Abstract 395: Effect of pantoprazole to enhance activity of docetaxel against human tumor xenografts by inhibiting autophagy

Abstract

Abstract Background: Autophagy allows recycling of cellular components and may facilitate cell survival after chemotherapy. Pantoprazole inhibits proton pumps, including that maintaining low pH in endosomes, and is reported to inhibit autophagy (1). Here we evaluate effects of pantoprazole to modify cytotoxicity of the anticancer drug docetaxel, and underlying mechanisms. Methods: Effects of docetaxel+/-pantoprazole were studied against wild-type and autophagy-deficient cultured PC3 cells derived by shRNA, and against four human xenografts. Effects of pantoprazole on autophagy in cultured cells were evaluated by quantifying LC3-I, LC3-II and p62 proteins in Western blots, and by fluorescent microscopy of cells transfected with the tandem sensor RFP-GFP-LC3. Since autophagy is known to be up-regulated in poorly-nourished tumor regions (2), the distribution of drug effects and of autophagy was quantified in tumor sections in relation to blood vessels and hypoxia by immunohistochemistry (IHC) using γH2AX, a marker of DNA damage, cleaved caspase-3, a marker of apoptosis, Ki67, a marker of proliferation and LC3 and p62 to quantify autophagy. Results: Pantoprazole increased toxicity of docetaxel for cultured cells, increased docetaxel-induced expression of γH2AX and cleaved caspase-3 and decreased Ki67 in tumor sections. Pantoprazole increased growth-delay of four human xenografts of low, moderate and high sensitivity to docetaxel, with minimal increase in toxicity. Docetaxel led to increased autophagy throughout tumor sections. Pantoprazole inhibited autophagy, and effects of pantoprazole were reduced against genetically-modified cells with decreased ability to undergo autophagy. Conclusions: Autophagy is a mechanism of resistance to docetaxel chemotherapy that may be modified by pantoprazole to improve therapeutic index. High-dose pantoprazole increases the therapeutic effectiveness of docetaxel in vitro and in vivo, and its main mechanism of action is via inhibition of autophagy. With this rationale, we are undertaking a phase II study of pantoprazole + docetaxel in men with castrate-resistant prostate cancer.