A cyclized berberine derivative and its preparation method and application

Abstract

The present invention provides a cyclized berberine derivative and a preparation method and application thereof. The cyclized berberine derivative exhibits excellent anticancer activity and can be used as an anticancer drug. The present invention discloses a preparation method thereof.

Description

Technical Field

[0001] The invention relates to a cyclized berberine derivative and application thereof in pharmacy, belonging to the technical field of medicine. Background Art

[0002] With the aging of the population, cancer has become a major cause of death in humans. As a classic cancer treatment, chemotherapy has the effects of reducing preoperative tumor volume, improving the patient's postoperative quality of life, reducing the chance of tumor recurrence, and prolonging patient survival. However, the efficacy of drugs often weakens as the duration of use increases, and even leads to drug resistance. In addition, the heterogeneity of tumor cells and the high toxic side effects of small molecule drugs greatly limit the clinical application of chemotherapy. Therefore, it is very necessary to develop new anti-cancer drugs with low toxicity.

[0003] Berberine is a natural isoquinoline alkaloid, which has been proven to have anticancer activity and low toxicity, and is selective for cancer cells and normal cells. Its combination with chemotherapy or radiotherapy drugs can reduce drug toxicity and enhance drug efficacy, but its poor fat solubility and water solubility and low oral bioavailability greatly limit its clinical application. The present invention uses berberine as a lead compound, introduces a group with antitumor activity into its molecular structure and cyclizes it, in order to improve the anticancer activity of berberine. Summary of the invention

[0004] The object of the present invention is to provide a cyclized berberine derivative having anticancer effect.

[0005] Another object of the present invention is to provide a method for preparing the cyclized berberine derivative.

[0006] Another object of the present invention is to provide the anti-cancer use of the cyclized berberine derivatives.

[0007] The present invention is described in detail below.

[0008] The cyclized berberine derivative provided by the present invention has the following general formula:

[0009]

[0010] In the formula, R is independently selected from , , , , , .

[0011] The cyclized berberine derivative has a representative example structure as follows:

[0012]

[0013] The preparation method of the cyclized berberine derivative is as follows:

[0014]

[0015] Where R is independently , , , , , .

[0016] The application of the cyclized berberine derivative in the preparation of anticancer drugs.

[0017] The present invention is further illustrated by the following examples, but it should be noted that the scope of the present invention is not limited by these examples. Implementation Example

[0018] Preparation of compound (1)

[0019] Berberine (1 g, 2.7 mmol) was dissolved in 25 mL of dry DMF, and 2 zeolites were added. The mixture was refluxed in a microwave (microwave power: 400 W) for 15 min. After the reaction was completed, 40 mL of water was added, and the mixture was cooled for crystallization, filtered, and dried to obtain the intermediate (I) with a yield of 89%. 1 H NMR (CD3OD, 400 MHz) δ: 9.22 (s, 1H), 7.96 (s, 1H), 7.48 (d,J = 8.2 Hz, 1H), 7.39 (s, 1H), 6.83 (d, J = 8.2 Hz, 1H), 6.81 (s, 1H), 6.02(s, 2H), 4.57 (t, J = 6.3 Hz, 2H), 3.86 (s, 3H), 3.10 (t, J = 6.3 Hz, 2H); ESI-MS (m / z): 322 [M-Cl]+.

[0020] The intermediate (I) (357 mg, 1.0 mmol) was dissolved in 10 mL of dry CH2Cl2, and Et3N (111 mg, 1.1 mmol) was added. After stirring, Me3SiCl (119 mg, 1.1 mmol) was slowly added dropwise. The reaction was continued for 6 h. The product was washed with water, dried over anhydrous Na2SO4, filtered, and evaporated to dryness under reduced pressure to obtain the intermediate (II), which was used directly without purification.

[0021] The intermediate (II) (430 mg, 1.0 mmol) was dissolved in 10 mL of 20% KOH solution, heated to reflux for 8 h, cooled to room temperature, extracted with ethyl acetate, dried over anhydrous Na2SO4, filtered, and concentrated under reduced pressure to obtain a crude product. The crude product was purified by silica gel column chromatography (CHCl3 / MeOH = 50:1) to obtain intermediate (III) with a yield of 59%. 1 H NMR (DMSO-d6, 300 MHz) δ:7.42 (s, 1H), 7.30 (d, J = 8.7 Hz, 1H), 7.26 (d, J = 8.7 Hz, 1H), 7.19 (s,1H), 6.70 (s, 1H), 5.98 (s, 2H), 4.27 (t, J = 6.0 Hz, 2H), 3.99 (s, 3H), 2.89 (t, J = 6.0 Hz, 2H), 0.28 (s, 9H); ESI-MS (m / z ) 409 [M] + .

