Lopinavir up-regulates expression of the antiviral protein ribonuclease L in human papillomavirus-positive cervical carcinoma cells
Lopinavir up-regulates expression of the antiviral protein ribonuclease L in human papillomavirus-positive cervical carcinoma cells
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DOI：
10.3851/imp1786
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发表时间：
2011-01-01
影响因子：
1.2
通讯作者：
Hampson, Ian N.
中科院分区：
文献类型：
--
作者：
Batman, Gavin;Oliver, Anthony W.;Hampson, Ian N.
背景资料：我们以前已经表明，HIV蛋白酶抑制剂洛匹那韦对人乳头瘤病毒（HPV）阳性的宫颈癌cells via a unknown mechanism.Methods：SiHa宫颈癌细胞稳定转染蛋白酶体传感器载体pZsProSensor-1，以确认洛匹那韦抑制这些细胞中的蛋白酶体。然后使用Panorama Xpress profiler 725抗体阵列分析洛匹那韦处理的Si Ha细胞与对照未处理的Si Ha细胞中蛋白质表达的特异性变化，随后进行PCR和蛋白质印迹。进行洛匹那韦处理的E6/E7永生化的人角质形成细胞与对照人角质形成细胞的比色生长测定。有针对性的小干扰RNA基因沉默，然后由洛匹那韦处理/未处理的Si Ha细胞的生长测定比较也used.Results：洛匹那韦诱导的pZsProSensor-1转染Si Ha细胞的荧光增加，指示蛋白酶体抑制。经PCR和western blot证实，经洛匹那韦处理的SiHa细胞中核糖核酸酶L（RNASEL）蛋白表达上调。靶向沉默RNASEL降低了SiHa细胞对洛匹那韦的敏感性。对E6/E7永生化角质形成细胞与对照细胞的选择性毒性也被视为与洛匹那韦和上调RNASEL expression.Conclusions：这些数据是一致的洛匹那韦对HPV阳性宫颈癌细胞的毒性有关，其能够阻断病毒蛋白酶体激活和诱导的抗病毒蛋白RNASEL的上调。这得到了药物的选择性毒性和E6/E7永生化角质形成细胞中RNASEL的上调以及在RNASEL基因表达沉默后SiHa细胞中观察到的对洛匹那韦的耐药性增加的支持。
Background: We have previously shown that the HIV protease inhibitor lopinavir has selective toxicity against human papillomavirus (HPV)-positive cervical carcinoma cells via an unknown mechanism.Methods: Si Ha cervical carcinoma cells were stably transfected with the proteasome sensor vector pZsProSensor-1 to confirm lopinavir inhibits the proteasome in these cells. The Panorama Xpress profiler 725 antibody array was then used to analyse specific changes in protein expression in lopinavir-treated versus control untreated Si Ha cells followed by PCR and western blotting. Colorimetric growth assays of lopinavir-treated E6/E7 immortalised versus control human keratinocytes were performed. Targeted small interfering RNA gene silencing followed by growth assay comparison of lopinavir-treated/untreated Si Ha cells was also used.Results: Lopinavir induced an increase in the fluorescence of pZsProSensor-1 transfected Si Ha cells, indicative of proteasomal inhibition. Ribonuclease L (RNASEL) protein was shown to be up-regulated in lopinavir-treated SiHa cells, which was confirmed by PCR and western blot. Targeted silencing of RNASEL reduced the sensitivity of SiHa cells to lopinavir. Selective toxicity against E6/E7 immortalised keratinocytes versus control cells was also seen with lopinavir and was associated with up-regulated RNASEL expression.Conclusions: These data are consistent with the toxicity of lopinavir against HPV-positive cervical carcinoma cells being related to its ability to block viral proteasome activation and induce an up-regulation of the antiviral protein RNASEL. This is supported by the drug's selective toxicity and up-regulation of RNASEL in E6/E7 immortalised keratinocytes combined with the increased resistance to lopinavir observed in SiHa cells following silencing of RNASEL gene expression.