劉 菲，劉 楠，何 菱

(四川大學(xué) 華西藥學(xué)院，四川 成都 610041)

·研究論文·

磺酰脒衍生物的合成及其抗腫瘤活性*

劉 菲，劉 楠，何 菱

(四川大學(xué) 華西藥學(xué)院，四川 成都 610041)

以氯化亞銅為催化劑，叔胺為底物，磺酰疊氮為氮源，經(jīng)“一鍋法”合成了11個(gè)磺酰脒衍生物(3a～3k,其中3c,3e～3i和3k為新化合物)，收率43%～96%，其結(jié)構(gòu)經(jīng)1H NMR，13C NMR和HR-ESI-MS表征。采用MTT法研究了3a～3j對(duì)人肺癌細(xì)胞(A549)，人結(jié)腸癌細(xì)胞(HCT116)和人肝癌細(xì)胞(HepG2)的抗腫瘤活性。結(jié)果表明：在用藥量為20μg·mL-1時(shí),3a～3k對(duì)A549，HCT116和HepG2的抑制率均優(yōu)于對(duì)照藥紫杉醇，其中N,N-二乙基-N′-對(duì)甲氧基苯磺酰脒(3b)對(duì)A549的抑制率為82%；N,N-二丙基-N′-對(duì)甲氧基苯磺酰脒(3c)對(duì)HCT116抑制率為80%，具有較好的抗腫瘤活性。

磺酰脒類衍生物；磺酰疊氮；叔胺；合成；抗腫瘤活性

磺胺衍生物因具有多種生物活性而受到人們的青睞，如抗菌、抗炎、抗血栓形成、抗?jié)儭⒖惯^敏、抗HIV、利尿、降血糖、降低血壓以及抗腫瘤活性[1-2]等，尤其能選擇性集聚腫瘤細(xì)胞[3]的特性使人們對(duì)磺胺衍生物的研究日益濃厚。因此，借助藥物化學(xué)最小修飾原則在磺胺衍生物上借助過渡金屬催化C-N鍵直接引入親水性的叔胺基團(tuán)，從而改善磺胺類衍生物水溶性差的缺陷，獲得副作用小活性更好的化合物具有研究意義。

為尋求一種簡(jiǎn)單、價(jià)廉、溫和、高效、快速、選擇性強(qiáng)的胺化方法，并探討其在藥物合成中的應(yīng)用，本文利用過渡金屬催化氮賓插入反應(yīng)，以疊氮為氮源，探討叔胺sp3C-H鍵插入胺化反應(yīng)[4]，為進(jìn)一步發(fā)掘磺胺藥物的生物活性及構(gòu)效關(guān)系奠定基礎(chǔ)。以磺酰疊氮(1a和1b)為氮源，親水性叔胺(2a，2c～2f,2h,2i,2k)為底物，氯化亞銅為催化劑，經(jīng)“一鍋法”合成了11個(gè)磺酰脒衍生物(3a～3k,其中3c,3e～3i和3k為新化合物，Scheme 1),收率43%～96%，其結(jié)構(gòu)經(jīng)1H NMR，13C NMR和HR-ESI-MS表征。采用MTT法研究了3a～3j對(duì)人肺癌細(xì)胞(A549)，人結(jié)腸癌細(xì)胞(HCT116)和人肝癌細(xì)胞(HepG2)的抗腫瘤活性。

1 實(shí)驗(yàn)部分

1．1 儀器與試劑

Varian INPVA 400型核磁共振儀(CDCl3為溶劑，TMS為內(nèi)標(biāo))；Bruker Daltonics Data Analysis 3.2型質(zhì)譜儀。

A549，HCT116，HepG220和MTT染料，Sigma；1和2按文獻(xiàn)[5-6]方法合成；其余所用試劑均為分析純。

反應(yīng)均在氮?dú)獗Ｗo(hù)下進(jìn)行，溶劑使用前經(jīng)干燥除水。

1.23a～3k的合成(以3a為例)[5-6]

