鄭玉國, 薛 偉, 熊 壯, 何 勇, 楊 濤, 盧 平
(1. 貴州大學(xué) 精細(xì)化工研究開發(fā)中心 教育部綠色農(nóng)藥與農(nóng)業(yè)生物工程重點(diǎn)實(shí)驗(yàn)室,貴州 貴陽 550025; 2. 興義民族師范學(xué)院 化學(xué)生物系,貴州 興義 562400)
雙酰胺類化合物具有抗癌活性[1~6];雜環(huán)化合物也成為新藥研究的重要對(duì)象,其中苯并噻唑類化合物具有抗真菌、抗結(jié)核、抗癌等藥理活性[7~15], 在醫(yī)藥研究領(lǐng)域引起廣泛關(guān)注。
本文采用活性拼接原理,以2,4-二氟苯甲酸(1a)或2-氟苯甲酸(1f)為起始原料,將苯并噻唑基引入雙酰胺結(jié)構(gòu)中,設(shè)計(jì)并合成了9個(gè)新型的N-[2-(取代苯并噻唑-2-氨基甲?;?苯基]取代氟基苯甲酰胺(6a~6i, Scheme 1),其結(jié)構(gòu)經(jīng)1H NMR,13C NMR, IR和元素分析表征。
采用MTT法對(duì)6a~6i進(jìn)行抑制PC3癌細(xì)胞體外活性測(cè)試, 結(jié)果表明6a~6i具有不同程度的抑制PC3癌細(xì)胞活性,其中10 μmol·L-16b對(duì)PC3的抑制率為69.9%。
X-5型熔點(diǎn)儀(溫度未校正);JEOL-ECX 500 NMR型核磁共振儀(DMSO-d6為溶劑,TMS為內(nèi)標(biāo));IR Prestige-21型紅外光譜儀(KBr壓片);Elementar Vario-Ⅲ型元素分析儀。
所用試劑均為分析純。
(1) 2-(2,4-二氟苯甲酰胺)苯甲酸(3a)和2-(2-氟苯甲酰胺)苯甲酸(3f)的合成
在單口圓底燒瓶中加入2,4-二氟苯甲酸(1a) 3 mmol和二氯亞砜6 mL,攪拌下回流反應(yīng)12 h。減壓蒸除過量的二氯亞砜制得2,4-二氟苯甲酰氯(2a)。
在三口圓底燒瓶中加入鄰氨基苯甲酸3 mmol和二氯甲烷40 mL,攪拌下滴加三乙胺2 mL,滴畢,冰浴冷卻下緩慢滴加2a的二氯甲烷(15 mL)溶液,滴畢,于室溫反應(yīng)至終點(diǎn)[TLC跟蹤,展開劑A:V(石油醚) ∶V(乙酸乙酯)=1 ∶1]。加飽和氯化鈉溶液40 mL,用乙酸乙酯(2×40 mL)萃取,合并萃取液,依次用稀鹽酸、飽和碳酸氫鈉溶液和飽和氯化鈉溶液洗滌,無水Na2SO4干燥,減壓脫溶得3a粗品,烘干,不純化直接用于下一步反應(yīng)。以2-氟苯甲酸(1f)代替1a,用類似方法合成3f。
(2) 2-(2,4-二氟苯基)-4(3H)-3,1-取代苯并噁嗪酮(4a)和2-(2-氟苯基)-4(3H)-3,1-取代苯并噁嗪酮(4f)的合成
在裝有分餾器(200 mm)和蒸餾裝置(裝無水氯化鈣干燥管)的反應(yīng)瓶中加入3a100 mmol,醋酐100 mL,控制回流速度(約每10秒1滴)反應(yīng)至終點(diǎn)(TLC跟蹤,展開劑A)。冷卻至室溫,抽濾,濾餅干燥得白色晶體4a,不純化直接用于下一步反應(yīng)。用類似方法合成4f。
(3)6的合成(以6a為例)
在反應(yīng)瓶中依次加入4a5 mmol,乙腈40 mL和K2CO30.7 g(5 mmol),攪拌下回流反應(yīng)5 min;慢慢加入6-甲氧基苯并噻唑胺(5a) 5 mmol的乙腈(15 mL)溶液,加畢,回流反應(yīng)反應(yīng)至終點(diǎn)(TLC跟蹤,展開劑A)。傾入冷水中靜置析晶,抽濾,濾餅經(jīng)柱層析[洗脫劑:V(石油醚) ∶V(乙酸乙酯)=2 ∶1]純化得6a。
用類似方法合成6b~6i。合成6的實(shí)驗(yàn)結(jié)果見表1,表征數(shù)據(jù)見表2。
表 1 合成6的實(shí)驗(yàn)結(jié)果Table 1 Experimental results of synthesizing 6
表 2 6的表征數(shù)據(jù)Table 2 Characteristic data of 6
續(xù)表2
Comp1H NMR δ(J/Hz)13C NMR δIR ν/cm-16e12.96(s, 1H, NH), 11.09(s, 1H, NH), 7.74(s, 2H, ArH), 7.39(s, 3H, ArH), 7.28(s, 2H, ArH), 7.05(d, J=8.0, 2H, ArH), 6.93(d, J=8.0, 2H, ArH)173.0, 169.4, 152.6, 132.3, 130,7, 130.2, 125.9, 124.43 479, 3 331, 1 680, 1 652, 1 614, 1 585, 1 550, 1 523, 1 496, 1 446, 1 280, 1 251, 1 2246f12.89(s, 1H, NH), 11.15(s, 1H, NH), 8.32(s, 1H, ArH), 8.01(s, 1H, ArH), 7.88(d, J=7.5, 1H, ArH), 7.80(d, J=7.5, 1H, ArH), 7.65(d, J=6.9, 3H, ArH), 7.45~7.36(m, 2H, ArH), 7.32~7.27(m, 2H, ArH), 2.42(s, 3H, CH3)162.4, 160.9, 158.9, 134.3, 134.2, 133.8, 133.3, 131.2, 130.2, 128.2, 125.5, 124.4, 123.6, 123.5, 123.4, 122.1, 117.2, 116.9, 21.53 360, 3 232, 3 064, 3 298, 1 653, 1 604, 1 585, 1 521, 1 452, 1 313, 1 292, 1 271, 921, 812, 7526g13.00(s, 1H, NH),11.16(s, 1H, NH), 8.29(s, 1H, ArH), 8.00(d, J=8.0, 1H, ArH), 7.88(t, J=8.0, 1H, ArH), 7.75(s, 1H, ArH), 7.66~7.63(m, 2H, ArH), 