim 1, which is selected from a group consisting of: ##STR00176## wherein, R.sub.1 is as defined in claim 1; R.sub.4 is as defined in claim 1; R.sub.5 is as defined in claim 1; and L is as defined in claim 1.
20. The compound, the isomer thereof or the pharmaceutically acceptable salt thereof according to claim 19, which is selected from a group consisting of: ##STR00177## ##STR00178## wherein, R.sub.1, R.sub.4 and L are as defined in claim 19.
21. The compound, the isomer thereof or the pharmaceutically acceptable salt thereof according to claim 1, which is selected from a group consisting of: ##STR00179## ##STR00180## ##STR00181## ##STR00182## ##STR00183## ##STR00184##
22. The compound, the isomer thereof or the pharmaceutically acceptable salt thereof according to claim 21, which is selected from a group consisting of: ##STR00185## ##STR00186## ##STR00187## ##STR00188## ##STR00189## ##STR00190## ##STR00191## ##STR00192## ##STR00193## ##STR00194##
23. A pharmaceutical composition, comprising a therapeutically effective amount of the compound, the isomer thereof or the pharmaceutically acceptable salt thereof according to claim 1 and a pharmaceutically acceptable carrier.
24. A method of treating a disease associated with an ROCK protein kinase, comprising administrating a therapeutically effective amount of the compound, the isomer thereof or the pharmaceutically acceptable salt thereof according to claim 1 to a subject.
25. The method of claim 24, wherein the disease is selected from glaucoma or ocular hypertension.
Description
DETAILED DESCRIPTION OF THE EMBODIMENTS
[0119] The following examples will describe the present disclosure in detail, but are not intended to impose any unfavorable limitation on the present disclosure. The present disclosure has been described in detail herein, and its specific examples are also disclosed. It will be obvious for those skilled in the art that various changes and modifications may be made to the examples of the present disclosure without departing from the spirit and scope of the present disclosure.
Synthesis of Fragments of the Technical Concentrate
[0120] ##STR00042##
Step 1
[0121] To a solution of compound 1a (15.0 g, 71.7 mmol) in thionyl chloride (164.0 g, 1.38 mol) was added N,N-dimethylformamide (0.55 mL), stirred for 12 hours at 80° C. After the reaction was completed, the reaction mixture was concentrated under reduced pressure to obtain a crude compound 1b. MS-ESI calculated value [M+H].sup.+ 228, actually measured value 228.
Step 2
[0122] At 0° C., to a solution of compound 1b (18.9 g, 62.0 mmol) in dichloromethane (150 mL) was added compound 1c (12.7 g, 68.2 mmol) and N,N-diisopropylethylamine (124 mL) under a nitrogen atmosphere, stirred for 0.5 hours at 25° C. After the reaction was completed, the reaction solution, after being concentrated under reduced pressure, was diluted with water (100 mL), and extracted with dichloromethane (60 mL×3), and the combined organic phase was washed with a saturated sodium chloride solution, dried with anhydrous sodium sulfate, and concentrated under reduced pressure after filtration. The residue was purified by silica gel column chromatography to obtain compound 1d. MS-ESI calculated value [M+H].sup.+ 378, actually measured value 378.
Step 3
[0123] At 0° C., to a solution of compound 1d (8 g, 20.3 mmol) in dichloromethane (100 mL) was added chloroperoxybenzoic acid (6.48 g, 30.4 mmol, purity: 81%) in batch, stirred for 2 hours at 25° C. After the reaction was completed, the reaction mixture was quenched with saturated sodium bicarbonate solution (100 mL), extracted with dichloromethane (60 mL×3), and the organic phase was dried by anhydrous sodium sulfate, then filtered, concentrated under reduced pressure to obtain compound 1e. MS-ESI calculated value [M+H].sup.+ 394, actually measured value 394.
Step 4
[0124] Compound 1e (11.1 g, 28.2 mmol) was dissolved in acetic anhydride (109 g, 1.07 mol), and after stirred for 2 hours at 130° C., acetic anhydride was removed by concentration. Tetrahydrofuran (100 mL) and 30% aqueous sodium hydroxide solution (150 mL) were added, stirred for 30 minutes at 25° C. After removal of tetrahydrofuran by concentration under reduced pressure, a 6 M hydrochloric acid solution was added dropwise to the system to adjust the pH of the reaction solution to 7, extracted with ethyl acetate (50 mL×3), and the combined organic phase dried with anhydrous sodium sulfate, filtered, and concentrated under reduced pressure. The residue was purified by silica gel column chromatography to obtain intermediate 1. .sup.1H NMR (400 MHz, DMSO-d.sub.6) δ=8.52 (d, J=7.8 Hz, 1H), 8.21 (dd, J=7.8, 1.3 Hz, 1H), 7.64 (t, J=7.8 Hz, 1H), 7.37 (d, J=7.8 Hz, 1H), 7.23 (d, J=7.6 Hz, 1H), 3.91 (d, J=5.4 Hz, 1H), 3.20-3.28 (m, 3H), 3.03 (dd, J=9.8, 5.0 Hz, 1H), 1.92-2.05 (m, 1H), 1.68-1.77 (m, 1H), 1.34 (s, 9H). MS-ESI calculated value [M+Na].sup.+ 416, actually measured value 416.
##STR00043##
Step 1
[0125] To a solution of compound 2a (5.0 g, 36.7 mmol) in concentrated hydrochloric acid (50 mL, 12 M) was added slowly the solution of sodium nitrite (2.29 g, 33.2 mmol) in water (8 mL) at −5° C., stirred for 1 hour at −5° C. To a solution of cuprous chloride (156 mg, 1.58 mmol) and cupric chloride (2.34 g, 17.4 mmol) in glacial acetic acid (80 mL) was introduced sulphur dioxide until saturation, and then the previous reaction solution was slowly added dropwise to the solution containing sulfur dioxide at −5° C., stirred for 1 hour at 20° C. After the reaction was completed, water (150 mL) was added to the reaction solution, extracted with dichloromethane (150 mL×4). The combined organic phase was washed with saturated brine (500 mL×1), dried with anhydrous sodium sulfate, filtered, the filtrate was concentrated under reduced pressure to obtain a residue, and the residue crude 2b was directly used for the next step without purification. MS-ESI calculated value [M+H].sup.+ 242, actually measured value 242.
Step 2
[0126] Compound 2c was obtained referring to step 2 of intermediate 1.
Step 3
[0127] Compound 2d was obtained referring to step 3 of intermediate 1.
Step 4
[0128] Intermediate 2 was obtained referring to step 4 of intermediate 1. MS-ESI calculated value [M+H].sup.+ 408, actually measured value 408.
##STR00044##
Step 1
[0129] Compound 3b was obtained referring to step 2 of intermediate 1. MS-ESI calculated value [M+H].sup.+ 406, actually measured value 406.
Step 2
[0130] Compound 3c was obtained referring to step 3 of intermediate 1. MS-ESI calculated value [M+H].sup.+ 422, actually measured value 422.
Step 3
[0131] Intermediate 3 was obtained referring to step 4 of intermediate 1. MS-ESI calculated value [M+H].sup.+ 422, actually measured value 422.
##STR00045##
Step 1
[0132] Compound 4b was obtained referring to step 2 of intermediate 1. MS-ESI calculated value [M+H].sup.+ 364, actually measured value 364.
Step 2
[0133] Compound 4c was obtained referring to step 3 of intermediate 1. MS-ESI calculated value [M+H].sup.+ 380, actually measured value 380.
Step 3
[0134] Intermediate 4 was obtained referring to step 4 of intermediate 1. MS-ESI calculated value [M+Na].sup.+ 402, actually measured value 402.
##STR00046##
Step 1
[0135] To a solution of diisopropylamine (36.4 g, 359 mmol) in tetrahydrofuran (1.5 mL) was added dropwise n-butyl lithium (2.5 M, 131.2 mL) at −78° C. under the nitrogen atmosphere; after stirred for 1 hour at −78° C., compound 5b (20.1 g, 299 mmol) was added dropwise to the reaction solution at −78° C. under the nitrogen atmosphere; after stirred for 1 hour at −78° C., compound 5a (50 g, 299 mmol) was added dropwise to the reaction solution at −78° C. under the nitrogen atmosphere, stirred for 1 hour at −78° C. under the nitrogen atmosphere. After the reaction was completed, quenched with saturated aqueous ammonium chloride solution (2 L) and extracted with ethyl acetate (500 mL×3). The combined organic phase was washed with saturated brine (1 L×1) and dried with anhydrous sodium sulfate, filtered, the filtrate was concentrated under reduced pressure, and the crude product was separated and purified by silica gel column chromatography (petroleum ether/ethyl acetate) to obtain compound 5c. .sup.1H NMR (400 MHz, CDCl.sub.3) δ=5.83-5.70 (m, 1H), 5.50 (dd, J=1.5, 16.9 Hz, 1H), 5.36-5.30 (m, 1H), 4.22-4.17 (m, 2H), 3.77-3.70 (m, 1H), 2.81-2.71 (m, 1H), 2.68-2.59 (m, 1H), 1.29-1.26 (m, 3H).
Step 2
[0136] To a solution of compound 5c (60.7 g, 396 mmol) in tetrahydrofuran (1.2 L) was added tetraisopropyltitanate (113 g, 396 mmol) under the nitrogen atmosphere, and added dropwise ethyl magnesium bromide (89.8 g, 674 mmol) at 20° C. under the nitrogen atmosphere, stirred for 3 hours at 20° C. After the reaction was completed, quenched with water (300 mL) and filtered, the filtrate was concentrated under reduced pressure, and the crude product was separated and purified by silica gel column chromatography (petroleum ether/ethyl acetate) to obtain compound 5d. MS-ESI calculated value [M+H].sup.+138, actually measured value 138.
Step 3
[0137] To a solution of compounds 5d (81.0 g, 590.5 mmol) in acetonitrile (500 mL) was added 4-dimethylaminopyridine (7.21 g, 59.1 mmol) and di-tert-butyl dicarbonate (155 g, 709 mmol) successively, stirred for 3 hours at 20° C. After the reaction was completed, quenched with water (1 L) and extracted with ethyl acetate (500 mL×3). The combined organic phase was washed with saturated brine (500 mL×1) and dried with anhydrous sodium sulfate, filtered, the filtrate was concentrated under reduced pressure, and the crude product was separated and purified by silica gel column chromatography (petroleum ether/ethyl acetate) to obtain compound 5e. MS-ESI calculated value [M+H].sup.+ 238, actually measured value 238.
Step 4
[0138] To a solution of compound 5e (10.8 g, 45.5 mmol) in methanol (48 mL) and water (72 mL) was added sodium periodate (29.2 g, 136.4 mmol) and osmium tetraoxide (173 mg, 682 μmol) at 20° C., stirred for 2 hours at 20° C. After the reaction was completed, water (100 mL) was added to the reaction solution and the reaction solution was extracted with ethyl acetate (100 mL×3). The combined organic phase was washed with saturated brine (100 mL×1) and dried with anhydrous sodium sulfate, filtered, the filtrate was concentrated under reduced pressure, and the crude product was separated and purified by silica gel column chromatography (petroleum ether/ethyl acetate) to obtain compound 5f.
Step 5
[0139] To a solution of compound 5f (4.00 g, 16.7 mmol) in tert-butanol (50 mL) and tetrahydrofuran (50 mL) was added isopentene (14.6 g, 208 mmol), sodium hypochlorite (1.66 g, 18.4 mmol), sodium phosphate monobasic monohydrate (4.61 g, 33.4 mmol) and water (35 mL) successively at 0° C., stirred for 12 hours at 20° C. After the reaction was completed, the pH of the reaction solution was adjusted to 4 with 1N aqueous hydrochloric acid solution and the reaction solution was extracted with ethyl acetate (50 mL×3). The combined organic phase was washed with saturated brine (100 mL×1) and dried with anhydrous sodium sulfate, and filtered, the filtrate was concentrated under reduced pressure to obtain a residual. Water (50 mL) was added to the residue, aqueous Na.sub.2CO.sub.3 solution was used to adjust the pH to 10, and ethyl acetate (50 mL×1) was used for extraction to collect aqueous phase. The pH of the aqueous phase was adjusted to 4 with the 1N aqueous hydrochloric acid solution and the aqueous phase was extracted with ethyl acetate (50 mL×3). The combined organic phase was washed with saturated brine (100 mL×1), dried with anhydrous sodium sulfate and filtered, and the filtrate was concentrated under reduced pressure to obtain compound 5g.
Step 6
[0140] To a solution of compound 5g (3.10 g, 12.1 mmol) in toluene (30 mL) was added N,N-diisopropylethylamine (2.04 g, 15.8 mmol), diphenylphosphorylazide (4.34 g, 15.8 mmol, 3.42 ml) and a 4A molecular sieve (2.00 g) at 0° C., stirred for 0.5 hours at 20° C., and stirred for another 0.5 hour at 90° C., then cooled to 20° C. Compound 5h (1.44 g, 13.4 mmol) was added to the reaction solution at 20° C., stirred the reaction solution for 12 hours at 20° C. After the reaction was completed, the reaction solution was filtered, water (30 mL) was added to the filtrate and extracted with ethyl acetate (30 mL×2). The combined organic phase was washed with saturated brine (50 mL×1) and dried with anhydrous sodium sulfate, filtered, the filtrate was concentrated under reduced pressure, and the crude product was separated and purified by silica gel column chromatography (petroleum ether/ethyl acetate) to obtain compound 5i.
Step 7
[0141] To a solution of compound 5i (500 mg, 1.39 mmol) in tetrahydrofuran (5 mL) was added borane-tetrahydrofuran (1 M, 13.9 mL) at 0° C. under the nitrogen atmosphere, stirred for 1 hour at 25° C. After the reaction was completed, quenched with saturated aqueous ammonium chloride solution (20 mL), and extracted with ethyl acetate (20 mL×3). The combined organic phase was washed with saturated brine (20 mL×1) and dried with anhydrous sodium sulfate, filtered, the filtrate was concentrated under reduced pressure, and the crude product was separated and purified by preparative thin layer chromatography (petroleum ether/ethyl acetate) to obtain compound 5j.
Step 8
[0142] To a solution of Compound 5j (295 mg, 852 μmol) in ethyl acetate (3 mL) was added ethyl acetate hydrochloride (4 M, 5 mL), stirred for 0.5 hour at 25° C. After the reaction was completed, the reaction solution was concentrated under reduced pressure to obtain compound 5k.
Step 9
[0143] To a solution of compound 5k (420 mg, 1.59 mmol) in dichloromethane (10 mL) was added compound 1b (268 mg, 948 μmol) and N,N-diisopropylethylamine (489.98 mg, 3.79 mmol) at 0° C. under the nitrogen atmosphere, stirred the reaction mixture for 12 hours at 25° C. After the reaction was completed, dichloromethane (10 mL) and water (15 mL) were added to the reaction solution, extracted with dichloromethane (10 mL×2); and the combined organic phase was washed with saturated brine (20 mL×1), dried with anhydrous sodium sulfate, filtered and concentrated under reduced pressure. The crude product was separated and purified by silica gel plate chromatography (petroleum ether/ethyl acetate) to obtain compound 5l. MS-ESI calculated value [M+H].sup.+ 438, actually measured value 438.
Step 10
[0144] Compound 5m was obtained referring to step 3 of intermediate 1. MS-ESI calculated value [M+H].sup.+ 454, actually measured value 454.
Step 11
[0145] Intermediate 5 was obtained referring to step 4 of intermediate 1. MS-ESI calculated value [M+H].sup.+ 454, actually measured value 454.
##STR00047##
Step 1
[0146] Compound 6b was obtained referring to step 2 of intermediate 1.
Step 2
[0147] Compound 6c was obtained referring to step 3 of intermediate 1.
Step 3
[0148] Intermediate 6 was obtained referring to step 4 of intermediate 1. MS-ESI calculated value [M+H].sup.+ 408, actually measured value 408.
Example 1: Compound 7
[0149] ##STR00048##
[0150] Synthesis Route:
##STR00049##
Step 1
[0151] To a solution of compound 7a (1 g, 4.99 mmol) in dichloromethane (30 mL) and water (30 mL) was added sodium bicarbonate (1.68 g, 12.0 mmol) and tetrabutylammonium fluoride (1 M, 499 μL), then a solution of compound 7b (824 mg, 4.99 mmol) in dichloromethane (10 mL) was added to the reaction system under the condition of stirring at 0° C., stirred for 16 hours at 30° C. The organic phase was separated, dried with anhydrous magnesium sulfate and filtered, concentrated under reduced pressure to obtain compound 7c.
Step 2
[0152] At 0° C., to a solution of intermediate 1 (150 mg, 324 μmol) in N,N-dimethylformamide (6 mL) was added sodium hydride (15.6 mg, 389 μmol, purity: 60%) under nitrogen atmosphere, then added compound 7c (113 mg, 454 μmol) after stirred for 0.5 hour, warmed to 25° C. stirred for 2 hours. Ice water (30 mL) was slowly added to the reaction solution, extracted with ethyl acetate (20 mL×3), and the organic phase was dried with anhydrous sodium sulfate, filtered, concentrated under reduced pressure. The residue was purified by silica gel plate to obtain compound 7d. MS-ESI calculated value [M+Na].sup.+ 628, actually measured value 628.
