Small interfering double-stranded RNAs as therapeutic molecules to restore chemosensitivity to thymidylate synthase inhibitor compounds
Small interfering double-stranded RNAs as therapeutic molecules to restore chemosensitivity to thymidylate synthase inhibitor compounds
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DOI：
10.1158/0008-5472.can-03-1203
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发表时间：
2004-02-15
期刊：
影响因子：
11.2
通讯作者：
Chu, E
中科院分区：
文献类型：
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作者：
Schmitz, JC;Chen, TM;Chu, E
RNA干扰是双链RNA特异性沉默相应基因表达的转录后机制。21-23个核苷酸的小干扰双链RNA（siRNA）可以诱导RNA干扰过程。本实验室的研究表明，胸苷酸合成酶（TS）mRNA的翻译受其自身蛋白终产物TS的负性自身调节。这一过程的破坏引起TS合成的增加，并导致细胞对TS靶向化合物的耐药性的发展。作为在mRNA水平抑制TS表达的策略，设计siRNA以靶向人TS mRNA上的核苷酸1058-1077。将TS 1058 siRNA转染入人结肠癌RKO细胞导致TS表达的剂量依赖性抑制，IC 50值为10 pM，但对α-微管蛋白或拓扑异构酶I的表达没有影响。TS 1058对TS表达的抑制在48小时达到最大，并保持抑制长达5天。用TS 1058 siRNA预处理RKO细胞抑制暴露于雷替曲塞后TS蛋白的诱导。另外。TS 1058恢复了耐药RKO-HTStet细胞系对各种TS抑制剂化合物的化学敏感性。TS 1058治疗后，雷替曲塞、1843089和5-氟-2 '-脱氧尿苷的IC 50值降低了15 -16倍。这些研究表明，TS-靶向siRNA是TS表达的有效抑制剂，并且其本身或与TS抑制剂化合物组合作为化学增敏剂可能具有治疗潜力。
RNA interference is a post-transcriptional mechanism by which double-stranded RNA specifically silence expression of a corresponding gene. Small interfering double-stranded RNA (siRNA) of 21-23 nucleotides can induce the process of RNA interference. Studies from our laboratory have shown that translation of thymidylate synthase (TS) mRNA is controlled by its own protein end-product TS in a negative autoregulatory manner. Disruption of this process gives rise to increased synthesis of TS and leads to the development of cellular drug resistance to TS-targeted compounds. As a strategy to inhibit TS expression at the mRNA level, siRNAs were designed to target nucleotides 1058-1077 on human TS mRNA. Transfection of TS1058 siRNA into human colon cancer RKO cells resulted in a dose-dependent inhibition of TS expression with an IC50 value of 10 pM but had no effect on the expression of a-tubulin or topoisomerase I. Inhibition of TS expression by TS1058 was maximal at 48 h and remained suppressed for up to 5 days. Pretreatment of RKO cells with TS1058 siRNA suppressed TS protein induction following exposure to raltitrexed. In addition. TS1058 restored chemosensitivity of the resistant RKO-HTStet cell line to various TS inhibitor compounds. On treatment with TS1058, IC50 values for raltitrexed, 1843089, and 5-fluoro-2'-deoxyuridine decreased by similar to15-16-fold. These studies suggest that TS-targeted siRNAs are effective inhibitors of TS expression and may have therapeutic potential by themselves or as chemosensitizers in combination with TS inhibitor compounds.