Abstract 2111: Regulation and localization of the cleaved form of PAR-4 in ovarian and endometrial cancers

Abstract

Abstract Prostate Apoptosis Response-4 (PAR-4) is a tumor suppressor protein whose expression level in cancer is frequently decreased; however, it is neither mutated nor suppressed. A unique feature of PAR-4 is that it can induce apoptosis selectively in cancer cells without destroying normal cells, giving it an interest in anti-cancer targeted therapy. PAR-4 is not only regulated at the expression level, but also post-translationally by phosphorylation and protein cleavage. Most recently, the caspase-3 cleavage of PAR-4 has been demonstrated by our laboratory and this cleaved fragment might be responsible for PAR–induced apoptosis. In the present study, we have investigated the mechanisms regulating PAR-4 expression/activity and its cleaved form in ovarian and endometrial cancer cell lines. Briefly, we used chemo-sensitive and resistant cancer cell lines in which we produced stable clones expressing the cleaved form of PAR-4 (cl.PAR-4) using lentiviral particles. Using these models, we studied the regulation of cl.PAR-4 at the protein and genomic level. Treated with cisplatin, sensitive cancer cells showed a decrease of PAR-4 and an important increase of endogenous cl.PAR-4. Using our stable clones, we observed that upon cisplatin treatment, the normally constitutively expressed cl.PAR-4 protein level was increased while the cDNA transgene was constitutively expressed, indicating a possible involvement of post-translational mechanisms. Our results indicate that degradation of our protein by the proteasome could be implicated because of the significant increase of cl.PAR-4 in the presence of a proteasome inhibitor (MG-132). PI3k could also be linked to this regulation because of an increase of cl.PAR-4 in the presence of PI3k inhibitors (LY294002, Wortmannin and BEZ235). Conversely, when using MAPK inhibitors (U0126 and PD98059), cl.PAR-4 was negatively regulated. We also investigated the localization of cl.PAR-4 using cytoplasmic/nuclear fractionationnation and immunofluorescence. The results indicated that cl.PAR-4 is mainly localized to the cytoplasm while being weakly present in the nucleus. Cisplatin treatment does not seem to influence its localization. This nuclear localization of cl.PAR-4 may have a role in its ability to induce apoptosis. Finally, bioinformatic analyses suggests possible ubiquitination and phosphorylation sites responsible for the regulation of cl.PAR-4. Further analyses of the post-translational mechanisms involved in cl.PAR-4 activity are still needed to better understand the regulation of this potential tumor suppressor. By better understanding the mechanisms of PAR-4 and its cleaved form, we may be able to target this protein to sensitize chemoresistant gynecological cancer cells to chemotherapy. Citation Format: Kevin Brasseur, Valérie Leblanc, Sophie Parent, Éric Asselin. Regulation and localization of the cleaved form of PAR-4 in ovarian and endometrial cancers. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 2111. doi:10.1158/1538-7445.AM2015-2111