CN105175277B - A kind of inhibitor of 3 GAPD and its preparation method and application - Google Patents

Abstract

The invention provides an inhibitor of 3-phosphate glyceraldehyde dehydrogenase, a preparation method thereof and an anticancer application. Pharmacological experiments of the present invention show that the compound has strong killing effect on human colon cancer cells, human gastric cancer cells and leukemia cells, and significantly induces cancer cell apoptosis. Further experiments showed that these compounds inhibited the production of ATP by inhibiting glyceraldehyde-3-phosphate dehydrogenase (GAPDH) in the glycolysis of cancer cells, thereby killing cancer cells.

Description

A kind of inhibitor of 3-phosphate glyceraldehyde dehydrogenase and its preparation method and application

technical field

The present invention relates to an inhibitor, more specifically, to an inhibitor of 3-phosphate glyceraldehyde dehydrogenase and its preparation method and application.

Background technique

Cancer has replaced cardiovascular disease as the disease with the largest number of deaths in the world. In recent decades, cancer drug treatment has made significant progress, and some anti-tumor drugs have been developed, which effectively prolong the survival of patients or improve the patient's quality of life. However, the drug resistance of tumor cells makes the drug treatment of cancer often less effective. The unlimited malignant proliferation ability and drug resistance of tumor cells depend on tumor stem cells with special biological characteristics in tumor tissues, and its existence has been confirmed in various tumors such as breast cancer. In most tumors, even under aerobic conditions, the activity of glycolysis is higher than that of normal cells, referred to as the Warburg effect (Science, 1956, 124, 269–270). Glyceraldehyde-3-phosphate dehydrogenase (GAPDH) is one of the key catalytic enzymes in the glycolysis process, which can catalyze the conversion of the substrate glyceraldehyde-3-phosphate to 1,3-bisphosphoglyceric acid. Therefore, inhibiting the activity of this enzyme can theoretically cut off the energy supply of tumor cells, thereby inhibiting or even killing tumor cells. The functions of GAPDH are diverse. It not only participates in cell energy metabolism, but also participates in various activities of cells, such as membrane fusion and transport, DNA replication and repair, etc., and GAPDH is used as a target to inhibit the sugar production of tumor cells. Metabolism, thereby cutting off its energy supply, is rare. Our team found in previous studies that GAPDH was overexpressed in colon cancer patient samples, and 3-bromo-2-oxopropionic acid could efficiently inhibit the activity of GAPDH (J. Bioenerg. Biomembr. 2012, 44, 117–125). The latest research of our team further reveals that compounds such as 3-bromo-2-oxopropionic acid, as glycolysis inhibitors, can effectively kill tumor cell side population cells with tumor stem cell properties, thereby improving the anti-tumor efficacy (Cell Death Differ .2014, 21, 124–135). These research results have extremely far-reaching impact on the development of anticancer drugs.

Contents of the invention

The purpose of the present invention is to provide a novel anticancer drug. In order to achieve this purpose, the present invention firstly provides an inhibitor of glyceraldehyde-3-phosphate dehydrogenase. The structural formula of the inhibitor is as follows:

The R ₂ is H or Br, the M is O or N,

The R ₁ is C1-5 alkane, benzyl, benzyl, phenethyl, benzhydryl, adamantyl or cyclohexane.

The R ₂ is preferably Br.

Said M is preferably O.

When said M is preferably N, said R ₁ is preferably two C1-5 alkanes.

The structure of the inhibitor is preferably,

Furthermore, a method for preparing the above-mentioned inhibitor of glyceraldehyde-3-phosphate dehydrogenase is provided, using the following reaction formula,

The protonic acid is concentrated sulfuric acid or perchloric acid; the solvent is tetrahydrofuran or dichloromethane; the solid particle carrier is magnesium sulfate, sodium sulfate, aluminum oxide, zeolite molecular sieve or silicon dioxide.

According to requirements, the application of the above-mentioned 3-phosphate glyceraldehyde dehydrogenase inhibitor in the preparation of anticancer drugs is provided.

