CD95-mediated apoptosis is impaired at receptor level by cellular FLICE-inhibitory protein (long form) in wild-type p53 human ovarian carcinoma
CD95-mediated apoptosis is impaired at receptor level by cellular FLICE-inhibitory protein (long form) in wild-type p53 human ovarian carcinoma
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DOI：
10.1158/1078-0432.ccr-03-0537
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发表时间：
2004-08-01
影响因子：
11.5
通讯作者：
Canevari, S
中科院分区：
文献类型：
--
作者：
Mezzanzanica, D;Balladore, E;Canevari, S
目的：卵巢癌是一种高度致命的恶性肿瘤，通常会对化疗产生耐药性。细胞凋亡信号和 p53 状态的改变会导致耐药性，而耐药细胞中也缺乏 CD95 介导的细胞凋亡。我们分析了 p53 状态不同的卵巢癌细胞系对 CD95 介导的细胞凋亡的抵抗机制。实验设计：通过激动性抗 CD95 抗体诱导 CD95 介导的细胞凋亡，并通过生化和功能分析监测细胞凋亡级联。结果：CD95 介导的细胞凋亡在人卵巢癌细胞中被阻断。在野生型 p53 细胞系中，用蛋白质合成抑制剂放线菌酮 (CHX) 与抗 CD95 一起治疗克服了耐药性，表明存在不稳定的抑制蛋白。事实上，不稳定的细胞 FLICE 抑制蛋白长形式 (c-FLIP,.) 被发现可以阻止 caspase-8 募集到死亡诱导信号复合物 (DISC)，并且 CHX 对细胞的致敏是由于 DISC 水平上的 c-FLIPL 下调。用反义寡核苷酸下调 c-FLIPL 会增加 CD95 介导的细胞凋亡，就像在 CHX 致敏的细胞中一样，这表明 c-FLIPL 直接参与细胞凋亡抵抗。在 DISC 水平去除 c-FLIPL 阻断可以完全激活线粒体途径，并最终导致野生型 p53 细胞凋亡，而在 p53 突变的细胞中，c-FLIPL 参与 CD95 介导的细胞凋亡抵抗似乎无关紧要。对卵巢肿瘤组织阵列的免疫组织化学分析显示，在没有 p53 积累的样本中 c-FLIPL 表达（P = 0.034），并且在 27 个已知 p53 状态的上皮性卵巢癌标本中也观察到 c-FLIPL 与 p53 表达水平之间存在显着（P = 0.037）的负相关关系。结论：抑制蛋白 c-FLIPL 参与野生型卵巢癌细胞对 CD95 介导的细胞凋亡的抵抗第 53 页。
Purpose: Ovarian carcinoma is a highly lethal malignancy that often becomes resistant to chemotherapy. Alterations in apoptotic signals and p53 status contribute to drug resistance, and CD95-mediated apoptosis is also deficient in resistant cells. We analyzed the mechanism of resistance to CD95-mediated apoptosis in ovarian carcinoma cell lines differing in p53 status.Experimental Design: CD95-mediated apoptosis was induced by agonistic anti-CD95 antibody, and the apoptotic cascade was monitored with biochemical and functional assays.Results: CD95-mediated apoptosis was blocked in human ovarian cancer cells. In cell lines with wild-type p53, treatment with the protein synthesis inhibitor cycloheximide (CHX) together with anti-CD95 overcame the resistance, suggesting the presence of a labile inhibiting protein. Indeed, the labile protein cellular FLICE-inhibitory protein long form (c-FLIP,.) was found to block caspase-8 recruitment to the death-inducing signaling complex (DISC), and sensitization of cells by CHX was due to c-FLIPL down-modulation at the DISC level. Down-regulation of c-FLIPL with antisense oligonucleotides increased CD95-mediated apoptosis as in cells sensitized by CHX, demonstrating the direct involvement of c-FLIPL in apoptosis resistance. Removal of c-FLIPL block at DISC level allowed full activation of the mitochondrial pathway and, eventually, apoptosis in wildtype p53 cells, whereas in cells with mutated p53, c-FLIPL involvement in CD95-mediated apoptosis resistance appeared to be irrelevant. Immunohistochemical analysis of an ovarian tumor tissue array revealed c-FLIPL expression in samples with no p53 accumulation (P = 0.034), and a significant (P = 0.037) inverse relationship between c-FLIPL and p53 expression levels was also observed in 27 epithelial ovarian cancer specimens with known p53 status.Conclusion: The inhibitory protein c-FLIPL is involved in resistance to CD95-mediated apoptosis in ovarian carcinoma cells with wild-type p53.