A synthetic paclitaxel-resistant cell line H460 TaxR Inhibitor Methods

Abstract

The present invention provides a synthetic paclitaxel-resistant cell line H460 TaxR The invention uses cheap and readily available 2-mercapto-3-aminopyridine, 2-fluorobenzaldehyde and 4-methylphenyl magnesium or lithium reagent as raw materials, 2,3-dichloro-5,6-dicyano-1,4-benzoquinone (DDQ) or manganese dioxide as an oxidative dehydrogenation reagent, and potassium peroxydisulfate complex salt #imgabs0# or persulfate as a sulfur atom oxidizing reagent to conveniently prepare paclitaxel-resistant cell line H460 TaxR Inhibitor 6-p-tolylbenzo[f]pyrido[2,3-b][1,4]thia#imgabs1#-11,11-dioxide. The synthesis method provided by the invention has simple steps, mild reaction conditions, simple operation, high synthesis efficiency, and has a positive effect in the fields of biology, medicine and fine chemicals.

Description

Technical Field

[0001] The invention belongs to the field of organic synthesis. 2-mercapto-3-aminopyridine, 4-bromophenol and 4-methylphenyl metal reagent are used as raw materials to prepare a paclitaxel-resistant cell line H460 with biological activity through a three-step reaction. TaxR Inhibitor 6-p-Tolylbenzo[f]pyrido[2,3-b][1,4]thia -11,11-dioxide. Background Art

[0002] Paclitaxel is a first-line broad-spectrum chemotherapy drug with significant efficacy. However, as the course of treatment increases, cancer cells inevitably develop clinical resistance to paclitaxel, which seriously affects the therapeutic effect (Hopper-Borge E.A.; Churchill, T.; Paulose, C. et al. Cancer Res. 2011, 71, 3649-3657). Therefore, it is urgent to develop new drugs that inhibit the anti-paclitaxel activity of cancer cells. 6-p-Tolylbenzo[f]pyrido[2,3-b][1,4]thia -11,11-dioxide (English system named 6-(p-tolyl)benzo[f]pyrido[2,3-b][1,4]thiazepine 11,11-dioxide) is a paclitaxel-resistant cell line H460 TaxR Inhibitors of EC 50<1.0μM), and has low toxicity to normal human cells (such as human fibroblasts) (Xiang, J.; Zhang, Z.; Mu, Y.; Xu, X.; Guo, S.; Liu, Y.; Russo, DP; Zhu, H.; Yan, B.; Bai, X. ACS Comb. Sci. 2016, 18, 230-235). Its unique drug activity allows it to be further applied in preclinical research and is expected to become a new clinical drug. In 2016, a literature first reported its total synthesis method (Xiang, J.; Zhang, Z.; Mu, Y.; Xu, X.; Guo, S.; Liu, Y.; Russo, DP; Zhu, H.; Yan, B.; Bai, X. ACS Comb. Sci. 2016, 18, 230-235). 2-Chloro-3-nitropyridine (Formula 7) was used as the raw material, and 2-phenylthio-3-aminopyridine (Formula 9) was prepared in a two-step yield of 43% through the aromatic nucleophilic substitution reaction of thiophenol and the iron reduction of the nitro group. Then, the corresponding 6-p-tolylbenzo[f]pyrido[2,3-b][1,4]thiapyridine was prepared through the condensation of 4-methylbenzoic acid and amino group mediated by polyphosphoric acid and phosphorus oxychloride and aromatic electrophilic cyclization. (Formula 6), and then oxidized with meta-chloroperbenzoic acid (m-CPBA) to successfully prepare the target product (Formula 1). The yield of the latter two steps is 25% in total, and the total yield of the entire route is 11%. The use of polyphosphoric acid-phosphorus oxychloride seriously limits the scale-up synthesis and potential industrial application of this method. The low synthesis efficiency has seriously affected the further research of the lead compound in the pharmaceutical chemistry community and has become a bottleneck restricting its development in the direction of clinical drugs.

