Abstract 2940: Identification of potent VCP/p97/CDC48 inhibitors with distinct biochemical mechanisms including a reversible, allosteric inhibitor that activates the unfolded protein response, induce autophagy and cancer cell death

Abstract

Abstract Valosin Containing Protein (VCP), also called p97 in mammals and cdc48 in yeast, is an ubiquitously expressed and essential AAA ATPase important in specific cellular processes including Endoplasmic Reticulum Associated Degradation (ERAD), Golgi reformation, membrane fusion and autophagy. VCP is a hexameric complex formed by six identical protomers, each composed of three domains: an N-terminal domain responsible for the interaction with co-factors and adaptor proteins, and two AAA ATPase domains, D1 and D2. VCP acts as a protein-directed molecular machine that converts energy derived from ATP hydrolysis into mechanical force to cause disassembly of multiprotein complexes or extraction of molecules from the membrane to be delivered to the proteasome for degradation. VCP also affects autophagy and aggresome formation processes. Overall, VCP plays a key role in cellular homeostasis. The clinical success of proteasome inhibitors, and recent advances in the preclinical development of molecules that interfere with protein folding and degradation has highlighted that cancer cells can be extremely sensitive to perturbation of protein homeostasis. The availability of small molecules that specifically inhibit VCP function would help to clarify whether VCP is a valid target for cancer therapy. We found that VCP silencing by siRNA induces cancer cell death in a variety of tumor cell lines, and examined the main cellular pathways modulated upon VCP ablation, thus identifying biomarkers suitable for characterizing the cellular activity of VCP inhibitors. We then performed a High Throughput Screening campaign using recombinant VCP and identified multiple compound classes that inhibit VCP function with distinct biochemical mechanisms. Initial hits included a compound that covalently modifies VCP, representatives of two classes of ATP-sensitive inhibitors, and an inhibitor class characterized by a novel allosteric mechanism of action. Chemical expansion of initial hits resulted in improvement of biochemical potency, yielding highly active derivatives for each class, suggesting that VCP is a druggable target. Notably, the binding site of the allosteric class of inhibitors was identified by photo-affinity labeling in combination with available structural data. We will present data on a potent (30 nM) and specific compound emerging from this class that displays anti-proliferative activity related to the modulation of direct VCP biomarkers, activation of the Unfolded Protein Response (UPR) and perturbation of autophagy, which ultimately result in cancer cell death. 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 2940. doi:1538-7445.AM2012-2940