A silybin derivative with anti-prostate cancer activity and its preparation method

Abstract

A silybin derivative with anti-prostate cancer activity and a preparation method thereof, relates to an anti-cancer silybin derivative and a preparation method thereof, the invention transforms and modifies the chemical structure of the natural product silybin, takes silybin as a lead compound, first uses propionic anhydride to esterify five hydroxyl groups, recrystallizes to obtain an intermediate 3,5,20,23-tetrapropionic acid silybin ester, and further reacts the intermediate with acyl chloride, halogenated hydrocarbon and sulfonyl chloride to synthesize 12 ester, ether and sulfonate silybin derivatives I1 to I12. The results of the MTT experiment show that the synthesized silybin derivatives exhibit inhibitory activity against human prostate cancer cells PC-3 and DU145, and the inhibitory activity is significantly higher than that of the parent silybin, among which the inhibitory activity of the derivative I3 is the best, which is better than the positive control drug Flutamide. Studies have shown that such compounds can enhance anti-tumor activity.

Description

Technical Field

[0001] The present invention relates to an anticancer derivative and a preparation method thereof, in particular to a silibinin derivative with anti-prostate cancer activity and a preparation method thereof. Background Art

[0002] Silybin is derived from the mature seeds of the plant milk thistle. It is the main active ingredient of silymarin and is composed of two diastereoisomers, silybin A and silybin B, in equal proportions. Molecular formula: C 25 H 22 O 10 , molecular weight: 482.436. Melting point: 164-174℃, density: 1.527g / cm 3 . Silybin is a pure off-white crystalline powder, odorless, slightly bitter, and hygroscopic. It is easily soluble in acetone, ethyl acetate, methanol, and ethanol, slightly soluble in chloroform, and almost insoluble in water. Silybin has pharmacological activities such as scavenging free radicals, resisting lipid peroxidation, protecting liver cell membranes, promoting the synthesis of DNA and structural proteins in damaged liver cells, and resisting fibrosis. It has good therapeutic effects on acute and chronic hepatitis, metabolic toxic liver damage, and cirrhosis.

[0003] The chemical structure of silybin is:

[0004] Summary of the invention

[0005] The purpose of the present invention is to provide a silibinin derivative with anti-prostate cancer activity and a preparation method thereof. The present invention uses silibinin as a lead compound, and the synthesized silibinin derivatives show inhibitory activity against human prostate cancer cells PC-3 and DU145, and the inhibitory activity is significantly higher than that of the parent silibinin, among which the inhibitory activity of derivative I3 is the best, which is better than the positive control drug Flutamide.

[0006] The objective of the present invention is achieved through the following technical solutions:

[0007] A silibinin derivative with anti-prostate cancer activity, wherein the derivative is obtained by structural modification of the C-7 hydroxyl group to obtain a series of silibinin derivatives I1 to I 12 , the silybin chemical modification is as follows:

[0008]

[0009] The group R introduced at the C-7 position is shown in the following table:

[0010]

[0011] A method for preparing a silibinin derivative having anti-prostate cancer activity, the method comprising the following preparation process:

[0012] (1) Silybin is used as a reaction raw material, dissolved in pyridine, propionic anhydride is added dropwise at room temperature, and DMAP is added as a catalyst to react to generate a compound 3,5,7,20,23-pentapropionic acid silybin ester;

[0013] (2) dissolving 3,5,7,20,23-pentapropionic acid silybin ester in pyridine under ice bath conditions, adding 3-aminopropanol, and then adding glacial acetic acid after 30 seconds to obtain 3,5,20,23-tetrapropionic acid silybin ester;

[0014] (3) The hydroxyl group at the C-7 position of the compound 3,5,20,23-tetrapropionic acid silybin ester was modified to form esters with different acyl chlorides, ethers with halogenated hydrocarbons, and sulfonates with sulfonyl chlorides to obtain a series of silybin derivatives I1~I 12 .