在干燥反應(yīng)瓶中依次加入三乙胺(2a)143mg(1mmol)，對(duì)甲苯磺酰疊氮(1a)591mg(3mmol)，氯化亞銅20mg(0.2mmol)和三乙基芐基氯化銨(TEBA)22.7mg(0.1mmol)的乙腈(5mL)溶液，攪拌下回流反應(yīng)8h。濃縮后經(jīng)硅膠柱層析[洗脫劑：V(石油醚)∶V(乙酸乙酯)=8∶1和6∶1]純化得淡黃色固體N,N-二乙基-N′-對(duì)甲基苯磺酰脒(3a)。

用類似的方法合成淡黃色固體3b～3k。

3a:產(chǎn)率96%；1H NMRδ：8.15(s,1H),7.76(d,J=8.4Hz,2H),7.25(d,J=8.4Hz,2H),3.47(q,J=7.2Hz,2H),3.38(q,J=7.2Hz,2H),2.40(s,3H),1.26(t,J=7.2Hz,3H),1.14(t,J=7.2Hz,3H)；13C NMRδ：158.1,142.3,139.8,129.3,26.4,47.1,40.9,21.5,14.5,12.1；HR-ESI-MSm/z:Calcd for C15H14N2O2SNa{[M+Na]+}277.0981,found 277.0976。

3b:產(chǎn)率88%；1H NMRδ：8.14(s,1H),7.80～7.83(m,2H),6.91～6.95(m,2H),3.85(s,3H),3.47(q,J=7.2Hz,2H),3.37(q,J=7.2Hz,2H),1.26(t,J=7.2Hz,3H)，1.14(t,J=7.2Hz,3H)；13C NMRδ：162.2,157.9,134.7,128.4,113.8,55.5,47.0,40.9,14.5,12.1；HR-ESI-MSm/z:Calcd for C12H18N2O3SNa{[M+Na]+}293.0936,found 293.0936。

3c:產(chǎn)率90%；1H NMRδ：8.14(s,1H),7.80(d,J=8.8Hz,2H),6.93(d,J=8.8Hz,2H),3.85(s,1H),3.36(t,J=7.6Hz,2H),3.26(t,J=7.6Hz,2H),1.53～1.68(m,4H),0.91(t,J=7.8Hz,3H),0.86(t,J=7.8Hz,3H)；13C NMRδ：162.2,158.8,134.7,128.3,113.8,55.5,54.2,47.8,21.9,20.0,11.2,10.9；HR-ESI-MSm/z:Calcd for C14H22N2O3SNa{[M+Na]+}321.1243,found 321.1244。

3d:產(chǎn)率54%；1H NMRδ：8.44(s,1H),7.84～7.88(m,2H),7.48(d,J=8.4Hz,1H),7.36(t,J=2.0Hz,1H),7.29(t,J=8.4Hz,1H),7.14(dd,J=2.0Hz,8.0Hz,1H),6.95～6.99(m,2H),3.95(q,J=7.2Hz,2H),3.87(s,3H),1.18(t,J=7.2Hz,3H)；13C NMRδ：162.6,157.6,143.3,133.5,131.1,130.7,128.7,126.5,123.4,122.0,114.0,55.6,44.0,12.4；HR-ESI-MSm/z:Calcd for C16H17N2O3SBrNa{[M+Na]+}419.0041,found 419.0022。

3e:產(chǎn)率58%；1H NMRδ：8.41(s,1H),7.84～7.87(m,2H),7.39～7.42(m,2H),7.12～7.16(m,2H),6.94～6.98(m,2H),3.91～3.97(q,J=7.2Hz,2H),3.86(s,3H),1.17(t,J=7.2Hz,3H)；13C NMRδ：162.6,157.6,140.5,133.6,133.4,130.0,128.7,124.8,114.0,55.6,44.1,12.3；HR-ESI-MSm/z:Calcd for C16H17N2O3SClNa{[M+Na]+}375.0541,found 375.0524。

3f:產(chǎn)率66%；1H NMRδ：8.34(s,1H),7.80(d,J=8.0Hz,2H),7.28(d,J=8.0Hz,2H),7.08～7.12(m,2H),6.90～6.94(m,2H),3.90(q,J=7.2Hz,2H),3.83(s,3H),2.41(s,3H),1.14(t,J=7.2Hz,3H)；13C NMRδ：159.1,158.3,142.6,139.4,134.8,129.4,126.6,125.6,114.9,55.6,44.5,21.5,12.3;HR-ESI-MSm/z:Calcd for C17H20N2O3SNa{[M+Na]+}355.1087,found 355.1077。

