Abstract 2609: Tissue transglutaminase (TG2) promotes resistance to HDAC inhibitor (HDI) vorinostat in cancer cells

Abstract Vorinostat has shown preclinical and clinical effects in human cancers and it is the first HDI approved by FDA for cutaneous T-cell-lymphoma treatment. TG2 is a multifunctional enzyme that catalyze a Ca2+-dependent transamidating reaction resulting in covalent cross-links between proteins. TG2 may also act as a G-protein in transmembrane signalling, as well as a cell surface adhesion mediator. TG2 upregulation has been demonstrated in several cancers and its expression levels correlate with resistance to chemotherapy and metastatic potential. In the present study, we demonstrated that the antiproliferative effects of vorinostat is paralleled by the induction of both TG2 mRNA and protein expression in breast, colorectal and oral cancer cell lines. This effect was also shared by other pan-HDIs and by class I HDIs but not by the specific inhibitor of HDAC-6. Vorinostat-induced TG2 upregulation correlated with an increased transamidating activity. Confocal microscopy analysis confirmed TG2 induction as well as TG2 cytosolic aggregation upon vorinostat treatment. Compartment analysis of TG2 protein expression revealed that vorinostat induces cytosolic, membrane and cytoskeleton but not nuclear upregulation of TG2. Apparently this effect seems confined to tumor cells, since same results were not observed in ex vivo vorinostat-treated peripheral blood lymphocytes from healthy donors. Notably high basal levels of TG2 protein correlated with lower antiproliferative activity of vorinostat. In order to confirm that TG2 play a role in vorinostat antitumor effect, we silenced by specific shRNA TG2 expression in high TG2 expressing HT29 colorectal cancer cells, and overexpressed full length TG2 in low TG2 expressing MCF7 breast cancer cells. We demonstrated that in TG2 silenced cells vorinostat antiproliferative and proapoptotic effects were enhanced whereas in TG2 overexpressed cells they were impaired, demonstrating that TG2 is directly involved in the mechanism of antitumor effect exerted by vorinostat. Moreover cotreatment of tumor cells with two specific inhibitors of TG2 transamidating activity (KCC009 and monodansylcadaverine) potentiated the antitumor effect of vorinostat, suggesting that TG2 cross-linking activity is important for vorinostat antitumor effect. Vorinostat also induced TG2 acetylation, however this posttranslational modification has no relevant effect on protein half-life as demonstrated by cotreatment with cycloheximide. Finally, vorinostat-resistant MCF-7 cell line selected by stepwise increasing concentrations of vorinostat up to 12 μM, significantly overexpressed TG2 protein compared with parental control cell line. Overall this study demonstrated that TG2 overexpression is a common mechanism of intrinsic or acquired resistance to vorinostat and that inhibition of TG2 transamidating activity may potentiate vorinostat antitumor effect. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 2609. doi:10.1158/1538-7445.AM2011-2609