The advantage of the present invention is that,

1. The obtained compound has good anticancer activity.

2. The present invention uses solid particles to support acid protons to catalyze different alcohols and 3-bromo-2-oxopropionic acid to prepare 3-bromo-2-oxopropionate.

Description of drawings

Figure 1 is the inhibition curve of compound 4 on gastric cancer cell HGC27.

Figure 2 is the inhibition curves of compounds 1, 3 and 4 on glyceraldehyde-3-phosphate dehydrogenase (GAPDH). It can be seen from the figure that all three compounds have obvious inhibitory effects.

Detailed ways

The present invention will be further described in detail below in conjunction with the accompanying drawings and specific embodiments. Unless otherwise specified, the reagents, equipment and methods used in the present invention are commercially available reagents, equipment and routinely used methods in this technical field.

Embodiment 1 Compound 1, the synthesis of phenylethyl ester of 3-bromo-2 oxopropionate

Add concentrated sulfuric acid (87μL, 1.64mmol) to the turbid solution of magnesium sulfate (788mg, 6.55mmol) in anhydrous dichloromethane (4mL), stir at room temperature for half an hour, then add 3-bromopyruvate (410mg, 2.46mmol) and phenylethyl alcohol (0.2 mL, 1.64 mmol). The reaction was carried out at room temperature, and the progress of the reaction was monitored by TLC and 1H-NMR. After the reaction, the reaction solution was diluted with dichloromethane, filtered under reduced pressure with a funnel covered with diatomaceous earth, and the filtrate was washed with saturated brine to remove excess 3-bromopyruvate, then dried with anhydrous sodium sulfate, and finally The solvent was evaporated under reduced pressure and dried in vacuo to obtain 1 as an orange liquid with a yield of 95%. ¹ H NMR (400MHz, CDCl ₃ ): δ7.36-7.26(m, 5H), 4.55(t, J=7.2Hz, 2H), 4.30(s, 2H), 3.10(t, J=6.8Hz, 2H ); ¹³ C NMR (100MHz, CDCl ₃ ): δ184.4, 159.2, 136.6, 128.9, 128.7, 127.0, 122.6, 67.4, 34.8, 30.7. IR (KBr, cm-1): υ 1741, 1457, 1240, 1056; LC-MALD-TOF for C ₁₁ H ₁₁ BrO ₃ : 271.073.

Embodiment 2, compound 2, the synthesis of 3-bromo-2 oxopropionic acid-1-phenylethyl-ester

Using the same conditions for the synthesis of phenylethyl 3-bromo-2-oxopropionate 1, 2 was successfully prepared as a green liquid with a yield of 93%. ¹ HNMR (400MHz, CDCl ₃ ): δ7.41-7.32 (m, 5H), 6.03 (q, J = 6.4Hz, 1H), 4.30 (s, 2H), 1.68 (d, J = 6.8Hz, 3H) ; ¹³ C NMR (100MHz, CDCl ₃ ): δ184.6, 158.7, 139.8, 128.7, 128.6, 127.0, 126.3, 76.0, 30.8, 21.9; IR (KBr, cm-1): υ1733, 1390, 1274, 1163, 1053.

Embodiment 3, the synthesis of compound 3,3-bromo-2 oxopropionic acid 3-pentyl ester

Using the same conditions for the synthesis of phenylethyl 3-bromo-2-oxopropionate 1, 3 was successfully prepared as a yellow liquid with a yield of 72%. ¹ H NMR (400MHz, CDCl ₃ ): δ4.97-4.91 (m, 1H), 4.29 (s, 2H), 1.73-1.66 (m, 4H), 0.954-0.910 (m, 6H); ¹³ C NMR ( 100MHz, CDCl ₃ ): δ185.1, 159.5, 80.9, 30.7, 26.3, 9.5; IR (KBr, cm-1): υ1736, 1399, 1265, 671.