7.48~7.33(m, 6H, ArH)162.4, 160.9, 158.9, 134.3, 134.2, 133.4, 131.2, 126.9, 125.5, 124.3, 117.2, 117.03 213, 3 188, 1 661, 1 653, 1 541, 1 496, 1 462, 1 442, 1 404, 1 340, 1 305, 1 271, 1 224, 1 097, 1 066, 914, 900, 7466h13.09(s, 1H, NH), 11.18(s, 1H, NH), 8.23(s, 1H, ArH), 7.93(dd, J=2.9, 2H, ArH), 7.87(t, J=8.0, 1H, ArH), 7.78(s, 1H, ArH), 7.66~7.65(m, 2H, ArH), 7.44~7.37(m, 2H, ArH), 7.35~7.30(m, 2H, ArH)162.4, 134.3, 134.2, 133.4, 131.2, 130.1, 125.4, 124.5, 123.6, 123.5, 117.1, 116.9, 114.9, 114.83 182, 3 064, 1 664, 1 606, 1 585, 1 550, 1 527, 1 456, 1 317, 1 290, 1 193, 1 074, 908, 852, 7546i14.41(s, 1H, NH), 13.10(s, 1H, NH), 8.64(d, J=8.1, 1H, ArH), 8.22~8.21(m, 2H, ArH), 7.84(s, 1H, ArH), 7.63~7.62(m, 1H, ArH), 7.53~7.36(m, 4H, ArH), 7.27(d, J=8.6, 1H, ArH), 7.17(t, J=8.1, 1H, ArH)170.7, 162.6, 161.4, 159.4, 151.9, 149.5, 133.6, 131.0, 130.9, 125.4, 125.3, 123.1, 120.9, 120.6, 117.3, 117.13 082, 3 030, 2 850, 1 687, 1 643, 1 589, 1 512, 1 463, 1 435, 1 404, 1 381, 1 273, 1 220, 1 134, 925, 759, 686
以6a為例,IR譜圖中兩組酰胺鍵由于所處的化學(xué)環(huán)境不同,受外部基團(tuán)的影響也不同,因此N-H和C=O峰都出現(xiàn)兩組伸縮振動(dòng)峰。由于酰胺基氮原子的孤對(duì)電子垂直于芳環(huán)的π軌道,這使得氮原子不能與芳環(huán)有效共軛,進(jìn)而酰胺氮原子可能會(huì)與羰基形成有效的共軛,降低羰基中C=O的能量,引起羰基的IR吸收波數(shù)降低(1 662 cm-1)。與噻唑環(huán)相鄰的酰胺鍵中的C=O受噻唑環(huán)及氨基的共同影響,伸縮振動(dòng)波數(shù)也較低(1 632 cm-1)。3 406 cm-1與3 323 cm-1為N-H伸縮振動(dòng),686 cm-1左右為C-S-C伸縮振動(dòng)。1H NMR譜圖上出現(xiàn)兩組NH質(zhì)子峰(12.87和11.17),說明6a中出現(xiàn)兩個(gè)-CONH基, 其中一個(gè)-CONH基受噻唑環(huán)的影響,向低場移動(dòng)(12.87)。6b~6i的譜圖也呈現(xiàn)類似規(guī)律。
6的抗癌活性測(cè)試以DMSO為參照物,阿霉素(ADM)為陽性對(duì)照藥劑。采用MTT比色法測(cè)定了6對(duì)人體前列腺癌細(xì)胞(PC3)的抑制率。將PC3細(xì)胞接種于96孔細(xì)胞培養(yǎng)板中,每孔2 200個(gè)細(xì)胞。于37 ℃在5%CO2飽和濕度下培養(yǎng)24 h,加入含處理因子的1 640培養(yǎng)基(200 μL/孔)。設(shè)空白對(duì)照和溶媒對(duì)照,每組4個(gè)平行,繼續(xù)培養(yǎng)72 h,加入5 mg·mL-1MTT 20 μL培養(yǎng)4 h,于酶標(biāo)儀上于570 nm處測(cè)定OD值。最后以溶媒對(duì)照組作參比計(jì)算抑制率,結(jié)果見表3。從表3可以看出,苯環(huán)上的取代基的電性對(duì)PC3癌細(xì)胞的抑制活性有一定的影響。當(dāng)R為2,4-F2時(shí)(6a~6e)對(duì)PC3癌細(xì)胞的抑制活性比R為2-F時(shí)(6f~6i)高;R′為6-F和6-Cl時(shí)(6b,6d,6h和6i)對(duì)PC3癌細(xì)胞的抑制活性比R′為其它取代基時(shí)高。6b和6d對(duì)PC3細(xì)胞的抑制活性相對(duì)較高,抑制率分別為69.6%和62.3%, 有進(jìn)一步優(yōu)化的潛力。這可能是取代基的吸電子誘導(dǎo)效應(yīng)引起了6分子上電荷分布的變化,使其更容易與受體結(jié)合。
表 3 6對(duì)PC3細(xì)胞的抑制活性*Table 3 Inhibition rates of 6 to PC3 cell
*c=10 μmmol·L-1
[1] Finerty M J, Bingham J P, Hartley J A,etal. Azinomycin bisepoxides containing rigid aromatic linkers:Synthesis, cytotoxicity and DNA interstrand cross-linking activity[J].Tetrahedron Letters,2009,50(26):3648-3650.
[2] Mahindroo N, Punchihewa C, Fujii N. Hedgehog-gli signaling pathway inhibitors as anticancer agents[J].Journal of Medicinal Chemestry,2009,52(13):3829-3845.