[0003]

[0004] In view of the above problems, the present invention aims to provide a simple and efficient synthetic paclitaxel-resistant cell line H460 TaxR Inhibitor 6-p-Tolylbenzo[f]pyrido[2,3-b][1,4]thia The method uses 2-mercapto-3-aminopyridine and 2-fluorobenzaldehyde as the initial raw materials, and benzo[f]pyrido[2,3-b][1,4]thiadiazole is prepared by condensation. (first step); then oxidized by addition of 4-methylphenylmagnesium bromide or 4-methylphenyllithium, 2,3-dichloro-5,6-dicyano-1,4-benzoquinone (DDQ) or manganese dioxide (MnO2) (second step), and then by potassium bisulfate complex salt or persulfate oxidation (the third step) to prepare the biologically active 6-p-tolylbenzo[f]pyrido[2,3-b][1,4]thia The overall yield of this method is high and the reaction conditions are relatively mild. Summary of the invention

[0005] The technical solution of the present invention is as follows:

[0006] As shown in the following reaction formula, 2-mercapto-3-aminopyridine (Formula 1) and 2-fluorobenzaldehyde are used as raw materials to first generate pyridobenzothiophene (Formula 4); then react with 4-methylphenylmagnesium bromide, 4-methylphenylmagnesium chloride or 4-methylphenyllithium, and oxidative dehydrogenation with DDQ or MnO2 to prepare 11-(4-methylphenyl)pyridobenzothia (Formula 6). Finally, potassium bisulfate Or persulfate oxidation to prepare the target product (Formula 1).

[0007]

[0008] In the above reaction formula:

[0009] The amount of 2-fluorobenzaldehyde (Formula 2) used is 1 to 20 molar equivalents relative to 2-mercapto-3-aminopyridine (Formula 1).

[0010] Relative to pyridobenzothiazole (Formula 4), the amount of 4-methylphenylmagnesium bromide, 4-methylphenylmagnesium chloride or 4-methylphenyllithium (Formula 5) is 1 to 20 molar equivalents, and the amount of DDQ or MnO2 is 1 to 20 molar equivalents.

[0011] Relative to 11-(4-methylphenyl)pyridobenzothiazole (Formula 6), potassium persulfate complex salt Or the amount of persulfate used is 2 to 20 molar equivalents.

[0012] In the embodiment, the first step product is 40%, the second step yield is 87%, the third step yield is 99%, and the total yield is 34.5%.

[0013] In the second step, the addition and dehydrogenation reactions can be carried out in a "one-pot process" (without separation of the addition product), or the addition product can be separated and then dehydrogenated ("two-pot process").

[0014] In the first step reaction, the reaction temperature is 0 to 250°C, preferably 100 to 150°C.

[0015] In the second step reaction, the reaction temperature is -78 to 100°C, with the optimum temperature range being 0 to 40°C.

[0016] In the third step reaction, the reaction temperature is 0 to 100°C, preferably 60 to 100°C.

[0017] In the first step, the solvent is usually PEG 400 , ethylene glycol, 1,2-propylene glycol, 1,3-propylene glycol, and glycerol can also be used. The reaction can also occur by adding one or more of the above solvents at the same time.

[0018] In the second step reaction, anhydrous tetrahydrofuran is usually used as the solvent; the reaction can also occur using anhydrous toluene, anhydrous 1,4-dioxane or a mixture thereof with tetrahydrofuran.

[0019] In the second step, after the addition reaction of 4-methylphenylmagnesium bromide, 4-methylphenylmagnesium chloride or 4-methylphenyllithium is completed, water is added to quench the reaction, and then DDQ or MnO2 is added for dehydrogenation.

[0020] In the third step reaction, the oxidant is preferably It can also be potassium persulfate (K2S2O8), sodium persulfate (Na2S2O8), ammonium persulfate ((NH4)2S2O8), or a mixture of one or more thereof. The solvent is methanol, ethanol, isopropanol, n-propanol, n-butanol, or a mixture of two or more thereof.

[0021] Advantages and positive effects of the present invention:

[0022] The invention provides a convenient method for preparing paclitaxel-resistant cell line H460 TaxR The present invention will have a positive effect in the fields of biology, medicine and fine chemical industry.

[0023] Compared with the previous methods (Xiang, J.; Zhang, Z.; Mu, Y.; Xu, X.; Guo, S.; Liu, Y.; Russo, DP; Zhu, H.; Yan, B.; Bai, X. ACS Comb. Sci. 2016, 18, 230-235), the synthesis method provided by the present invention is simple to operate, has mild reaction conditions and high synthesis efficiency. BRIEF DESCRIPTION OF THE DRAWINGS

[0024] Figure 1 Benzo[f]pyrido[2,3-b][1,4]thia H NMR spectrum (400 MHz, CDCl3).

[0025] Figure 2 Benzo[f]pyrido[2,3-b][1,4]thia C NMR spectrum (100 MHz, CDCl3).

[0026] Figure 3 Benzo[f]pyrido[2,3-b][1,4]thia High-resolution mass spectrometry.

[0027] Figure 4 6-p-Tolylbenzo[f]pyrido[2,3-b][1,4]thia H NMR spectrum (400 MHz, CDCl3).

