Preparation of novel indole camptothecin derivative and application of novel indole camptothecin derivative in anti-tumor aspect

Abstract

The invention relates to preparation of a novel indole camptothecin derivative and an application of the novel indole camptothecin derivative in an anti-tumor aspect. The structural general formula of the compound is shown in the specification. In-vitro cytotoxic activity screening results show that the novel indole camptothecin derivative has broad-spectrum antitumor activity, and shows relatively strong inhibitory activity on human liver cancer cells (HepG2), human non-small cell lung cancer cells (A549), human colon cancer cells (SW480), human intrahepatic bile duct cancer cells (RBE), human breast cancer cells (MCF-7), human pancreatic cancer cells (BxPC3), human bladder cancer cells (T24) and human bladder cancer cells (umuc3). Wherein the anti-tumor activity of the compound D-18 is the most prominent, the compound D-18 shows a relatively strong inhibition effect on the tested 8 tumor cell lines, the IC50 range is 0.2585-10.69 mu M, the compound D-18 is obviously superior to that of a contrast drug irinotecan, and the activity of the compound D-18 is equivalent to that of topotecan. Therefore, the indole camptothecin derivative is expected to be developed into a novel antitumor drug. # imgabs0 #

Description

Technical Field

[0001] The invention belongs to the field of natural drug chemistry and relates to the preparation of a novel indole camptothecin derivative and its use in anti-tumor. Background Art

[0002] The indole ring is one of the heterocyclic compounds widely found in nature. This skeleton exhibits good antitumor activity and is commonly found in active ingredients and natural products, such as vinblastine, vincristine, and koumine. As some indoles and azaindoles have been successfully developed as anticancer drugs, the design and synthesis of indole and azaindole derivatives with antitumor activity have received increasing attention and made significant progress. Therefore, the hybridization of indole with other active substances is an effective and commonly used strategy for finding and designing new anticancer drugs.

[0003] Camptothecin is a pentacyclic quinoline alkaloid isolated from the bark and stem of Camptotheca acuminata. It exerts anti-tumor activity by acting on DNA topoisomerase I and has become one of the important lead compounds in the development of anti-cancer drugs. To date, camptothecin drugs such as irinotecan, topotecan, and belotecan have been approved for marketing for tumor treatment. The structure-activity relationship of camptothecin shows that modification of the AB ring structure will significantly affect its anti-tumor activity and is the best modification position for further development of new anti-cancer drugs.

[0004] Therefore, we adopted the skeleton transition design idea, replaced the quinoline AB ring of the camptothecin nucleus with an indole molecular fragment, constructed a series of novel indole camptothecin derivatives, and tested their inhibitory activity against various tumor cells. The experimental results showed that the novel indole camptothecin derivatives had a broad spectrum of anti-tumor activity, among which the highly active compound D-18 had better in vitro inhibitory activity against various tumor cells than the clinical drug irinotecan, and had comparable activity to topotecan, and could be developed as a new type of anti-tumor drug. Summary of the invention

[0005] The present invention provides a novel indole camptothecin derivative preparation and its anti-tumor use

[0006] The structural formula of the novel indole camptothecin derivatives of the present invention is as follows:

[0007]

[0008] The preparation method of the novel indole camptothecin derivatives of the present invention is carried out according to the following chemical formula 1:

[0009]

[0010] By adopting the Fischer indole synthesis reaction, using differently substituted phenylhydrazine or phenylhydrazine hydrochloride as raw materials, heating and reflux with tricyclic ketone for 18-20 hours in an acidic environment, a series of novel indole camptothecin derivatives can be obtained.

[0011] The present invention relates to a novel indole camptothecin derivative that can play a role in the preparation of anti-tumor drugs, and more specifically, can be used in the preparation of drugs for treating human liver cancer, non-small cell lung cancer, colon cancer, bile duct cancer, breast cancer, pancreatic cancer, and bladder cancer. The results of in vitro anti-tumor activity screening show that the novel indole camptothecin derivative has a broad spectrum of anti-tumor activity, and exhibits strong inhibitory activity against human liver cancer cells (HepG2), human non-small cell lung cancer cells (A549), human colon cancer cells (SW480), human intrahepatic bile duct cancer cells (RBE), human breast cancer cells (MCF-7), human pancreatic cancer cells (BxPC3), human bladder cancer cells (T24), and human bladder cancer cells (umuc3). Among them, the anti-tumor activity of compound D-18 is the most prominent, and it exhibits strong inhibitory effects on the 8 tumor cell lines tested, IC 50 The range was 0.2585~10.69μM, which was significantly better than the control drug irinotecan and had the same activity as topotecan.

