CN103664903A - Benzoazepine compounds and preparation method and application thereof - Google Patents

Abstract

The present invention relates to a class of benzodiazepine compounds represented by the following general formula I or pharmaceutically acceptable salts or solvates thereof, a preparation method thereof, a pharmaceutical composition comprising the compound, and the preparation of these compounds for the prevention or treatment of The use of drugs for the treatment of diseases related to abnormal cell proliferation, morphological changes, and hyperkinesia related to progressive lymphoma enzymes in vivo, as well as drugs for diseases related to angiogenesis or cancer metastasis, especially in the preparation of drugs for the treatment of Or use in medicines for preventing tumor growth and metastasis.

Description

Benzozepine compounds and their preparation method and use

technical field

The present invention relates to a class of benzodiazepine compounds or their pharmaceutically acceptable salts or solvates having progressive lymphoma enzyme (ALK) inhibitory activity, a preparation method thereof, a pharmaceutical composition containing the compound, and these compounds Use in the preparation of medicines for preventing or treating diseases related to aberrant cell proliferation, morphological changes, and hyperkinesia related to progressive lymphoma enzymes in vivo, as well as medicines for diseases related to angiogenesis or cancer metastasis , especially for use in medicines for treating or preventing tumor growth and metastasis.

Background technique

1. Biological functions and inhibitors of ALK

Aplastic lymphoma kinase (ALK) is a receptor tyrosine kinase that belongs to the insulin receptor superfamily. It was first discovered in progressive large cell lymphoma (ALCL), and ALK is expressed in about 60%-85% of ALCL, while normal ALK is exclusively expressed in the nervous system, especially in the brain of newborns. The expression level of ALK gene in human body decreases with the development and maturation of the brain, the amount in mature brain tissue is very low, and the expression exists in certain regions; the expression of ALK is not found in other systems, especially the hematopoietic system. ALK gene lacks expression in most non-hematopoietic tumors and normal tissues, indicating that the distribution range of ALK protein is extremely narrow.

The ALK gene is located at chromosome 2p23. Under normal circumstances, human ALK can be transcribed to produce a 6222bp mRNA, which consists of 29 exons and encodes a 1620 amino acid sequence of 200KDa type I transmembrane protein ALK. The ALK gene is usually in a dormant state, and due to fusion with other genes, the cells deteriorate and develop into malignant tumors. However, there are many genes that can fuse with it. In non-small cell lung cancer (NSCLC), it is mainly fused with the EML4 gene (echinoderm microtubule-associated protein-like 4), which is related to echinoderm microtubules. The incidence of protein-like 4-anaplastic lymphoma kinase (EML4-ALK) fusion gene in NSCLC is 2% to 7%.

With the continuous deepening of molecular biology research on non-small cell lung cancer (NSCLC), individualized therapy based on molecular markers has moved from the laboratory to the clinic, and significant clinical progress has been made in the treatment of advanced NSCLC patients. It is also important that, in addition to the traditional histopathological classification, NSCLC can also be classified according to the expression of various molecular markers, and the driver genes related to tumorigenesis and development can be targeted. Research and develop new drugs, carry out targeted and individualized molecular targeted therapy, and improve the prognosis of patients. Ideally, all NSCLC patients should be tested for relevant molecular markers before treatment, and targeted treatment should be implemented on the basis of fully understanding the molecular expression characteristics of the patient's tumor, so as to improve the treatment effect. In this context, ALK small-molecule inhibitors and c-Met/ALK dual-target inhibitors have become hot molecular targets.

At present, the small molecule inhibitor PF02341066 (compound 1) developed by Pfizer has been approved for marketing by the US FDA on August 26, 2011. This is also the only small molecule inhibitor of ALK that has been marketed. However, existing clinical studies have shown that drug resistance to PF02341066 has emerged, and the in vivo bioavailability of PF02341066 needs to be further improved. In addition, Japan's AF-802 (compound 2) is in clinical phase 2, Novartis' LDK-378, Astellas' ASP-3026 and ARIAD's AP-26113 are in clinical phase 1. Another 9 compounds are in the discovery stage.

Contents of the invention

One object of the present invention is to provide a class of benzodiazepine bicyclic structure compounds or pharmaceutically acceptable salts or solvates thereof, said compounds have the structure shown in the following general formula I, which is a class of compounds with ALK inhibitory activity compound of.

Another object of the present invention is to provide the preparation method of the benzodiazepine compound represented by the above-mentioned general formula I.

The benzodiazepine compound represented by the general formula I provided by the present invention or its pharmaceutically acceptable salt or solvate can inhibit the tumor by inhibiting the abnormal proliferation, morphological changes and hyperkinetic function of cells related to anaplastic lymphoma enzymes. effect on cell growth. These compounds also have the effect of inhibiting angiogenesis or inhibiting the metastasis of cancer cells.

Therefore, another object of the present invention is to provide the benzodiazepine compound shown in general formula I or its pharmaceutically acceptable salt or solvate in the preparation for preventing or treating organisms with progressive lymphoma enzyme overexpression Related diseases related to abnormal cell proliferation, morphological changes and hyperkinesia, as well as diseases related to angiogenesis or cancer metastasis, especially in the preparation of drugs for treating or preventing tumor growth and metastasis.

Another object of the present invention is to provide a pharmaceutical composition comprising the benzodiazepine compound represented by general formula I or a pharmaceutically acceptable salt or solvate or a mixture thereof as an active ingredient.

Another object of the present invention is to provide a method for treating diseases related to abnormal cell proliferation, morphological changes and hyperkinesia associated with the overexpression of progressive lymphoma enzymes in vivo, as well as diseases related to angiogenesis or cancer metastasis. The method includes administering to a patient a therapeutically effective amount of a pharmaceutical composition comprising a compound represented by general formula I, a pharmaceutically acceptable salt or a pharmaceutically acceptable solvate thereof, or a mixture thereof as an active ingredient.