[0028] Figure 5 6-p-Tolylbenzo[f]pyrido[2,3-b][1,4]thia C NMR spectrum (100 MHz, CDCl3).

[0029] Figure 6 6-p-Tolylbenzo[f]pyrido[2,3-b][1,4]thia High-resolution mass spectrometry.

[0030] Figure 7 6-p-Tolylbenzo[f]pyrido[2,3-b][1,4]thia -11,11-dioxide hydrogen nuclear magnetic resonance spectrum (400MHz,CDCl3).

[0031] Figure 8 6-p-Tolylbenzo[f]pyrido[2,3-b][1,4]thia -11,11-dioxide carbon NMR spectrum (100 MHz, CDCl3). DETAILED DESCRIPTION

[0032] The present invention is further illustrated by the following examples, but the present invention is not limited to the scope of the examples.

[0033] first step:

[0034] 2-Mercapto-3-aminopyridine (5mmol, 631mg), 2-fluorobenzaldehyde (5mmol, 621mg), potassium carbonate (5mmol, 0.690g) were added to 20mL PEG400 solvent and heated at 100°C for 6h. After cooling to room temperature, 150mL water was added. The mixture was extracted with ethyl acetate, washed twice with saturated brine, and the organic phase was dried over anhydrous sodium sulfate. After the solvent was removed by rotary evaporation, a mixed solution of petroleum ether (abbreviated as PE) and ethyl acetate (abbreviated as EA) was used for column chromatography to obtain the product shown in Formula 4.

[0035] Benzo[f]pyrido[2,3-b][1,4]thia

[0036] Light yellow solid, 424 mg, yield 40%, mp 128–130°C, R f =0.20(PE / EA=5:1,v / v,2%ofEt3N). 1 H NMR (400MHz, CDCl3, δ): 8.92 (s, 1H), 8.34 (dd, J=4.6, 1.7Hz, 1H), 7.60 (dd, J=7.9, 1.7Hz, 1H ),7.52(d,J=7.4Hz,1H),7.47–7.40(m,2H),7.39–7.34(m,1H),7.26(dd,J=7.9,4.7Hz,1H).

[0037] 13 C{ 1 H}NMR (101MHz, CDCl3, δ): 163.6, 148.0, 147.5, 144.6, 137.3, 136.6, 134.8, 132.7, 132.2, 129.5, 128.5, 124.0.

[0038] HRMS(ESI)m / z:[M+H] + Calculate for C 12 H9N2S + ,213.0481;found,213.0488.

[0039] Step 2:

[0040] In a 25 mL round bottom flask equipped with a magnetic rod, add benzo[f]pyrido[2,3-b][1,4]thia 9 (0.65mmol), the flask was sealed with a rubber stopper, and after nitrogen replacement three times, 4mL of dry tetrahydrofuran was added. The flask was placed in an ice-water bath, and the pre-prepared tetrahydrofuran solution of 4-methylphenylmagnesium bromide (1.5equiv., 1.6mL, 0.6M in THF) was slowly added to the flask with a syringe. After the addition was completed, the reaction was carried out at room temperature for 3 hours. After TLC detection, after the substrate was completely consumed, a small amount of water was added to quench the reaction. The stopper was removed, and DDQ (0.48mol, 109mg) was added under open conditions, and the reaction was carried out at 100°C for 10 minutes. Cool to room temperature, add 10 ml of 5% sodium hydroxide solution, extract three times with ethyl acetate, and dry the organic phase with anhydrous sodium sulfate. After rotary evaporation to remove the solvent, a mixed solution of petroleum ether (abbreviated as PE) and ethyl acetate (abbreviated as EA) was used for column chromatography to obtain the product shown in formula 6

[0041] 6-p-Tolylbenzo[f]pyrido[2,3-b][1,4]thia

[0042] The shape of the compound and the following data are consistent with those reported in the literature (Xiang, J.; Zhang, Z.; Mu, Y.; Xu, X.; Guo, S.; Liu, Y.;

[0043] Russo, DP; Zhu, H.; Yan, B.; Bai, X. ACS Comb. Sci. 2016, 18, 230-235).

[0044] Yellow solid, 106 mg, yield 87%, mp 156–158 °C, R f =0.30 (PE / EA=5:1, v / v, 4% of Et3N).

[0045] 1 H NMR (400MHz, CDCl3, δ): 8.26 (dd, J=4.6, 1.7Hz, 1H), 7.69 (d, J=8.2Hz, 2H), 7.67 (dd, J=8.0, 0.8Hz, 1H), 7.61 (dd, J= 8.0,1.7Hz,1H),7.44(td,J=7.6,1.6Hz,1H),7.29(td,J=7.5,1.2Hz,1H),7.23(dd,J=8.0,4.6Hz,4H),2.41(s,3H).