[0012] Therefore, the novel indole camptothecin derivatives described in the present invention can be used to prepare anti-tumor drugs, and have novel structures, cheap and available raw materials, high product purity, and exhibit strong inhibitory effects on the proliferation of various tumor cell lines, and have good application prospects.

[0013] The above contents of the present invention are further described in detail below through specific implementation modes, but this should not be construed as limiting the present invention. DETAILED DESCRIPTION

[0014] In order to better understand the present invention, the above contents of the present invention are further described in detail below through specific implementation methods. The experimental methods described in the following examples are conventional methods unless otherwise specified.

[0015] Example 1: Synthesis of target compound D-1

[0016]

[0017] The synthesis method of compound D-1 of the present invention is carried out according to chemical formula 2:

[0018]

[0019] Synthesis of compound D-1: Dissolve tricyclic ketone (200 mg, 0.76 mmol, 1 equivalent) in 15 ml of glacial acetic acid, add phenylhydrazine (288 mg, 2.66 mmol, 3.5 equivalents), monitor by TLC, and reflux for 18 to 20 hours. After the reaction is completed, cool to room temperature, pour the reaction solution into 100 ml of water, precipitate a large amount of solid, filter, and wash with water. After the filter cake is dried, it is chromatographed on a silica gel column and eluted with a dichloromethane / methanol system to obtain a yellow solid D-1.

[0020] The test data of the product obtained by the reaction are as follows: yield: 66%; yellow solid; 1 HNMR(400MHz,DMSO)δ12.43(s,1H),7.58(dd,J=9.3,2.3Hz,1H),7.44–7.35(m,1H),7.12(s,1H),6.43(s,1H) ,5.45–5.28(m,2H),4.99(s,2H),1.83(m,J=14.3,7.2Hz,2H),0.88(t,J=7.4Hz,3H); ESI-MSm / z:337.1[M+H] + .

[0021] Example 2: Synthesis of target compound D-2

[0022]

[0023] The experimental steps were the same as those in Example 1, except that 4-fluorophenylhydrazine hydrochloride was used instead of phenylhydrazine. The test data of the product obtained by the reaction were as follows: Yield: 58%; yellow solid; 1 HNMR(400MHz,DMSO)δ12.06(s,1H),7.50(td,J=8.7,3.5Hz,2H),7.09(td,J=9.2,2.7Hz,1H),6.89(s,1H) ,6.38(s,1H),5.40–5.26(m,2H),4.94(s,2H),1.79(m,2H),0.83(t,J=7.4Hz,3H); ESI-MSm / z:355.1[M+H] + .

[0024] Example 3: Synthesis of target compound D-3

[0025]

[0026] The experimental steps were the same as those in Example 1, except that 3-fluorophenylhydrazine hydrochloride was used instead of phenylhydrazine. The test data of the product obtained by the reaction were as follows: Yield: 36%; yellow solid; 1HNMR (400MHz, DMSO) δ12.11(s,1H),7.55(td,J=9.2,3.7Hz,2H),6.96(d,J=14.6Hz,1H),6.46(s,1H), 5.38(d,J=3.8Hz,2H),4.98(s,2H),1.87–1.83(m,2H),0.88(t,J=7.3Hz,3H); ESI-MSm / z:355.1[M+H] + .

[0027] Example 4: Synthesis of target compound D-4

[0028]

[0029] The experimental steps were the same as those in Example 1, except that 2-fluorophenylhydrazine hydrochloride was used instead of phenylhydrazine. The test data of the product obtained by the reaction were as follows: Yield: 55%; yellow solid; 1 HNMR(600MHz,DMSO)δ12.45(s,1H),7.60–7.53(m,1H),7.18–7.10(m,2H),6.99(s,1H),6.44(s ,1H),5.43–5.34(m,2H),5.08–4.98(m,2H),1.84(m,2H),0.90(t,3H); ESI-MSm / z:355.1[M+H] + .

[0030] Example 5: Synthesis of target compound D-5

[0031]

[0032] The experimental steps were the same as those in Example 1, except that 4-bromophenylhydrazine hydrochloride was used instead of phenylhydrazine. The test data of the product obtained by the reaction were as follows: Yield: 55%; orange solid; 1 HNMR (400MHz, DMSO) δ12.21(s,1H),7.98(d,J=2.0Hz,1H),7.53(d,J=8.8Hz,1H),7.39(dd,J=8.8,2.0Hz,1H),6.94(s,1H) ,6.44(s,1H),5.45–5.31(m,2H),4.99(s,2H),1.91–1.74(m,J=7.2Hz,2H),0.87(t,J=7.3Hz,3H); ESI-MSm / z:415.1[M+H] + .