Step 3
[0153] To a solution of compound 7d (96.0 mg, 154 μmol) in dichloromethane (20 mL) was added trimethylsllytrifluoromethanesulphonate (68.6 mg, 309 μmol), then stirred for 1 hour at 0-5° C. 2,6-dimethylpyridine (49.6 mg, 462 μmol) was added dropwise, stirred for another 1 hour at 0-5° C. Water (30 mL) was added to the reaction solution, extracted with dichloromethane (20 mL×3), and the organic phase was dried with anhydrous sodium sulfate, filtered, concentrated under reduced pressure. The residue was purified by preparative high performance liquid (formic acid condition) to obtain a formate of compound 7. .sup.1H NMR (400 MHz, CD.sub.3OD) δ=8.70 (d, J=7.8 Hz, 1H), 8.52 (s, 1H), 8.39 (dd, J=7.8, 1.3 Hz, 1H), 7.74 (t, J=7.8 Hz, 1H), 7.67 (d, J=7.8 Hz, 1H), 7.51 (d, J=7.8 Hz, 1H), 6.00 (s, 2H), 3.81-3.90 (m, 1H), 3.57-3.66 (m, 1H), 3.49-3.56 (m, 1H), 3.42 (dd, J=10.8, 4.1 Hz, 1H), 3.30 (d, J=6.2 Hz, 1H), 2.39 (t, J=7.4 Hz, 2H), 2.26-2.36 (m, 1H), 1.92-2.03 (m, 1H), 1.55-1.69 (m, 1H), 1.55-1.69 (m, 2H), 1.20-1.34 (m, 16H), 0.91 (t, J=6.8 Hz, 3H). MS-ESI calculated value [M+H].sup.+ 506, actually measured value 506.
[0154] The formate of compound 7 can be adjusted the pH of the reaction solution to 8-9 by adding a saturated aqueous sodium carbonate solution to the system, and then, after the extraction with ethyl acetate, the reaction solution was concentrated to obtain compound 7.
Example 2: Compound 8
[0155] ##STR00050##
Synthesis Route:
[0156] ##STR00051##
Step 1
[0157] Compound 8b was obtained referring to step 1 of Example 1. .sup.1H NMR (400 MHz, CDCl.sub.3) δ=8.27 (d, J=1.6 Hz, 1H), 7.94 (dd, J=8.8, 1.7 Hz, 1H), 7.60 (d, J=2.4 Hz, 1H), 7.44 (d, J=8.8 Hz, 1H), 6.74 (dd, J=2.2, 0.7 Hz, 1H), 5.89 (s, 2H).
Step 2
[0158] Compound 8c was obtained referring to step 2 of Example 1. MS-ESI calculated value [M+Na].sup.+ 590, actually measured value 590.
Step 3
[0159] A formate of compound 8 was obtained referring to step 3 of Example 1. .sup.1H NMR (400 MHz, CD.sub.3OD) δ=8.72 (d, J=7.8 Hz, 1H), 8.48 (s, 1H), 8.37-8.42 (m, 2H), 8.04 (dd, J=8.8, 1.7 Hz, 1H), 7.88 (d, J=2.2 Hz, 1H), 7.82 (d, J=7.8 Hz, 1H), 7.73 (t, J=7.8 Hz, 1H), 7.58 (dd, J=19.8, 8.4 Hz, 2H), 6.97 (dd, J=2.2, 0.7 Hz, 1H), 6.27 (s, 2H), 3.72-3.83 (m, 1H), 3.49-3.63 (m, 2H), 3.35 (d, J=1.4 Hz, 1H), 2.20-2.33 (m, 1H), 1.85-1.97 (m, 1H). MS-ESI calculated value [M+H].sup.+ 468, actually measured value 468.
[0160] The formate of compound 8 can be adjusted the pH of the reaction solution to 8-9 by adding a saturated aqueous sodium carbonate solution to the system, and then, after the extraction with ethyl acetate, the reaction solution was concentrated to obtain compound 8.
Example 3: Compound 9
[0161] ##STR00052##
[0162] Synthesis Route:
##STR00053##
Step 1
[0163] Compound 9b was obtained referring to step 1 of Example 1. .sup.1H NMR (400 MHz, CDCl.sub.3) δ=5.72 (s, 2H), 4.63-4.77 (m, 1H), 1.93 (dd, J=12.5, 3.9 Hz, 2H), 1.75 (dd, J=9.2, 3.9 Hz, 2H), 1.45-1.58 (m, 3H), 1.23-1.42 (m, 3H).
Step 2
[0164] Compound 9c was obtained referring to step 2 of Example 1. MS-ESI calculated value [M+Na].sup.+ 572, actually measured value 572.
Step 3
[0165] A formate of compound 9 was obtained referring to step 3 of Example 1. .sup.1H NMR (400 MHz, CD.sub.3OD) δ=8.68 (d, J=8.0 Hz, 1H), 8.46 (s, 1H), 8.39 (dd, J=7.8, 1.2 Hz, 1H), 7.73 (t, J=7.8 Hz, 1H), 7.67 (d, J=7.8 Hz, 1H), 7.51 (d, J=7.8 Hz, 1H), 6.00 (s, 2H), 4.62-4.71 (m, 1H), 3.77-3.87 (m, 1H), 3.50-3.64 (m, 2H), 3.34-3.40 (m, 2H), 2.24-2.36 (m, 1H), 1.86-2.03 (m, 3H), 1.75 (dt, J=6.4, 3.1 Hz, 2H), 1.24-1.61 (m, 6H). MS-ESI calculated value [M+H].sup.+ 450, actually measured value 450.
[0166] The formate of compound 9 can be adjusted the pH of the reaction solution to 8-9 by adding saturated aqueous sodium carbonate solution to the system, and then, after the extraction with ethyl acetate, the reaction solution was concentrated to obtain compound 9.
Example 4: Compound 10
[0167] ##STR00054##
[0168] Synthesis Route:
##STR00055##
Step 1
[0169] Compound 10b was obtained referring to step 1 of Example 1. .sup.1H NMR (400 MHz, CDCl.sub.3) δ=7.99 (dd, J=8.2, 1.3 Hz, 1H), 7.47 (td, J=7.4, 1.4 Hz, 1H), 7.25-7.32 (m, 2H), 5.96 (s, 2H), 2.65 (s, 3H).
Step 2
[0170] Compound 10c was obtained referring to step 2 of Example 1. MS-ESI calculated value [M+Na].sup.+ 564, actually measured value 564.
Step 3
[0171] A formate of compound 10 was obtained referring to step 3 of Example 1. .sup.1H NMR (400 MHz, CD.sub.3OD) δ=8.69 (d, J=7.8 Hz, 1H), 8.37 (dd, J=7.6, 1.2 Hz, 1H), 7.91 (d, J=7.8 Hz, 1H), 7.79 (d, J=7.8 Hz, 1H), 7.69-7.76 (m, 1H), 7.53 (d, J=7.8 Hz, 1H), 7.45 (td, J=7.4, 1.2 Hz, 1H), 7.22-7.33 (m, 2H), 6.21 (s, 2H), 3.81-3.92 (m, 1H), 3.51-3.64 (m, 2H), 3.41 (dd, J=10.8, 4.2 Hz, 1H), 3.32-3.33 (m, 1H), 2.57 (s, 3H), 2.24-2.40 (m, 1H), 1.90-2.04 (m, 1H). MS-ESI calculated value [M+H].sup.+ 442, actually measured value 442.
[0172] The formate of compound 10 can be adjusted the pH of the reaction solution to 8-9 by adding a saturated aqueous sodium carbonate solution to the system, and then, after the extraction with ethyl acetate, the reaction solution was concentrated to obtain compound 10.
Example 5: Compound 11
[0173] ##STR00056##
[0174] Synthesis Route:
##STR00057##
Step 1
[0175] Compound 11b was obtained referring to step 1 of Example 1. .sup.1H NMR (400 MHz, CDCl.sub.3) δ=7.94-7.98 (m, 2H), 7.25 (d, J=8.4 Hz, 2H), 5.93 (s, 2H), 2.40 (s, 3H).
Step 2
[0176] Compound 11c was obtained referring to step 2 of Example 1. MS-ESI calculated value [M+Na].sup.+ 564, actually measured value 564.
Step 3
[0177] A formate of compound 11 was obtained referring to step 3 of Example 1. .sup.1H NMR (400 MHz, CD.sub.3OD) δ=8.72 (d, J=8.6 Hz, 1H), 8.39 (d, J=7.6 Hz, 1H), 7.94 (d, J=8.0 Hz, 2H), 7.68-7.84 (m, 2H), 7.54 (d, J=7.8 Hz, 1H), 7.31 (d, J=7.8 Hz, 2H), 6.23 (s, 2H), 3.84 (s, 1H), 3.49-3.64 (m, 2H), 3.39 (d, J=15.2 Hz, 2H), 2.42 (s, 3H), 2.30 (s, 1H), 1.97 (s, 1H). MS-ESI calculated value [M+H].sup.+ 442, actually measured value 442.
[0178] The formate of compound 11 can be adjusted the pH of the reaction solution to 8-9 by adding a saturated aqueous sodium carbonate solution to the system, and then, after the extraction with ethyl acetate, the reaction solution was concentrated to obtain compound 11.
Example 6: Compound 12
[0179] ##STR00058##
[0180] Synthesis Route:
##STR00059##
Step 1
[0181] Compound 12b was obtained referring to step 1 of Example 1. .sup.1H NMR (400 MHz, CDCl.sub.3) δ=7.92 (d, J=7.8 Hz, 1H), 7.05-7.15 (m, 2H), 5.95 (s, 2H), 2.63 (s, 3H), 2.39 (s, 3H).
Step 2
[0182] Compound 12c was obtained referring to step 2 of Example 1. MS-ESI calculated value [M+Na].sup.+ 578, actually measured value 578.
Step 3
[0183] Compound 12 was obtained referring to step 3 of Example 1. .sup.1H NMR (400 MHz, CD.sub.3OD) δ=8.73 (d, J=7.8 Hz, 1H), 8.40 (dd, J=7.8, 1.3 Hz, 1H), 7.83 (dd, J=18.2, 8.0 Hz, 2H), 7.74 (t, J=7.8 Hz, 1H), 7.55 (d, J=7.8 Hz, 1H), 7.14 (s, 1H), 7.09 (d, J=8.4 Hz, 1H), 6.20 (s, 2H), 3.79-3.89 (m, 1H), 3.50-3.65 (m, 2H), 3.35-3.45 (m, 2H), 2.56 (s, 3H), 2.35 (s, 3H), 2.24-2.34 (m, 1H), 1.90-2.03 (m, 1H). MS-ESI calculated value [M+H].sup.+ 456, actually measured value 456.
Example 7: Compound 13
[0184] ##STR00060##
[0185] Synthesis Route:
##STR00061##
Step 1
[0186] Compound 13b was obtained referring to step 1 of Example 1. .sup.1H NMR (400 MHz, CDCl.sub.3) δ=7.79-7.85 (m, 1H), 7.59 (dd, J=4.8, 1.2 Hz, 1H), 7.04-7.10 (m, 1H), 5.79-5.90 (m, 2H)
Step 2
[0187] Compound 13c was obtained referring to step 2 of Example 1. MS-ESI calculated value [M+Na].sup.+ 556, actually measured value 556.
Step 3
[0188] A formate of compound 13 was obtained referring to step 3 of Example 1. .sup.1H NMR (400 MHz, CD.sub.3OD) δ=8.71 (d, J=7.8 Hz, 1H), 8.40 (s, 1H), 8.39 (dd, J=7.8, 1.3 Hz, 1H), 7.89 (dd, J=3.8, 1.2 Hz, 1H), 7.83 (dd, J=5.0, 1.2 Hz, 1H), 7.69-7.79 (m, 2H), 7.54 (d, J=7.8 Hz, 1H), 7.19 (dd, J=5.0, 3.9 Hz, 1H), 6.21 (s, 2H), 3.70-3.83 (m, 1H), 3.48-3.65 (m, 2H), 3.33-3.35 (m, 2H), 2.21-2.35 (m, 1H), 1.84-1.99 (m, 1H). MS-ESI calculated value [M+H].sup.+ 434, actually measured value 434.
[0189] The formate of compound 13 can be adjusted the pH of the reaction solution to 8-9 by adding a saturated aqueous sodium carbonate solution to the system, and then, after the extraction with ethyl acetate, the reaction solution was concentrated to obtain compound 13.
Example 8: Compound 14
[0190] ##STR00062##
[0191] Synthesis Route:
##STR00063##
Step 1
[0192] Compound 14b was obtained referring to step 1 of Example 1. .sup.1H NMR (400 MHz, CDCl.sub.3) δ=7.98-8.05 (m, 1H), 7.37-7.42 (m, 1H), 6.71 (dd, J=1.8, 0.7 Hz, 1H), 5.81 (s, 2H).
Step 2
[0193] Compound 14c was obtained referring to step 2 of Example 1. MS-ESI calculated value [M+Na].sup.+ 540, actually measured value 540.
Step 3
[0194] A formate of compound 14 was obtained referring to step 3 of Example 1. .sup.1H NMR (400 MHz, CD.sub.3OD) δ=8.70 (d, J=7.8 Hz, 1H), 8.49 (s, 1H), 8.39 (dd, J=7.8, 1.3 Hz, 1H), 8.23-8.26 (m, 1H), 7.69-7.78 (m, 2H), 7.62 (t, J=1.8 Hz, 1H), 7.53 (d, J=7.8 Hz, 1H), 6.80 (dd, J=1.8, 0.7 Hz, 1H), 6.17 (s, 2H), 3.74-3.83 (m, 1H), 3.50-3.62 (m, 2H), 3.34-3.38 (m, 1H), 3.29-3.32 (m, 1H), 2.20-2.34 (m, 1H), 1.85-1.99 (m, 1H). MS-ESI calculated value [M+H].sup.+ 418, actually measured value 418.
[0195] The formate of compound 14 can be adjusted the pH of the reaction solution to 8-9 by adding a saturated aqueous sodium carbonate solution to the system, and then, after the extraction with ethyl acetate, the reaction solution was concentrated to obtain compound 14.
Example 9: Compound 15
[0196] ##STR00064##
[0197] Synthesis Route:
##STR00065##
Step 1
[0198] Compound 15b was obtained referring to step 2 of Example 1. MS-ESI calculated value [M+Na].sup.+ 516, actually measured value 516.
Step 2
[0199] A formate of compound 15 was obtained referring to step 3 of Example 1. .sup.1H NMR (400 MHz, CD.sub.3OD) δ=8.67 (d, J=7.8 Hz, 1H), 8.39 (s, 1H), 8.37 (dd, J=7.8, 1.3 Hz, 1H), 7.72 (t, J=7.8 Hz, 1H), 7.66 (d, J=7.8 Hz, 1H), 7.50 (d, J=7.8 Hz, 1H), 5.99 (s, 2H), 3.76-3.89 (m, 1H), 3.50-3.65 (m, 2H), 3.33 (d, J=1.6 Hz, 2H), 2.62 (spt, J=7.0 Hz, 1H), 2.22-2.37 (m, 1H), 1.90-2.02 (m, 1H), 1.17 (d, J=7.0 Hz, 6H), 0.00-0.00 (m, 1H). MS-ESI calculated value [M+H].sup.+ 394, actually measured value 394.
[0200] The formate of compound 15 can be adjusted the pH of the reaction solution to 8-9 by adding a saturated aqueous sodium carbonate solution to the system, and then, after the extraction with ethyl acetate, the reaction solution was concentrated to obtain compound 15.
Example 10: Compound 16
[0201] ##STR00066##
[0202] Synthesis Route:
##STR00067##
Step 1
[0203] Compound 16b was obtained referring to step 2 of Example 1. .sup.1H NMR (400 MHz, CDCl.sub.3) δ=8.72 (dd, J=8.0, 1.2 Hz, 1H), 8.32 (dd, J=7.8, 1.2 Hz, 1H), 7.59 (t, J=8.0 Hz, 1H), 7.34-7.48 (m, 2H), 5.95 (s, 2H), 4.66 (s, 1H), 4.18 (s, 1H), 3.37-3.61 (m, 2H), 3.27 (s, 2H), 2.09-2.18 (m, 4H), 1.84 (d, J=5.6 Hz, 1H), 1.42 (s, 9H).
Step 2
[0204] A formate of compound 16 was obtained referring to step 3 of Example 1. .sup.1H NMR (400 MHz, CD.sub.3OD) δ=8.68 (d, J=7.6 Hz, 1H), 8.44 (s, 1H), 8.37 (dd, J=7.6, 1.3 Hz, 1H), 7.72 (t, J=8.0 Hz, 1H), 7.65 (d, J=8.0 Hz, 1H), 7.50 (d, J=8.0 Hz, 1H), 5.97 (s, 2H), 3.75-3.88 (m, 1H), 3.46-3.63 (m, 2H), 3.32-3.43 (m, 2H), 2.21-2.36 (m, 1H), 2.09 (s, 3H), 1.86-2.01 (m, 1H). MS-ESI calculated value [M+H].sup.+ 366, actually measured value 366.