In order to achieve the above object, one aspect of the present invention provides a benzodiazepine compound represented by the following general formula I or a pharmaceutically acceptable salt or solvate thereof,

Wherein, R ₁ is unsubstituted or phenyl-substituted C1-C8 alkyl; or -TG;

where T is -C(=O)-, -S(=O) ₂ -, -C(=O)N(R _a )-, -BC(=O)-, -BS(=O) ₂ -, or -BC(=O)N(R _a )-; wherein R _a is hydrogen or C1-C4 alkyl; B is C1-C4 alkylene;

G is C1-C8 alkyl; C3-C8 cycloalkyl; C6-C10 aryl; 4-8 membered heterocyclic group containing 1-3 selected from N, O and S atoms; or containing 1-3 optional 5-8 membered heteroaryls from N, O and S atoms;

R ₂ and R ₂ ' are each independently hydrogen, C1-C8 alkyl or C6-C10 aryl;

其中，R_b和R_c各自独立地为氢或C1-C4烷基；Alternatively, _R2 and _R2 ' form the group

Wherein, R _b and R _c are each independently hydrogen or C1-C4 alkyl;

R and R are _each independently hydrogen, halogen, cyano, nitro _, hydroxyl, amino or C1-C8 alkyl;

Or, R ₃ and R ₄ form a C4-C8 aliphatic ring together with the connected carbon atoms; a C6-C10 aromatic ring; a 4-8 membered heterocyclic ring containing 1-3 selected from N, O and S atoms; or Containing 1-3 5-8 membered heteroaromatic rings selected from N, O and S atoms;

R ₅ and R ₆ are each independently hydrogen, C1-C8 alkyl or

Wherein L is -C(=O)-, -S(=O) ₂ -, -C(=O)N(R _d )- or -S(=O) ₂ N(R _d )-; wherein, R _d is hydrogen or C1-C4 alkyl;

M is hydrogen or C1-C8 alkyl.

Preferably, in compounds of general formula I,

Wherein, R ₁ is unsubstituted or phenyl-substituted C1-C6 alkyl; or -TG;

where T is -C(=O)-, -S(=O) ₂ -, -C(=O)N(R _a )-, -BC(=O)-, -BS(=O) ₂ -, or -BC(=O)N(R _a )-; wherein, R _a is hydrogen or C1-C4 alkyl; B is C1-C4 alkylene;

G is C1-C6 alkyl; C3-C6 cycloalkyl; C6-C8 aryl; 4-6 membered heterocyclyl containing 1-3 selected from N, O and S atoms; or containing 1-3 optional 5-7 membered heteroaryls from N, O and S atoms;

R ₂ and R ₂ ' are each independently hydrogen, C1-C6 alkyl or C6-C8 aryl;

其中，R_b和R_c各自独立地为氢或C1-C4烷基；Alternatively, _R2 and _R2 ' form the group

Wherein, R _b and R _c are each independently hydrogen or C1-C4 alkyl;

R and R are _each independently hydrogen, halogen, cyano, nitro _, hydroxyl, amino or C1-C6 alkyl;

Or, R ₃ and R ₄ form a C4-C6 aliphatic ring together with the connected carbon atoms; a C6-C8 aromatic ring; a 4-6 membered heterocyclic ring containing 1-3 selected from N, O and S atoms; or Containing 1-3 5-7 membered heteroaromatic rings selected from N, O and S atoms;

R ₅ and R ₆ are each independently hydrogen, C1-C6 alkyl or

L is -C(=O)-, -S(=O) ₂ -, -C(=O)N(R _d )- or -S(=O) ₂ N(R _d )-; wherein, R _d is hydrogen or C1-C4 alkyl;

M is hydrogen or C1-C6 alkyl.

More preferably, in the compound of general formula I,

Wherein, R ₁ is unsubstituted or phenyl substituted C1-C4 alkyl; or -TG;

where T is -C(=O)-, -S(=O) ₂ -, -C(=O)N(R _a )-, -BC(=O)-, -BS(=O) ₂ -, or -BC(=O)N(R _a )-; wherein, R _a is hydrogen or C1-C2 alkyl; B is C1-C2 alkylene;

G is C1-C4 alkyl; C3-C6 cycloalkyl; C6-C8 aryl; 4-6 membered heterocyclyl containing 1-3 selected from N, O and S atoms; or containing 1-3 optional 5-7 membered heteroaryls from N, O and S atoms;

R ₂ and R ₂ ' are each independently hydrogen, C1-C4 alkyl or C6-C8 aryl;

其中，R_b和R_c各自独立地为氢或C1-C4烷基；Alternatively, _R2 and _R2 ' form the group

Wherein, R _b and R _c are each independently hydrogen or C1-C4 alkyl;

R and R are _each independently hydrogen, halogen, cyano, nitro _, hydroxyl, amino or C1-C4 alkyl;

Or, R ₃ and R ₄ form a C4-C6 aliphatic ring together with the connected carbon atoms; a C6-C8 aromatic ring; a 4-6 membered heterocyclic ring containing 1-3 selected from N, O and S atoms; or Containing 1-3 5-7 membered heteroaromatic rings selected from N, O and S atoms;

R ₅ and R ₆ are each independently hydrogen, C1-C4 alkyl or

L is -C(=O)-, -S(=O) ₂ -, -C(=O)N(R _d )- or -S(=O) ₂ N(R _d )-; wherein, R _d is hydrogen or C1-C2 alkyl;

M is hydrogen or C1-C4 alkyl.

Further preferably, in the compound of general formula I,

Wherein, R ₁ is methyl, benzyl or -TG;

where T is -C(=O)-, -S(=O) ₂ -, -C(=O)N(R _a )-, -BC(=O)-, -BS(=O) ₂ -, or -BC(=O)N(R _a )-; wherein, R _a is hydrogen or methyl; B is methylene;

G is methyl, ethyl, cyclopropyl, phenyl or morpholinyl;

_R2 and _R2 ' are each independently hydrogen, ethyl, n-propyl, isopropyl or phenyl;

Alternatively, R ₂ and R ₂ ' form methylene, 1,1-ethylene or 2,2-propylene;

R and R are _each independently hydrogen, halogen, _cyano , nitro, hydroxyl or amino;

Alternatively, R ₃ and R ₄ form a pyridine ring together with the carbon atoms connected to each other;

R _{and R} are each independently hydrogen or

L is -C(=O)-, -S(=O) ₂ -, -C(=O)N(R _d )- or -S(=O) ₂ N(R _d )-; wherein, R _d Is hydrogen or methyl; M is methyl or isopropyl.