[0033] Example 6: Synthesis of target compound D-6

[0034]

[0035] The experimental steps were the same as those in Example 1, except that 3-bromophenylhydrazine hydrochloride was used instead of phenylhydrazine. The test data of the product obtained by the reaction were as follows: Yield: 33%; brown solid; 1 HNMR(400MHz,DMSO)δ12.19(s,1H),7.70(s,1H),7.47(m,1H),7.27(m,1H),6.99(d,1H), 6.46(s,1H),5.39(d,2H),5.02(s,2H),1.87(m,2H),0.87(t,3H); ESI-MSm / z:415.1[M+H] + .

[0036] Example 7: Synthesis of target compound D-7

[0037]

[0038] The experimental steps were the same as those in Example 1, except that 2-bromophenylhydrazine hydrochloride was used instead of phenylhydrazine. The test data of the product obtained by the reaction were as follows: Yield: 56%; yellow solid; 1 HNMR (400MHz, DMSO) δ12.21(s,1H),7.75(d,J=8.0Hz,1H),7.50(d,J=7.6Hz,1H),7.18(s,1H),7.11(t,J=7.8Hz,1H),6 .43(s,1H),5.45–5.31(m,2H),5.02(s,2H),1.94–1.75(m,J=7.1Hz,2H),0.89(t,J=7.3Hz,3H); ESI-MSm / z:415.1[M+H] + .

[0039] Example 8: Synthesis of target compound D-8

[0040]

[0041] The experimental steps were the same as those in Example 1, except that 4-chlorophenylhydrazine hydrochloride was used instead of phenylhydrazine. The test data of the product obtained by the reaction were as follows: Yield: 45%; brown solid; 1 HNMR(600MHz,CD3OD)δ7.81(d,J=2.0Hz,1H),7.56(d,J=8.8Hz,1H),7.31(dt,J=8.8,1.7Hz,1H),7.09(s, 1H),5.60–5.53(m,1H),5.41–5.35(m,2H),5.09(s,2H),1.94(m,2H),1.03(t,3H); ESI-MSm / z:371.1[M+H] + .

[0042] Example 9: Synthesis of target compound D-9

[0043]

[0044] The experimental steps were the same as those in Example 1, except that 4-methylphenylhydrazine hydrochloride was used instead of phenylhydrazine. The test data of the product obtained by the reaction were as follows: Yield: 61%; yellow solid; 1 HNMR(600MHz,DMSO)δ11.85(s,1H),7.52–7.48(m,1H),7.43(d,J=8.4Hz,1H),7.11(dd,J=8.5,1.7Hz,1H),6.89(s,1H),6.41(s,1 H),5.40–5.32(m,2H),4.97(d,J=1.8Hz,2H),2.41(s,3H),1.83(m,J=21.4,7.1Hz,2H),0.88–0.85(t,3H); ESI-MSm / z:351.1[M+H] + .

[0045] Example 10: Synthesis of target compound D-10

[0046]

[0047] The experimental steps were the same as those in Example 1, except that 3-methylphenylhydrazine hydrochloride was used instead of phenylhydrazine. The test data of the product obtained by the reaction were as follows: Yield: 37%; yellow solid; 1 HNMR(400MHz,DMSO)δ11.78(s,1H),7.52(d,1H),7.28(s,1H),6.95–6.79(m,2H),6.37(s,1H) ),5.31(m,2H),4.89(d,2H),2.53(s,3H),1.79(m,2H),0.82(t,3H); ESI-MSm / z:351.1[M+H] + .

[0048] Example 11: Synthesis of target compound D-11

[0049]

[0050] The experimental steps were the same as those in Example 1, except that 4-methoxyphenylhydrazine hydrochloride was used instead of phenylhydrazine. The test data of the product obtained by the reaction were as follows: Yield: 45%; brown solid; 1HNMR(400MHz,DMSO)δ11.88(s,1H),7.44(s,1H),7.24(s,1H),6.92(m,2H),6.43(s,1H), 5.38(d,2H),4.97(s,2H),3.80(s,3H),1.84(m,2H),0.88(t,3H); ESI-MSm / z:367.1[M+H] + .