[0205] The formate of compound 16 can be adjusted the pH of the reaction solution to 8-9 by adding a saturated aqueous sodium carbonate solution to the system, and then, after the extraction with ethyl acetate, the reaction solution was concentrated to obtain compound 16.
Example 11: Compound 17
[0206] ##STR00068##
[0207] Synthesis Route:
##STR00069##
Step 1
[0208] Compound 17b was obtained referring to step 2 of Example 1. .sup.1H NMR (400 MHz, CDCl.sub.3) δ=8.66-8.78 (m, 1H), 8.31 (dd, J=7.8, 1.4 Hz, 1H), 7.59 (t, J=7.8 Hz, 1H), 7.36-7.48 (m, 2H), 5.96 (s, 2H), 4.68 (s, 1H), 4.14-4.30 (m, 1H), 3.36-3.64 (m, 2H), 3.27 (s, 2H), 2.36 (t, J=7.4 Hz, 2H), 2.14 (td, J=13.8, 6.8 Hz, 1H), 1.78-1.88 (m, 1H), 1.63-1.70 (m, 2H), 1.42 (s, 9H), 0.94 (t, J=7.4 Hz, 3H).
Step 2
[0209] A formate of compound 17 was obtained referring to step 3 of Example 1. .sup.1H NMR (400 MHz, CD.sub.3OD) δ=8.67 (d, J=7.8 Hz, 1H), 8.48 (s, 1H), 8.37 (dd, J=7.8, 1.2 Hz, 1H), 7.71 (t, J=7.8 Hz, 1H), 7.65 (d, J=7.8 Hz, 1H), 7.49 (d, J=7.8 Hz, 1H), 5.98 (s, 2H), 3.73-3.89 (m, 1H), 3.46-3.64 (m, 2H), 3.32-3.41 (m, 2H), 2.36 (t, J=7.3 Hz, 2H), 2.18-2.30 (m, 1H), 1.84-1.98 (m, 1H), 1.64 (sxt, J=7.4 Hz, 2H), 0.93 (t, J=7.4 Hz, 3H). MS-ESI calculated value [M+H].sup.+ 394, actually measured value 394.
[0210] The formate of compound 17 can be adjusted the pH of the reaction solution to 8-9 by adding a saturated aqueous sodium carbonate solution to the system, and then, after the extraction with ethyl acetate, the reaction solution was concentrated to obtain compound 17.
Example 12: Compound 18
[0211] ##STR00070##
##STR00071##
Step 1
[0212] Compound 18b was obtained referring to step 1 of Example 1.
Step 2
[0213] Compound 18c was obtained referring to step 2 of Example 1. .sup.1H NMR (400 MHz, CDCl.sub.3) δ=8.72 (d, J=7.8 Hz, 1H), 8.28-8.35 (m, 1H), 7.59 (t, J=7.8 Hz, 1H), 7.37-7.48 (m, 2H), 5.90-6.03 (m, 1H), 5.96 (s, 1H), 4.64 (s, 1H), 4.17 (d, J=9.4 Hz, 1H), 3.38-3.60 (m, 2H), 3.15-3.36 (m, 3H), 2.13-2.37 (m, 5H), 1.81-2.02 (m, 3H), 1.43 (s, 9H).
Step 3
[0214] A formate of compound 18 was obtained referring to step 3 of Example 1. .sup.1H NMR (400 MHz, CD.sub.3OD) δ=8.67 (d, J=7.8 Hz, 1H), 8.38 (s, 1H), 8.37 (dd, J=7.8, 1.2 Hz, 1H), 7.71 (t, J=7.8 Hz, 1H), 7.65 (d, J=8.0 Hz, 1H), 7.49 (d, J=8.0 Hz, 1H), 5.98 (s, 2H), 3.74-3.87 (m, 1H), 3.45-3.64 (m, 2H), 3.32-3.38 (m, 2H), 3.14-3.27 (m, 1H), 2.14-2.36 (m, 5H), 1.83-2.07 (m, 3H). MS-ESI calculated value [M+H].sup.+ 406, actually measured value 406.
[0215] The formate of compound 18 can be adjusted the pH of the reaction solution to 8-9 by adding a saturated aqueous sodium carbonate solution to the system, and then, after the extraction with ethyl acetate, the reaction solution was concentrated to obtain compound 18.
Example 13: Compound 19
[0216] ##STR00072##
[0217] Synthesis Route:
##STR00073##
Step 1
[0218] Compound 19b was obtained referring to step 1 of Example 1.
Step 2
[0219] Compound 19c was obtained referring to step 2 of Example 1. MS-ESI calculated value [M+Na].sup.+ 542, actually measured value 542.
Step 3
[0220] A formate of compound 19 was obtained referring to step 3 of Example 1. .sup.1H NMR (400 MHz, CD.sub.3OD) δ=8.68 (d, J=7.6 Hz, 1H), 8.37 (dd, J=7.6, 1.4 Hz, 2H), 7.72 (t, J=8.0 Hz, 1H), 7.65 (d, J=7.8 Hz, 1H), 7.49 (d, J=8.4 Hz, 1H), 5.97 (s, 2H), 4.62 (s, 1H), 3.78-3.87 (m, 1H), 3.55-3.65 (m, 1H), 3.51 (dd, J=10.8, 6.4 Hz, 1H), 3.38 (dd, J=10.8, 4.0 Hz, 1H), 2.76-2.86 (m, 1H), 2.19-2.35 (m, 1H), 1.84-2.01 (m, 3H), 1.73-1.83 (m, 2H), 1.53-1.72 (m, 4H). MS-ESI calculated value [M+H].sup.+ 420, actually measured value 420.
[0221] The formate of compound 19 can be adjusted the pH of the reaction solution to 8-9 by adding a saturated aqueous sodium carbonate solution to the system, and then, after the extraction with ethyl acetate, the reaction solution was concentrated to obtain compound 19.
Example 14: Compound 20
[0222] ##STR00074##
[0223] Synthesis Route:
##STR00075##
Step 1
[0224] Compound 20b was obtained referring to step 1 of Example 1.
Step 2
[0225] Compound 20c was obtained referring to step 2 of Example 1.
Step 3
[0226] A formate of compound 20 was obtained referring to step 3 of Example 1. .sup.1H NMR (400 MHz, CD.sub.3OD) δ=8.69 (d, J=7.8 Hz, 1H), 8.40 (s, 1H), 8.39 (dd, J=7.8, 1.2 Hz, 1H), 7.73 (t, J=7.8 Hz, 1H), 7.66 (d, J=8.0 Hz, 1H), 7.51 (d, J=8.0 Hz, 1H), 5.99 (s, 2H), 4.58 (s, 2H), 3.71-3.83 (m, 1H), 3.46-3.65 (m, 2H), 2.40 (tt, J=11.0, 3.6 Hz, 1H), 2.21-2.33 (m, 1H), 1.85-2.00 (m, 3H), 1.71-1.82 (m, 2H), 1.65 (d, J=8.8 Hz, 1H), 1.22-1.50 (m, 5H). MS-ESI calculated value [M+H].sup.+ 434, actually measured value 434.
[0227] The formate of compound 20 can be adjusted the pH of the reaction solution to 8-9 by adding a saturated aqueous sodium carbonate solution to the system, and then, after the extraction with ethyl acetate, the reaction solution was concentrated to obtain compound 20.
Example 15: Compound 21
[0228] ##STR00076##
[0229] Synthesis Route:
##STR00077##
Step 1
[0230] Compound 21b was obtained referring to step 1 of Example 1.
Step 2
[0231] Compound 21c was obtained referring to step 2 of Example 1. .sup.1H NMR (400 MHz, CDCl.sub.3) δ=8.70 (dd, J=8.0, 1.1 Hz, 1H), 8.26-8.35 (m, 1H), 7.59 (t, J=7.8 Hz, 1H), 7.34-7.45 (m, 2H), 5.97 (s, 2H), 4.59-4.83 (m, 1H), 4.06-4.29 (m, 1H), 3.94 (dt, J=11.4, 3.5 Hz, 2H), 3.48-3.54 (m, 1H), 3.34-3.42 (m, 2H), 3.16-3.33 (m, 2H), 2.56-2.69 (m, 1H), 2.08-2.20 (m, 1H), 1.74-1.84 (m, 6H), 1.34-1.43 (m, 9H).
Step 3
[0232] A formate of compound 21 was obtained referring to step 3 of Example 1. .sup.1H NMR (400 MHz, CD.sub.3OD) δ=8.67 (d, J=8.2 Hz, 1H), 8.45 (s, 1H), 8.37 (d, J=7.8 Hz, 1H), 7.71 (t, J=8.0 Hz, 1H), 7.65 (d, J=7.8 Hz, 1H), 7.49 (d, J=8.0 Hz, 1H), 7.45-7.53 (m, 1H), 6.00 (s, 2H), 3.89 (dt, J=11.7, 3.5 Hz, 2H), 3.72-3.82 (m, 1H), 3.48-3.63 (m, 2H), 3.43 (td, J=11.4, 2.4 Hz, 2H), 3.35 (d, J=3.6 Hz, 1H), 2.59-2.73 (m, 1H), 2.18-2.33 (m, 1H), 1.92 (td, J=13.5, 5.8 Hz, 1H), 1.61-1.87 (m, 5H). MS-ESI calculated value [M+H].sup.+ 436, actually measured value 436.
[0233] The formate of compound 21 can be adjusted the pH of the reaction solution to 8-9 by adding a saturated aqueous sodium carbonate solution to the system, and then, after the extraction with ethyl acetate, the reaction solution was concentrated to obtain compound 21.
Example 16: Compound 22
[0234] ##STR00078##
##STR00079##
Step 1
[0235] Compound 22b was obtained referring to step 1 of Example 1.
Step 2
[0236] Compound 22c was obtained referring to step 2 of Example 1. .sup.1H NMR (400 MHz, CDCl.sub.3) δ=8.72 (dd, J=8.0, 0.8 Hz, 1H), 8.32 (dd, J=7.8, 1.4 Hz, 1H), 7.59 (t, J=7.8 Hz, 1H), 7.34-7.48 (m, 2H), 5.96 (s, 2H), 4.67 (s, 1H), 4.18 (s, 1H), 3.36-3.64 (m, 2H), 3.27 (s, 2H), 2.37 (t, J=7.6 Hz, 2H), 2.14 (dq, J=14.4, 6.6 Hz, 1H), 1.75-1.91 (m, 1H), 1.65-1.68 (m, 1H), 1.59-1.62 (m, 1H), 1.42 (s, 9H), 1.27-1.33 (m, 4H), 0.78-0.89 (m, 3H).
Step 3
[0237] A formate of compound 22 was obtained referring to step 3 of Example 1. .sup.1H NMR (400 MHz, CD.sub.3OD) δ=8.67 (d, J=8.0 Hz, 1H), 8.48 (s, 1H), 8.37 (d, J=7.4 Hz, 1H), 7.71 (t, J=7.8 Hz, 1H), 7.64 (d, J=8.0 Hz, 1H), 7.49 (d, J=7.8 Hz, 1H), 5.98 (s, 2H), 4.57 (s, 1H), 3.71-3.74 (m, 1H), 3.46-3.64 (m, 2H), 2.37 (t, J=7.6 Hz, 2H), 2.23-2.24 (m, 1H), 1.88-1.89 (m, 1H), 1.53-1.71 (m, 2H), 1.27-1.30 (m, 5H), 0.85 (t, J=6.8 Hz, 3H). MS-ESI calculated value [M+H].sup.+ 422, actually measured value 422.
[0238] The formate of compound 22 can be adjusted the pH of the reaction solution to 8-9 by adding a saturated aqueous sodium carbonate solution to the system, and then, after the extraction with ethyl acetate, the reaction solution was concentrated to obtain compound 22.
Example 17: Compound 23
[0239] ##STR00080##
[0240] Synthesis Route:
##STR00081##
Step 1
[0241] To a solution of compound 23b (1.0 g, 7.76 mmol) and pyridine (920 mg, 11.6 mmol) in dichloromethane (10 mL) was added compound 23a (466 mg, 7.76 mmol) dropwise at 0° C., stirred for 16 hours at 30° C. Hydrochloric acid (1M, 10 mL) was added to the reaction solution, stirred for 5 minutes; the organic phase was separated, washed with water (10 mL), dried with anhydrous magnesium sulfate, filtered and concentrated to obtain compound 23c. .sup.1H NMR (400 MHz, CDCl.sub.3) δ=5.73 (s, 2H), 4.19 (t, J=6.6 Hz, 2H), 1.73 (sxt, J=7.2 Hz, 2H), 0.97 (t, J=7.2 Hz, 3H).
Step 2
[0242] Compound 23d was obtained referring to step 2 of Example 1. MS-ESI calculated value [M+Na].sup.+ 532, actually measured value 532.
Step 3
[0243] A formate of compound 23 was obtained referring to step 3 of Example 1. .sup.1H NMR (400 MHz, CD.sub.3OD) δ=8.67 (d, J=8.0 Hz, 1H), 8.38 (s, 1H), 8.37 (d, J=7.8 Hz, 1H), 7.71 (t, J=7.8 Hz, 1H), 7.65 (d, J=8.0 Hz, 1H), 7.50 (d, J=8.0 Hz, 1H), 5.99 (s, 2H), 4.58 (s, 1H), 4.13 (t, J=6.6 Hz, 2H), 3.76-3.88 (m, 1H), 3.46-3.64 (m, 2H), 3.34-3.40 (m, 1H), 2.20-2.34 (m, 1H), 1.94 (td, J=13.6, 5.8 Hz, 1H), 1.68 (sxt, J=7.2 Hz, 2H), 0.94 (t, J=7.6 Hz, 3H). MS-ESI calculated value [M+H].sup.+ 410, actually measured value 410.
[0244] The formate of compound 23 can be adjusted the pH of the reaction solution to 8-9 by adding a saturated aqueous sodium carbonate solution to the system, and then, after the extraction with ethyl acetate, the reaction solution was concentrated to obtain compound 23.
Example 18: Compound 24
[0245] ##STR00082##
[0246] Synthesis Route:
##STR00083##
Step 1
[0247] Compound 24b was obtained referring to step 1 of Example 1.
Step 2
[0248] Compound 24c was obtained referring to step 2 of Example 1. .sup.1H NMR (400 MHz, CDCl.sub.3) δ ppm 8.72 (d, J=7.2 Hz, 1H), 8.32 (dd, J=7.8, 1.4 Hz, 1H), 7.60 (t, J=7.8 Hz, 1H), 7.33-7.43 (m, 3H), 7.27-7.32 (m, 4H), 5.97 (s, 2H), 4.53-4.75 (m, 1H), 4.18 (s, 1H), 3.70 (s, 2H), 3.37-3.60 (m, 2H), 3.25 (s, 2H), 2.08-2.24 (m, 1H), 1.77-1.91 (m, 1H), 1.43 (s, 9H).
Step 3
[0249] A formate of compound 24 was obtained referring to step 3 of Example 1. .sup.1H NMR (400 MHz, CD.sub.3OD) δ=8.66 (d, J=7.8 Hz, 1H), 8.45 (s, 1H), 8.37 (dd, J=7.8, 1.2 Hz, 1H), 7.71 (t, J=8.0 Hz, 1H), 7.60 (d, J=8.0 Hz, 1H), 7.46 (d, J=7.8 Hz, 1H), 7.16-7.32 (m, 4H), 6.00 (s, 2H), 3.74-3.85 (m, 1H), 3.70 (s, 2H), 3.46-3.61 (m, 2H), 3.33-3.38 (m, 1H), 3.25-3.30 (m, 1H), 2.18-2.33 (m, 1H), 1.83-2.02 (m, 1H). MS-ESI calculated value [M+Na].sup.+ 464, actually measured value 464.
[0250] The formate of compound 24 can be adjusted the pH of the reaction solution to 8-9 by adding a saturated aqueous sodium carbonate solution to the system, and then, after the extraction with ethyl acetate, the reaction solution was concentrated to obtain compound 24.
Example 19: Compound 25
[0251] ##STR00084##
[0252] Synthesis Route:
##STR00085##
Step 1
[0253] Compound 25a was obtained referring to step 1 of Example 17.
Step 2
[0254] Compound 25b was obtained referring to step 2 of Example 1. MS-ESI calculated value [M+Na].sup.+ 504, actually measured value 504.
Step 3
[0255] A formate of compound 25 was obtained referring to step 3 of Example 1. .sup.1H NMR (400 MHz, CD.sub.3OD) δ=8.68 (d, J=7.8 Hz, 1H), 8.38 (dd, J=7.8, 1.2 Hz, 1H), 7.72 (t, J=8.0 Hz, 1H), 7.65 (d, J=7.8 Hz, 1H), 7.50 (d, J=8.0 Hz, 1H), 5.99 (s, 2H), 3.83-3.90 (m, 1H), 3.80 (s, 3H), 3.59-3.68 (m, 1H), 3.44-3.59 (m, 2H), 3.37-3.42 (m, 1H), 2.23-2.37 (m, 1H), 1.97 (td, J=13.6, 6.0 Hz, 1H). MS-ESI calculated value [M+H].sup.+ 382, actually measured value 382.
[0256] The formate of compound 25 can be adjusted the pH of the reaction solution to 8-9 by adding a saturated aqueous sodium carbonate solution to the system, and then, after the extraction with ethyl acetate, the reaction solution was concentrated to obtain compound 25.