3g:產(chǎn)率70%；1H NMRδ：8.33(s,1H),7.83～7.87(m,2H),7.08～7.11(m,2H),6.91～6.97(m,4H),3.89(q,J=7.2Hz,2H),3.85(s,3H),3.82(s,3H),1.14(t,J=7.2Hz,3H)；13C NMRδ：162.4,159.1,158.1,134.8,134.1,128.6,125.6,114.9,113.9,55.6,55.5,44.5,12.3；HR-ESI-MSm/z:Calcd for C17H20N2O4SNa{[M+Na]+}371.1041,found 371.1063。

3h:產(chǎn)率55%；1H NMRδ：8.12(s,1H),7.78～7.82(m,2H),6.91～6.95(m,2H),3.85(s,3H),3.39(t,J=7.6Hz,2H),3.28(t,J=7.6Hz,2H),1.47～1.61(m,4H),1.23～1.33(m,4H),0.94(t,J=7.2Hz,2H),0.87(t,J=7.2Hz,2H)；13C NMRδ：162.1,158.6,134.8,128.3,113.8,55.5,52.3,45.9,30.7,28.7,19.9,19.6,13.6,13.5；HR-ESI-MSm/z:Calcd for C16H26N2O3SNa{[M+Na]+}349.1562,found 349.1568。

3i:產(chǎn)率65%；1H NMRδ：8.37(s,1H),7.84～7.87(m,2H),7.10～7.19(m,4H),6.94～6.98(m,2H),3.92(q,J=7.2Hz,2H),3.86(s,1H),1.16(t,J=7.2Hz,3H)；13C NMRδ：162.5,157.9,138.1,138.0,133.8,128.7,125.9,125.8,116.9,116.7,114.0,55.6,44.4,12.3；HR-ESI-MSm/z:Calcd for C16H17N2O3SFNa{[M+Na]+}359.0836,found 359.0819。

3j:產(chǎn)率88%；1H NMRδ：8.14(s,1H),7.74(d,J=8.0Hz,2H),7.24(d,J=8.0Hz,2H),3.35(t,J=7.6Hz,2H),3.25(t,J=7.6Hz,2H),2.39(s,3H),1.54～1.67(m,4H),0.90(t,J=7.6Hz,3H),0.85(t,J=7.6Hz,3H)；13C NMRδ：158.9,142.2,139.8,129.3,126.3,54.2,47.8,21.8,21.4,19.9,11.2,10.8；HR-ESI-MSm/z:Calcd for C14H22N2O2SNa{[M+Na]+}305.1294,found 305.1284。

3k:產(chǎn)率43%；1H NMRδ：7.82(d,J=8.8Hz,2H),7.27～7.33(m,3H),7.20～7.24(m,2H),6.87～6.92(m,2H),4.69(s,2H),3.84(s,3H),3.30(bro,2H),3.17(bro,2H),1.74～1.80(m,4H)；13C NMRδ：165.9,161.8,136.3,135.8,128.7,128.2,128.1,127.8,113.6,55.5,53.3,48.3,28.7,22.2,19.6；HR-ESI-MSm/z:Calcd for C19H22N2O3SNa{[M+Na]+}381.1243,found 381.1234。

1.3 體外抗腫瘤活性測(cè)定[7]

取對(duì)數(shù)生長期癌細(xì)胞A549，HCT116和HepG2，用完全培養(yǎng)液調(diào)整細(xì)胞濃度為2．0×104/mL接種到96孔板上，每孔板100μL，放入培養(yǎng)箱中培養(yǎng)24h后更換不同濃度藥物的完全培養(yǎng)基繼續(xù)培養(yǎng)48h(以0.1%DMSO溶液做空白對(duì)照，紫杉醇作為陽性對(duì)照藥)。加入MTT 20μL (5mg·mL-1)，孵育4h；除去培養(yǎng)基，加入DMSO(150μL)，振蕩15min后用酶標(biāo)于570nm處測(cè)定吸光度，取平均值，計(jì)算相對(duì)細(xì)胞抑制生長率。

