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Förster, Florian (2014): Targeting the actin cytoskeleton with natural compounds: elucidating the underlying mechanism of apoptosis induction. Dissertation, LMU München: Fakultät für Chemie und Pharmazie



Targeting the cytoskeleton (CSK) of cancer cells offers a valuable strategy in cancer therapy. Whereas drugs which address microtubule CSK such as vinca alkaloids or taxanes are well established in the clinic, compounds binding to the actin CSK are still far away from their therapeutical application. One reason might be the lacking knowledge on their mode of cytotoxicity and moreover their tumor specific mechanism of action. We used the myxobacterial compound Chondramide as a tool to first elucidate the mechanisms of cytotoxicity by actin targeting in different breast cancer cells, namely MCF7 and MDA-MB-231. Chondramide inhibits actin filament assembly and dynamics shown by a fluorescence-based analysis (FRAP) in whole cells and leads to apoptosis characterized by phosphatidylserine exposure, release of cytochrome C from mitochondria and finally activation of caspases (-9 and -3). Detailed analysis revealed, that Chondramide induces apoptosis by enhancing the occurrence of mitochondrial permeability transition (MPT). Known MPT-modulators were found to be affected by Chondramide: Hexokinase II (HkII) bound to the voltage dependent anion channel (VDAC) translocated from the outer mitochondrial membrane to the cytosol and the proapoptotic protein Bad was recruited to the mitochondria. Importantly, PKCε, a prosurvival serine/threonine kinase possessing an actin-binding site and known to regulate the HkII/VDAC interaction as well as Bad phosphoylation was identified as the link between actin CSK and apoptosis induction. PKCε which was found overexpressed in breast cancer cells accumulated in actin bundles induced by Chondramide and lost its activity. The second goal of our work was to inform on a potential tumor specific action of actin binding agents such as Chondramide. As the nontumor breast epithelial cell line MCF-10A in fact shows resistance to Chondramide induced apoptosis and notably express very low level of PKCε we claim that trapping PKCε via Chondramide induced actin hyperpolymerization displays tumor cell specificity. Our work provides a link between targeting the ubiquitously occurring actin CSK and selective inhibition of pro-tumorigenic PKCε, thus setting the stage for actin-stabilizing agents as innovative cancer drugs. This is moreover supported by the in vivo efficacy of Chondramide triggered by abrogation of PKCε signaling shown in a xenograft breast cancer model. For the actin targeting compound Doliculide we could show that Doliculide impairs the dynamics of the actin CSK similar to Chondramide. Moreover, it reduces the proliferation rate and migration of cancer cells and also leads to the induction of apoptosis, thus Doliculide is also an interesting lead structure for further preclinical investigations.