[0015] The advantages and effects of the present invention are:

[0016] The present invention uses silybin as a lead compound, firstly uses propionic anhydride to esterify five hydroxyl groups, and recrystallizes to obtain 3,5,7,20,23-pentapropionic acid silybin ester, and then uses 3-aminopropanol reagent to selectively aminolyze the C-7 ester group into a hydroxyl group, and recrystallizes to obtain an intermediate 3,5,20,23-tetrapropionic acid silybin ester, and further reacts the intermediate with acyl chloride, halogenated hydrocarbon and sulfonyl chloride to synthesize 12 ester, ether and sulfonate ester silybin derivatives I1 to I2. 12 The results of MTT experiments showed that the synthesized silibinin derivatives exhibited inhibitory activity against human prostate cancer cells PC-3 and DU145, and the inhibitory activity was significantly higher than that of the parent silibinin. Among them, the inhibitory activity of derivative I3 was the best, which was better than the positive control drug Flutamide. DETAILED DESCRIPTION

[0017] The present invention is described in detail below with reference to the embodiments.

[0018] 1. After pre-defatting the silybum seed meal with n-hexane, use anhydrous ethanol as the extraction solvent, heat and dissolve, decolorize with activated carbon, let the filtrate stand to precipitate white crystals, and recrystallize from ethyl acetate-methanol to obtain silybin.

[0019] 2. Dissolve silybin in pyridine, mix and stir at room temperature, add propionic anhydride and DMAP, and continue to react the mixed organic matter at room temperature for 5 minutes. TLC is used to detect the end point of the reaction. Pour ice water into the reaction mixture, recrystallize, filter, wash with petroleum ether, and dry to obtain 3,5,7,20,23-pentapropionic acid silybin ester.

[0020]

[0021] 3. The intermediate 3,5,7,20,23-pentapropionic acid silybin ester was dissolved in pyridine, and 3-aminopropanol was quickly added under ice bath conditions. After 28 seconds of reaction, glacial acetic acid was quickly added to quench the reaction. Ice water was added to the reaction mixture, and the mixture was recrystallized, filtered, washed with petroleum ether, and dried to obtain 3,5,20,23-tetrapropionic acid silybin ester.

[0022]

[0023] 4. The hydroxyl group at the C-7 position of the compound 3,5,20,23-tetrapropionic acid silybin ester was modified to form esters with different acyl chlorides, ethers with halogenated hydrocarbons, and sulfonates with sulfonyl chlorides to obtain a series of silybin derivatives I1~I 12 .

[0024]

[0025] Wherein R is isobutyryl, cyclopropylformyl, dimethylaminoformyl, pivaloyl, acetyl, chloroacetyl, p-cyanobenzoyl, benzenesulfonyl, methylsulfonyl, benzoyl, (3-methyl-4-nitro)benzoyl, or benzyl.

[0026]

[0027] Flutamide is an anti-tumor drug for the treatment of prostate cancer. It is mainly suitable for patients with advanced prostate cancer who have not been treated before or who are ineffective or failed to respond to hormone control therapy. Flutamide was used as a positive control and the MTT colorimetric method was used to conduct a preliminary in vitro anti-tumor activity test on the synthesized compound. The in vitro experimental results of the compound are as follows:

[0028]

[0029] Note: a. Inhibition rate measured when the compound concentration is 10 μM. b. IC 50 It indicates the half effective inhibitory concentration.

[0030] The synthetic silibinin derivatives were tested for biological activity using the MTT assay with human prostate cancer cells PC-3 and DU145 as target cells. The results showed that the synthetic silibinin derivatives exhibited certain inhibitory activity against human prostate cancer cells PC-3 and DU145, and the inhibitory activity was significantly higher than that of the parent silibinin. Among them, derivative I3 had the best inhibitory activity against human prostate cancer cells PC-3 and DU145, and the inhibition rate was better than that of the positive control Flutamide.

[0031] Here are some examples:

[0032] Example 1

[0033] Synthesis of 3,5,7,20,23-pentapropionic acid silybin ester:

[0034] Silybin (2g, 4mmol) was dissolved in 10mL Py, mixed and stirred at room temperature, propionic anhydride (4mL) and DMAP (7mg) were added, and the mixed organic matter was continued to react for 5min at room temperature, and the reaction end point was detected by TLC (chloroform: acetone = 10:1). After the reaction was completed, 150mL of ice water was poured into the reaction mixture, and a white solid was recrystallized and precipitated, which was filtered, washed with petroleum ether, and dried to obtain a white powder solid 3,5,7,20,23-pentapropionic acid silybin ester, with a yield of 95% and mp 123-128°C.