即通式I所示的化合物优选为下述通式V所示的化合物：Preferably, in the above general formula I, one of R _{and R} is hydrogen, and the other is

That is, the compound represented by the general formula I is preferably a compound represented by the following general formula V:

Wherein, the definitions of R ₁ , R ₂ , R ₂ ′, R ₃ , R ₄ , L and M are the same as those of the compound represented by the above general formula I. Typical compounds of the present invention include but are not limited to the compounds shown in Table 1:

Table I

Another aspect of the present invention provides a preparation method of the benzodiazepine compound represented by the general formula I. The benzodiazepine compound represented by the general formula I can be obtained from the compound II and the compound III by coupling or substitution.

Aniline II and pyrimidine III are coupled to obtain a compound represented by general formula I in the presence of a catalyst;

For example, mix 1 equivalent of substituted aniline II and 1 equivalent of substituted pyrimidine III in a microwave tube, add 2 equivalents of cesium carbonate, 0.25 equivalents of palladium acetate, 0.5 equivalents of 4,5-bisdiphenylphosphine-9, 9-Dimethylxanthene and 3 mL of solvent 1,4-1-dioxane were stirred at 150 watts and 120 degrees for 2 hours. The reaction liquid was extracted with ethyl acetate, washed with saturated brine, and dried over anhydrous sodium sulfate. Filter, spin dry, and pass through a silica gel column to obtain the target product.

or,

Aniline II and pyrimidine III undergo a substitution reaction under the catalysis of camphorsulfonic acid to obtain a compound represented by general formula I;

For example, mix 1 equivalent of substituted aniline II and 1 equivalent of pyrimidine III in a microwave tube, add 2 equivalents of camphorsulfonic acid and 3 mL of isopropanol, heat at 150 watts at 80°C for one hour, and wash the reaction solution with ethyl acetate Extract, wash with saturated sodium bicarbonate, wash with saturated brine, and dry over anhydrous sodium sulfate. Filter, spin dry, and pass through a silica gel column to obtain the target product.

Wherein, the definitions of R ₁ , R ₂ , R ₂ ', R ₃ , R ₄ , R ₅ and R ₆ are the same as those in the above general formula I;

X is fluorine, chlorine, bromine or iodine.

Another aspect of the present invention also provides the preparation of the benzodiazepine compound represented by general formula I or its pharmaceutically acceptable salt or its solvate for preventing or treating the overexpression of progressive lymphoma enzymes in vivo Related diseases related to abnormal cell proliferation, morphological changes and hyperkinesia, as well as diseases related to angiogenesis or cancer metastasis, especially in the preparation of drugs for treating or preventing tumor growth and metastasis.

Another aspect of the present invention provides a pharmaceutical composition, which comprises a therapeutically effective amount of one or more benzodiazepine compounds represented by general formula I or pharmaceutically acceptable salts or solvates thereof, and Optionally, a pharmaceutically acceptable carrier or excipient may be further included.

Another aspect of the present invention provides a method for treating diseases related to abnormal cell proliferation, morphological changes and hyperkinesia related to the overexpression of progressive lymphoma enzymes in vivo, as well as diseases related to angiogenesis or cancer metastasis, so The method includes administering to a patient a therapeutically effective amount of a pharmaceutical composition comprising a benzodiazepine compound represented by general formula I, a pharmaceutically acceptable salt or a pharmaceutically acceptable solvate thereof, or a mixture thereof as an active ingredient.

Description of drawings

Fig. 1 is the HPLC spectrogram of racemate S1;

Fig. 2 is the HPLC spectrogram of optical isomer S2;

Fig. 3 is the HPLC spectrogram of optical isomer S3.

Detailed ways

The present invention will be further elaborated below in conjunction with specific examples. These examples are for illustrative purposes only, and do not limit the scope and spirit of the present invention.

Preparation Example

¹ H-NMR is measured with a Varian MercuryAMX300 instrument; 2,4,5-trichloropyrimidine, isatoic anhydride, p-nitrophenylethanol, N-methylbenzylamine, 2,4-dichloropyrido[3,2 -d] pyrimidine, 2-(isopropylsulfonyl)aniline, palladium acetate, 2-chloro-1-morpholin-1-one, 2-chloro-N,N-dimethylacetamide, 4,5 -Bisdiphenylphosphine-9,9-dimethylxanthene was purchased from J&KChemica Bailingwei Chemical Reagent Company, and the rest of the reagents were produced by China Pharmaceutical Reagent Co., Ltd. All solvents were re-distilled before use, and the anhydrous solvents used were obtained by drying according to standard methods; except for the instructions, all reactions were carried out under nitrogen protection and followed by TLC. Washing with aqueous sodium solution and drying with anhydrous sodium sulfate; silica gel (200-300 mesh) column chromatography is used for purification of products unless specified; silica gel (200-300 mesh) is produced by Qingdao Ocean Chemical Factory, GF-254 thin layer The silica gel plate is produced by Yantai Jiangyou Silicone Development Co., Ltd.

1. Synthesis of compound S1

The synthesis reference of raw material 1-1: WO2008080891. The synthesis reference of raw material 1-2: Bioorganic & Medicinal Chemistry Letters, 21(2), 660-663; 2011.

Compounds 1-1, 1-2, palladium acetate, ligand 4,5-bisdiphenylphosphine-9,9-dimethylxanthene, and cesium carbonate were mixed in a microwave tube, and 2ml 1,4- Dioxane, microwave at 110 degrees, 1 hour, 150 watts. Cool to room temperature, add ethyl acetate to wash with water, combine the organic layers, wash with saturated sodium bicarbonate, dry over anhydrous sodium sulfate, filter and spin dry, put on a silica gel column, dichloromethane: methanol: ammonia water = 30:1:0.1 to obtain the product S1 .

Compound S1: ¹ H NMR (300MHz, CDCl ₃ ) δ11.05(s,1H),8.64(d,J=8.4Hz,1H),8.13–8.03(m,1H),7.50–7.35(m,3H) ,7.16(s,1H),7.04(dd,J=12.7,7.1Hz,3H),6.25(s,1H),3.17–2.62(m,8H),2.36(s,5H),1.30(d,J =7.2Hz,3H).

2. Synthesis of Compound S2

The synthesis reference of raw material 2-1: WO2008080891.

The synthesis method of compound S2 is the same as that of S1.