Example 20: Compound 26
[0257] ##STR00086##
[0258] Synthesis Route:
##STR00087##
Step 1
[0259] Compound 26b was obtained referring to step 1 of Example 17.
Step 2
[0260] Compound 26c was obtained referring to step 2 of Example 1. .sup.1H NMR (400 MHz, CDCl.sub.3) δ=8.72 (d, J=8.0 Hz, 1H), 8.32 (d, J=7.8 Hz, 1H), 7.59 (t, J=8.0 Hz, 1H), 7.43 (s, 2H), 5.98 (s, 2H), 4.65 (s, 1H), 4.18 (t, J=6.8 Hz, 2H), 3.46 (d, J=18.0 Hz, 2H), 3.28 (s, 2H), 2.14 (dq, J=13.6, 7.2 Hz, 1H), 1.84 (s, 1H), 1.64-1.72 (m, 2H), 1.43 (s, 9H), 1.27-1.37 (m, 6H), 0.88 (t, J=6.8 Hz, 3H).
Step 3
[0261] A formate of compound 26 was obtained referring to step 3 of Example 1. .sup.1H NMR (400 MHz, CD.sub.3OD) δ=8.67 (d, J=8.0 Hz, 1H), 8.45 (s, 1H), 8.37 (dd, J=7.8, 1.2 Hz, 1H), 7.71 (t, J=8.0 Hz, 1H), 7.65 (d, J=7.8 Hz, 1H), 7.50 (d, J=8.0 Hz, 1H), 5.99 (s, 2H), 4.17 (t, J=6.6 Hz, 2H), 3.76-3.85 (m, 1H), 3.48-3.63 (m, 2H), 3.32-3.40 (m, 2H), 2.22-2.39 (m, 1H), 1.89-1.99 (m, 1H), 1.60-1.72 (m, 2H), 1.30-1.40 (m, 6H), 0.83-0.94 (m, 3H). MS-ESI calculated value [M+H].sup.+ 452, actually measured value 452.
[0262] The formate of compound 26 can be adjusted the pH of the reaction solution to 8-9 by adding a saturated aqueous sodium carbonate solution to the system, and then, after the extraction with ethyl acetate, the reaction solution was concentrated to obtain compound 26.
Example 21: Compound 27
[0263] ##STR00088##
[0264] Synthesis Route:
##STR00089##
Step 1
[0265] Compound 27b was obtained referring to step 2 of Example 1. MS-ESI calculated value [M+Na].sup.+ 550, actually measured value 550.
Step 2
[0266] A formate of compound 27 was obtained referring to step 3 of Example 1. .sup.1H NMR (400 MHz, CDCl.sub.3) δ=8.73 (d, J=8.0 Hz, 1H), 8.35 (d, J=7.8 Hz, 1H), 8.09 (d, J=7.4 Hz, 2H), 7.54-7.64 (m, 3H), 7.45 (dt, J=7.8, 3.8 Hz, 3H), 6.22 (s, 2H), 3.62 (s, 1H), 3.46-3.55 (m, 2H), 3.31-3.42 (m, 1H), 3.10 (dd, J=9.9, 4.1 Hz, 1H), 2.03-2.22 (m, 2H). MS-ESI calculated value [M+H].sup.+ 428, actually measured value 428.
[0267] The formate of compound 27 can be adjusted the pH of the reaction solution to 8-9 by adding a saturated aqueous sodium carbonate solution to the system, and then, after the extraction with ethyl acetate, the reaction solution was concentrated to obtain compound 27.
Example 22: Compound 28
[0268] ##STR00090##
[0269] Synthesis Route:
##STR00091##
Step 1
[0270] Compound 28b was obtained referring to step 2 of Example 1. MS-ESI calculated value [M+Na].sup.+ 518, actually measured value 518.
Step 2
[0271] A formate of compound 28 was obtained referring to step 3 of Example 1. .sup.1H NMR (400 MHz, deuterated acetone) 6 ppm 8.63 (d, J=7.4 Hz, 1H), 8.43 (dd, J=7.8, 1.4 Hz, 1H), 8.12 (s, 1H), 7.64-7.74 (m, 2H), 7.49 (d, J=8.0 Hz, 1H), 6.03 (d, J=1.0 Hz, 2H), 4.22 (q, J=7.0 Hz, 2H), 4.16 (t, J=4.6 Hz, 1H), 3.59 (dd, J=9.8, 5.8 Hz, 1H), 3.45-3.52 (m, 2H), 3.22 (dd, J=9.8, 3.9 Hz, 1H), 2.11-2.16 (m, 1H), 1.76 (td, J=12.4, 5.2 Hz, 1H), 1.27 (t, J=7.0 Hz, 3H). MS-ESI calculated value [M+H].sup.+ 396, actually measured value 396.
[0272] The formate of compound 28 can be adjusted the pH of the reaction solution to 8-9 by adding a saturated aqueous sodium carbonate solution to the system, and then, after the extraction with ethyl acetate, the reaction solution was concentrated to obtain compound 28.
Example 23: Compound 29
[0273] ##STR00092##
[0274] Synthesis Route:
##STR00093##
Step 1
[0275] Compound 29b was obtained referring to step 2 of Example 1. MS-ESI calculated value [M+Na].sup.+ 532, actually measured value 532.
Step 2
[0276] A formate of compound 29 was obtained referring to step 3 of Example 1. .sup.1H NMR (400 MHz, CD.sub.3OD) δ=8.69 (d, J=7.8 Hz, 1H), 8.49 (s, 1H), 8.39 (dd, J=7.8, 1.3 Hz, 1H), 7.73 (t, J=7.8 Hz, 1H), 7.67 (d, J=7.8 Hz, 1H), 7.51 (d, J=7.8 Hz, 1H), 5.99 (s, 2H), 3.76-3.88 (m, 1H), 3.49-3.66 (m, 2H), 3.28-3.40 (m, 3H), 2.22-2.38 (m, 1H), 1.87-2.01 (m, 1H), 1.30 (d, J=6.4 Hz, 7H). MS-ESI calculated value [M+H].sup.+ 410, actually measured value 410.
[0277] The formate of compound 29 can be adjusted the pH of the reaction solution to 8-9 by adding a saturated aqueous sodium carbonate solution to the system, and then, after the extraction with ethyl acetate, the reaction solution was concentrated to obtain compound 29.
Example 24: Compound 30
[0278] ##STR00094##
[0279] Synthesis Route:
##STR00095##
Step 1
[0280] Compound 30b was obtained referring to step 2 of Example 1. MS-ESI calculated value [M+Na].sup.+ 530, actually measured value 530.
Step 2
[0281] A formate of compound 30 was obtained referring to step 3 of Example 1. .sup.1H NMR (400 MHz, CDCl.sub.3) δ ppm 8.63 (d, J=7.8 Hz, 1H), 8.25 (d, J=7.6 Hz, 1H), 8.21 (s, 1H), 7.51 (t, J=7.8 Hz, 1H), 7.34-7.40 (m, 1H), 7.29-7.34 (m, 1H), 5.87 (s, 2H), 3.69 (s, 1H), 3.50 (s, 1H), 3.42 (s, 1H), 3.26 (s, 2H), 2.10 (s, 1H), 1.85 (s, 1H), 1.13 (s, 9H). MS-ESI calculated value [M+H].sup.+ 408, actually measured value 408.
[0282] The formate of compound 30 can be adjusted the pH of the reaction solution to 8-9 by adding a saturated aqueous sodium carbonate solution to the system, and then, after the extraction with ethyl acetate, the reaction solution was concentrated to obtain compound 30.
Example 25: Compound 31
[0283] ##STR00096##
##STR00097##
Step 1
[0284] Compound 31a was obtained referring to step 2 of Example 1. MS-ESI calculated value [M+H].sup.+ 554, actually measured value 554.
Step 2
[0285] Trifluoroacetic acid (1.0 mL) was added to a microwave tube and compound 31a (79 mg, 143 μmol) was added to the microwave tube, microwaving for 1 hour at 60° C. The solvent was removed by concentration under reduced pressure, and the crude product was purified by high performance liquid chromatography (acid, formic acid system) to obtain a formate of compound 31. .sup.1H NMR (400 MHz, CD.sub.3OD) δ=8.66 (d, J=8.1 Hz, 1H), 8.50 (s, 1H), 8.45-8.39 (m, 1H), 7.73-7.63 (m, 2H), 7.17 (d, J=7.9 Hz, 1H), 5.97 (s, 2H), 4.12-4.01 (m, 1H), 3.66-3.55 (m, 1H), 3.41 (br d, J=4.5 Hz, 1H), 2.65-2.57 (m, 1H), 2.56-2.46 (m, 1H), 2.13-2.02 (m, 1H), 1.23-1.17 (m, 1H), 1.15 (d, J=7.0 Hz, 7H), 0.80 (t, J=8.9 Hz, 2H). MS-ESI calculated value [M+H].sup.+ 420, actually measured value 420.
[0286] The formate of compound 31 can be adjusted the pH of the reaction solution to 8-9 by adding a saturated aqueous sodium carbonate solution to the system, and then, after the extraction with ethyl acetate, the reaction solution was concentrated to obtain compound 31.
Example 26: Compound 32
[0287] ##STR00098##
[0288] Synthesis Route:
##STR00099##
Step 1
[0289] Compound 32a was obtained referring to step 2 of Example 1. MS-ESI calculated value [M+Na].sup.+ 590, actually measured value 590.
Step 2
[0290] A formate of compound 32 was obtained referring to step 2 of Example 25. .sup.1H NMR (400 MHz, CD.sub.3OD) δ=8.67 (d, J=7.6 Hz, 1H), 8.47 (s, 1H), 8.45-8.41 (m, 1H), 7.74-7.61 (m, 2H), 7.19 (d, J=7.8 Hz, 1H), 5.97 (s, 2H), 4.12-3.99 (m, 1H), 3.68-3.54 (m, 1H), 3.36-3.33 (m, 1H), 2.59-2.42 (m, 1H), 2.08-1.94 (m, 1H), 1.19 (s, 9H), 1.18-1.06 (m, 2H), 0.88-0.74 (m, 2H). MS-ESI calculated value [M+H]+ 434, actually measured value 434.
[0291] The formate of compound 32 can be adjusted the pH of the reaction solution to 8-9 by adding a saturated aqueous sodium carbonate solution to the system, and then, after the extraction with ethyl acetate, the reaction solution was concentrated to obtain compound 32.
Example 27: Compound 33
[0292] ##STR00100##
[0293] Synthesis Route:
##STR00101##
Step 1
[0294] Compound 33a was obtained referring to step 2 of Example 1. MS-ESI calculated value [M+H].sup.+ 588, actually measured value 588.
Step 2
[0295] A formate of compound 33 was obtained referring to step 2 of Example 25. .sup.1H NMR (400 MHz, CD.sub.3OD) δ=8.66 (d, J=7.9 Hz, 1H), 8.48 (s, 1H), 8.43-8.37 (m, 1H), 8.05-7.99 (m, 2H), 7.78 (d, J=7.9 Hz, 1H), 7.68 (t, J=7.9 Hz, 1H), 7.64-7.57 (m, 1H), 7.50-7.43 (m, 2H), 7.19 (d, J=7.8 Hz, 1H), 6.22 (s, 2H), 4.12-4.00 (m, 1H), 3.65-3.55 (m, 1H), 3.47-3.39 (m, 1H), 2.59-2.44 (m, 1H), 2.15-2.03 (m, 1H), 1.26-1.10 (m, 2H), 0.83-0.74 (m, 2H). MS-ESI calculated value [M+H].sup.+ 454, actually measured value 454.
[0296] The formate of compound 33 can be adjusted the pH of the reaction solution to 8-9 by adding a saturated aqueous sodium carbonate solution to the system, and then, after the extraction with ethyl acetate, the reaction solution was concentrated to obtain compound 33.
Example 28: Compound 34
[0297] ##STR00102##
[0298] Synthesis Route:
##STR00103##
Step 1
[0299] Compound 34a was obtained referring to step 2 of Example 1. MS-ESI calculated value [M+Na].sup.+ 592, actually measured value 592.
Step 2
[0300] A formate of compound 34 was obtained referring to step 2 of Example 25. .sup.1H NMR (400 MHz, CD.sub.3OD) δ=8.66 (d, J=8.1 Hz, 1H), 8.50 (br s, 1H), 8.46-8.40 (m, 1H), 7.72-7.63 (m, 2H), 7.23 (d, J=7.8 Hz, 1H), 5.97 (s, 2H), 4.92-4.90 (m, 1H), 4.02-3.92 (m, 1H), 3.66-3.55 (m, 1H), 3.29-3.20 (m, 1H), 2.51-2.36 (m, 1H), 2.00-1.88 (m, 1H), 1.28 (d, J=6.4 Hz, 6H), 1.18-1.02 (m, 2H), 0.95-0.87 (m, 1H), 0.79-0.72 (m, 1H). MS-ESI calculated value [M+H].sup.+ 436, actually measured value 436.
[0301] The formate of compound 34 can be adjusted the pH of the reaction solution to 8-9 by adding a saturated aqueous sodium carbonate solution to the system, and then, after the extraction with ethyl acetate, the reaction solution was concentrated to obtain compound 34.
Example 29: Compound 35
[0302] ##STR00104##
[0303] Synthesis Route:
##STR00105##
Step 1
[0304] Compound 35a was obtained referring to step 2 of Example 1. MS-ESI calculated value [M+Na].sup.+ 600, actually measured value 600.
Step 2
[0305] A formate of compound 35 was obtained referring to step 2 of Example 25. .sup.1H NMR (400 MHz, CD.sub.3OD) δ=8.68 (d, J=7.9 Hz, 1H), 8.46 (br s, 1H), 8.44-8.41 (m, 1H), 8.23 (s, 1H), 7.78-7.67 (m, 2H), 7.60 (t, J=1.7 Hz, 1H), 7.20 (d, J=7.8 Hz, 1H), 6.79 (d, J=1.2 Hz, 1H), 6.15 (s, 2H), 4.12-4.02 (m, 1H), 3.65-3.55 (m, 1H), 3.43-3.35 (m, 1H), 2.59-2.45 (m, 1H), 2.11-2.00 (m, 1H), 1.26-1.08 (m, 2H), 0.87-0.74 (m, 2H). MS-ESI calculated value [M+H].sup.+ 444, actually measured value 444.
[0306] The formate of compound 35 can be adjusted the pH of the reaction solution to 8-9 by adding a saturated aqueous sodium carbonate solution to the system, and then, after the extraction with ethyl acetate, the reaction solution was concentrated to obtain compound 35.
Example 30: Compound 36
[0307] ##STR00106##
[0308] Synthesis Route:
##STR00107##
Step 1
[0309] Compound 36a was obtained referring to step 2 of Example 1. MS-ESI calculated value [M+H].sup.+ 554, actually measured value 554.
Step 2
[0310] A formate of compound 36 was obtained referring to step 2 of Example 25. .sup.1H NMR (400 MHz, CD.sub.3OD) δ=8.67 (d, J=7.8 Hz, 1H), 8.53 (br s, 1H), 8.49-8.43 (m, 1H), 7.74-7.63 (m, 2H), 7.25 (d, J=7.9 Hz, 1H), 5.99 (s, 2H), 4.02-3.89 (m, 1H), 3.68-3.57 (m, 1H), 3.27-3.19 (m, 1H), 2.48-2.40 (m, 1H), 2.38 (t, J=7.3 Hz, 2H), 1.99-1.88 (m, 1H), 1.72-1.60 (m, 2H), 1.17-1.02 (m, 2H), 0.99-0.91 (m, 4H), 0.80-0.72 (m, 1H). MS-ESI calculated value [M+H].sup.+ 420, actually measured value 420.
[0311] The formate of compound 36 can be adjusted the pH of the reaction solution to 8-9 by adding saturated aqueous sodium carbonate solution to the system, and then, after the extraction with ethyl acetate, the reaction solution was concentrated to obtain compound 36.
Example 31: Compound 37
[0312] ##STR00108##
[0313] Synthesis Route:
##STR00109##
Step 1
[0314] Compound 37a was obtained referring to step 2 of Example 1. MS-ESI calculated value [M+H].sup.+ 580, actually measured value 580.
Step 2
[0315] A formate of compound 37 was obtained referring to step 2 of Example 25. .sup.1H NMR (400 MHz, CD.sub.3OD) δ=8.66 (d, J=8.0 Hz, 1H), 8.46 (br s, 1H), 8.43-8.40 (m, 1H), 7.74-7.62 (m, 2H), 7.16 (d, J=7.8 Hz, 1H), 5.97 (s, 2H), 4.14-4.04 (m, 1H), 3.64-3.55 (m, 1H), 3.48-3.41 (m, 1H), 2.86-2.76 (m, 1H), 2.61-2.47 (m, 1H), 2.16-2.05 (m, 1H), 1.94-1.84 (m, 2H), 1.83-1.73 (m, 2H), 1.72-1.53 (m, 4H), 1.25-1.11 (m, 2H), 0.85-0.73 (m, 2H). MS-ESI calculated value [M+H].sup.+ 446, actually measured value 446.
