A novel inhibitor of DNA polymerase beta enhances the ability of temozolomide to impair the growth of colon cancer cells.
A novel inhibitor of DNA polymerase beta enhances the ability of temozolomide to impair the growth of colon cancer cells.
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DOI：
10.1158/1541-7786.mcr-09-0309
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发表时间：
2009-12
期刊：
影响因子：
--
通讯作者：
Narayan S
中科院分区：
文献类型：
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作者：
Jaiswal AS;Banerjee S;Panda H;Bulkin CD;Izumi T;Sarkar FH;Ostrov DA;Narayan S
最近出现的概念是通过抑制碱基切除修复（BER）途径中的各种蛋白质来使癌细胞对DNA烷基化药物敏感。在本研究中，我们使用DNA聚合酶β（Pol-β）的基于结构的分子对接，并确定了一种有效的小分子量抑制剂（SMI），NSC-666715。我们通过使用APE 1、Fen 1、DNA连接酶I和Pol-β定向的单核苷酸（SN）-和长补丁（LP）-BER的体外活性来确定该SMI对Pol-β的特异性。还通过使用荧光各向异性测定了NSC-666715与Pol-β的结合特异性。NSC-666715对DNA烷基化药物替莫唑胺（TMZ）对结肠癌细胞的细胞毒性的影响通过体外克隆形成和体内异种移植试验测定。相对于未治疗或单药治疗，联合治疗中肿瘤生长的减少更高。NSC-666715显示出对阻断Pol-β活性的高度特异性。它阻断Pol-β介导的SN-和LP-BER，而不影响APE 1、Fen 1和DNA连接酶I的活性。荧光各向异性数据表明，NSC-666715直接特异性地与Pol-β相互作用，并干扰与受损DNA的结合。在体外和体内测定中，NSC-666715显著诱导TMZ对结肠癌细胞的敏感性。结果进一步表明，NSC-666715对BER的破坏否定了其对耐药性的贡献，并绕过了其他耐药因素，如错配修复缺陷。我们的研究结果为开发高度特异性且因此更安全的基于结构的抑制剂以预防肿瘤进展和/或治疗结直肠癌提供了“概念验证”。
The recent emerging concept to sensitize cancer cells to DNA-alkylating drugs is by inhibiting various proteins in the base excision repair (BER) pathway. In the present study, we used structure-based molecular docking of DNA polymerase β (Pol-β) and identified a potent small molecular weight inhibitor (SMI), NSC-666715. We determined the specificity of this SMI for Pol-β by using in vitro activities of APE1, Fen1, DNA ligase I, and Pol-β-directed single nucleotide (SN)- and long-patch (LP)-BER. The binding specificity of NSC-666715 with Pol-β was also determined by using fluorescence anisotropy. The effect of NSC-666715 on the cytotoxicity of the DNA-alkylating drug, Temozolomide (TMZ), to colon cancer cells was determined by in vitro clonogenic and in vivo xenograft assays. The reduction in tumor growth was higher in the combination treatment relative to untreated or monotherapy treatment. NSC-666715 showed a high specificity for blocking Pol-β activity. It blocked Pol-β-directed SN- and LP-BER without affecting the activity of APE1, Fen1 and DNA ligase I. Fluorescence anisotropy data suggested that NSC-666715 directly and specifically interacts with Pol-β and interferes with binding to damaged DNA. NSC-666715 drastically induces the sensitivity of TMZ to colon cancer cells both in vitro and in vivo assays. The results further suggest that the disruption of BER by NSC-666715 negates its contribution to drug-resistance and bypasses other resistance factors, such as mismatch repair defects. Our findings provide the “proof-of-concept” for the development of highly specific and thus safer structure-based inhibitors for the prevention of tumor progression and/or treatment of colorectal cancer.