G-四聯(lián)體的小分子配體的研究進(jìn)展

鮑偉偉

（溫州大學(xué)，浙江 溫州 310015）

Davies[1]于 1962年首次提出G-四聯(lián)體。G-四聯(lián)體是由富G序列在一定濃度的Na+、K+條件下所形成一種DNA二級(jí)結(jié)構(gòu)[2-4]。G-四聯(lián)體結(jié)構(gòu)具有多態(tài)性，主要是因?yàn)殒湹臄?shù)量、鏈的結(jié)構(gòu)、鏈的取向以及環(huán)的結(jié)構(gòu)形態(tài)等存在著多樣性。G-四聯(lián)體的基本單元是4個(gè)鳥嘌呤通 過氫鍵圍繞形成平面得到G-四分體，其中的氫鍵包括Hoogste en (N1-O6)(N2-N7)和Watson-Crick，而 G-四分體之間通過π-π堆積得到G-四聯(lián)體。G-四聯(lián)體主要有平行結(jié)構(gòu)和反平行結(jié)構(gòu)，主要受到鳥嘌呤中的堿基和糖之間的syn和anti這2種糖苷鍵的影響。E Gavathiotis等對(duì)于平行的G-四聯(lián)體帶 有anti糖苷鍵的闡述非常明白[5]。其中最影響拓?fù)浣Y(jié)構(gòu)的應(yīng)當(dāng)是一條鏈中帶有對(duì)角環(huán)、側(cè)邊環(huán)以及螺旋槳環(huán)等多種環(huán)形結(jié)構(gòu)的G-四聯(lián)體，因?yàn)檫@樣形成的結(jié)構(gòu)中存在著4種形態(tài)的溝槽，比如對(duì)角環(huán)和對(duì)角環(huán)所形成的簡(jiǎn)單環(huán)，但存在螺旋槳環(huán)就會(huì)很復(fù)雜。

在20世紀(jì)80年代初期，只有生物物理學(xué)家對(duì)其感興趣，而到80年代末期時(shí)，位于端粒末端的富G序列 能夠形成G-四聯(lián)體結(jié)構(gòu)的這個(gè)事實(shí)[6-7]以及在1991年時(shí)Zahl er[8]與其同伴證實(shí)了經(jīng)K+穩(wěn)定G-四聯(lián)體結(jié)構(gòu)具有限制端粒酶活性的功能后，G-四聯(lián)體就成為了抑制端粒酶的靶點(diǎn)[9-10]。隨著對(duì)G-四聯(lián)體研究的不斷深入，大量事實(shí)證實(shí)G-四聯(lián)體確實(shí)存在于活的生物體中[11-14]并且發(fā)揮著重要的生物性作用[15]。在真核細(xì)胞的基因組中，有很多關(guān)鍵的富G區(qū)域有望形成G-四聯(lián)體，像端粒[16]、免疫球蛋白開關(guān)區(qū)域[17]、致癌基因啟動(dòng)子區(qū)域[18]以及核糖體DNA[19]。G-四聯(lián)體與小分子配體結(jié)合時(shí)能夠發(fā)揮重要作用，如限制端粒酶的活性[20]，干擾端粒的活性，影響端粒的維持[21-22]，進(jìn)一步影響細(xì)胞的生長(zhǎng)與增殖[23]，甚至影響癌細(xì)胞的發(fā) 生與發(fā)展。

本文綜述了藥物及其衍生物作為G-四聯(lián)體的配體以及顯 示其優(yōu)越的抗腫瘤活性。

1 蒽醌類化合物

蒽醌類化合物廣泛存在于蓼科、豆科等眾多的植物中。早期因其對(duì)腫瘤細(xì)胞具有毒性而作為DNA嵌入劑。分子模擬研究表明蒽醌類化合物可能可以通過嵌插 模式與G-四聯(lián)體相互作用[24]，Sun等[25]于1997首先提出G-四聯(lián)體的配體BSU-1051，且抑制端粒酶活性的50%的有效濃度（teIIC50）為23μM。隨后大量報(bào)道經(jīng)取代反應(yīng)得到蒽醌類衍生物，其中包括1,4-, 1,5-, 1,8-, 2,6-,和2,7-等，其中的側(cè)鏈變化趨勢(shì)是氨基到特異性氨基酸官能團(tuán)，并且考察了側(cè)鏈的大小和長(zhǎng)度之間的關(guān)系以及活性[26-30]。

近年來，研究者發(fā)現(xiàn)經(jīng)肽基取代的蒽醌類衍生物對(duì)G-四聯(lián)體表現(xiàn)出極為卓越的選擇性。他們通過加入疏水性殘基苯丙氨酸到賴氨酸側(cè)鏈的2,6 或者 2,7上，表現(xiàn)出的選擇性要遠(yuǎn)遠(yuǎn)大于雙鏈DNA，并且賴氨酸被苯丙氨酸所取代的蒽醌類衍生物能夠在更低的濃度下使得細(xì)胞凋亡[31]。

2 喹叨啉類衍生物

喹叨啉于1977年第一次從南非的一種 植株中提取出來[32]，它具有許多優(yōu)良的性質(zhì)，包括抗菌、抗瘧疾、抗炎癥等。SYUIQ-5是一種廣為研究的喹叨啉類衍生物，一系列的抗癌研究表明SYUIQ-5具有抑制端粒酶的活性，可縮短端粒的長(zhǎng)度，誘導(dǎo)細(xì)胞的衰老和生長(zhǎng)的停止。另外，這種化合物能夠和c-myc啟動(dòng)子區(qū)域上的G-四聯(lián)體相互結(jié)合并且在細(xì)胞的增殖和衰老的過程中占據(jù)重要作用[33]。