[0051] Example 12: Synthesis of target compound D-12

[0052]

[0053] The experimental steps were the same as those in Example 1, except that 4-trifluoromethoxyphenylhydrazine hydrochloride was used instead of phenylhydrazine. The test data of the product obtained by the reaction were as follows: Yield: 44%; white solid; 1 H NMR(600MHz,CD3OD)δ7.69(s,1H),7.58(d,1H),7.22(d,1H),7.06(s,1H),5.52(d ,1H),5.34(d,1H),5.07(s,2H),1.92(m,2H),0.98(t,3H); ESI-MSm / z:421.1[M+H] + .

[0054] Example 13: Synthesis of target compound D-13

[0055]

[0056] The experimental steps were the same as those in Example 1, except that 2,4-difluorophenylhydrazine hydrochloride was used instead of phenylhydrazine. The test data of the product obtained by the reaction were as follows: Yield: 47%; orange solid; 1 HNMR(600MHz,DMSO)δ12.57(s,1H),7.45(dd,J=9.2,2.3Hz,1H),7.22(m,1H),6.99(s,1H),6.45(s,1H), 5.38(d,J=6.6Hz,2H),5.00(t,J=3.3Hz,2H),1.83(m,2H),0.87(t,J=7.4Hz,3H); ESI-MSm / z:373.1[M+H] + .

[0057] Example 14: Synthesis of target compound D-14

[0058]

[0059] The experimental steps were the same as those in Example 1, except that 2-chloro-4-fluorophenylhydrazine hydrochloride was used instead of phenylhydrazine. The test data of the product obtained by the reaction were as follows: Yield: 52%; yellow solid; 1 HNMR(400MHz,DMSO)δ12.43(s,1H),7.58(dd,J=9.3,2.3Hz,1H),7.44–7.35(m,1H),7.12(s,1H),6.4 3(s,1H),5.45–5.28(m,2H),4.99(s,2H),1.83(m,2H),0.88(t,J=7.4Hz,3H); ESI-MSm / z:389.1[M+H] + .

[0060] Example 15: Synthesis of target compound D-15

[0061]

[0062] The experimental steps were the same as those in Example 1, except that 2,4-dichlorophenylhydrazine hydrochloride was used instead of phenylhydrazine. The test data of the product obtained by the reaction were as follows: Yield: 48%; yellow solid; 1 HNMR (400MHz, DMSO) δ12.55(s,1H),7.86(d,J=1.8Hz,1H),7.48(d,J=1.9Hz,1H),7.13(s,1H),6.44(s,1 H),5.39(d,J=4.2Hz,2H),5.01(s,2H),1.87–1.80(m,2H),0.88(t,J=7.2Hz,3H); ESI-MSm / z:405.1[M+H] + .

[0063] Example 16: Synthesis of target compound D-16

[0064]

[0065] The experimental steps were the same as those in Example 1, except that 2,5-dichlorophenylhydrazine hydrochloride was used instead of phenylhydrazine. The test data of the product obtained by the reaction were as follows: Yield: 38%; yellow solid; 1 HNMR(400MHz, CDCl3)7.71(d,1H),7.58(d,1H),7.06(s,1H),6.81(s,1H),5 .60(d,2H),5.13(s,2H),1.87(m,2H),0.88(t,3H); ESI-MSm / z:405.1[M+H] + .

[0066] Example 17: Synthesis of target compound D-17

[0067]

[0068] The experimental steps were the same as those in Example 1, except that 3-chloro-4-fluorophenylhydrazine hydrochloride was used instead of phenylhydrazine. The test data of the product obtained by the reaction were as follows: Yield: 31%; orange solid; 1 HNMR (400MHz, DMSO) δ12.24 (s, 1H), 7.75 (d, J = 8.9Hz, 1H), 7.37 (d, 1H), 6.99 (s, 1H), 6. 44(s,1H),5.39(m,2H),5.13(s,2H),1.84(m,2H),0.96(t,3H); ESI-MSm / z:389.1[M+H] + .

[0069] Example 18: Synthesis of target compound D-18

[0070]

[0071] The experimental steps were the same as those in Example 1, except that 3,4-dichlorophenylhydrazine hydrochloride was used instead of phenylhydrazine. The test data of the product obtained by the reaction were as follows: Yield: 36%; brown solid; 1 HNMR(400MHz,DMSO)δ12.31(s,1H),8.06(s,1H),7.88(s,1H),7.00(s,1H),6.44(s,1H), 5.46–5.35(m,2H),4.99(s,2H),1.87–1.80(m,2H),0.92(t,3H); ESI-MSm / z:405.1[M+H] + .

