Department of Medical Oncology, University Medical Center Groningen, University of Groningen, Groningen, The Netherlands.
Mol Pharmacol. 2012 May;81(5):701-9. doi: 10.1124/mol.111.076539. Epub 2012 Feb 10.
In cervical cancer, the p53 and retinoblastoma (pRb) tumor suppressor pathways are disrupted by the human papilloma virus (HPV) E6 and E7 oncoproteins, because E6 targets p53 and E7 targets pRb for rapid proteasome-mediated degradation. We have investigated whether E6 suppression with small interfering RNA (siRNA) restores p53 functionality and sensitizes the HPV16-positive cervical cancer cell line SiHa to apoptosis by cisplatin, irradiation, recombinant human tumor necrosis factor-related apoptosis-inducing ligand (rhTRAIL), or agonistic anti-Fas antibody. E6 siRNA resulted in decreased E6 mRNA levels and enhanced p53 and p21 expression, demonstrating the restoration of p53 functionality in SiHa cells, without inducing high levels of apoptosis (<10%). Cell surface expression of the proapoptotic death receptors (DRs) DR4, DR5, and Fas was not affected by E6 suppression. E6 suppression conferred susceptibility to cisplatin-induced apoptosis but not to irradiation-, rhTRAIL-, or anti-Fas antibody-induced apoptosis. Combining cisplatin with rhTRAIL or anti-Fas antibody induced even higher apoptosis levels in E6-suppressed cells. At the molecular level, cisplatin treatment resulted in elevated p53 levels, enhanced caspase-3 activation, and reduced p21 levels in E6-suppressed cells. Cisplatin in combination with death receptor ligands enhanced caspase-8 and caspase-3 activation and reduced X-linked inhibitor-of-apoptosis protein (XIAP) levels in these cells. We showed using siRNA that the enhanced apoptosis in E6-supressed cells was related to reduced XIAP levels and not due to reduced p21 levels. In conclusion, targeting E6 or XIAP in combination with cisplatin can efficiently potentiate rhTRAIL-induced apoptosis in HPV-positive cervical cancer cells.
在宫颈癌中，人乳头瘤病毒（HPV）的 E6 和 E7 癌蛋白破坏了 p53 和视网膜母细胞瘤（pRb）肿瘤抑制途径，因为 E6 靶向 p53，E7 靶向 pRb 以进行快速蛋白酶体介导的降解。我们研究了用小干扰 RNA（siRNA）抑制 E6 是否能恢复 p53 的功能，并通过顺铂、辐射、重组人肿瘤坏死因子相关凋亡诱导配体（rhTRAIL）或激动性抗 Fas 抗体使 HPV16 阳性宫颈癌细胞系 SiHa 敏感化以诱导细胞凋亡。E6 siRNA 导致 E6 mRNA 水平降低，并增强了 p53 和 p21 的表达，表明 SiHa 细胞中 p53 功能得到了恢复，而没有诱导高水平的凋亡（<10%）。促凋亡死亡受体（DR）DR4、DR5 和 Fas 的细胞表面表达不受 E6 抑制的影响。E6 抑制赋予了对顺铂诱导的细胞凋亡的敏感性，但对辐射、rhTRAIL 或抗 Fas 抗体诱导的细胞凋亡没有敏感性。将顺铂与 rhTRAIL 或抗 Fas 抗体联合使用可使 E6 抑制的细胞中的凋亡水平更高。在分子水平上，顺铂处理导致 E6 抑制的细胞中 p53 水平升高， caspase-3 激活增强，p21 水平降低。顺铂与死亡受体配体联合使用可增强这些细胞中 caspase-8 和 caspase-3 的激活，并降低 X 连锁凋亡抑制蛋白（XIAP）的水平。我们使用 siRNA 表明，E6 抑制的细胞中增强的凋亡与 XIAP 水平降低有关，而不是由于 p21 水平降低所致。总之，靶向 E6 或 XIAP 与顺铂联合使用可以有效地增强 HPV 阳性宫颈癌细胞中 rhTRAIL 诱导的凋亡。