[0316] The formate of compound 37 can be adjusted the pH of the reaction solution to 8-9 by adding a saturated aqueous sodium carbonate solution to the system, and then, after the extraction with ethyl acetate, the reaction solution was concentrated to obtain compound 37.
Example 32: Compound 38
[0317] ##STR00110##
##STR00111##
Step 1
[0318] Compound 38a was obtained referring to step 2 of Example 1. MS-ESI calculated value [M+H].sup.+ 594, actually measured value 594.
Step 2
[0319] A formate of compound 38 was obtained referring to step 3 of Example 25. .sup.1H NMR (400 MHz, CD.sub.3OD) δ=8.64 (d, J=8.0 Hz, 1H), 8.47-8.40 (m, 1H), 7.74-7.59 (m, 2H), 7.29 (d, J=7.9 Hz, 1H), 5.97 (s, 2H), 3.88-3.77 (m, 1H), 3.67-3.56 (m, 1H), 3.08-3.02 (m, 1H), 2.44-2.26 (m, 2H), 1.88 (d, J=13.4 Hz, 2H), 1.82-1.68 (m, 3H), 1.63 (br d, J=9.5 Hz, 1H), 1.49-1.38 (m, 2H), 1.34-1.27 (m, 3H), 1.09-0.94 (m, 3H), 0.75-0.67 (m, 1H). MS-ESI calculated value [M+H].sup.+ 460, actually measured value 460.
[0320] The formate of compound 38 can be adjusted the pH of the reaction solution to 8-9 by adding a saturated aqueous sodium carbonate solution to the system, and then, after the extraction with ethyl acetate, the reaction solution was concentrated to obtain compound 38.
Example 33: Compound 39
[0321] ##STR00112##
[0322] Synthesis Route:
##STR00113##
Step 1
[0323] Compound 39a was obtained referring to step 2 of Example 1. MS-ESI calculated value [M+H].sup.+ 582, actually measured value 582.
Step 2
[0324] Compound 39 was obtained referring to step 2 of Example 25. .sup.1H NMR (400 MHz, CD.sub.3OD) δ=8.66-8.59 (m, 1H), 8.47-8.39 (m, 1H), 7.72-7.60 (m, 2H), 7.37-7.28 (m, 1H), 5.97 (s, 2H), 3.83-3.57 (m, 2H), 3.01-2.93 (m, 1H), 2.37 (t, J=7.4 Hz, 2H), 2.32-2.21 (m, 1H), 1.80-1.67 (m, 1H), 1.65-1.56 (m, 2H), 1.27 (dd, J=3.6, 7.3 Hz, 4H), 1.16-1.06 (m, 1H), 1.03-0.95 (m, 1H), 0.95-0.89 (m, 1H), 0.84 (t, J=7.0 Hz, 3H), 0.73-0.63 (m, 1H). MS-ESI calculated value [M+H].sup.+ 448, actually measured value 448.
[0325] The formate of compound 39 can be adjusted the pH of the reaction solution to 8-9 by adding a saturated aqueous sodium carbonate solution to the system, and then, after the extraction with ethyl acetate, the reaction solution was concentrated to obtain compound 39.
Example 34: Compound 40
[0326] ##STR00114##
[0327] Synthesis Route:
##STR00115##
Step 1
[0328] Compound 40a was obtained referring to step 2 of Example 1. MS-ESI calculated value [M+H].sup.+ 602, actually measured value 602.
Step 2
[0329] A formate of compound 40 was obtained referring to step 2 of Example 25. .sup.1H NMR (400 MHz, CD.sub.3OD) δ=8.64 (d, J=7.8 Hz, 1H), 8.48 (br s, 1H), 8.44-8.39 (m, 1H), 7.69 (t, J=7.9 Hz, 1H), 7.60 (d, J=7.9 Hz, 1H), 7.29-7.17 (m, 4H), 7.13 (d, J=7.9 Hz, 1H), 5.99 (s, 2H), 4.12-4.00 (m, 1H), 3.70 (s, 2H), 3.64-3.52 (m, 1H), 3.44-3.36 (m, 1H), 2.57-2.45 (m, 1H), 2.16-1.99 (m, 1H), 1.21-1.09 (m, 2H), 0.83-0.73 (m, 2H). MS-ESI calculated value [M+H].sup.+ 468, actually measured value 468.
[0330] The formate of compound 40 can be adjusted the pH of the reaction solution to 8-9 by adding a saturated aqueous sodium carbonate solution to the system, and then, after the extraction with ethyl acetate, the reaction solution was concentrated to obtain compound 40.
Example 35: Compound 41
[0331] ##STR00116##
[0332] Synthesis Route:
##STR00117##
Step 1
[0333] Compound 41a was obtained referring to step 2 of Example 1. MS-ESI calculated value [M+H].sup.+ 566, actually measured value 566.
Step 2
[0334] A formate of compound 41 was obtained referring to step 2 of Example 25. .sup.1H NMR (400 MHz, CD.sub.3OD) δ=8.66 (d, J=7.9 Hz, 1H), 8.50 (br s, 1H), 8.45-8.39 (m, 1H), 7.73-7.63 (m, 2H), 7.18 (d, J=7.9 Hz, 1H), 5.97 (s, 2H), 4.10-3.99 (m, 1H), 3.67-3.55 (m, 1H), 3.39-3.33 (m, 1H), 3.27-3.18 (m, 1H), 2.57-2.44 (m, 1H), 2.33-2.14 (m, 4H), 2.09-1.95 (m, 2H), 1.94-1.81 (m, 1H), 1.24-1.07 (m, 2H), 0.90-0.71 (m, 2H). MS-ESI calculated value [M+H].sup.+ 432, actually measured value 432.
[0335] The formate of compound 41 can be adjusted the pH of the reaction solution to 8-9 by adding a saturated aqueous sodium carbonate solution to the system, and then, after the extraction with ethyl acetate, the reaction solution was concentrated to obtain compound 41.
Example 36: Compound 42
[0336] ##STR00118##
[0337] Synthesis Route:
##STR00119##
Step 1
[0338] Compound 42a was obtained referring to step 2 of Example 1. MS-ESI calculated value [M+H].sup.+ 594, actually measured value 594.
Step 2
[0339] A formate of compound 42 was obtained referring to step 2 of Example 25. .sup.1H NMR (400 MHz, CD.sub.3OD) δ=8.68 (d, J=7.6 Hz, 1H), 8.46 (s, 1H), 8.44-8.41 (m, 1H), 7.90-7.86 (m, 1H), 7.83-7.79 (m, 1H), 7.76 (d, J=8.1 Hz, 1H), 7.70 (t, J=7.9 Hz, 1H), 7.23-7.14 (m, 2H), 6.18 (s, 2H), 4.11-4.01 (m, 1H), 3.65-3.56 (m, 1H), 3.40-3.36 (m, 1H), 2.59-2.45 (m, 1H), 2.12-1.98 (m, 1H), 1.27-1.06 (m, 2H), 0.88-0.72 (m, 2H). MS-ESI calculated value [M+H].sup.+ 460, actually measured value 460.
[0340] The formate of compound 42 can be adjusted the pH of the reaction solution to 8-9 by adding a saturated aqueous sodium carbonate solution to the system, and then, after the extraction with ethyl acetate, the reaction solution was concentrated to obtain compound 42.
Example 37: Compound 43
[0341] ##STR00120##
[0342] Synthesis Route:
##STR00121##
Step 1
[0343] Compound 43a was obtained referring to step 2 of Example 1. MS-ESI calculated value [M+H].sup.+ 628, actually measured value 628.
Step 2
[0344] A formate of compound 43 was obtained referring to step 2 of Example 25. .sup.1H NMR (400 MHz, CD.sub.3OD) δ=8.69 (d, J=7.9 Hz, 1H), 8.49 (s, 1H), 8.45-8.41 (m, 1H), 8.38 (d, J=1.6 Hz, 1H), 8.05-8.00 (m, 1H), 7.86 (d, J=2.3 Hz, 1H), 7.81 (d, J=7.9 Hz, 1H), 7.70 (t, J=7.9 Hz, 1H), 7.58 (d, J=8.8 Hz, 1H), 7.23 (d, J=7.9 Hz, 1H), 6.95 (d, J=1.5 Hz, 1H), 6.24 (s, 2H), 4.10-3.96 (m, 1H), 3.68-3.54 (m, 1H), 3.36-3.32 (m, 1H), 2.55-2.42 (m, 1H), 2.08-1.94 (m, 1H), 1.26-1.04 (m, 2H), 0.89-0.72 (m, 2H). MS-ESI calculated value [M+H].sup.+ 494, actually measured value 494.
[0345] The formate of compound 43 can be adjusted the pH of the reaction solution to 8-9 by adding a saturated aqueous sodium carbonate solution to the system, and then, after the extraction with ethyl acetate, the reaction solution was concentrated to obtain compound 43.
Example 38: Compound 44
[0346] ##STR00122##
[0347] Synthesis Route:
##STR00123##
Step 1
[0348] Compound 44a was obtained referring to step 2 of Example 1. MS-ESI calculated value [M+H].sup.+ 602, actually measured value 602.
Step 2
[0349] A formate of compound 44 was obtained referring to step 2 of Example 25. .sup.1H NMR (400 MHz, CD.sub.3OD) δ=8.69 (d, J=8.0 Hz, 1H), 8.50 (s, 1H), 8.46-8.40 (m, 1H), 7.91 (d, J=7.9 Hz, 1H), 7.80 (d, J=7.9 Hz, 1H), 7.74-7.67 (m, 1H), 7.49-7.41 (m, 1H), 7.32-7.19 (m, 3H), 6.20 (s, 2H), 4.15-4.05 (m, 1H), 3.67-3.57 (m, 1H), 3.49-3.43 (m, 1H), 2.60-2.50 (m, 4H), 2.17-2.05 (m, 1H), 1.28-1.13 (m, 2H), 0.86-0.75 (m, 2H). MS-ESI calculated value [M+H].sup.+ 468, actually measured value 468.
[0350] The formate of compound 44 can be adjusted the pH of the reaction solution to 8-9 by adding a saturated aqueous sodium carbonate solution to the system, and then, after the extraction with ethyl acetate, the reaction solution was concentrated to obtain compound 44.
Example 39: Compound 45
[0351] ##STR00124##
[0352] Synthesis Route:
##STR00125##
Step 1
[0353] Compound 45a was obtained referring to step 2 of Example 1. MS-ESI calculated value [M+H].sup.+ 602, actually measured value 602.
Step 2
[0354] A formate of compound 45 was obtained referring to step 2 of Example 25. .sup.1H NMR (400 MHz, CD.sub.3OD) δ=8.69 (d, J=8.0 Hz, 1H), 8.46 (s, 1H), 8.44-8.40 (m, 1H), 7.92 (d, J=8.3 Hz, 2H), 7.78 (d, J=8.0 Hz, 1H), 7.70 (t, J=7.9 Hz, 1H), 7.29 (d, J=8.2 Hz, 2H), 7.20 (d, J=8.0 Hz, 1H), 6.20 (s, 2H), 4.12-4.01 (m, 1H), 3.65-3.54 (m, 1H), 3.43-3.36 (m, 1H), 2.58-2.46 (m, 1H), 2.39 (s, 3H), 2.11-1.99 (m, 1H), 1.26-1.08 (m, 2H), 0.86-0.74 (m, 2H). MS-ESI calculated value [M+H].sup.+ 468, actually measured value 468.
[0355] The formate of compound 45 can be adjusted the pH of the reaction solution to 8-9 by adding a saturated aqueous sodium carbonate solution to the system, and then, after the extraction with ethyl acetate, the reaction solution was concentrated to obtain compound 45.
Example 40: Compound 46
[0356] ##STR00126##
[0357] Synthesis Route:
##STR00127##
Step 1
[0358] Compound 46a was obtained referring to step 2 of Example 1. MS-ESI calculated value [M+H].sup.+ 556, actually measured value 556.
Step 2
[0359] A formate of compound 46 was obtained referring to step 2 of Example 25. .sup.1H NMR (400 MHz, CD.sub.3OD) δ=8.66 (d, J=7.9 Hz, 1H), 8.52 (s, 1H), 8.46-8.40 (m, 1H), 7.74-7.65 (m, 2H), 7.18 (d, J=7.9 Hz, 1H), 5.99 (s, 2H), 4.27-4.19 (m, 2H), 4.11-4.02 (m, 1H), 3.67-3.56 (m, 1H), 3.45-3.39 (m, 1H), 2.59-2.47 (m, 1H), 2.14-2.02 (m, 1H), 1.29 (t, J=7.1 Hz, 3H), 1.24-1.11 (m, 2H), 0.88-0.76 (m, 2H). MS-ESI calculated value [M+H].sup.+ 422, actually measured value 422.
[0360] The formate of compound 46 can be adjusted the pH of the reaction solution to 8-9 by adding a saturated aqueous sodium carbonate solution to the system, and then, after the extraction with ethyl acetate, the reaction solution was concentrated to obtain compound 46.
Example 41: Compound 47
[0361] ##STR00128##
##STR00129##
Step 1
[0362] Compound 47a was obtained referring to step 2 of Example 1. MS-ESI calculated value [M+H].sup.+ 570, actually measured value 570.
Step 2
[0363] A formate of compound 47 was obtained referring to step 2 of Example 25. .sup.1H NMR (400 MHz, CD.sub.3OD) δ=8.66 (d, J=7.9 Hz, 1H), 8.49 (s, 1H), 8.44-8.38 (m, 1H), 7.74-7.63 (m, 2H), 7.17 (d, J=7.8 Hz, 1H), 6.01-5.94 (m, 2H), 4.13 (t, J=6.6 Hz, 2H), 4.10-4.02 (m, 1H), 3.65-3.56 (m, 1H), 3.45-3.39 (m, 1H), 2.58-2.47 (m, 1H), 2.13-2.03 (m, 1H), 1.73-1.63 (m, 2H), 1.24-1.10 (m, 2H), 0.94 (t, J=7.5 Hz, 3H), 0.80 (t, J=9.0 Hz, 2H). MS-ESI calculated value [M+H].sup.+ 436, actually measured value 436.
[0364] The formate of compound 47 can be adjusted the pH of the reaction solution to 8-9 by adding a saturated aqueous sodium carbonate solution to the system, and then, after the extraction with ethyl acetate, the reaction solution was concentrated to obtain compound 47.
Example 42: Compound 48
[0365] ##STR00130##
[0366] Synthesis Route:
##STR00131##
Step 1
[0367] Compound 48a was obtained referring to step 2 of Example 1. MS-ESI calculated value [M+H].sup.+ 666, actually measured value 666.
Step 2
[0368] A formate of compound 48 was obtained referring to step 2 of Example 25. .sup.1H NMR (400 MHz, CD.sub.3OD) δ=8.65 (d, J=7.6 Hz, 1H), 8.48 (s, 1H), 8.43-8.39 (m, 1H), 7.74-7.60 (m, 2H), 7.16 (d, J=7.9 Hz, 1H), 5.97 (s, 2H), 4.14-4.02 (m, 1H), 3.65-3.54 (m, 1H), 3.47-3.38 (m, 1H), 2.61-2.47 (m, 1H), 2.37 (t, J=7.3 Hz, 2H), 2.14-2.03 (m, 1H), 1.66-1.54 (m, 2H), 1.30-1.15 (m, 18H), 0.89 (t, J=7.0 Hz, 3H), 0.83-0.73 (m, 2H). MS-ESI calculated value [M+H].sup.+ 532, actually measured value 532.
[0369] The formate of compound 48 can be adjusted the pH of the reaction solution to 8-9 by adding a saturated aqueous sodium carbonate solution to the system, and then, after the extraction with ethyl acetate, the reaction solution was concentrated to obtain compound 48.
Example 43: Compound 49
[0370] ##STR00132##
[0371] Synthesis Route:
##STR00133##
Step 1
[0372] Compound 49a was obtained referring to step 2 of Example 1. MS-ESI calculated value [M+H].sup.+ 596, actually measured value 596.
Step 2
[0373] A formate of compound 49 was obtained referring to step 2 of Example 25. .sup.1H NMR (400 MHz, CD.sub.3OD) δ=8.65 (d, J=7.6 Hz, 1H), 8.49 (s, 1H), 8.43-8.40 (m, 1H), 7.75-7.61 (m, 2H), 7.16 (d, J=7.8 Hz, 1H), 5.99 (s, 2H), 4.12-4.02 (m, 1H), 3.92-3.83 (m, 2H), 3.66-3.54 (m, 1H), 3.47-3.37 (m, 3H), 2.73-2.62 (m, 1H), 2.59-2.45 (m, 1H), 2.15-2.01 (m, 1H), 1.88-1.78 (m, 2H), 1.75-1.61 (m, 2H), 1.24-1.10 (m, 2H), 0.85-0.75 (m, 2H). MS-ESI calculated value [M+H].sup.+ 462, actually measured value 462.
[0374] The formate of compound 49 can be adjusted the pH of the reaction solution to 8-9 by adding a saturated aqueous sodium carbonate solution to the system, and then, after the extraction with ethyl acetate, the reaction solution was concentrated to obtain compound 49.
Example 44: Compound 50
[0375] ##STR00134##
[0376] Synthesis Route:
##STR00135##
Step 1
[0377] Compound 50a was obtained referring to step 2 of Example 1. MS-ESI calculated value [M+H].sup.+ 542, actually measured value 542.