[0022] Add 10 mL of POCl3 to intermediate (III) (409 mg, 1.0 mmol) and heat under reflux for 2 h. Cool to room temperature, filter, and wash the filter cake with CH2Cl2 to obtain intermediate (IV), which is used directly without purification.

[0023] Add intermediate (IV) (392 mg, 1.0 mmol) to 15 mL of acetonitrile, stir, pass ammonia, react at room temperature for 2 days, evaporate to dryness under reduced pressure to obtain intermediate (V). Add 10 mL of ethyl acetate and N , N '-Dicyclohexylcarboximide (DCC, 618 mg, 3.0 mmol) was added to the intermediate (V), stirred, cooled to 0°C, and chlorambucil (319 mg, 1.05 mmol) was added. The reaction was continued for 24 hours, filtered, and the filtrate was washed with dilute sodium bicarbonate and dilute citric acid solution in turn, dried over anhydrous Na2SO4, filtered, and concentrated under reduced pressure to obtain a crude product. The crude product was purified by silica gel column chromatography (CHCl3 / MeOH = 50:1) to obtain compound (1) with a yield of 81.7%. 1HNMR (DMSO-d6, 300 MHz) δ: 7.72 (s, 1H), 7.08 (d, J = 8.6 Hz, 2H), 6.82 (s,1H), 6.71 (d, J = 8.7 Hz, 1H), 6.64 (d, J = 8.6 Hz, 2H), 6.62 (d, J = 8.7 Hz,1H), 6.59 (s, 1H), 5.90 (s, 2H), 3.87 (m, 4H), 3.74 (s, 3H), 3.60 (m, 4H), 3.52 (m, 2H), 2.85 (m, 2H), 2.58 (t, J = 7.5 Hz, 2H), 2.37 (t, J = 7.4 Hz,2H), 1.95-1.88 (m, 2H); ESI-MS (m / z): 604 [M-Cl] + . Example

[0024] Preparation of compound (2)

[0025] use N -Boc-melphalan (425 mg, 1.05 mmol) was used to replace chlorambucil (319 mg, 1.05 mmol) to react with intermediate (IV). After the reaction was completed, trifluoroacetic acid was used to remove the Boc group at room temperature. The other operations were the same as those in Example 1 to obtain compound (2) with a yield of 78.1%. 1 H NMR (DMSO-d6, 300 MHz) δ: 7.71 (s, 1H), 7.08 (d, J =8.5 Hz, 2H), 7.01(s, 1H), 6.71 (d, J = 8.7 Hz, 1H), 6.64 (d, J = 8.6 Hz, 1H), 6.63 (d, J = 8.5 Hz, 2H), 6.61 (s, 1H), 5.93 (s, 2H), 3.88 (m, 1H), 3.74 (s,3H), 3.72-3.60 (m, 8H), 3.53 (m, 2H), 3.00 (dd, J = 13.7, 5.1 Hz, 1H), 2.86(m, 2H), 2.78 (dd, J = 13.7, 7.8 Hz, 1H), 2.66 (s, 2H); ESI-MS (m / z ) : 605[M-Cl] + . Example

[0026] Preparation of compound (3)

[0027] Chlorambucil (319 mg, 1.05 mmol) was replaced with methyl nitrogen (349 mg, 1.05 mmol). Other operations were the same as in Example 1 to obtain compound (3) with a yield of 83.3%. 1 H NMR (DMSO-d6, 300 MHz) δ: 9.51 (s, 1H),7.77 (s, 1H), 7.08 (d, J = 8.4 Hz, 2H), 7.01 (s, 1H), 6.72 (d, J = 8.7 Hz,1H), 6.65 (d, J = 8.6 Hz, 1H), 6.63 (d, J = 8.5 Hz, 2H), 6.60 (s, 1H), 5.93(s, 2H), 3.89 (m, 1H), 3.73 (s, 3H), 3.71-3.60 (m, 8H), 3.53 (m, 2H), 3.00(dd, J = 13.7, 5.1 Hz, 1H), 2.86 (m, 2H), 2.78 (dd, J = 13.7, 7.8 Hz, 1H), 2.69 (s, 1H); ESI-MS (m / z ): 633 [M-Cl] + . Example

[0028] Preparation of compound (4)