Compound S2: ¹ H NMR (300MHz, CDCl ₃ ) δ11.05(s, 1H), 8.64(d, J=8.4Hz, 1H), 8.13-8.03(m, 1H), 7.50-7.35(m, 3H) ,7.16(s,1H),7.04(dd,J=12.7,7.1Hz,3H),6.25(s,1H),3.17–2.62(m,8H),2.36(s,5H),1.30(d,J =7.2Hz,3H).

3. Synthesis of compound S3

The synthesis reference of raw material 3-1: WO2008080891.

The synthesis method of compound S3 is the same as that of S1.

Compound S3: ¹ H NMR (300MHz, CDCl ₃ ) δ11.05 (s, 1H), 8.64 (d, J=8.4Hz, 1H), 8.13–8.03 (m, 1H), 7.50–7.35 (m, 3H) ,7.16(s,1H),7.04(dd,J=12.7,7.1Hz,3H),6.25(s,1H),3.17–2.62(m,8H),2.36(s,5H),1.30(d,J =7.2Hz,3H).

4. Synthesis of compound S4

Synthesis reference of starting material 4-1: WO2009143389.

The synthesis method of compound S4 is the same as that of S1.

Compound S4: ¹ H NMR (300MHz, CDCl ₃ ) δ9.56(s, 1H), 8.53(d, J=8.4Hz, 1H), 8.11(d, J=1.4Hz, 1H), 7.87(d, J =7.9Hz,1H),7.53(t,J=7.9Hz,1H),7.36(d,J=8.1Hz,1H),7.23–7.13(m,2H),7.00(d,J=8.0Hz,1H ),3.29-3.17(m,1H),3.14-2.94(m,2H),2.92-2.71(m,2H),2.64(d,J=12.2Hz,1H),2.34(s,4H),2.24( d,J=9.1Hz,1H),1.29(d,J=6.0Hz,6H),1.22(d,J=7.2Hz,3H).

5. Synthesis of Compound S5

The synthesis of intermediate 5-3 is the same as that of 1-2.

The synthesis of compound S5 is the same as that of S1.

Compound S5: ¹ H NMR (300MHz, CDCl ₃ ) δ12.37(s, 1H), 8.97(d, J=7.8Hz, 1H), 8.64(d, J=4.1Hz, 1H), 7.82(d, J =8.4Hz,1H),7.59–7.46(m,4H),7.07(d,J=7.9Hz,1H),6.87(d,J=7.9Hz,1H),6.52(s,1H),6.47(d ,J=7.8Hz,2H),3.32(ddd,J=45.7,24.5,12.4Hz,5H),3.05(d,J=4.8Hz,3H),2.92(d,J=11.5Hz,2H),2.50 (s,3H),1.36(d,J=7.3Hz,3H).

6. Synthesis of Compound S6

Reference patent for the synthesis of raw material 6-1: WO8500808.

The synthesis of compound S6 is the same as that of S1.

Compound S6: ¹ H NMR (300MHz, CDCl ₃ ) δ11.00(s, 1H), 8.59(d, J=8.0Hz, 1H), 8.00(s, 1H), 7.58(d, J=7.5Hz, 1H ),7.46(d,J=8.2Hz,1H),7.33(dd,J=16.2,8.9Hz,4H),7.17(d,J=7.0Hz,2H),7.11–7.03(m,2H),6.80 (s,1H),6.58(s,1H),6.21(s,1H),4.30(s,1H),3.06(s,2H),3.02(d,J=4.9Hz,3H),2.87(s, 4H), 2.41(s, 3H).

7. Synthesis of compound S7

The synthesis of raw material 7-3 is the same as that of 1-2, the synthesis reference: WO2008080891.

The synthesis of compound S7 is the same as that of S1.

Compound S7: ¹ H NMR (300MHz, CDCl ₃ ) δ11.14(s, 1H), 8.51(d, J=8.2Hz, 1H), 7.52–7.30(m, 6H), 7.29(d, J=7.2Hz ,1H),7.23(d,J=7.5Hz,1H),7.13(d,J=8.2Hz,1H),7.04(t,J=7.5Hz,1H),6.93(s,1H),6.79(s ,1H),6.30(s,1H),4.45(d,J=8.9Hz,1H),3.19(d,J=11.3Hz,2H),3.01(d,J=4.8Hz,3H),2.99–2.81 (m,3H),2.45(s,4H).

8. Synthesis of compound S8

Synthesis reference of starting material 8-1: WO2008080891.

Synthesis of intermediate 8-2: Dissolve raw material 8-1 in DMF, slowly add DMF diluent of allylbenzylamine dropwise under ice bath, move to room temperature and stir for 1 hour after dropping, pour the reaction solution into ice water, Add ethyl acetate for extraction, combine the organic phases, wash with saturated sodium bicarbonate and saturated sodium chloride, and dry over anhydrous sodium sulfate. Filter and spin dry to obtain 8-2.

Synthesis of intermediate 8-3: raw material 8-2, tris(dibenzylideneacetone) dipalladium, tri-tert-butylphosphine tetrafluoroborate and triethylamine mixed in 1,4-dioxane, 90 degrees Stir overnight. After cooling, it was poured into water and extracted with ethyl acetate. After the organic phase was dried, column chromatography gave 8-3.

Synthesis of intermediate 8-4: raw material 8-3 was added to DCE, potassium carbonate and 1-chloroethyl chloroformate were added, refluxed for 2 hours, potassium carbonate was filtered, filtrate was spin-dried and methanol was added, reflux was continued for 5 hours, methanol was spin-dried and methanol was added Extracted with dichloromethane, dried and passed through the column to obtain the intermediate 8-4. ¹ H NMR (300MHz, CDCl ₃ ) δ8.13-7.97 (m, 2H), 7.25 (d, J=3.4Hz, 1H), 5.32 (s,1H),5.23(s,1H),3.58(s,2H),3.13-2.90(m,4H).

Synthesis of intermediate 8-5: raw material 8-4, 2-chloro-1-morpholin-1-one and potassium carbonate were mixed in acetonitrile, heated to 45 degrees, stirred overnight, extracted product with ethyl acetate, column layer The intermediate 8-5. ¹ H NMR (300MHz, CDCl ₃ ) δ8.12-7.98(m, 2H), 7.25(d, J=2.7Hz, 1H), 5.43-5.25(m, 2H), 3.61 (d,J=7.1Hz,4H),3.52(s,2H),3.48(s,4H),3.39(s,2H),2.94(dd,J=10.7,3.4Hz,4H).

