P74

Abstract

The Hsp70 chaperone is one of the major components of tumor microenvironment displaying multiple not yet established functions. It was shown to stimulate innate and adaptive anti-tumor immunity in a variety of cancer models. These effects were due to active release of endogenous Hsp70 to an extracellular matrix and therefore the extracellular chaperone can be a trigger of multiple events in the whole tumor. Factors inducing Hsp70 release are heat stress, inhibitors of important signaling proteins, anticancer drugs and X-ray treatment. Recently we have shown that delivery of Hsp70 can be efficient pusher of its intracellular analogue to extracellular milieu. In this study we show that intracellular cycling of exo- and endogenous Hsp70s increases the sensitivity of cancer cells to cytotoxic lymphocytes. Moreover, the results of in vivo studies employing B16 mouse melanoma and C6 rat glioblastoma as targets proved the therapeutic relevance of exogenous Hsp70 in intra-tumoral application. To uncover the mechanisms of the anticancer effect we used inhibitors and markers of intra- and extracellular protein transport. The data of these studies showed multiplicity of pathways using which exo-Hsp70 reaches cytosol and more usable ones was endocytosis. To be exported intracellular Hsp70 employs vesicular structures as well as intra-membrane lipid structures. Analyzing Hsp70 molecule we found the domain with potential vector activity, i.g. cell penetrating function. This peptide was synthesized and shown to cross a cellular membrane with efficacy exceeding that the whole Hsp70 molecule. Furthermore, the new peptide was used as a carrier for the delivery inside living cells antibody to Hsp70. The resulting construct was found to reduce the resistance of tumor cells to pro-apoptotic effect of staurosporin. Taking together these data, we can suggest that Hsp70 and its fragments can be effective players in communication between cells assembling functionally active tumor. The work was supported by Grant of Russian Scientific Foundation (N 14-50-00068).