CD95-mediated apoptosis is impaired at receptor level by cellular FLICE-inhibitory protein (long form) in wild-type p53 human ovarian carcinoma

Delia Mezzanzanica, Emanuela Balladore, Fabio Turatti, Elena Luison, Paola Alberti, Marina Bagnoli, Mariangela Figini, Alessandra Mazzoni, Francesco Raspagliesi, Maria Oggionni, Silvana Pilotti, Silvana Canevari

Research output: Contribution to journalArticle

Abstract

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-FLIPL) 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 mhochondrial pathway and, eventually, apoptosis in wild-type 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-FLIP L is involved in resistance to CD95-mediated apoptosis in ovarian carcinoma cells with wild-type p53.

Original languageEnglish
Pages (from-to)5202-5214
Number of pages13
JournalClinical Cancer Research
Volume10
Issue number15
DOIs
Publication statusPublished - Aug 1 2004

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ASJC Scopus subject areas

  • Cancer Research
  • Oncology

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