[3] Suzuki N, Suzuki T, Ota Y,etal. Design,synthesis, and biological activity of boronic acid-based histone deacetylase inhibitors[J].Journal of Medicinal Chemestry,2009,52(9):2909-2922.
[4] Castellano S, Milite C, Ragno R,etal. Design,synthesis and biological evaluation of carboxy analogues of arginine methyltransferase inhibitor 1(AMI-1)[J].Journal of Medicinal Chemestry,2010,5(3):398-414.
[5] Yu M, Lizarzaburu M, Beckmann H,etal. Identification of piperazine-bisamide GHSR antagonists for the treatment of obesity[J].Bioorganic & Medicinal Chemistry Letters,2010,20(5):1758-1762.
[6] Descoteaux C, Leblanc V, Brasseur K,etal. Synthesis of D- and L-tyrosine-chlorambucil analogs active against breast cancer cell lines[J].Bioorganic & Medicinal Chemistry Letters,2010,20(24):7388-7392.
[7] Koci J, Klimesova V, Waisser K,etal. Heterocyclic benzazole derivatives with antimycobacterial in vitro activity[J].Bioorganic & Medicinal Chemistry Letters,2002,12(22):3275-3278.
[8] Hutchinson I, Chua M S, Browne H L,etal. Synthesis and in vitro biological properties of fluorinated 2-(4-aminophenyl)benzothiazoles[J].Journal of Medicinal Chemestry,2001,44(9):1446-1455.
[9] Bradshaw T D, Westwell A D. The development of the antitumor benzothiazole prodrug, phortress, as a clinical candidate[J].Current Topics in Medicinal Chemistry,2004,11(13):1009-1021.
[10] Zhang L, Biamonte M, Busch D. 7-Substituted benzothiazolothio and pyridinothiazolothio purines as potent heat shock protein 90 inhibitors[J].Journal of Medicinal Chemestry,2006,49(17):5352-5362.
[11] Vu C B, Milne J C, Carney D P,etal. Discovery of benzothiazole derivatives as efficacious and enterocyte-specific MTP inhibitors[J].Bioorganic & Medicinal Chemistry Letters,2009,19(5):1416-1420.
[12] Wang Z, Shi X H, Wang J,etal. Synthesis,structure-activity relationships and preliminary antitumor evaluation of benzothiazole-2-thiol derivatives as novel apoptosis inducers[J].Bioorganic & Medicinal Chemistry Letters,2011,21(4):1097-1101.
[13] Soni B, Ranawat M S, Sharma R,etal. Synthesis and evaluation of some new benzothiazole derivatives as potential antimicrobial agents[J].European Journal of Medical Chemistry,2010,45(7):2938-2942.
[14] Saeed S, Rashid N, Jones P G,etal. Synthesis,characterization and biological evaluation of some thiourea derivatives bearing benzothiazole moiety as potential antimicrobial and anticancer agents[J].European Journal of Medical Chemistry,2010,45(4):1323-1331.
[15] Bondock S, Fadaly W, Metwally M A. Synthesis and antimicrobial activity of some new thiazole, thiophene and pyrazole derivatives containing benzothiazole moiety[J].European Journal of Medical Chemistry,2010,45(9):3692-3701.