Synthesis of intermediate 8-6: raw material 8-5 was dissolved in a mixture of ethanol and water, iron powder and ammonium chloride were added, heated to reflux for 1 h, filtered, the filtrate was extracted with ethyl acetate, the organic layer was dried, and the column layer The intermediate 8-6. ¹ H NMR (300MHz, CDCl ₃ ) δ6.83(d, J=7.9Hz, 1H), 6.58(d, J=2.4Hz, 1H), 6.48(dd, J= 7.9,2.5Hz,1H),5.22(d,J=1.7Hz,1H),5.12–5.07(m,1H),3.67–3.47(m,11H),3.40(s,2H),3.35(d,J =1.7Hz,2H),2.79(tt,J=9.5,3.0Hz,4H).

Synthesis of compound S8: Dissolve raw materials 1-2 and 8-6 in isopropanol, add 4A molecular sieves, camphorsulfonic acid, microwave at 80 degrees, 1h, 150w, add sodium bicarbonate and ethyl acetate to extract, and dry the organic layer Compound S8 was obtained by direct column chromatography.

Compound S8: ¹ H NMR (300MHz, CDCl ₃ ) δ11.07(s, 1H), 8.62(d, J=8.6Hz, 1H), 8.08(s, 1H), 7.54–7.28(m, 4H), 7.09 –6.95(m,3H),6.31(s,1H),5.22(s,1H),5.12(s,1H),3.67–3.46(m,12H),3.02(d,J=4.8Hz,3H), 2.86(s,4H).

9. Synthesis of Compound S9

The synthesis of intermediate 9-2 is the same as that of 8-6.

The synthesis of compound S9 is the same as S8.

Compound S9: ¹ H NMR (300MHz, DMSO) δ11.62(s, 1H), 9.44(s, 1H), 8.73(t, J=6.9Hz, 2H), 8.22(s, 1H), 7.73(d, J=7.9Hz,1H),7.61(d,J=7.0Hz,2H),7.46(t,J=7.8Hz,1H),7.11(dd,J=11.5,8.1Hz,2H),5.21(d, J=11.3Hz,2H),4.17(s,2H),3.40(d,J=5.8Hz,2H),2.89(d,J=5.9Hz,2H),2.79(d,J=4.5Hz,3H) ,2.72(s,3H).

10. Synthesis of Compound S10

The synthesis of compound S10 is the same as S8.

Compound S10: ¹ H NMR (400MHz, CDCl ₃ ) δ9.60(s, 1H), 8.52(d, J=8.3Hz, 1H), 8.16(s, 1H), 7.92(dd, J=8.0, 1.4Hz ,1H),7.66–7.57(m,1H),7.48(dd,J=6.2,2.2Hz,2H),7.38(s,1H),7.09(d,J=8.9Hz,1H),5.25(s, 2H),4.28(s,2H),3.65–3.57(m,2H),3.30–3.21(m,1H),3.00–2.94(m,2H),2.58(s,3H),1.32(d,J= 6.9Hz,6H).

11. Synthesis of Compound S11

Synthesis of raw material 11-1 Reference: WO2008080891

The synthesis of intermediates 11-2 and 11-3 is the same as that of 9-1 and 9-2.

Intermediate 11-3: ¹ HNMR (300MHz, CDCl ₃ ) δ6.86(t, J=7.3Hz, 1H), 6.56-6.37(m, 2H), 3.55(ddd, J=23.1, 15.2, 10.3Hz, 6H),3.09-2.87(m,2H),2.84-2.67(m,1H),2.37(dd,J=14.5,7.2Hz,2H),1.27(dd,J=9.0,5.5Hz,3H),1.14 (dd,J=13.6,7.4Hz,3H).

The synthesis of compound S11 is the same as that of S1.

Compound S11: ¹ H NMR (400MHz, CDCl ₃ ) δ11.13(d, J=20.9Hz, 1H), 8.64(t, J=8.4Hz, 1H), 8.10(s, 1H), 7.53(d, J =6.9Hz,1H),7.46-7.35(m,2H),7.31(d,J=13.3Hz,1H),7.23(s,0H),7.14–6.94(m,2H),6.59(s,1H) ,3.96–3.42(m,4H),3.05(dd,J=15.7,5.8Hz,5H),2.84(dd,J=15.3,6.2Hz,1H),2.41(q,J=7.4Hz,2H), 1.33–1.26(m,2H),1.23(d,J=7.2Hz,2H),1.16(td,J=7.4,4.6Hz,3H).

12. Synthesis of Compound S12

The synthesis of compound S12 is the same as S8.

Compound S12: ¹ H NMR (300MHz, CDCl ₃ ) δ11.09(s, 1H), 8.63(d, J=8.4Hz, 1H), 8.06(s, 1H), 7.48(d, J=7.6Hz, 1H ),7.43–7.31(m,3H),7.15(s,1H),7.00(dd,J=13.9,7.7Hz,2H),6.57(d,J=4.3Hz,1H),3.24(d,J= 2.1Hz, 2H), 3.13(s, 3H), 2.99(d, J=4.7Hz, 3H), 2.95(s, 3H), 2.92(s, 2H), 2.66(d, J=11.9Hz, 5H) ,1.27(d,J=7.3Hz,3H).

13. Synthesis of Compound S13

Intermediate 13-2: ¹ H NMR (300MHz, CDCl ₃ ) δ6.87(d, J=7.8Hz, 1H), 6.60(d, J=2.1Hz, 1H), 6.55–6.48(m, 1H), 5.21(d,J=16.3Hz,2H),3.57(s,2H),3.29(s,2H),2.78(d,J=4.1Hz,4H),2.43(s,3H).

The synthesis of compound S13 is the same as S8.

Compound S13: ¹ H NMR (300MHz, CDCl ₃ ) δ11.06(s, 1H), 8.62(d, J=8.5Hz, 1H), 8.07(s, 1H), 7.49–7.33(m, 3H), 7.28 (s,1H),7.02(t,J=8.2Hz,2H),6.36(s,1H),5.18(d,J=13.8Hz,2H),3.31(s,2H),2.99(d,J= 4.8Hz,3H),2.83(d,J=15.9Hz,4H),2.42(s,3H).