雙取代以及11號(hào)位上取代的喹叨啉衍生物都表現(xiàn)為使得G-四聯(lián)體更為穩(wěn)定，以及端粒酶抑制作用，但是它們對(duì)G-四聯(lián)體的選擇性與雙鏈DNA相比卻顯得差強(qiáng)人意。不過11位取代的喹叨啉衍生物表現(xiàn)出了更強(qiáng)的端粒酶活性，其中喹叨啉telIC50＞138μM；11位取代衍生物telIC50= 0.44～12.3μM[34]。另外5-N-甲基化喹叨啉因?yàn)檎姾芍行呐cG-四聯(lián)體的負(fù)離子通道之間的相互作用，提高了其與反平行G-四聯(lián)體結(jié)構(gòu)之間的選擇性以及穩(wěn)定性[35]。

Jixun Dai等[36]通過了解溶液中的喹叨啉與G-四聯(lián)體（2∶1）復(fù)合物的結(jié)構(gòu)，表明藥物能夠誘導(dǎo)側(cè)翼序列形成一個(gè)新的綁定位點(diǎn)，并且強(qiáng)調(diào)了堆垛相互作用以及靜電相互作用間的重要性。他們首先強(qiáng)調(diào)了藥物的形狀以及2個(gè)側(cè)翼堿基在決定藥物綁定特異性方面的 重要作用。

3 小檗堿類衍生物

小檗堿是從中藥黃連中提取出來的一種藥物，在中 醫(yī)中主要是用來醫(yī)治濕熱癥。在早期，小檗堿因?yàn)槠浔旧硎且环N抗生素使得其不是一種良好的抗腫瘤藥品，后來因獲得了一種混合小檗堿的新藥，使其成為 了抗腫瘤藥物[37]，引起了廣泛關(guān)注。小檗堿能夠抑制各種腫瘤細(xì)胞的生長(zhǎng)[38-39]，其中包括乳腺癌細(xì)胞[40]、胰腺癌細(xì)胞[41]以及胃癌細(xì)胞[42-43]。后來，Tsuruo和其同伴發(fā)現(xiàn)小檗堿能夠抑制端粒酶的活性并且限制端粒的延長(zhǎng)，同時(shí)也報(bào)道能夠和G-四聯(lián)體相互作用。Zhang等[44]發(fā)現(xiàn)9-取代小檗堿衍生物對(duì)G-四聯(lián)體結(jié)構(gòu)具有超強(qiáng)的相互結(jié)合能力并且能夠抑制端粒酶的活性。為了開發(fā)更具活性的端粒酶抑制劑，研究者開發(fā)了一系列的9-N-取代小檗堿衍生物，研究證實(shí)這些衍生物都能夠誘導(dǎo)并且穩(wěn)定c-myc上啟動(dòng)子區(qū)域平行G-四聯(lián)體，還能夠抑制c-myc表達(dá)[45]。

4 卟啉類衍生物及其相關(guān)的類似物

眾所周知的卟啉是紅細(xì)胞中的色素— —亞鐵血紅素?；衔颰MPyP4是最具代 表性的陽(yáng)離子卟啉類化合物。Hurley課題組對(duì)此進(jìn)行了深入而廣泛的研究[46-56]。TMPyP4對(duì)G-四聯(lián)體表現(xiàn)出較強(qiáng)的相互作用（ΔT1/2= 17℃），能夠有效抑制端粒酶的活性（IC50=6μM）。但問題是其對(duì)于平行以及反平行的G-四聯(lián)體缺乏明顯的選擇性[57-58]，甚至它對(duì)所有的核酸都具有選擇性(包括單鏈、雙鏈、三鏈以及四鏈)[59]。Parkinson等[60]報(bào)道了端粒G-四聯(lián)體與TMPyP4復(fù)合物的X單晶衍射結(jié)構(gòu)，表明TMPyP4不直接與G-四分體相互結(jié)合。

盡管TMPyP4表現(xiàn)出來的選擇性不盡人意， 但是研究者們對(duì)其的熱情卻不見消退，如Gerald等通過ITC、CD以及ESI-MS等方法來考察卟啉經(jīng)N-甲基-4-吡啶基取代的取代數(shù)目(如 5-N-甲基-4-吡啶基卟啉)、有效性以及取代后的結(jié)合能的大小，KTMPyP4，KP(5,15)＞ KP(5,10,15)＞ KP(5,10)，KP(5)，其中認(rèn)為P(5,15)和G-四聯(lián)體采用的是嵌插模式，而P(5,10,15)和G-四聯(lián)體采用的是底端堆垛模式[61]。

5 展望

自然界本身存在的藥物對(duì)新藥物的發(fā)現(xiàn)以及發(fā)展是一筆巨大寶貴的財(cái)富。其多態(tài) 性和結(jié)構(gòu)的復(fù)雜性，使其成為抗腫瘤藥物選更具選擇性，通過對(duì)G-四聯(lián)體的結(jié)構(gòu)特征以及功能的深入了解，我們期望在未來尋找到更多更好更具針對(duì)性的配體。

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