Step 2
[0378] A formate of compound 50 was obtained referring to step 2 of Example 25. .sup.1H NMR (400 MHz, CD.sub.3OD) δ=8.64 (d, J=7.9 Hz, 1H), 8.44 (s, 1H), 8.42-8.38 (m, 1H), 7.72-7.64 (m, 2H), 7.14 (d, J=7.8 Hz, 1H), 5.98 (s, 2H), 4.14-4.05 (m, 1H), 3.80 (s, 3H), 3.64-3.56 (m, 1H), 3.48 (d, J=5.1 Hz, 1H), 2.61-2.50 (m, 1H), 2.18-2.07 (m, 1H), 1.26-1.14 (m, 2H), 0.85-0.73 (m, 2H). MS-ESI calculated value [M+H]+408, actually measured value 408.
[0379] The formate of compound 50 can be adjusted the pH of the reaction solution to 8-9 by adding a saturated aqueous sodium carbonate solution to the system, and then, after the extraction with ethyl acetate, the reaction solution was concentrated to obtain compound 50.
Example 45: Compound 51
[0380] ##STR00136##
[0381] Synthesis Route:
##STR00137##
Step 1
[0382] Compound 51a was obtained referring to step 2 of Example 1. MS-ESI calculated value [M+H].sup.+ 616, actually measured value 616.
Step 2
[0383] A formate of compound 51 was obtained referring to step 2 of Example 25. .sup.1H NMR (400 MHz, CD.sub.3OD) δ=8.64 (d, J=7.9 Hz, 1H), 8.49 (br s, 1H), 8.42-8.35 (m, 1H), 7.77 (t, J=8.1 Hz, 2H), 7.70-7.62 (m, 1H), 7.18 (d, J=7.8 Hz, 1H), 7.09-6.98 (m, 2H), 6.15 (s, 2H), 4.10-3.98 (m, 1H), 3.63-3.53 (m, 1H), 3.45-3.37 (m, 1H), 2.57-2.44 (m, 4H), 2.30 (s, 3H), 2.14-2.01 (m, 1H), 1.26-1.08 (m, 2H), 0.79 (t, J=9.0 Hz, 2H). MS-ESI calculated value [M+H].sup.+ 482, actually measured value 482.
[0384] The formate of compound 51 can be adjusted the pH of the reaction solution to 8-9 by adding a saturated aqueous sodium carbonate solution to the system, and then, after the extraction with ethyl acetate, the reaction solution was concentrated to obtain compound 51.
Example 46: Compound 52
[0385] ##STR00138##
[0386] Synthesis Route:
##STR00139##
Step 1
[0387] Compound 52a was obtained referring to step 2 of Example 1. MS-ESI calculated value [M+Na].sup.+ 634, actually measured value 634.
Step 2
[0388] A formate of compound 52 was obtained referring to step 2 of Example 25. .sup.1H NMR (400 MHz, CD.sub.3OD) δ=8.66 (d, J=7.8 Hz, 1H), 8.61 (s, 1H), 8.42 (d, J=7.8 Hz, 1H), 7.74-7.64 (m, 2H), 7.14 (d, J=7.8 Hz, 1H), 6.03-5.92 (m, 2H), 4.22-4.05 (m, 3H), 3.59-3.61 (m, 1H), 3.50 (d, J=4.8 Hz, 1H), 2.54-2.57 (m, 1H), 2.20-2.08 (m, 1H), 1.71-1.59 (m, 2H), 1.41-1.12 (m, 8H), 0.87 (t, J=6.8 Hz, 3H), 0.84-0.67 (m, 2H). MS-ESI calculated value [M+H].sup.+ 478, actually measured value 478.
[0389] The formate of compound 52 can be adjusted the pH of the reaction solution to 8-9 by adding a saturated aqueous sodium carbonate solution to the system, and then, after the extraction with ethyl acetate, the reaction solution was concentrated to obtain compound 52.
Example 47: Compound 53
[0390] ##STR00140##
[0391] Synthesis Route:
##STR00141##
Step 1
[0392] Compound 53b was obtained referring to step 1 of Example 17.
Step 2
[0393] Compound 53c was obtained referring to step 2 of Example 1. MS-ESI calculated value [M+Na].sup.+ 606, actually measured value 606.
Step 3
[0394] A formate of compound 53 was obtained referring to step 2 of Example 25. .sup.1H NMR (400 MHz, CD.sub.3OD) δ=8.66 (d, J=8.0 Hz, 1H), 8.48 (s, 1H), 8.42 (dd, J=1.1, 7.8 Hz, 1H), 7.75-7.62 (m, 2H), 7.17 (d, J=7.8 Hz, 1H), 6.04-5.91 (m, 2H), 4.17 (t, J=6.6 Hz, 2H), 4.05-4.07 (m, 1H), 3.65-3.53 (m, 1H), 3.41 (dd, J=3.2, 7.6 Hz, 1H), 2.51-2.53 (m, 1H), 2.13-1.99 (m, 1H), 1.69-1.56 (m, 2H), 1.35-1.40 (m, 2H), 1.25-1.08 (m, 2H), 0.92 (t, J=7.4 Hz, 3H), 0.85-0.75 (m, 2H). MS-ESI calculated value [M+H].sup.+ 450, actually measured value 450.
[0395] The formate of compound 53 can be adjusted the pH of the reaction solution to 8-9 by adding a saturated aqueous sodium carbonate solution to the system, and then, after the extraction with ethyl acetate, the reaction solution was concentrated to obtain compound 53.
Example 48: Compound 54
[0396] ##STR00142##
##STR00143##
Step 1
[0397] Compound 54a was obtained referring to step 2 of Example 1. MS-ESI calculated value [M+Na].sup.+ 610, actually measured value 610.
Step 2
[0398] A formate of compound 54 was obtained referring to step 2 of Example 25. .sup.1H NMR (400 MHz, CD.sub.3OD) δ=8.66-8.64 (d, J=7.8 Hz, 1H), 8.50 (s, 1H), 8.43-8.41 (d, J=7.8 Hz, 1H), 8.34 (m, 1H), 7.76-7.63 (m, 2H), 7.14 (d, J=7.8 Hz, 1H), 5.96-5.93 (m, 2H), 4.66-4.61 (m, 1H), 3.94-3.59 (m, J=6.4, 9.0, 11.8 Hz, 1H), 3.58-3.56 (m, J=4.8 Hz, 1H), 3.56-3.24 (m, J=4.8 Hz, 1H), 3.24-3.22 (m, J=4.8 Hz, 1H), 2.41-1.89 (dt, J=8.4, 14.0 Hz, 2H), 1.87-1.72 (m, 2H), 1.71-1.36 (m, 7H), 1.36-1.39 (m, 2H), 1.14-1.12 (m, J=6.8 Hz, 1H), 0.90-0.75 (m, 1H). MS-ESI calculated value [M+H].sup.+ 476, actually measured value 476.
[0399] The formate of compound 54 can be adjusted the pH of the reaction solution to 8-9 by adding a saturated aqueous sodium carbonate solution to the system, and then, after the extraction with ethyl acetate, the reaction solution was concentrated to obtain compound 54.
Example 49: Compound 55
[0400] ##STR00144##
[0401] Synthesis Route:
##STR00145##
Step 1
[0402] Compound 3 (150 mg, 231 μmol) and potassium carbonate (63.8 mg, 462 μmol) were dissolved in anhydrous tetrahydrofuran (5 mL), stirred for 1 hour at 60° C. Then compound 15b (70.4 mg, 462 μmol) was added to the reaction system at 60° C., stirred for 12 hours at 60° C. The solvent was removed by concentration under reduced pressure, and the crude product was purified by silica gel plate chromatography (ethyl acetate: ethanol) and concentrated under reduced pressure to obtain compound 55a. MS-ESI calculated value [M+H].sup.+ 538, actually measured value 538.
Step 2
[0403] Compound 55 was obtained referring to step 3 of Example 1 (purified by neutral preparative high performance liquid chromatography). .sup.1H NMR (400 MHz, CD.sub.3OD) δ=8.68 (dd, J=1.4, 8.1 Hz, 1H), 8.07 (dd, J=1.5, 7.8 Hz, 1H), 7.63 (t, J=7.9 Hz, 1H), 7.42 (d, J=0.9 Hz, 1H), 5.93 (s, 2H), 4.93-4.89 (m, 1H), 3.84 (d, J=12.9 Hz, 2H), 3.18-3.09 (m, 2H), 2.99-2.87 (m, 1H), 2.68 (d, J=0.8 Hz, 3H), 2.01-1.90 (m, 2H), 1.55-1.42 (m, 2H), 1.28 (d, J=6.3 Hz, 6H). MS-ESI calculated value [M+H].sup.+ 438, actually measured value 438.
Example 50: Compound 56
[0404] ##STR00146##
[0405] Synthesis Route:
##STR00147##
Step 1
[0406] To a solution of compound 2 (435 mg, 1.07 mmol) in N,N-dimethylformamide (10 mL) was added potassium tert-butoxide (359 mg, 3.20 mmol) at 0° C., stirred for 1 hour at 0° C. Compound 15a (226 mg, 1.17 mmol) was added to the reaction solution at 0° C., stirred for 13 hours at 25° C. Water (4 mL) was added to the reaction solution which was extracted with ethyl acetate (3 mL×3), the combined organic phase was washed with saturated brine (5 mL×3), dried with anhydrous sodium sulfate, filtered, concentrated under reduced pressure, and the crude product was separated and purified by silica gel column chromatography (petroleum ether/ethyl acetate) to obtain compound 56a.
Step 2
[0407] To a solution of compound 56a (150 mg, 0.266 mmol) in anhydrous ethyl acetate (2 mL) added ethyl acetate hydrochloride (4M, 16.0 mmol), stirred for 3 hours at 20° C. The solvent was removed by concentration under reduced pressure, and the crude product was prepared by high performance liquid chromatography (hydrochloride system) to obtain a hydrochloride of compound 56. .sup.1H NMR (400 MHz, CD.sub.3OD) δ8.68 (d, J=8.0 Hz, 1H), 8.46 (s, 1H), 8.39 (dd, J=7.8, 1.2 Hz, 1H), 7.73 (t, J=7.8 Hz, 1H), 7.67 (d, J=7.8 Hz, 1H), 7.51 (d, J=7.8 Hz, 1H), 6.00 (s, 2H), 4.62-4.71 (m, 1H), 3.77-3.87 (m, 1H), 3.50-3.64 (m, 2H), 3.34-3.40 (m, 2H), 2.24-2.36 (m, 1H), 1.86-2.03 (m, 3H), 1.75 (dt, J=6.4, 3.1 Hz, 2H), 1.24-1.61 (m, 6H). MS-ESI calculated value [M+H].sup.+ 450, actually measured value 450.
[0408] The hydrochloride of compound 56 can be adjusted the pH of the reaction solution to 8-9 by adding a saturated aqueous sodium carbonate solution to the system, and then, after the extraction with ethyl acetate, the reaction solution was concentrated to obtain compound 56.
Example 51: Compound 57
[0409] ##STR00148##
[0410] Synthesis Route:
##STR00149##
Step 1
[0411] Compound 57a was obtained referring to step 1 of Example 50. MS-ESI calculated value [M+H].sup.+ 510, actually measured value 510.
Step 2
[0412] A formate of compound 57 was obtained from crude product by high performance liquid chromatography (formic acid system) referring to step 3 of Example 1. .sup.1H NMR (400 MHz, CDCl.sub.3) δ 8.69-8.63 (m, 1H), 8.25-8.20 (m, 1H), 7.64-7.61 (m, 1H), 7.59-7.40 (m, 1H), 5.96-5.74 (m, 2H), 4.24-4.20 (m, 2H), 3.70-3.66 (m, 3H), 3.54 (s, 1H), 3.20-3.18 (m, 1H), 2.68-2.66 (m, 3H), 2.28-2.27 (m, 1H), 1.92-1.91 (m, 1H), 1.31-1.26 (m, 3H). MS-ESI calculated value [M+H].sup.+ 410, actually measured value 410.
[0413] The formate of compound 57 can be adjusted the pH of the reaction solution to 8-9 by adding a saturated aqueous sodium carbonate solution to the system, and then, after the extraction with ethyl acetate, the reaction solution was concentrated to obtain compound 57.
Example 52: Compound 58
[0414] ##STR00150##
[0415] Synthesis Route:
##STR00151##
Step 1
[0416] Compound 58a was obtained referring to step 2 of Example 1. MS-ESI calculated value [M+H].sup.+ 556, actually measured value 556.
Step 2
[0417] Compound 58 was obtained from crude product by high performance liquid chromatography (neutral system) referring to step 3 of Example 1. .sup.1H NMR (400 MHz, CD.sub.3OD) δ=8.72 (d, J=8.1 Hz, 1H), 8.10 (d, J=7.7 Hz, 1H), 8.04 (d, J=8.1 Hz, 2H), 7.68-7.59 (m, 2H), 7.56 (s, 1H), 7.51-7.45 (m, 2H), 6.20 (s, 2H), 3.85 (d, J=13.0 Hz, 2H), 3.14 (t, J=11.2 Hz, 2H), 2.99-2.89 (m, 1H), 2.72 (s, 3H), 1.95 (d, J=10.9 Hz, 2H), 1.55-1.42 (m, 2H). MS-ESI calculated value [M+H].sup.+ 456, actually measured value 456.
Example 53: Compound 59
[0418] ##STR00152##
[0419] Synthesis Route:
##STR00153##
Step 1
[0420] Compound 59a was obtained referring to step 1 of Example 49. MS-ESI calculated value [M+H].sup.+ 538, actually measured value 538.
Step 2
[0421] A formate of compound 59 was obtained referring to step 3 of Example 1. .sup.1H NMR (400 MHz, CD.sub.3OD) δ=8.73 (d, J=7.7 Hz, 1H), 8.55 (s, 1H), 8.10 (d, J=7.6 Hz, 1H), 7.66 (t, J=7.8 Hz, 1H), 7.47 (s, 1H), 5.96 (s, 2H), 4.59 (m, 1H), 3.99 (d, J=12.8 Hz, 2H), 3.46-3.37 (m, 1H), 3.23 (t, J=12.3 Hz, 2H), 2.70 (s, 3H), 2.14 (d, J=11.2 Hz, 2H), 1.85-1.61 (m, 2H), 1.30 (d, J=6.1 Hz, 6H). MS-ESI calculated value [M+H].sup.+ 438, actually measured value 438.
[0422] The formate of compound 59 can be adjusted the pH of the reaction solution to 8-9 by adding a saturated aqueous sodium carbonate solution to the system, and then, after the extraction with ethyl acetate, the reaction solution was concentrated to obtain compound 59.
Example 54: Compound 60
[0423] ##STR00154##
[0424] Synthesis Route:
##STR00155##
Step 1
[0425] Compound 60a was obtained referring to step 1 of Example 49. MS-ESI calculated value [M+H].sup.+ 542, actually measured value 542.
Step 2
[0426] Compound 60 was obtained from crude product by high performance liquid chromatography (neutral system) referring to step 3 of Example 1. MS-ESI calculated value [M+H].sup.+ 442, actually measured value 442. .sup.1H NMR (400 MHz, CD.sub.3OD) δ=8.72 (dd, J=1.41, 8.01 Hz, 1H), 8.25 (dd, J=1.47, 7.70 Hz, 1H), 7.98-8.11 (m, 2H), 7.58-7.71 (m, 2H), 7.56 (d, J=0.86 Hz, 1H), 7.43-7.52 (m, 2H), 6.21 (s, 2H), 3.63-3.76 (m, 3H), 3.51-3.60 (m, 1H), 3.21 (dd, J=4.65, 9.17 Hz, 1H), 2.72 (d, J=0.86 Hz, 3H), 2.24-2.35 (m, 1H), 1.92 (dd, J=6.97, 13.33 Hz, 1H).
Example 55: Compound 61
[0427] ##STR00156##
[0428] Synthesis Route:
##STR00157##
Step 1
[0429] Compound 61a was obtained referring to step 1 of Example 49. MS-ESI calculated value [M+Na].sup.+ 502, actually measured value 502.
Step 2
[0430] A formate of compound 61 was obtained referring to step 3 of Example 1. MS-ESI calculated value [M+Na].sup.+ 402, actually measured value 402. .sup.1H NMR (400 MHz, CDCl.sub.3) δ=8.84-8.56 (m, 1H), 8.30 (d, J=6.5 Hz, 1H), 8.13 (s, 1H), 7.56 (s, 1H), 7.45 (d, J=7.5 Hz, 1H), 7.28-7.23 (m, 1H), 5.94 (s, 2H), 4.10-3.96 (m, 2H), 3.85 (s, 2H), 3.79 (s, 2H), 3.72-3.37 (m, 1H), 2.74-2.46 (m, 1H), 1.25-1.04 (m, 6H).
[0431] The formate of compound 61 can be adjusted the pH of the reaction solution to 8-9 by adding a saturated aqueous sodium carbonate solution to the system, and then, after the extraction with ethyl acetate, the reaction solution was concentrated to obtain compound 61.