14. Synthesis of Compound S14

The synthesis of intermediate 14-2 is the same as 8-3.

The synthesis of intermediate 14-3 is the same as that of 8-6.

The synthesis of compound S14 is the same as S8.

Compound S14: ¹ H NMR (300MHz, DMSO) δ11.60(s,1H),9.34(s,1H),8.72(s,2H),8.19(s,1H),7.72(d,J=8.0Hz, 1H),7.46(d,J=8.0Hz,1H),7.40(s,1H),7.34(d,J=7.9Hz,1H),7.28(d,J=4.4Hz,5H),7.25-7.17( m,1H),7.08(t,J=7.5Hz,1H),6.96(d,J=8.1Hz,1H),5.53(d,J=7.0Hz,1H),3.64(s,2H),3.25( s,2H),2.79(d,J=4.4Hz,3H),2.71(s,4H),1.52(d,J=6.9Hz,3H).

15. Synthesis of Compound S15

The synthesis of intermediate 15-3 is the same as that of 12-2.

The synthesis of compound S15 is the same as S8.

Compound S15: ¹ H NMR (300MHz, DMSO) δ11.57(s,1H),9.39(s,1H),8.74(s,2H),8.20(s,1H),7.74(d,J=7.9Hz, 1H),7.48(t,J=8.1Hz,2H),7.39(s,1H),7.14(t,J=7.6Hz,1H),7.01(d,J=8.3Hz,1H),3.68(s, 2H),3.17–2.93(m,2H),2.86–2.76(m,8H),2.69(d,J=12.4Hz,2H),1.63(d,J=24.1Hz,2H),0.83(s,3H ).

16. Synthesis of Compound S16

The synthesis of compound S16 is the same as S8.

Compound S16: ¹ H NMR (300MHz, CDCl ₃ ) δ9.56(s, 1H), 8.53(d, J=8.4Hz, 1H), 8.13(s, 1H), 7.90(dd, J=8.0, 1.5Hz ,1H),7.59(t,J=7.9Hz,1H),7.44-7.36(m,1H),7.31(s,1H),7.19(d,J=1.8Hz,1H),7.01(d,J= 8.2Hz,1H),3.82(d,J=12.5Hz,2H),3.22(dt,J=20.0,9.8Hz,3H),3.00-2.78(m,2H),2.74(s,4H),1.83- 1.66(m,2H),1.30(dd,J=6.8,3.4Hz,7H),0.88(t,J=7.3Hz,3H).

17. Synthesis of Compound S17

The synthesis of compound S17 is the same as that of S1.

Compound S17: ¹ H NMR (300MHz, CDCl ₃ ) δ11.10(d, J=10.7Hz, 1H), 8.64(dd, J=8.2, 3.9Hz, 1H), 8.09(s, 1H), 7.55-7.37 (m,3H),7.23(s,1H),7.15-6.95(m,3H),6.53(d,J=4.8Hz,1H),4.33-3.64(m,2H),3.43(d,J=13.2 Hz,1H),3.14(dd,J=27.2,15.4Hz,1H),3.01(d,J=4.8Hz,3H),2.96–2.66(m,2H),1.91–1.47(m,4H),0.81 (qd,J=18.3,8.2Hz,7H).

18. Synthesis of Compound S18

Synthesis reference of intermediate 18-1: WO2011140338.

The synthesis of intermediate 18-2 is the same as that of 11-3.

Intermediate 18-2: ¹ H NMR (300MHz, CDCl ₃ ) δ6.86(d, J=8.5Hz, 1H), 6.45(d, J=5.1Hz, 2H), 3.81(s, 2H), 3.57( s,1H),3.20–3.03(m,2H),2.91(d,J=12.0Hz,1H),2.81–2.60(m,5H),1.78(m,1H),0.93(d,6H).

The synthesis of compound S18 is the same as S8.

Compound S18: ¹ H NMR (300MHz, DMSO) δ11.57(s,1H),9.39(s,1H),8.74(s,2H),8.20(s,1H),7.74(d,J=7.9Hz, 1H),7.48(t,J=8.1Hz,2H),7.39(s,1H),7.14(t,J=7.6Hz,1H),7.01(d,J=8.3Hz,1H),3.68(s, 2H),3.17–2.93(m,2H),2.86–2.76(m,7H),2.69(s,3H),0.83(d,6H).

19. Synthesis of Compound S19

The synthesis of intermediate 19-1 is the same as that of 13-2.

Intermediate 19-1: ¹ H NMR (300MHz, CDCl ₃ ) δ6.87(d, J=7.8Hz, 1H), 6.60(d, J=2.1Hz, 1H), 6.55–6.48(m, 1H), 5.21(d, J=16.3Hz, 2H), 3.57(s, 2H), 3.29(s, 2H), 2.78(d, J=4.1Hz, 4H), 2.43(s, 3H), 1.80(s, 6H ).

The synthesis of compound S19 is the same as that of S8.

Compound S19: ¹ H NMR (300MHz, CDCl ₃ ) δ11.06(s, 1H), 8.62(d, J=8.5Hz, 1H), 8.07(s, 1H), 7.49-7.33(m, 3H), 7.28 (s,1H),7.02(t,J=8.2Hz,2H),6.36(s,1H),5.18(d,J=13.8Hz,2H),3.31(s,2H),2.99(d,J= 4.8Hz, 3H), 2.83(d, J=15.9Hz, 4H), 2.42(s, 3H), 1.82(s, 6H).

2. Experimental example

1. Molecular level activity evaluation of receptor tyrosine kinase ALK

Molecular level screening of receptor tyrosine kinase ALK activity inhibition of representative compounds, the method used:

Enzyme-linked immunosorbent assay (ELISA)

(1) Enzyme reaction substrate Poly(Glu,Tyr) 4:1 was diluted to 20μg/mL with potassium ion-free PBS (10mM sodium phosphate buffer, 150mMNaCl, pH7.2-7.4), and 125μL/well was coated with enzyme labeling plate and react at 37°C for 12-16 hours. Discard the liquid in the well. Wash the plate, wash the plate three times with 200 μL/well T-PBS (potassium-free PBS containing 0.1% Tween-20), 5 minutes each time. Dry the plate in a 37°C oven for 1-2 hours.

