Abstract 3793: Celastrol inhibits Hsp90 and triggers ER stress-induced cell death in non-small cell lung cancer cells

Abstract

Abstract Celastrol, a plant-derived triterpenoid, is a known inhibitor of Hsp90 and NF-kB activation pathways and has recently been suggested to be of therapeutic importance in various cancers. However, the molecular mechanisms of celastrol-mediated effects in lung cancer are not known. Inhibition of Hsp90, commonly linked with the suppression of multiple cell-survival pathways, is also associated with induction of maladaptive endoplasmic reticulum (ER)-stress response. Resistance to ER-stress response is essential for the growth and survival of cancer cells. In response to cellular stress from cytotoxic agents an initial protective ER-stress response is initiated within the cell with the goal of rescuing the cell; however, sustained ER stress overwhelms the cellular-protective mechanisms, triggering cell death through specific programmed cell death pathways. The role of ER-stress outcomes in mediating celastrol-induced cell death is not known. In this study, we have examined the response of two NSCLC cell lines to celastrol and explore the molecular mechanisms to determine the proteins governing the cellular responses. We observed that celastrol inhibited the proliferation of A549 and H1299 non-small lung carcinoma cells in a time- and dose-dependent manner as indexed by MTT assay. Evaluation of downstream targets of celastrol-induced effects in H1299 cells by immunoblotting revealed an increase in Hsp70 and Hsp90 protein expression indicating cellular stress, a hallmark response of inhibiton of Hsp90 function. Disruption of Hsp90 function has also been shown to inhibit TNFα-mediated NF-kB activation as was also observed in our studies where a 4 h celastrol pre-treatment of H1299 cells completely abrogated TNFα-induced NF-kB activation. Celastrol treatment also up regulated the expression of ER-stress chaperones Grp78, and pPERK. These changes in ER-stress mediators were paralleled by an increase in apoptotic response as evidenced by increased annexin-V/PI staining evaluated by FACS and immunoblotting which showed a time- and dose-dependent upregulation of the ER stress specific pro-apoptotic transcription factor, GADD153/CHOP, alteration of Bax/Bcl2 ratio and enhanced cleavage of PARP (a downstream effector of caspase-3 activation). Interestingly, however, celastrol treatment also induced the activation of pro-survival Erk1/2 and Akt pathways’, suggesting that celastrol concomitantly activates pro-survival and pro-apoptotic mechanisms, with the pro-apoptotic pathways taking precedence resulting in cell death. Together, our data demonstrate the chemotherapeutic potential of celastrol in lung cancer via the inhibition of Hsp90 function and induction of chronic ER-stress-mediated cell death. (Supported from KLCRP grant and Agnes Brown Duggan Endowment). Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 3793.