Example 56: Compound 62
[0432] ##STR00158##
[0433] Synthesis Route:
##STR00159##
Step 1
[0434] To a solution of dimethylamine hydrochloride (332 mg, 4.07 mmol) and potassium carbonate (1.23 g, 8.92 mmol) in anhydrous dichloromethane (5 mL) was added dropwise chloromethylchloroformate 23b (500 mg, 3.88 mmol) at −78° C., stirred for 3 hours at −78° C. The reaction solution was filtered when being cold, and concentrated to obtain the crude product compound 62a which was directly used for the next step of the reaction without purification. .sup.1H NMR (400 MHz, CDCl.sub.3) δ=5.71 (s, 2H), 2.89 (d, J=8.8 Hz, 6H).
Step 2
[0435] Compound 62b was obtained referring to step 1 of Example 49. MS-ESI calculated value [M+H].sup.+ 481, actually measured value 481.
Step 3
[0436] A formate of compound 62 was obtained referring to step 3 of Example 1. MS-ESI calculated value [M+H].sup.+ 381, actually measured value 381. .sup.1H NMR (400 MHz, CD.sub.3OD) δ=8.66 (d, J=7.6 Hz, 1H), 8.37 (s, 1H), 8.36 (dd, J=1.3, 7.7 Hz, 1H), 7.74-7.61 (m, 2H), 7.32 (d, J=7.9 Hz, 1H), 5.96 (s, 2H), 4.13-4.06 (m, 2H), 3.99-3.87 (m, 1H), 3.87-3.77 (m, 2H), 2.91 (d, J=2.7 Hz, 6H).
[0437] The formate of compound 62 can be adjusted the pH of the reaction solution to 8-9 by adding a saturated aqueous sodium carbonate solution to the system, and then, after the extraction with ethyl acetate, the reaction solution was concentrated to obtain compound 62.
Example 57: Compound 63
[0438] ##STR00160##
##STR00161##
Step 1
[0439] Compound 63a was obtained referring to step 1 of Example 49. MS-ESI calculated value [M+H].sup.+ 570, actually measured value 570.
Step 2
[0440] Compound 63 was obtained from crude product by high performance liquid chromatography (neutral system) referring to step 3 of Example 50. MS-ESI calculated value [M+H].sup.+ 470, actually measured value 470. .sup.1H NMR (400 MHz, CD.sub.3OD) δ=8.71 (dd, J=1.4, 8.0 Hz, 1H), 8.24 (dd, J=1.5, 7.8 Hz, 1H), 7.81 (d, J=7.9 Hz, 1H), 7.64 (t, J=7.9 Hz, 1H), 7.54 (d, J=0.9 Hz, 1H), 7.09 (s, 1H), 7.05 (d, J=7.9 Hz, 1H), 6.14 (s, 2H), 3.77-3.62 (m, 3H), 3.59-3.50 (m, 1H), 3.20 (dd, J=4.9, 9.5 Hz, 1H), 2.70 (d, J=0.9 Hz, 3H), 2.53 (s, 3H), 2.32 (s, 3H), 2.30-2.23 (m, 1H), 1.96-1.86 (m, 1H).
Example 58: Compound 64
[0441] ##STR00162##
##STR00163##
Step 1
[0442] Compound 64a was obtained referring to step 1 of Example 49. MS-ESI calculated value [M+H].sup.+ 542, actually measured value 542.
Step 2
[0443] Compound 64 was obtained from crude product by high performance liquid chromatography (neutral system) referring to step 3 of Example 1. MS-ESI calculated value [M+H].sup.+ 442, actually measured value 442. .sup.1H NMR (400 MHz, CD.sub.3OD) δ=8.69-8.63 (m, 1H), 8.23-8.16 (m, 1H), 8.04-7.96 (m, 2H), 7.64-7.54 (m, 2H), 7.51 (s, 1H), 7.46-7.39 (m, 2H), 6.17 (s, 2H), 3.75-3.59 (m, 3H), 3.57-3.48 (m, 1H), 3.23-3.13 (m, 1H), 2.67 (s, 3H), 2.33-2.20 (m, 1H), 1.96-1.81 (m, 1H).
Example 59: Control compound 1
[0444] ##STR00164##
Step 1
[0445] To a solution of intermediate 2 (360 mg, 0.82 mmol) in ethyl acetate (5 mL) was added dropwise an ethyl acetate hydrochloride solution (1.86 mL, 4M). After the reaction was completed, the precipitated solid was filtered, then washed with ethyl acetate (5 mL×3), and dried to obtain a hydrochloride of control compound 1. MS-ESI calculated value [M+H].sup.+ 308, actually measured value 308.
[0446] The hydrochloride of control compound 1 can be adjusted the pH of the reaction solution to 8-9 by adding a saturated aqueous sodium carbonate solution to the system, and then, after the extraction with ethyl acetate, the reaction solution was concentrated to obtain control compound 1.
Example 60: Control compound 2
[0447] ##STR00165##
Step 1
[0448] To a solution of intermediate 6 (4 g, 9.57 mmol) in tetrahydrofuran (40 mL) was added concentrated sulfuric acid (1.88 g, 19.15 mmol) dropwise at 0° C. After the reaction was completed, the precipitated solid was filtered, and then control compound 2 was prepared by high performance liquid chromatography (neutral system). MS-ESI calculated value [M+H].sup.+ 308, actually measured value 308. .sup.1H NMR (400 MHz, CD.sub.3OD) δ 8.68 (d, 1H), 8.24-8.26 (m, 1H), 7.61-7.65 (m, 1H), 7.13 (s, 1H), 3.67-3.73 (m, 3H), 3.57-3.66 (m, 1H), 3.19-3.22 (m, 1H), 2.69 (s, 3H), 2.27-2.30 (m, 1H), 1.91-1.95 (m, 1H).
BIOASSAY EXPERIMENTS
Experimental Example 1. Pharmacokinetic Test in Aqueous Humor
[0449] Experiment Objective:
[0450] The compounds are prodrug molecules containing ester functional groups, which can be hydrolyzed into active drug molecules (technical concentrate) by the action of abundant ester hydrolases in the ocular tissue during eye-drop administration. In this experiment, the production rate of the active drug ingredients and the exposure amount of the active drug ingredients were detected.
[0451] Experiment Materials:
[0452] Male New Zealand rabbits, aged 3-6 months, weighted 2.0-5.0 kg, purchased from PizhouDongfang Breeding Co. Ltd.
[0453] Preparation of Eye-Drop Sample:
[0454] Solvents used were 1.2% hydroxypropyl methyl cellulose E5/20.5% Poloxamer P407/1.6% Poloxamer P188.
[0455] Experiment Operations:
[0456] The dose of eye-drop administration was 0.5 mg/eye and eye-drop administration was performed on both eyes. Aqueous humor was collected 0.25 h. 0.5 h, 2 h, 4 h, 8 h, and 24 h after administration to prepare the aqueous humor sample. All samples were quantitatively detected for the content of the administered compound in aqueous humor of the experiment animals by liquid chromatography-coupled mass spectrometry (LC-MS) and mass spectrometry (MS) technology. The detected concentration values were subject to WinNonlin non-compartment model. According to the aqueous humor concentration-time data, parameters of the half-life period, peak concentration of the drug in the aqueous humor, peak time of the drug in the aqueous humor, unit exposure and the like were calculated.
TABLE-US-00001 TABLE 1 Pharmacokinetic test results in aqueous humor of New Zealand rabbits Peak Peak time Test sample concentration of drug in (compounds of drug in Half-life aqueous Unit obtained in aqueous humor period humor exposure various examples) (nM) (h) (h) (nM.h) Formate of 1222 3.39 2.00 8322 compound 30 Formate of 3505 2.39 0.5 15911 compound 31 Formate of 3810 1.91 2.0 14826 compound 42 Hydrochloride of 3345 2.47 2.00 14124 compound 56 Compound 60 2600 2.83 0.5 14332 Compound 63 7470 2.72 2.0 36581 Hydrochloride of 425 3.9 0.5 2508 control compound 1 Control compound 2 868 — 2.0 3555 “—”: not detected.
[0457] Conclusion: Experiment results showed that the test sample compounds (prodrug molecules) were not detected in the aqueous humor, and what were mainly detected were their active metabolites (technical concentrate molecules) after the ester hydrolysis. Compared with control compound 1 and control compound 2, the compounds of the present disclosure significantly increase the exposure amount of the active drugs and meanwhile significantly increase the peak blood concentration and the action duration.
Experimental Example 2. Intraocular Pressure Reduction Experiment in New Zealand Rabbits with Normal Intraocular Pressure
[0458] Experiment Objective:
[0459] Rabbits with normal intraocular pressure were adopted to screen intraocular pressure reduction actions of the potential compounds by eye-drop administration.
[0460] Experiment Materials:
[0461] Male New Zealand rabbits, aged 97-127 days, weighted 2.65-3.5 kg, purchased from PizhouDongfang Breeding Co. Ltd.
[0462] Experiment Operations:
[0463] The male New Zealand rabbits were randomly divided into 8 rabbits/per group using a computer-generated randomization method Animals in each group were eye-dropped with different test samples in the right eye, and normal saline or solvent in the left eye, and the volume of the administration was both 50 μL/eye. The intraocular pressure of the animals was measured before administration and in 1, 2, 4, 6, 8 and 10 hours after the administration, respectively. Experimental results are shown in Table 2:
TABLE-US-00002 TABLE 2 Change results of the intraocular pressure of both eyes (Mean +/− SEM) of the New Zealand rabbits before and after administration of different test samples Time point of measurement 1 hour 2 hours 4 hours 6 hours 8 hours 10 hours Measured Before after after after after after after intraocular admini- admini- admini- admini- admini- admini- admini- pressure stration stration stration stration stration stration stration K-115 Intraocular 12.29 11.42 11.67 12.08 12.38 12.83 15.25 20 pressure in mg/mL the left eye (mmHg) Intraocular 12.92 7.54 8.63 10.83 11.83 12.63 14.92 pressure in the right eye (mmHg) Reduction −0.63 3.88 3.04 1.25 0.55 0.2 0.33 magnitude of intraocular pressure (mmHg) Com- Intraocular 12.63 12.42 12.96 13.17 12.54 13.17 13.71 pound pressure in 60 the left eye 20 (mmHg) mg/mL Intraocular 12.42 10.21 7.17 7.92 7.63 8.83 10.83 pressure in the right eye (mmHg) Reduction 0.21 2.21 5.79 5.25 4.91 4.34 2.88 magnitude of intraocular pressure (mmHg) Com- Intraocular 11.29 11.96 9.79 11.50 11.54 11.75 11.88 pound pressure in 64 the left eye 20 (mmHg) mg/mL Intraocular 12.04 11.54 7.42 7.13 7.38 7.63 9.71 pressure in the right eye (mmHg) Reduction −0.75 0.42 2.37 4.37 4.16 4.12 2.17 magnitude of intraocular pressure (mmHg) Com- Intraocular 12.13 11.63 11.58 12.83 12.38 12.29 14.33 pound pressure in 60 the left eye 2.5 (mmHg) mg/mL Intraocular 12.38 8.38 7.58 9.04 9.88 11.33 13.71 pressure in the right eye (mmHg) Reduction −0.25 3.25 4 3.79 2.5 0.96 0.62 magnitude of intraocular pressure (mmHg) Com- Intraocular 11.79 11.33 11.42 12.63 12.75 13.29 14.08 pound pressure in 60 the left eye 5.0 (mmHg) mg/mL Intraocular 11.75 7.83 7.00 7.79 8.54 10.13 12.54 pressure in the right eye (mmHg) Reduction 0.04 3.5 4.42 4.84 4.21 3.16 1.54 magnitude of intraocular pressure (mmHg) Com- Intraocular 11.96 12.08 12.08 12.96 12.38 13.04 13.38 pound pressure in 60 the left eye 10.0 (mmHg) mg/mL Intraocular 11.33 7.96 6.42 7.13 8.00 9.42 10.96 pressure in the right eye (mmHg) Reduction 0.63 4.12 5.66 5.83 4.38 3.62 2.42 magnitude of intraocular pressure (mmHg)
[0464] Conclusion: Compared with K-115, the compound of the present disclosure exhibited a larger reduction magnitude of intraocular pressure and a longer intraocular pressure reduction action duration. Moreover, compound 60 witnessed a remarkably significant reduction of intraocular pressure at the concentrations of 2.5, 5 and 10 mg/mL.
Experimental Example 3. Intraocular Pressure Reduction Experiment in New Zealand Rabbits with Acute Ocular Hypertension
[0465] Experiment Objective:
[0466] Acute ocular hypertension of rabbits were induced by anterior chamber injection of a viscoelastic agent and the intraocular pressure reduction action of compound 60 and compound 63 at different concentrations were explored by eye-drop administration.
[0467] Experiment Materials:
[0468] Male New Zealand rabbits, aged 97-127 days, weighted 2.5-3.4 kg, purchased from PizhouDongfang Breeding Co. Ltd.
[0469] Experiment Operation 1:
[0470] Male New Zealand rabbits were randomly divided into 8 rabbits/per group according to body weight using a computer-generated randomization method. The animals in each group were injected with medical sodium hyaluronate gel in the anterior chamber of right eye in a single dose, 100 μL/eye, to induce ocular hypertension. In 5-15 minutes, 3 hours and 6 hours after right eye modeling, both eyes were dropped with a solvent, K-115 or test samples (compound 60 at different concentrations) at a volume of 50 μL/eye and intraocular pressure of the animals was measured before administration and in 1, 2, 4, 6, 8 and 10 hours after administration, respectively. Experimental results are shown in Table 3:
TABLE-US-00003 TABLE 3 Change results of the intraocular pressure of both eyes (Mean +/− SEM) of animals in each group before and after modeling and administration Time point of measurement 1 hour 2 hours 4 hours 6 hours 8 hours 10 hours Measured Before after after after after after after Intraocular admini- admini- admini- admini- admini- admini- admini- pressure stration stration stration stration stration stration stration Negative Intra- 12.46 5.21 6.25 9.04 11.63 12.63 12.46 control ocular pressure in the left eye (mmHg) Intra- 12.00 31.14 25.05 26.33 24.38 23.14 12.00 ocular pressure in the right eye (mmHg) K-115 Intra- 11.79 5.88 5.96 8.29 10.54 11.88 11.79 10 mg/mL ocular pressure in the left eye (mmHg) Intra- 11.92 28.79 22.67 26.08 23.25 23.25 11.92 ocular pressure in the right eye (mmHg) Compound Intra- 12.00 5.00 6.63 8.29 10.71 12.17 12.00 60 ocular 2.5 mg/mL pressure in the left eye (mmHg) Intra- 12.54 22.33 19.75 20.29 17.00 14.54 12.54 ocular pressure in the right eye (mmHg) Compound Intra- 11.75 4.96 5.96 8.29 11.21 11.46 11.75 60 ocular 5.0 mg/mL pressure in the left eye (mmHg) Intra- 11.25 21.00 16.29 16.75 14.88 13.08 11.25 ocular pressure in the right eye (mmHg) Compound Intra- 12.13 4.63 5.50 9.25 10.79 11.75 12.13 60 ocular 10.0 mg/mL pressure in the left eye (mmHg) Intra- 12.08 19.83 17.46 15.00 12.33 11.25 12.08 ocular pressure in the right eye (mmHg)
[0471] Experiment Operation 2:
[0472] 50 male New Zealand rabbits were randomly divided into 5 groups according to body weight, with 10 rabbits/per group Animals in 1-5 groups were injected with medical sodium hyaluronate gel in the anterior chamber of right eye in a single dose, 100 μL/eye, to induce ocular hypertension. In 5-15 minutes after modeling, the right eye was respectively dropped with a solvent, K-115 and test samples (compound 63 at different concentrations), the left eye was dropped with a solvent, the volume of the administration was all 50 μL/eye, and intraocular pressure in both eyes of the animals was measured before administration and in 2, 4, 6, 8 and 10 hours after administration, respectively. Experimental results are shown in Table 4:
TABLE-US-00004 TABLE 4 Change results of the intraocular pressure of both eyes (Mean +/− SEM) of animals in each group before and after modeling and administration Time point of measurement 2 hours 4 hours 6 hours 8 hours 10 hours Measured Before after after after after after Intraocular admini- admini- admini- admini- admini- admini- pressure stration stration stration stration stration stration Negative Intra- 9.93 6.40 6.97 9.83 12.03 13.63 control ocular pressure in the left eye (mmHg) Intra- 10.57 8.57 14.00 17.67 16.30 15.87 ocular pressure in the right eye (mmHg) K-115 Intra- 11.80 6.50 8.54 11.40 13.20 13.37 4 mg/mL ocular pressure in the left eye (mmHg) Intra- 11.77 8.20 11.27 15.07 15.57 16.47 ocular pressure in the right eye (mmHg) Compound Intra- 10.53 6.07 6.60 9.10 11.50 12.43 63 ocular 0.25 mg/mL pressure in the left eye (mmHg) Intra- 11.43 8.47 8.60 11.20 13.37 13.10 ocular pressure in the right eye (mmHg) Compound Intra- 11.27 6.60 7.04 11.23 12.84 13.10 63 ocular 2.0 mg/mL pressure in the left eye (mmHg) Intra- 11.23 8.52 11.56 13.81 12.96 11.96 ocular pressure in the right eye (mmHg) Compound Intra- 11.20 5.93 6.43 10.10 11.87 13.19 63 ocular 8.0 mg/mL pressure in the left eye (mmHg) Intra- 11.53 7.40 7.97 8.84 8.60 8.78 ocular pressure in the right eye (mmHg)
[0473] Conclusion: In the model of acute ocular hypertension, the compounds of the present disclosure all exhibited good hypotensive effects at different test doses, showed certain dose correlation, and the hypotensive amplitude and the action duration were both superior to K-115.