(2) Add 49 μL of ATP solution diluted with reaction buffer (50mM HEPES pH 7.4, 50mM MgCl ₂ , 0.5mM MnCl ₂ , 0.2mM Na ₃ VO ₄ , 1mM DTT) into each well, add 1 μL of compound to each well, and add The test compound was then added with 50 μL of each kinase domain recombinant protein diluted with reaction buffer to start the reaction, and two wells without ATP control wells were required for each experiment. Put it on a shaker (100rpm) at 37°C for 1 hour. The liquid in the well was discarded, and the plate was washed three times with T-PBS.

(3) Add antibody PY99 100 μL/well (the antibody is diluted 1:500 with T-PBS containing BSA 5 mg/mL), and react on a shaker at 37°C for 0.5 hours. The liquid in the well was discarded, and the plate was washed three times with T-PBS.

(4) Add 100 μL/well of horseradish peroxidase-labeled goat anti-mouse secondary antibody (the antibody is diluted 1:2000 with T-PBS containing 5 mg/ml BSA), and react on a shaker at 37°C for 0.5 hours. The liquid in the well was discarded, and the plate was washed three times with T-PBS.

(5) Add 100 μL/well of 2 mg/ml OPD chromogenic solution (diluted with 0.1M citric acid-sodium citrate buffer (pH=5.4) containing 0.03% H ₂ O ₂ ), and react in the dark at 25°C for 1 -10 minutes.

(6) Add 50 μL/well of 2M H ₂ SO ₄ to stop the reaction, and read with VERSAmax on a wavelength-tunable microplate microplate reader with a wavelength of 490 nm.

(7) Analysis of results

The test results of representative compounds are shown in Table 2 below:

Table II

The test results of representative compounds show that the inhibitory rate of this type of compound on ALK is above 80% at a concentration of 10 μM, and the inhibition rate is above 50% at a concentration of 1 μM. The activity of some compounds is equivalent to that of the positive control or Better and therefore a very potential ALK inhibitor.

In addition, we found that some of the compounds have obvious inhibitory activity on the mutant ALK (L1196M) enzyme. The test method is the same as above.

Table 3. Inhibition rate of compounds on receptor tyrosine kinase ALK (L1196M) enzyme activity (%)

2. Compound chiral resolution

Since most of the compounds have at least one chiral center at the 1-position, we resolved them by chiral preparative liquid chromatography to obtain the corresponding optical isomers. For example, the two enantiomers S2 and S3 of compound S1 both showed high ALK inhibitory activity, and the activity of S3 was twice as high as that of S2, indicating that the combination of S3 and the enzyme was better.

(1) Prepare the implementation plan

The chiral isomers were separated by HPLC using a chiral column, the corresponding components were collected, and the solvent was removed by rotary evaporation to obtain pure optical isomers.

Split condition:

Chiral column: CHIRALCEL OJ-H

Chiral column size: 0.46cmI.D.×15cmL

Mobile phase: MeOH/DEA=100/0.1 Flow rate: 1ml/min

Detection wavelength: UV254nm

preparation results

1) Table 4: Optical purity values

2) Table 5: Quality

2) Table 6: Drying balance process

Wb: weight of bottle and stopper; Ws+b: total weight of bottle, stopper and sample; Wsample: final weight of sample

(2) Please refer to Figure 1-3 for the chiral HPLC spectrum.

(3) Enantiomer ALK enzyme inhibitory activity:

Table 7: ALK enzyme inhibitory activity of S1 and its corresponding isomers S2 and S3

3. Evaluation of cell level activity of receptor tyrosine kinase ALK

(1) Test method:

KARPAS 299 cells in the logarithmic growth phase were inoculated into 96-well microculture plates at 5000/well, 100 μL per well, and after culturing overnight, compounds with different concentrations (1, 0.1,) were added for 72 hours, and each concentration was set at three Duplicate the wells, and set the corresponding concentration of normal saline vehicle control and no cells to zero wells. After the effect, the culture medium was discarded, and 10% (w/v) trichloroacetic acid (100 μL/well) was added to fix at 4°C for 1 hour, then washed five times with distilled water, and after drying at room temperature, each well was added SRB solution (4mg/mL, dissolved in 1% glacial acetic acid) 100μL, incubate at room temperature for 15 minutes, wash with 1% glacial acetic acid five times to wash off unbound SRB, dry at room temperature, add 10mM Tris solution to each well 100 μL, the optical density (OD value) at a wavelength of 515 nm was measured with a VERSMax microplate reader. Experiments were repeated twice.

(2) The test results are shown in Table 8 below:

table eight

From the above results, it can be seen that the new compound not only has high activity at the level of ALK enzyme, but also shows obvious activity on KARPAS 299 cells, which are directly related to ALK, and some compounds are more active than the positive compound PF02341066. Therefore, these compounds have better anti-tumor R&D potential

Claims (9)

1. A class of benzodiazepine compounds represented by the following general formula I or pharmaceutically acceptable salts or solvates thereof,

Wherein, R ₁ is unsubstituted or phenyl-substituted C1-C8 alkyl; or -TG; where T is -C(=O)-, -S(=O) ₂ -, -C(=O)N(R _a )-, -BC(=O)-, -BS(=O) ₂ -, or -BC(=O)N(R _a )-; wherein R _a is hydrogen or C1-C4 alkyl; B is C1-C4 alkylene; G is C1-C8 alkyl; C3-C8 cycloalkyl; C6-C10 aryl; 4-8 membered heterocyclic group containing 1-3 selected from N, O and S atoms; or containing 1-3 optional 5-8 membered heteroaryls from N, O and S atoms; R ₂ and R ₂ ' are each independently hydrogen, C1-C8 alkyl or C6-C10 aryl; Alternatively, _R2 and _R2 ' form the group

Wherein, R _b and R _c are each independently hydrogen or C1-C4 alkyl; R and R are _each independently hydrogen, halogen, cyano, nitro _, hydroxyl, amino or C1-C8 alkyl; Or, R ₃ and R ₄ form a C4-C8 aliphatic ring together with the connected carbon atoms; a C6-C10 aromatic ring; a 4-8 membered heterocyclic ring containing 1-3 selected from N, O and S atoms; or Containing 1-3 5-8 membered heteroaromatic rings selected from N, O and S atoms; R ₅ and R ₆ are each independently hydrogen, C1-C8 alkyl or