[0046] 13 C{ 1 H}NMR (101MHz, CDCl3, δ): 169.6, 147.6, 146.3, 144.9, 141.6, 138.9, 137.2, 137.0, 133.1, 131.5, 130.5, 129.9, 129.1, 127.9, 123.9, 21.6.

[0047] HRMS(ESI)m / z:[M+H] + Calculate for C 19 H 15 N2S + ,303.0950;found,303.0956.

[0048] Step 3:

[0049] 6-p-Tolylbenzo[f]pyrido[2,3-b][1,4]thia (0.28mmol, 86mg) and ethanol (5mL) were added to a dry round-bottom flask in sequence. Solid powder was added to the flask at 0°C. (Potassium persulfate complex salt) oxidant (515 mg, 0.84 mmol), reflux for 5 hours. After the reaction is completed, ethanol is removed by rotary evaporation, ethyl acetate (10 mL) is added, and the mixture is washed with water three times to remove inorganic salts. The organic phase is dried over anhydrous sodium sulfate, and the solvent is removed by rotary evaporation. Then, a mixed solution of petroleum ether (abbreviated as PE) and ethyl acetate (abbreviated as EA) is used for column chromatography separation to obtain the product shown in Formula 1.

[0050] 6-p-Tolylbenzo[f]pyrido[2,3-b][1,4]thia -11,11-dioxide

[0051] The shape of the compound and the following data are consistent with those reported in the literature (Xiang, J.; Zhang, Z.; Mu, Y.; Xu, X.; Guo, S.; Liu, Y.;

[0052] Russo, DP; Zhu, H.; Yan, B.; Bai, X. ACS Comb. Sci. 2016, 18, 230-235).

[0053] Yellow solid, 94 mg, yield 99%, mp 229–231 °C, R f =0.25 (PE / EA=2:1, v / v, 4% of Et3N).

[0054] 1 H NMR (400 MHz, CDCl3, δ): 8.53 (dd, J = 4.5, 1.5 Hz, 1H), 8.26 (dd, J = 7.8, 1.4 Hz, 1H), 7.90 (dd, J = 8.2, 1.5 Hz, 1H), 7.75 (d, J = 8.2 Hz, 2H), 7.72 (d, J = 1.4 Hz,1H),7.68(td,J=7.5,1.5 Hz,1H),7.56(dd,J=8.2,4.4 Hz,1H),7.51(dd,J=7.7,1.3 Hz,1H),7.28(d,J=8.2 Hz,2H),2.44(s,3H).

[0055] 13 C{ 1 H}NMR(101 MHz, CDCl3, δ): 168.6, 147.9, 146.3, 143.6, 142.6, 141.5, 136.8, 135.8, 133.2, 131.8, 131.1, 130.3, 129.4, 129.3, 128.1, 125.8, 21.7.

Claims

1. A synthetic paclitaxel-resistant cell line H460 TaxR The method of producing an inhibitor, characterized in that The method comprises the following three steps: the first step is to use 2-mercapto-3-aminopyridine (Formula 2) and 2-fluorobenzaldehyde (Formula 3) as raw materials to prepare an intermediate benzo[f]pyrido[2,3-b][1,4]thia (Formula 4), the second step, reaction with 4-methylphenyl metal reagent (Formula 5) and subsequent oxidative dehydrogenation of 2,3-dichloro-5,6-dicyano-1,4-benzoquinone or manganese dioxide to prepare 6-p-tolylbenzo[f]pyrido[2,3-b][1,4]thia (Formula 6); Step 3, using potassium peroxybisulfate complex salt or persulfate to oxidize sulfur atoms to prepare paclitaxel-resistant cell line H460 TaxR Inhibitor 6-p-Tolylbenzo[f]pyrido[2,3-b][1,4]thia -11,11-dioxide (Formula 1) 2. The method according to claim 1, characterized in that 2-mercapto-3-aminopyridine (Formula 2) and 2-fluorobenzaldehyde (Formula 3) are used as raw materials.

3. The method according to claim 1, characterized in that The 4-methylphenyl metal reagent (Formula 5) is 4-methylphenyl magnesium chloride, 4-methylphenyl magnesium bromide or 4-methylphenyl lithium.

4. The method according to claim 1, characterized in that: 2,3-dichloro-5,6-dicyano-1,4-benzoquinone or manganese dioxide is used as the oxidative dehydrogenation reagent in the second step, and potassium peroxydisulfate complex salt or persulfate is used as the oxidant in the third step.

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