[0474] Experimental example 4. Experiment on reduction of intraocular pressure and ocular toxicity of New Zealand rabbits with normal intraocular pressure by repeated eye-drop administration for 14 days
[0475] Experiment Objective:
[0476] The intraocular pressure reduction action and the potential ocular toxicity of compound 63 were explored by 14 days of repeated eye-drop administration in rabbits with normal intraocular pressure.
[0477] Experiment Materials:
[0478] Male New Zealand rabbits, aged 97-127 days, weighted 2.6-3.5 kg, purchased from PizhouDongfang Breeding Co. Ltd.
[0479] Experiment Operation 1:
[0480] The male New Zealand rabbits were randomly divided into 7 groups, with 6 rabbits per group. Animals were randomly divided according to body weight Animals in 1-7 groups were eye-dropped with solvent/control sample/test sample in both eyes, the volume of the administration was all 50 μL/eye, once per day for 14 consecutive days, and the day of the administration was recorded as Day 1. On Day 1, intraocular pressure of the animals was measured before the administration and in 1, 2, 4, 6, 8 and 10 hours after the administration, respectively; on Days 2-14, intraocular pressure of the animals was measured in 1 hour after daily administration for the group administered with K-115, and intraocular pressure of the animal was measured in 4 hours after daily administration for the other groups. Experiment results are shown in Tables 5, 6 and 7:
TABLE-US-00005 TABLE 5 Changes of the intraocular pressure of both eyes (Mean +/− +/− SEM) of animals in each group before and after Day 1 administration Time point of measurement 1 hour 2 hours 4 hours 6 hours 8 hours 10 hours Measured Before after after after after after after intraocular admini- admini- admini- admini- admini- admini- admini- pressure stration stration stration stration stration stration stration Negative Mean 9.89 10.31 10.97 11.31 11.94 12.97 14.42 control intraocular pressure of both eyes (mmHg) K-115 Mean 10.83 7.72 9.00 9.67 10.56 10.28 12.81 4 mg/mL intraocular pressure of both eyes (mmHg) Change of −0.94 2.59 1.97 1.64 1.38 2.69 1.61 intraocular pressure (mmHg) Compound Mean 10.11 8.83 7.56 7.11 9.61 9.06 9.97 63 intraocular 0.5 mg/mL pressure of both eyes (mmHg) Change of −0.22 1.48 3.41 4.2 2.33 3.91 4.45 intraocular pressure (mmHg) Compound Mean 9.94 7.08 6.67 6.44 8.00 9.22 10.56 63 intraocular 1.0 mg/mL pressure of both eyes (mmHg) Change of −0.05 3.23 4.3 4.87 3.94 3.75 3.86 intraocular pressure (mmHg) Compound Mean 9.67 8.56 7.47 7.47 7.25 8.44 9.67 63 intraocular 2.0 mg/mL pressure of both eyes (mmHg) Change of 0.22 1.75 3.5 3.84 4.69 4.53 4.75 intraocular pressure (mmHg) Compound Mean 9.67 10.44 7.81 6.47 6.22 7.33 8.64 63 intraocular 4.0 mg/mL pressure of both eyes (mmHg) Change of 0.22 −0.13 3.16 4.84 5.72 5.64 5.78 intraocular pressure (mmHg) Compound Mean 10.17 7.69 6.39 7.06 7.11 7.22 7.97 63 intraocular 8.0 mg/mL pressure of both eyes (mmHg) Change of −0.28 2.62 4.58 4.25 4.83 5.75 6.45 intraocular pressure (mmHg)
TABLE-US-00006 TABLE 6 Changes of the intraocular pressure of both eyes (Mean +/− SEM) of animals in each group before and after Days 2-7 repeated administration Measured intraocular Time point of measurement pressure Day 2 Day 3 Day 4 Day 5 Day 6 Day 7 Negative Mean 11.08 10.39 10.61 10.97 9.31 10.06 control intraocular pressure of both eyes (mmHg) K-115 Mean 6.83 6.81 6.92 6.67 6.75 7.39 4 mg/mL intraocular pressure of both eyes (mmHg) Change of 4.25 3.58 3.69 4.3 2.56 2.67 intraocular pressure (mmHg) Compound Mean 7.94 7.31 7.78 7.19 6.61 6.00 63 intraocular 0.5 mg/mL pressure of both eyes (mmHg) Change of 3.14 3.08 2.83 3.78 2.7 4.06 intraocular pressure (mmHg) Compound Mean 7.64 7.50 7.33 7.44 6.92 7.00 63 intraocular 1.0 mg/mL pressure of both eyes (mmHg) Change of 3.44 2.89 3.28 3.53 2.39 3.06 intraocular pressure (mmHg) Compound Mean 7.03 7.25 6.86 6.47 5.92 5.78 63 intraocular 2.0 mg/mL pressure of both eyes (mmHg) Change of 4.05 3.14 3.75 4.5 3.39 4.28 intraocular pressure (mmHg) Compound Mean 6.53 5.58 5.53 6.61 5.47 5.56 63 intraocular 4.0 mg/mL pressure of both eyes (mmHg) Change of 4.55 4.81 5.08 4.36 3.84 4.5 intraocular pressure (mmHg) Compound Mean 6.33 5.78 5.97 6.42 5.56 5.28 63 intraocular 8.0 mg/mL pressure of both eyes (mmHg) Change of 4.75 4.61 4.64 4.55 3.75 4.78 intraocular pressure (mmHg)
TABLE-US-00007 TABLE 7 Changes of the intraocular pressure of both eyes (Mean +/− SEM) of animals in each group before and after Days 8-14 repeated administration Measured intraocular Time point of measurement pressure Day 8 Day 9 Day 10 Day 11 Day 12 Day 13 Day 14 Negative Mean 11.31 9.44 9.75 9.69 9.53 11.39 10.97 control intraocular pressure of both eyes (mmHg) K-115 Mean 7.53 7.78 7.94 7.94 7.89 8.25 8.50 4 mg/mL intraocular pressure of both eyes (mmHg) Change of 3.78 1.66 1.81 1.75 1.64 3.14 2.47 intraocular pressure (mmHg) Compound Mean 7.11 6.19 6.75 6.44 6.25 8.08 7.25 63 intraocular 0.5 mg/mL pressure of both eyes (mmHg) Change of 4.2 3.25 3 3.25 3.28 3.31 3.72 intraocular pressure (mmHg) Compound Mean 7.28 6.94 7.31 7.50 6.89 8.06 7.44 63 intraocular 1.0 mg/mL pressure of both eyes (mmHg) Change of 4.03 2.5 2.44 2.19 2.64 3.33 3.53 intraocular pressure (mmHg) Compound Mean 6.78 5.72 5.94 6.03 6.03 7.11 7.06 63 intraocular 2.0 mg/mL pressure of both eyes (mmHg) Change of 4.53 3.72 3.81 3.66 3.5 4.28 3.91 intraocular pressure (mmHg) Compound Mean 6.78 5.47 5.08 5.83 5.89 7.03 6.33 63 intraocular 4.0 mg/mL pressure of both eyes (mmHg) Change of 4.53 3.97 4.67 3.86 3.64 4.36 4.64 intraocular pressure (mmHg) Compound Mean 5.72 5.19 5.56 5.53 5.56 6.67 6.94 63 intraocular 8.0 mg/mL pressure of both eyes (mmHg) Change of 5.59 4.25 4.19 4.16 3.97 4.72 4.03 intraocular pressure (mmHg)
[0481] Conclusion: A single dose of compound 63 exhibited superior efficacy (highest hypotensive effect and action duration) at all test doses (0.5-8.0 mg/mL), significantly superior to K-115. After 14 days of continuous administration, compound 63 can maintain a significant hypotensive effect at the dose of 0.5 mg/mL and is still significantly superior to K-115 in evaluation of peak hypotensive effect (Cmax).
[0482] Experiment Operation 2:
[0483] 42 male New Zealand rabbits were randomly divided into 7 groups, with 6 rabbits per group. Animals were randomly divided according to body weight Animals in 1-7 groups were eye-dropped with normal saline in the left eye and with solvent/control sample/test sample in the right eye respectively by a volume of 50 μL/eye, once per day for 14 consecutive days, and the day of administration was recorded as Day 1. The intraocular pressure of the animals was measured before Day 1 administration and in 1, 2, 4, 6, 8 and 10 hours after Day 1 administration, respectively (Table 8).
[0484] Before the experiment (Day 2/Day 1), before daily administration during the administration (Days 1-14), and in 1, 2, 4, 24, 48 and 72 hours after the last administration (Day 14), the eyes of animals were examined for ocular stimulation response and fluorescein sodium by a hand-held slit lamp (scoring by referring to the scoring criteria).
TABLE-US-00008 TABLE 8 Changes of the intraocular pressure of both eyes (Mean +/− SEM) of animals in each group before and after Day 1 administration Time point of measurement 1 hour 2 hours 4 hours 6 hours 8 hours 10 hours Measured Before after after after after after after intraocular admini- admini- admini- admini- admini- admini- admini- pressure stration stration stration stration stration stration stration Negative 9.89 10.31 10.97 11.31 11.94 12.97 14.42 12.17 control intraocular pressure of both eyes (mmHg) K-115 Mean 10.17 5.50 7.06 9.00 9.17 10.61 12.33 4 mg/mL intraocular pressure of both eyes (mmHg) Change of 0.83 −3.39 −2.50 −0.89 −1.61 0.23 0.17 intraocular pressure (mmHg) Compound Mean 9.84 6.22 6.17 6.67 8.06 9.72 10.95 63 intraocular 0.25 mg/mL pressure of both eyes (mmHg) Change of 0.50 −2.67 −3.39 −3.22 −2.72 −0.66 −1.22 intraocular pressure (mmHg) Compound Mean 9.22 5.39 5.50 6.95 8.94 9.89 11.56 63 intraocular 0.5 mg/mL pressure of both eyes (mmHg) Change of −0.11 −3.50 −4.05 −2.94 −1.84 −0.50 −0.61 intraocular pressure (mmHg) Compound Mean 9.17 5.56 5.39 6.67 7.33 10.50 10.89 63 intraocular 1.0 mg/mL pressure of both eyes (mmHg) Change of −0.17 −3.33 −4.17 −3.22 −3.45 0.11 −1.28 intraocular pressure (mmHg) Compound Mean 9.72 8.11 5.84 6.06 7.11 9.00 10.05 63 intraocular 2.0 mg/mL pressure of both eyes (mmHg) Change of 0.39 −0.78 −3.72 −3.83 −3.67 −1.39 −2.11 intraocular pressure (mmHg) Compound Mean 10.00 6.61 5.84 5.72 6.61 8.39 10.67 63 intraocular 4.0 mg/mL pressure of both eyes (mmHg) Change of 0.67 −2.28 −3.72 −4.17 −4.17 −2.00 −1.50 intraocular pressure (mmHg)
[0485] Conclusion: A single dose of compound 63 exhibited superior efficacy (highest hypotensive effect and action duration) at all test doses (0.25-4.0 mg/mL), significantly superior to K-115.
[0486] Before the experiment (Day 2/Day 1), before the first daily administration (Days 1-14) during the administration, and in 1, 2, 4, 24, 48 and 72 hours after the last administration, the eyes of animals were examined for ocular stimulation response by a hand-held slit lamp. The scoring criteria were as follows.
TABLE-US-00009 Ocular stimulation response Score Cornea No turbidity 0 Scattered or diffuse turbidity, clear and visible iris 1 Semi-transparent region easy to be recognized, obscure iris 2 Gray semi-transparent region, details of the iris being 3 obscure, marginally visible pupils Opaque cornea, iris beyond recognition 4 Iris Normal 0 Remarkably deepened, congested and swollen crease, slight 1 congestion around cornea, pupils still responsive to light Bleeding/visible necrosis/not responsive to light 2 (or at least one of them) Conjunctiva Congestion (palpebral conjunctiva and bulbar conjunctiva) Normal blood vessels 0 Congested and bright red blood vessels 1 Congested and dark red blood vessels, beyond recognition 2 Diffuse congestion and being magenta 3 Edema No edema 0 Slight edema (eyelid included) 1 Obvious edema accompanied by partial eyelid ectropion 2 Edema to near eyelid semi-closure 3 Edema to beyond eyelid semi-closure 4 Secretion No secretion 0 A small amount of secretion 1 Secretion making eyelids and eyelashes moist or adhered 2 Secretion making the entire eye region moist or adhered 3 Maximum total score 16
[0487] Evaluation of ocular stimulation response: adding the maximum score of cornea, iris, conjunctiva, edema, and secretion to obtain the total score of ocular stimulation symptoms for each animal eye at each time point. For the score of ocular stimulation symptom, calculate the integral mean value of each group of animals at each observation time point, and determine the degree of ocular stimulation of each group of animals at each time point according to the following table.
[0488] Evaluation Standard of Ocular Stimulation
TABLE-US-00010 Score Evaluation 0-3 No stimulation 4-8 Slight stimulation 9-12 Mild stimulation 13-16 Severe stimulation
[0489] Fluorescein sodium examination: after each ocular stimulation response examination, a hand-held slit lamp was used for fluorescein sodium examination, and the scoring criteria were as follows:
TABLE-US-00011 Fluorescent staining Fluorescein was used to aid the diagnosis of corneal epithelial injury. Fluorescent staining areas can be divided into grades 0-4 according to the degree of corneal opacity: No fluorescent staining. 0 Local small-area slight fluorescent staining. Observed under 1 diffuse light, clear and visible eye structures under cornea (clear pupil edge, observation not affected by fluorescent staining). Local small-area mild fluorescent staining Observed under 2 diffuse light, although some details of the eye under the cornea missed, structure being clear and visible. Large obvious fluorescent staining in the staning area 3 Observed under diffuse light, eye structures under cornea marginally recognizable. Serious fluorescent staining Observed under diffuse light, eye 4 structures under cornea beyond recognition.
[0490] The results of the experiment were as follows:
[0491] According to the evaluation criteria of ocular stimulation, the total score of ocular stimulation response of each group at each time point was less than 3, and all were classified as no stimulation according to the standard.
[0492] During the experiment, the scores of fluorescein sodium examination on eyes (with left eyes treated with normal saline, and right eyes treated with the solvent, K-115 and compound 63) were all lower than 1. Corneal fluorescence staining score of 1 was observed in each group of animals at each treatment and individual time point, and biological staining was considered. There was no corneal epithelial injury in each group at each time point.
[0493] Conclusion: Under the conditions of this experiment, K-115 was eye-dropped at the concentration of 4 mg/mL for 14 consecutive days by 50 μL/eye/day, showing no stimulation. Compound 63 was eye-dropped in the concentration range of 0.25-4 mg/mL for 14 consecutive days by 50 μL/eye/day, showing no stimulation.
Experimental example 5. Toxicokinetics Experiment
[0494] Experiment Objective:
[0495] The production rate of active drug ingredients and the exposure amount of active drug ingredients of the compound in plasma were detected after 14 days of continuous administration.
[0496] Experiment Materials:
[0497] Male New Zealand rabbits, aged 3-6 months, weighted 2.0-5.0 kg, purchased from PizhouDongfang Breeding Co. Ltd.
[0498] Experiment Operations:
[0499] After 14 days of continuous administration, blood samples from the group administrated with compound 63 (8.0 mg/mL) were collected at 0 hour (before administration) and in 0.5, 1, 2, 4, 8, and 24 hours after administration on Days 14-15. About 0.8 mL of whole blood was collected from the auricular artery or the saphenous vein of the hind limb (or other appropriate sites) of toxicokinetics experiment animals, and was placed in labeled collection tubes using ethylenediaminetetraacetic acid dipotassium (K.sub.2EDTA) as an anticoagulant. Plasma was obtained by centrifugation for 10 minutes at 3000 rpm and under the condition of 2° C. to 8° C. in 60 minutes after blood collection. All samples were quantitatively detected for the content of the administered compounds in plasma of the experimental animals by liquid chromatography-coupled mass spectrometry (LC-MS) and mass spectrometry (MS) technology.
TABLE-US-00012 TABLE 9 Test results of active compounds in plasma of New Zealand rabbits after 14 days of continuous administration Concentration (ng/mL) Compound No. Time (hour) Group 1 Group 2 Compound 63 0.00 BQL BQL 0.50 3.63 3.44 1.00 2.87 2.20 2.00 1.80 1.66 4.00 0.934 0.818 8.00 BQL BQL 24.0 BQL BQL Note: BQL indicates below the detection limit.
[0500] Conclusion: The metabolite concentration of compound 63 was 0.934 ng/mL in 4 hours after high administration dose of 8 mg/mL; in 8 hours after administration, the concentration of metabolite of the compound was below the detection limit, indicating high system safety.