Wherein L is -C(=O)-, -S(=O) ₂ -, -C(=O)N(R _d )- or -S(=O) ₂ N(R _d )-; wherein, R _d is hydrogen or C1-C4 alkyl; M is hydrogen or C1-C8 alkyl. 2. The benzodiazepine compound or its pharmaceutically acceptable salt or solvate according to claim 1, wherein, _R is an unsubstituted or phenyl substituted C1-C6 alkyl group; or -TG; where T is -C(=O)-, -S(=O) ₂ -, -C(=O)N(R _a )-, -BC(=O)-, -BS(=O) ₂ -, or -BC(=O)N(R _a )-; wherein, R _a is hydrogen or C1-C4 alkyl; B is C1-C4 alkylene; G is C1-C6 alkyl; C3-C6 cycloalkyl; C6-C8 aryl; 4-6 membered heterocyclyl containing 1-3 selected from N, O and S atoms; or containing 1-3 optional 5-7 membered heteroaryls from N, O and S atoms; R ₂ and R ₂ ' are each independently hydrogen, C1-C6 alkyl or C6-C8 aryl; Alternatively, _R2 and _R2 ' form the group Wherein, R _b and R _c are each independently hydrogen or C1-C4 alkyl; R and R are _each independently hydrogen, halogen, cyano, nitro _, hydroxyl, amino or C1-C6 alkyl; Or, R ₃ and R ₄ form a C4-C6 aliphatic ring together with the connected carbon atoms; a C6-C8 aromatic ring; a 4-6 membered heterocyclic ring containing 1-3 selected from N, O and S atoms; or Containing 1-3 5-7 membered heteroaromatic rings selected from N, O and S atoms; R ₅ and R ₆ are each independently hydrogen, C1-C6 alkyl or

L is -C(=O)-, -S(=O) ₂ -, -C(=O)N(R _d )- or -S(=O) ₂ N(R _d )-; wherein, R _d is hydrogen or C1-C4 alkyl; M is hydrogen or C1-C6 alkyl. 3. The benzodiazepine compound or its pharmaceutically acceptable salt or solvate according to claim 2, wherein, R ₁ is unsubstituted or phenyl substituted C1-C4 alkyl; or -TG; where T is -C(=O)-, -S(=O) ₂ -, -C(=O)N(R _a )-, -BC(=O)-, -BS(=O) ₂ -, or -BC(=O)N(R _a )-; wherein, R _a is hydrogen or C1-C2 alkyl; B is C1-C2 alkylene; G is C1-C4 alkyl; C3-C6 cycloalkyl; C6-C8 aryl; 4-6 membered heterocyclyl containing 1-3 selected from N, O and S atoms; or containing 1-3 optional 5-7 membered heteroaryls from N, O and S atoms; R ₂ and R ₂ ' are each independently hydrogen, C1-C4 alkyl or C6-C8 aryl; Alternatively, _R2 and _R2 ' form the group

Wherein, R _b and R _c are each independently hydrogen or C1-C4 alkyl; R and R are _each independently hydrogen, halogen, cyano, nitro _, hydroxyl, amino or C1-C4 alkyl; Or, R ₃ and R ₄ form a C4-C6 aliphatic ring together with the connected carbon atoms; a C6-C8 aromatic ring; a 4-6 membered heterocyclic ring containing 1-3 selected from N, O and S atoms; or Containing 1-3 5-7 membered heteroaromatic rings selected from N, O and S atoms; R ₅ and R ₆ are each independently hydrogen, C1-C4 alkyl or

L is -C(=O)-, -S(=O) ₂ -, -C(=O)N(R _d )- or -S(=O) ₂ N(R _d )-; wherein, R _d is hydrogen or C1-C2 alkyl; M is hydrogen or C1-C4 alkyl. 4. The benzodiazepine compound or its pharmaceutically acceptable salt or solvate according to claim 3, wherein, _R is methyl, benzyl or -TG; where T is -C(=O)-, -S(=O) ₂ -, -C(=O)N(R _a )-, -BC(=O)-, -BS(=O) ₂ -, or -BC(=O)N(R _a )-; wherein, R _a is hydrogen or methyl; B is methylene; G is methyl, ethyl, cyclopropyl, phenyl or morpholinyl; _R2 and _R2 ' are each independently hydrogen, ethyl, n-propyl, isopropyl or phenyl; Alternatively, R ₂ and R ₂ ' form methylene, 1,1-ethylene or 2,2-propylene; R and R are _each independently hydrogen, halogen, _cyano , nitro, hydroxyl or amino; Alternatively, R ₃ and R ₄ form a pyridine ring together with the carbon atoms connected to each other; R _{and R} are each independently hydrogen or

L is -C(=O)-, -S(=O) ₂ -, -C(=O)N(R _d )- or -S(=O) ₂ N(R _d )-; wherein, R _d is hydrogen or methyl; M is methyl or isopropyl. 5. The benzodiazepine compound or its pharmaceutically acceptable salt or solvate according to claim 1, wherein the benzoazepine compound is one of the following compounds:

6. a method for preparing the benzodiazepine compound described in claim 1, Aniline II and pyrimidine III are coupled to obtain a compound represented by general formula I in the presence of a catalyst; or, Aniline II and pyrimidine III undergo a substitution reaction under the catalysis of camphorsulfonic acid to obtain a compound represented by general formula I;

Wherein, the definitions of R ₁ , R ₂ , R ₂ ′, R ₃ , R ₄ , R ₅ and R ₆ are the same as those in claim 1; X is fluorine, chlorine, bromine or iodine. 7. The benzodiazepine compound or its pharmaceutically acceptable salt or solvate described in any one of claims 1-5 is used for the prevention or treatment of cell abnormalities associated with progressive lymphoma enzymes in organisms Use in a drug for diseases related to proliferation, morphological changes, and hyperkinesia, and a drug for diseases related to angiogenesis or cancer metastasis. 8. Use of the benzoazepine compound or its pharmaceutically acceptable salt or solvate according to any one of claims 1-5 in the preparation of medicines for treating or preventing tumor growth and metastasis. 9. A pharmaceutical composition comprising one or more therapeutically effective doses of the benzoazepine compounds or pharmaceutically acceptable salts or solvates thereof according to any one of claims 1-5, And may optionally further comprise a pharmaceutically acceptable carrier or excipient.

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