Embodiment 4, the synthesis of compound 4,3-bromo-2 oxopropionate cyclohexyl

Using the same conditions for the synthesis of phenylethyl 3-bromo-2-oxopropionate 1, 4 was successfully prepared as a yellow liquid with a yield of 81%. ¹ H NMR (400MHz, CDCl ₃ )δ5.17-4.80(m,1H),4.30(s,1H),1.94-1.86(m,2H),1.80-1.72(m,2H),1.64-1.51(m ,4H),1.46-1.34(m,2H), ¹³ CNMR(100MHz,CDCl ₃ ):δ185.1,158.8,76.4,31.2,30.9,25.1,23.5; IR(KBr,cm-1):υ2939,2861,1737 _, 1263, 1050; LC-MALD-TOF _{for C9H13BrO3} : 249.978.

Embodiment 5, compound 5, the synthesis of adamantyl 3-bromo-2 oxopropionate

Using the same conditions for the synthesis of phenylethyl 3-bromo-2-oxopropionate 1, 5 was successfully prepared as a yellow liquid with a yield of 98%. ¹ H NMR (400MHz, CDCl ₃ ): δ4.27(s, 2H), 2.20 (s, 9H), 1.70-1.63 (m, 6H); ¹³ C NMR (100MHz, CDCl ₃ ): δ185.7, 158.1, 85.5 , 45.2, 41.0, 35.9, 31.0; IR (KBr, cm-1): υ2915, 2857, 1724, 1455, 1278, 1162, 1047; LC-MALD-TOF for C ₁₃ H ₁₇ BrO ₃ : 301.151.

Embodiment 6, the synthesis of compound 6,3-bromo-N,N-diethyl-2-oxopropionamide

Under an ice-water bath, add HOBt (1.02g, 7.52mmol), EDCI (1.96g, 10.25mmol) and triethylamine (1.91mL, 13.67mmol) to a solution of pyruvic acid (662mg, 7.52mmol) in dichloromethane (25mL). ), after stirring for half an hour, diethylamine (0.704 mL, 6.84 mmol) was added dropwise. Then, the reaction solution was stirred at room temperature for 18 hours. When the reaction was terminated, ethyl acetate (50 mL) was added for dilution and water (6 mL). After the two phases were separated, the aqueous phase was extracted with ethyl acetate (5 mL×3). The organic phases were combined, washed with saturated brine (5mL×1), dried over anhydrous sodium sulfate, and separated by silica gel column chromatography (petroleum ether/ethyl acetate 6/1-4/1) to obtain an intermediate, light yellow Oil (410 mg, 42% yield). This intermediate (300mg, 2.1mmol) was dissolved in dichloromethane (5mL), bromine (0.13mL, 2.51mmol) in dichloromethane (1mL) was slowly added dropwise, and stirred at room temperature for 7h. When the reaction was terminated, add ethyl acetate (30mL) for dilution, water (5mL), after separation, the aqueous phase was extracted with ethyl acetate (5mL×3), the organic phases were combined, and the organic phase was washed with saturated brine (5mL×1 ), dried over anhydrous sodium sulfate, and separated by silica gel column chromatography (petroleum ether/ethyl acetate 15/1-10/1) to obtain 6 (122 mg) as a yellow liquid with a yield of 27%. ¹ HNMR (400MHz, CDCl ₃ ) δ4.26(s, 2H), 3.45(q, J=7.1Hz, 2H), 3.36(q, J=7.1Hz, 2H), 1.25(t, J=7.1Hz, 3H), 1.19 (t, J=7.1Hz, 3H); ¹³ C NMR (100MHz, CDCl ₃ ) δ190.1, 164.3, 42.6, 40.0, 32.0, 14.5, 12.6.

Embodiment 7, the synthesis of compound 7,3-bromo-N-phenethyl 2-oxygen-phenylpropanamide

7 was successfully prepared using the same conditions as for the synthesis of 3-bromo-N-ethyl-2-oxo-4-phenylbutyramide 6.

¹ H NMR (400MHz, CDCl ₃ ) δ7.33(t, J=7.3Hz, 2H), 7.24(d, J=7.5Hz, 1H), 7.19(d, J=7.3Hz, 2H), 6.95(s ,1H),4.49(s,2H),3.60(t,J=7.1Hz,2H),2.87(t,J=7.1Hz,2H).