[0035] Example 2

[0036] Synthesis of 3,5,20,23-tetrapropionic acid silybin ester:

[0037] The intermediate 3,5,7,20,23-pentapropionic acid silybin ester (0.7620g, 1mmol) was dissolved in 5mL Py, and 3-aminopropanol (230μL) was quickly added under ice bath conditions. After 28s of reaction, glacial acetic acid (230μL) was quickly added to quench the reaction. The reaction end point was detected by TLC (chloroform: acetone = 10:1). 250mL of ice water was added to the reaction mixture, and a white solid was recrystallized and precipitated. The solid was filtered, washed with petroleum ether 2 to 3 times, and dried to obtain 0.7006g of light yellow powder solid 3,5,20,23-tetrapropionic acid silybin ester, with a yield of 99.2% and mp 135-140°C. 1HNMR(400MHz,DMSO-d6)δ11.10(s,1H),7.29–6.21(m,8H),5.74(dd,J=12.1,6.4Hz,1 H),5.51(d,J=12.0Hz,1H),5.10(t,J=8.04Hz,1H),4.66–4.54(m,1H),4.16(dd,J=12.4 ,3.2Hz,1H),4.00(dd,J=12.4,5.0Hz,1H),3.78(s,3H),2.63–2.54(m,4H),2.34–2.18( m,4H),1.14(td,J=7.5,4.5Hz,6H),1.00(t,J=7.5Hz,3H),0.90(td,J=7.5,1.1Hz,3H).

[0038] Example 3

[0039] Compound I1: Synthesis of 7-isobutyryloxy-3,5,20,23-tetrapropionic acid silybin ester:

[0040] 3,5,20,23-tetrapropionic acid silybin ester (0.7065 g, 1 mmol) was dissolved in 4 mL THF, isobutyryl chloride (114 μL, 1.1 eq) and triethylamine (556 μL, 4 eq) were added, and the reaction was stirred continuously at room temperature for 5 min. The reaction endpoint was detected by TLC (chloroform: acetone = 10:1). After the reaction, the triethylamine hydrochloride precipitate was filtered, the filtrate was collected, the solvent was dried under reduced pressure, 2 mL of methanol was added at 60 ° C, and then 5 mL of dilute hydrochloric acid with a volume fraction of 5% was added, a large amount of white solid was precipitated by recrystallization, filtered, and dried to obtain 0.6309 g of light yellow powdery solid derivative I1: 7-isobutyryloxy-3,5,20,23-tetrapropionic acid silybin ester, with a yield of 81.24%, mp99-105 ° C. 1H NMR (400 MHz, DMSO-d6) δ7.27-6.76 (m, 8H), 5.96 (dd, J = 12.3, 6.3 Hz, 1H), 5.69 (d, J = 12.3 Hz, 1 H),5.11(t,J=7.8Hz,1H),4.67–4.56(m,1H),4.17(dd,J=12.4,3.2Hz,1H),4.00(dd,J=12.4,5.0Hz,1H),3.78(s,3H),2.87-2.76(m,1H), 2.66–2.55(m,4H),2.35–2.17(m,4H),1.25–1.19(m,6H),1.15(td,J=7.5,4.4Hz,6H),1.00(t,J=7.5Hz,3H),0.92(td,J=7.6,1.2Hz,3H).

[0041] Example 4

[0042] Compound I2: Synthesis of 7-cyclopropylformyloxy-3,5,20,23-tetrapropionic acid silybin ester:

[0043] According to the synthesis method of derivative I1, 3,5,20,23-tetrapropionic acid silybin ester was reacted with cyclopropylcarbonyl chloride (100 μL, 1.1 eq) and filtered to obtain a crude product. The crude product was purified by recrystallization, and the reaction was subjected to a series of post-treatments to obtain 0.7200 g of light yellow powder solid derivative I2: 7-cyclopropylcarbonyloxy-3,5,20,23-tetrapropionic acid silybin ester, with a yield of 93.90%, mp 123-128 ° C. 1HNMR (400 MHz, DMSO-d6) δ 7.29–6.73 (m, 8H), 5.95 (dd, J = 12.3, 6.2 Hz, 1H), 5.67 (d, J = 12.3 Hz, 1H), 5.11 (t, J = 7.6 Hz, 1H), 4.67–4. 56(m,1H),4.16(dd,J=12.3,3.1Hz,1H),3.99(dd,J=12.4,5.0Hz,1H),3.78(s,3H),2.67–2.54(m,4H),2.35–2.14(m,4H ),1.94–1.85(m,1H),1.14(td,J=7.5,3.0Hz,6H),1.11–1.02(m,4H),1.00(t,J=7.5Hz,3H),0.91(td,J=7.5,1.0Hz,3H).

[0044] Example 5

[0045] Compound I3: Synthesis of 7-dimethylaminoformyloxy-3,5,20,23-tetrapropionic acid silybin ester:

[0046] According to the synthetic method of derivative I1, 3,5,20,23-tetrapropionic acid silybin ester was reacted with dimethylaminoformyl chloride (119 μL, 1.3 eq) and filtered to obtain a crude product. The crude product was purified by recrystallization, and the reaction was subjected to a series of post-treatments to obtain 0.5826 g of light yellow powdered solid derivative I3 with a yield of 75%, mp114-117°C. 1HNMR (400 MHz, DMSO-d6) δ7.26–6.78 (m, 8H), 5.96 (dd, J=12.32, 6.32 Hz, 1H), 5.69 (d, J=12.2 Hz, 1H), 5.11 (t, J=7.92 Hz, 1H), 4.67–4.58 (m, 1H), 4.17 (dd, J=12.5 6,3.36Hz,1H),4.00(dd,J=12.44,5.04Hz,1H),3.78(s,3H),3.08(t,J=7.52Hz,3H),3.01(t,J=7.56Hz,3H),2.65-2 .56(m,4H),2.35–2.16(m,4H),1.15(td,J=7.4,4.36Hz,6H),1.00(t,J=7.52Hz,3H),0.92(td,J=7.52,0.96Hz,3H).

Claims

1. A method for preparing a silibinin derivative having anti-prostate cancer activity, characterized in that: The method comprises the following preparation process: Synthesis of 3,5,7,20,23-pentapropionic acid silybin ester: 2 g, 4 mmol of silybin was dissolved in 10 mL of pyridine, mixed and stirred at room temperature, 4 mL of propionic anhydride and 7 mg of DMAP were added, and the mixed organic matter was reacted for 5 min at room temperature, and the reaction end point was detected by TLC; wherein, the detection solvent was a mixed solvent of chloroform: acetone = 10:1; after the reaction, 150 mL of ice water was poured into the reaction mixture, and a white solid was recrystallized and precipitated, which was filtered, washed with petroleum ether, and dried to obtain a white powdery solid 3,5,7,20,23-pentapropionic acid silybin ester with a yield of 95% and mp123-128°C; Synthesis of 3,5,20,23-tetrapropionic acid silybin ester: 0.7620 g, 1 mmol of the intermediate 3,5,7,20,23-pentapropionic acid silybin ester was dissolved in 5 mL of pyridine, and 230 µL of 3-aminopropanol was quickly added under ice bath conditions. After 28 s of reaction, 230 µL of glacial acetic acid was quickly added to quench the reaction, and the reaction endpoint was detected by TLC; wherein, the detection solvent was a mixed solvent of chloroform: acetone = 10:1; 250 mL of ice water was added to the reaction mixture, and a white solid was recrystallized and precipitated, which was filtered, washed with petroleum ether 2-3 times, and dried to obtain 0.7006 g of light yellow powdery solid 3,5,20,23-tetrapropionic acid silybin ester, with a yield of 99.2% and mp 135-140°C; Compound I3: Synthesis of 7-dimethylaminoformyloxy-3,5,20,23-tetrapropionic acid silybin ester: 119 µL, 1.3 eq of 3,5,20,23-tetrapropionic acid silybin ester was reacted with dimethylcarbamoyl chloride and filtered to obtain a crude product; the crude product was purified by recrystallization, and the reaction was subjected to a series of post-treatments to obtain 0.5826 g of light yellow powdery solid derivative I3 with a yield of 75%.

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