[0029] Compound (2) (642 mg, 1.0 mmol) was dissolved in 10 mL of acetonitrile, 1,2-dichloroethane (118.2 mg, 1.2 mmol) and K2CO3 (165.6 mg, 1.2 mmol) were added, and the mixture was refluxed for 3 h. The mixture was cooled to room temperature and filtered. The filtrate was evaporated to dryness under reduced pressure and purified by silica gel column chromatography (CHCl3 / MeOH = 50:1) to obtain compound (4) with a yield of 76.1%. 1H NMR (DMSO-d6, 300 MHz) δ: 7.77 (s, 1H), 7.01(s, 1H), 6.96 (d, J = 8.6 Hz, 2H), 6.74 (d,J = 8.7 Hz, 1H), 6.66 (d, J = 8.6 Hz, 1H), 6.63 (d, J = 8.7 Hz, 2H), 6.60 (s,1H), 5.93 (s, 2H), 3.89 (m, 1H), 3.73 (s, 3H), 3.63-3.60 (m, 8H), 3.53 (m,2H), 3.00 (dd, J = 13.7, 5.1 Hz, 1H), 2.86 (m, 2H), 2.76 (dd, J = 13.7, 7.8Hz, 1H), 2.69 (m, 4H); ESI-MS (m / z): 631 [M-Cl] + . Example

[0030] Preparation of compound (5)

[0031] use N , N -Bis(2-chloroethyl)phenylalanine (304.5 mg, 1.05 mmol) was used to replace chlorambucil (319 mg, 1.05 mmol). Other operations were carried out in the same manner as in Example 1 to obtain compound (5) with a yield of 74.6%. 1 H NMR (DMSO-d6, 300MHz) δ: 7.71 (s, 1H), 7.21-7.08 (m, 5H), 7.02 (s, 1H), 6.80 (d, J = 8.7 Hz, 1H), 6.73 (d, J = 8.6 Hz, 1H), 6.63 (s, 1H), 5.91 (s, 2H), 3.86 (m, 1H), 3.73(s, 3H), 3.56 (m, 2H), 3.48 (m, 4H), 3.01 (dd, J = 13.7, 5.1 Hz, 1H), 2.86(m, 2H), 2.76 (dd, J = 13.7, 7.7 Hz, 1H), 2.64 (m, 4H); ESI-MS (m / z ): 590[M-Cl] + . Example

[0032] Preparation of compound (6)

[0033] useN , N -Bis(2-chloroethyl)glycine (304.5 mg, 1.05 mmol) was used to replace chlorambucil (319 mg, 1.05 mmol). Other operations were the same as in Example 1 to obtain compound (6) with a yield of 75.5%. 1 H NMR (DMSO-d6, 300MHz) δ: 7.74 (s, 1H), 7.00 (s, 1H), 6.80 (d, J = 8.7 Hz, 1H), 6.74 (d, J =8.6 Hz, 1H), 6.61 (s, 1H), 5.93 (s, 2H), 3.73 (s, 3H), 3.53 (m, 2H), 3.48 (m,4H), 3.30 (s, 2H), 2.86 (m, 2H), 2.64 (m, 4H); ESI-MS (m / z ): 500 [M-Cl] + . Example

[0034] Antitumor activity of cyclized berberine derivatives

[0035] The experiment selected human chronic myeloid leukemia cells (K562), human lung cancer cells (A549), and human liver cancer cells (HepG2) in logarithmic growth phase, and the concentration of these cells was adjusted to 5×10 5 / mL, 100μL per well was inoculated in a 96-well culture plate. After culturing in a 37℃, 5% CO2 incubator for 24h, 10μmol / L of chlorambucil, melphalan, berberine and cyclized berberine derivatives were added respectively, and the blank control group was added with the same amount of culture medium. The culture plate was moved into a CO2 incubator and cultured for 48 hours at 37℃, 5% CO2 and saturated humidity. 20μL MTT solution (5 g / L) was added to each well of the 96-well culture plate, and the culture plate was moved into a CO2 incubator. The culture was continued for 4 hours at 37℃, 5% CO2 and saturated humidity. The culture was terminated and the culture supernatant in the well was carefully aspirated and discarded. For cells growing in suspension, the culture medium in the well should be discarded after centrifugation. 100μL dimethyl sulfoxide was added to each well and shaken for 10 min to fully dissolve the purple-blue crystals. The absorbance A value of each group was measured at a wavelength of 570 nm using an ELISA reader. IC was calculated using the Mosmann method. 50 Values ​​(Table 1).

[0036]

Claims

1. A cyclized berberine derivative, characterized in that: The structural formula is as follows:

2. A berberine derivative according to claim 1, characterized in that The preparation method is as follows: In the formula, R is independently selected from 3. Use of a cyclized berberine derivative according to claim 1 in the preparation of anti-tumor drugs.

Images