Embodiment 8, the synthesis of compound 8,3-bromo-2-oxo-4-phenylbutyric acid methyl ester

Acetyl chloride (0.4 mL, 5.61 mmol) was slowly added dropwise to anhydrous methanol solution (8 mL) under ice-cooling, and stirred at room temperature for 1 hour to prepare anhydrous hydrogen chloride methanol solution. The newly prepared anhydrous hydrogen chloride methanol solution was slowly added dropwise to a solution of 2-oxo-4-phenylbutyric acid (500mg, 2.81mmol) in methanol (2mL), heated to reflux for 3.5 hours, the reaction solution was concentrated to dryness, and then Dissolve in dichloromethane (5mL), add water (0.2mL) and trifluoroacetic acid (2mL) successively, stir at room temperature for 2.5 hours, then concentrate and dry the reaction solution to obtain the intermediate 2-oxyl-4-phenylbutyric acid Methyl ester as pale yellow oil (540 mg, 100%). This intermediate 2-oxyl-4-phenylbutyric acid methyl ester (150 mg, 0.78 mmol) was dissolved in chloroform (1 mL), and bromine (1.72 mmol, 2.2 equiv) in chloroform (1 mL) was slowly added dropwise, Stir at room temperature for 24 hours. When the reaction was terminated, ethyl acetate (30 mL) was added for dilution, water (5 mL), and after separation, the aqueous phase was extracted with ethyl acetate (3 mL×3), the organic phases were combined, and the organic phase was washed with saturated brine (3 mL). It was dried over anhydrous sodium sulfate and separated by silica gel column chromatography (petroleum ether/ethyl acetate 12/1-8/1) to obtain 8 (97 mg) as a light yellow liquid with a yield of 36%. ¹ H NMR (400MHz, CDCl ₃ )δ7.34-7.28(m,3H),7.26-7.23(m,2H),5.30-5.25(m,1H),3.90(s,3H),3.57-3.51(m ,1H),3.29-3.23(m,1H); ¹³ CNMR(100MHz,CDCl ₃ )δ185.3,160.7,136.6,129.5,128.9,127.6,53.6,47.5,38.4; IR(KBr,cm-1):υ2955, 1735, 1443, 1261, 1027.

Embodiment 9, the synthesis of compound 9,3,3-dibromo-N,N-diethyl-2-oxopropionamide

Using the reaction conditions for the synthesis of 3-bromo-N-ethyl-2-oxo-4-phenylbutyramide 6, compound 9 was also obtained as a yellow oil with an isolated yield of 25%. ¹ H NMR (400MHz, CDCl ₃ ) δ6.90(s, 1H), 3.48(q, J=7.2Hz2H), 3.40(q, J=7.2Hz, 2H), 1.30(t, J=7.2Hz, 3H ), 1.21 (t, J=7.2, 3H); ¹³ C NMR (100MHz, CDCl ₃ ) δ182.0, 162.4, 42.9, 42.3, 40.3, 14.4, 12.4; IR (KBr, cm-1): υ1735, 1643, 1397 , 1048; LC-Mass for C ₇ H ₁₁ Br ₂ NO ₂ [M+1]: 301.1.

Embodiment 10, the synthesis of compound 10,3,3,-dibromo-2 oxopropionic acid isopropyl ester

Using the same conditions for the synthesis of phenylethyl 3-bromo-2-oxopropionate 1, 11 was successfully prepared as a yellow liquid with a yield of 60%. ¹ HNMR (400MHz, CDCl ₃ ): δ6.64 (s, 1H), 5.35-5.08 (m, 1H), 1.36-1.34 (d, J＝1.6Hz, 6H). ¹³ C NMR (100MHz, CDCl ₃ ) δ178.50, 157.53, 72.58, 38.74, 21.48; IR (KBr, cm-1): υ1651, 1399, 1150, 672.