[0072] Example 19: Test method and results of antitumor activity of novel indole camptothecin derivatives

[0073] The standard MTT method was used for in vitro anti-tumor test. Irinotecan and topotecan were used as positive control drugs to test the inhibitory activity of the novel indole camptothecin derivatives on human hepatoma cells (HepG2), human non-small cell lung cancer cells (A549), human colon cancer cells (SW480), human intrahepatic cholangiocarcinoma cells (RBE), human breast cancer cells (MCF-7), human pancreatic cancer cells (BxPC3), human bladder cancer cells (T24), and human bladder cancer cells (umuc3). The compound was dissolved in DMSO to prepare a mother solution with a concentration of 10mM, and diluted to a suitable concentration with different culture media. The DMSO concentration in the dilution should be less than 0.01% (v / v) to reduce the toxicity of DMSO to cells and reduce the test error. Tumor cells of different cell lines were cultured in RPMI-1640 or DMEM high-glucose medium containing 10% fetal bovine serum (FBS), and various cancer cells growing logarithmically were collected, digested with trypsin / EDTA digestion solution and prepared into a suitable cell suspension. Add 100uL of cell suspension to a 96-well plate (usually 5000 cells per well) and place in a 37°C incubator containing 5% CO2 for 24 hours. Then add solutions of the compound to be tested at different concentrations. After 48 hours of culture, discard the old culture medium and wash the cells twice with PBS. Add 25uLMTT (5mg / mL) of fresh culture medium and continue to culture for 4 hours. Thereafter, discard the culture medium and add 200uLDMSO, shake on a shaker for 15 minutes, and wait for the formazan to completely dissolve. Finally, measure the absorbance at 490nm using an enzyme reader and calculate the IC 50 All experiments were performed in three parallel groups or repeated three times. The results of the cytotoxic activity test of the novel indole camptothecin derivatives are shown in Table 1.

[0074] Table 1 In vitro cytotoxic activity of novel indole camptothecin derivatives

[0075]

[0076]

[0077] Note: (1) Screening method: standard MTT colorimetric method; (2) Action time: 48 hours; (3) Compounds D-1 to D-18 are the products obtained in the aforementioned Examples 1 to 18.

[0078] The results of in vitro cytotoxic activity screening showed that the new indole camptothecin derivatives have a broad spectrum of anti-tumor activity, and showed strong inhibitory activity against human liver cancer cells (HepG2), human non-small cell lung cancer cells (A549), human colon cancer cells (SW480), human intrahepatic cholangiocarcinoma cells (RBE), human breast cancer cells (MCF-7), human pancreatic cancer cells (BxPC3), human bladder cancer cells (T24), and human bladder cancer cells (umuc3). Among them, the anti-tumor activity of compound D-18 is the most prominent, showing strong inhibitory effects on all 8 tumor cell lines tested, with IC 50 The range is 0.2585~10.69μM, which is significantly better than the control drug irinotecan and has the same activity as topotecan. Therefore, indole camptothecin derivatives are expected to be developed into a new type of anti-tumor drug.

Claims

1. A novel indole camptothecin derivative, such as compounds D-1 to D-18 shown in the formula.

2. A method for preparing a novel indole camptothecin derivative according to claim 1: By adopting the Fischer indole synthesis reaction, using differently substituted phenylhydrazine or phenylhydrazine hydrochloride as raw materials, heating and reflux with tricyclic ketone for 18-20 hours in an acidic environment, a series of novel indole camptothecin derivatives can be obtained.

3. Use of a novel indole camptothecin derivative according to claim 1 in the preparation of a drug for treating human liver cancer (HepG2).

4. Use of a novel indole camptothecin derivative according to claim 1 in the preparation of a drug for treating human non-small cell lung cancer (A549).

5. Use of a novel indole camptothecin derivative according to claim 1 in the preparation of a drug for treating human colon cancer (SW480).

6. Use of a novel indole camptothecin derivative according to claim 1 in the preparation of a drug for treating human intrahepatic cholangiocarcinoma cells (RBE).

7. Use of the novel indole camptothecin derivative according to claim 1 in the preparation of a drug for treating human breast cancer (MCF-7).

8. Use of the novel indole camptothecin derivative according to claim 1 in the preparation of a medicament for treating human pancreatic cancer (BxPC3).

9. Use of a novel indole camptothecin derivative according to claim 1 in the preparation of a drug for treating human bladder cancer (T24).

10. Use of a novel indole camptothecin derivative according to claim 1 in the preparation of a drug for treating human bladder cancer (UMUC3).

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