2 結(jié)果與討論

2．1 反應(yīng)條件優(yōu)化

以合成3j為模板，考察金屬催化劑及其用量對(duì)反應(yīng)的影響，結(jié)果見表1。由表1可見，在用量(20mol%)相同時(shí)，以CuCl為催化劑時(shí)，反應(yīng)效果最好，產(chǎn)率最高(88%)。由表1還可見，在催化劑(CuCl)相同情況下，隨著其用量的增加，產(chǎn)率增加;當(dāng)用量為20mol%時(shí)，產(chǎn)率達(dá)到頂峰(88%)；再增加用量，產(chǎn)率無變化。

較佳催化劑為CuCl,用量為20mol%。

2．2 抗腫瘤活性

3的抗腫瘤活性結(jié)果見表2。由表2可見，3a～3c對(duì)A549的抑制率超過70%,3b達(dá)82%，優(yōu)于紫杉醇(31%)。從結(jié)構(gòu)上看，叔胺取代基較小，有利于增加對(duì)A549的抑制活性，3c對(duì)HCT116的抑制率達(dá)80%。R為甲氧基比R為甲基的抗腫瘤活性更好；3g和3i對(duì)三種腫瘤細(xì)胞均具有一定抑制活性；R4為芳香基團(tuán)時(shí)(3d～3i和3k),對(duì)HepG2的抑制活性更有利。

表1 催化劑及其用量對(duì)3j產(chǎn)率的影響*Table1 Effect of catalysts and their amount on the yield of 3j

*反應(yīng)條件同1．2

表2 3a～3k的抗腫瘤活性*Table2 Antitumor activities of 3a～3k

*c(3)20μg·mL-1；A549：人肺癌細(xì)胞；HCT116：人結(jié)腸癌細(xì)胞；HepG2：人肝癌細(xì)胞

3 結(jié)論

以叔氨為底物，磺酰疊氮為氮源，氯化亞銅為催化劑(用量20mol%)，經(jīng)胺化反應(yīng)合成了一系列磺酰脒衍生物(3a～3k)。該方法具有原料簡(jiǎn)單易得、催化劑價(jià)廉、操作簡(jiǎn)單和產(chǎn)率較高等優(yōu)點(diǎn)。

初步活性篩選結(jié)果表明：3a～3k對(duì)A549，HCT116和HepG220腫瘤細(xì)胞表現(xiàn)出一定抑制活性，其中N,N-二乙基-N′-對(duì)甲氧基苯磺酰脒(3b)對(duì)A549的抑制率為82%；N，N-二丙基-N′-對(duì)甲氧基苯磺酰脒(3c)對(duì)人結(jié)腸癌細(xì)胞HCT116的抑制率為80%，均表現(xiàn)出較好的抗腫瘤活性?；酋ｋ哐苌锏倪M(jìn)一步結(jié)構(gòu)修飾與活性篩選的研究正在進(jìn)行中。

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SynthesisandAntitumorActivitiesofSulfonylAmidineDerivatives

LIU Fei, LIU Nan，HE Ling

(West China School of Pharmacy,Sichuan University,Chengdu 610041,China)

Eleven sulfonyl amidine derivatives(3a～3k)were synthesized by “one-pot” using tertiaryamines as the substrate,sulfonyl azides as nitrogen source and CuCl as the catalyst.The structures were characterized by1H NMR,13C NMR and HR-ESI-MS.3c,3e～3iand3kwere new compounds.Theinvitroantitumor activities of3a～3kwere tested by MTT method.The results showed that3a～3kexhibited better antitumor activities agast A549，HCT116and HepG220cells at 20μg·mL-1．The inhibition ofN,N-dipropyl-N′-tosylformimidamide(3b)against A549cells was 82%,andN′[(4-methoxyphenyl)sulfonyl]-N,N-dipropylformimidamide(3c)against HCT116cells was 80%.

sulfonyl amidine;sulfonyl azide;tertiaryamine;synthesis;antitumor activity

2014-03-11；

2014-04-10

國家自然科學(xué)基金資助項(xiàng)目(21072131)

劉菲(1988-)，女，漢族，四川內(nèi)江人，碩士研究生，主要從事藥物化學(xué)的合成研究。

何菱，教授，Tel.028-88503365,E-mail:lhe2001@sina.com

O623．626；O621．3

A

1005-1511(2014)04-0440-04