Embodiment 11, the synthesis of compound 11,3,3,-tert-butyl dibromo-2 oxopropionate

Add concentrated sulfuric acid (43μL, 0.81mmol) to the turbid solution of magnesium sulfate (489.6mg, 4.07mmol) in anhydrous dichloromethane (4mL), stir at room temperature for half an hour, then add 3-bromopyruvate (200mg, 0.81mmol ) and tert-butanol (0.3 mL, 3.25 mmol). The reaction was carried out at room temperature, and the progress of the reaction was monitored by TLC. After the reaction, the reaction solution was diluted with dichloromethane, filtered with a funnel covered with diatomaceous earth under reduced pressure, and the filtrate was washed with saturated brine to remove excess 3,3-dibromopyruvate, and then washed with anhydrous sodium sulfate After drying, the solvent was distilled off under reduced pressure and dried in vacuo to obtain compound 11 as a yellow liquid with a reaction yield of 42.3%. ¹ HNMR (400MHz, CDCl ₃ ): δ6.64(s, 1H), 1.58(s, 9H).

Embodiment 12, the synthesis of compound 12,3,3,-cyclohexyl dibromo-2 oxopropionate

Compound 12 was successfully prepared under the same conditions as for the synthesis of tert-butyl 3,3,-dibromo-2-oxopropionate as a yellow liquid with a reaction yield of 61.8%. ¹ H NMR (400MHz, CDCl ₃ ): δ7.40-7.33 (m, 10H), 7.00 (s, 1H), 4.33 (s, 2H). ¹ H NMR (400MHz, CDCl ₃ )δ6.65(s,1H),4.16(s,1H),1.99-1.86(m,3H),1.84-1.72(m,3H),1.48-1.27(m,4H ).

Embodiment 13, the synthesis of compound 13,3,3,-dibromo-2-oxopropionic acid 2-butyl ester

Compound 13 was successfully prepared under the same conditions as for the synthesis of tert-butyl 3,3,-dibromo-2-oxopropionate as a yellow liquid with a reaction yield of 61.8%. ¹ H NMR (400MHz, CDCl ₃ ) δ6.66(s, 1H), 5.07(dd, J=12.6, 6.3Hz, 1H), 1.80-1.70(m, 2H), 1.36-1.34(d, J=6.3 Hz,3H),0.97-0.94(t,J＝9.2Hz,3H).

Embodiment 14, the synthesis of compound 14,3,3,-dibromo-2 oxopropionic acid diphenylmethyl ester

Compound 14 was successfully prepared under the same conditions as for the synthesis of tert-butyl 3,3,-dibromo-2-oxopropionate as a white solid with a reaction yield of 40%. ¹ HNMR (400MHz, CDCl ₃ )δ7.56- 7.21(m,10H),6.96(s,1H),5.95(s,1H).

Example 15, compound 15, 3,3,-dibromo-2 oxopropionic acid-1-phenylethyl ester synthesis

Compound 15 was successfully prepared under the same conditions as for the synthesis of tert-butyl 3,3,-dibromo-2-oxopropionate as a white solid with a reaction yield of 56%. ¹ H NMR (400MHz, CDCl ₃ ) δ 7.38-7.24 (m, 4H), 6.57 (s, 1H), 6.03-5.95 (m, 1H), 1.63 (d, J=6.6Hz, 3H).

Embodiment 16, the synthesis of compound 16, 3,3,-dibromo-2 oxopropionate adamantyl ester

Compound 16 was successfully prepared under the same conditions as for the synthesis of tert-butyl 3,3,-dibromo-2-oxopropionate as a white solid with a reaction yield of 52%. ¹ H NMR (400MHz, CDCl ₃ )δ5.85(s,1H),2.23-2.22(m,3H),2.18-2.17(m,5H),1.69-1.68(m,5H),1.55(s,2H ).

Embodiment 17 Effect

Through the cell experiment of 3 kinds of cancer cells, the statistics of the drug effect are as follows.

The above table proves that the compound provided by the present invention has a very good effect. This type of compound has a strong killing effect on human colon cancer cells, human gastric cancer cells and leukemia cells, and significantly induces cancer cell apoptosis.

Claims (2)

1. A kind of 1. application of inhibitor of glyceraldehyde 3-phosphate dehydro-genase in cancer therapy drug is prepared, it is characterised in that the 3- The inhibitor of GAPD is the application in resisting human gastric cancer medicine is prepared, and the structural formula of described inhibitor is as follows It is shown：
2. 2. a kind of inhibitor of glyceraldehyde 3-phosphate dehydro-genase, it is characterised in that the structural formula of described inhibitor is as follows：