Abstract
Abstract Our goal was to investigate the effects of the Novartis compound NVP-BEZ235, a dual PI3K/mTOR inhibitor, on hypoxia-inducible factor-1 alpha (HIF-1α) expression. NVP-BEZ235 was much more potent than either the mTOR inhibitor rapamycin or the PI3K inhibitor LY294002 in blocking the hypoxic induction of HIF-1α in vitro, due to decreased protein translation. 7-methyl GTP chromatography showed that NVP-BEZ235 led to a robust recruitment of 4E-BP1 to eIF4E and a near absence of binding of eIF4G, consistent with inhibition of protein translation. NVP-BEZ235 also decreased expression of several proteins regulated by eIF4E including cyclin B1 and D1, survivin, and vascular endothelial growth factor. NVP-BEZ235 specifically decreased eIF4G but not eIF4E expression. As HIF-1α has been associated with adaptation under hypoxia, we examined the effect of NVP-BEZ235 on cell survival in low pO2 conditions. The drug increased killing of cells under hypoxia as measured by both short-term (MTT) and long-term (clonogenic) assays. To understand the mechanism behind this observation, we performed immunoblotting for factors associated with cell survival. In normoxia, Akt S473 phosphorylation decreased within an hour of NVP-BEZ235 treatment, but then increased by 24 hours. In contrast, under hypoxia, NVP-BEZ235 caused a prolonged suppression of Akt phosphorylation. Furthermore, we found a greater increase in PARP cleavage in hypoxic cells treated 1-16 hours after drug treatment than in normoxic cells, consistent with increased apoptosis. This was confirmed using an ELISA assay that measures cytoplasmic oligonucleosomes. We also found that NVP-BEZ235 increases autophagy as measured by (i) LC3-I to LC3-II conversion on immunoblotting, (ii) degradation of p62 and (iii) GFP-LC3 subcellular localization detected by immunofluorescence. Treatment of cells with the drug under hypoxia further increased autophagy, although it is unclear at this point whether this actually contributes to cell death or is a pro-survival mechanism. In conclusion, we have found that NVP-BEZ235 blocks HIF-1α induction and increases cell killing under hypoxia, which may have important implications for the use of this drug in cancer therapy. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 2800. doi:1538-7445.AM2012-2800
Published Version
Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have
Schedule a call