Philippe Guedat

Small-molecule inhibitor of USP7/HAUSP ubiquitin protease stabilizes and activates p53 in cells

Deregulation of the ubiquitin/proteasome system has been implicated in the pathogenesis of many human diseases, including cancer. Ubiquitin-specific proteases (USP) are cysteine proteases involved in the deubiquitination of protein substrates. Functional connections between USP7 and essential viral proteins and oncogenic pathways, such as the p53/Mdm2 and phosphatidylinositol 3-kinase/protein kinase B networks, strongly suggest that the targeting of USP7 with small-molecule inhibitors may be useful for the treatment of cancers and viral diseases. Using high-throughput screening, we have discovered HBX 41,108, a small-molecule compound that inhibits USP7 deubiquitinating activity with an IC50 in the submicromolar range. Kinetics data indicate an uncompetitive reversible inhibition mechanism. HBX 41,108 was shown to affect USP7-mediated p53 deubiquitination in vitro and in cells. As RNA interference-mediated USP7 silencing in cancer cells, HBX 41,108 treatment stabilized p53, activated the transcription of a p53 target gene without inducing genotoxic stress, and inhibited cancer cell growth. Finally, HBX 41,108 induced p53-dependent apoptosis as shown in p53 wild-type and null isogenic cancer cell lines. We thus report the identification of the first lead-like inhibitor against USP7, providing a structural basis for the development of new anticancer drugs.[Mol Cancer Ther 2009;8(8):2286–95]

Patented small molecule inhibitors in the ubiquitin proteasome system

Deregulation of the ubiquitin proteasome system (UPS) has been implicated in the pathogenesis of many human diseases, including cancer and neurodegenerative disorders. The recent approval of the proteasome inhibitor Velcade® (bortezomib) for the treatment of multiple myeloma and mantle cell lymphoma establishes this system as a valid target for cancer treatment. We review here new patented proteasome inhibitors and patented small molecule inhibitors targeting more specific UPS components, such as E3 ubiquitin ligases and deubiquitylating enzymes.Publication history: Republished from Current BioDatas Targeted Proteins database (TPdb; http://www.targetedproteinsdb.com).

Synthesis and Biological Evaluation of 9-Oxo-9H-indeno[1,2-b]pyrazine-2,3-dicarbonitrile Analogues as Potential Inhibitors of Deubiquitinating Enzymes

High‐throughput screening highlighted 9‐oxo‐9H‐indeno[1,2‐b]pyrazine‐2,3‐dicarbonitrile (1) as an active inhibitor of ubiquitin‐specific proteases (USPs), a family of hydrolytic enzymes involved in the removal of ubiquitin from protein substrates. The chemical behavior of compound 1 was examined. Moreover, the synthesis and in vitro evaluation of new compounds, analogues of 1, led to the identification of potent and selective inhibitors of the deubiquitinating enzyme USP8.

Abstract 2642: Identification and characterization of selective inhibitors of ubiquitin specific protease 7

Regulated protein turnover is primarily controlled by the ubiquitin-proteasome system. The only marketed drug related to the ubiquitin-proteasome system, bortezomib, is acting as a proteasome inhibitor and has been approved for the treatment of some hematological cancers. Targeting the upstream ubiquitin conjugation/deconjugation system carries out promises of therapeutics with increased specificity and selectivity. Ubiquitin-specific proteases (USP) are involved in the deubiquitination of specific target substrates regulating their stability, subcellular localization and/or activation status. USP represent a drugable target class due to their thiol-protease catalytic core which is amenable to pharmacological inhibition by small molecules. A genome-wide RNAi screen of the catalytically active human USPs in cancer-relevant cellular models and phenotypic assays allowed us to identify USP7/HAUSP as promising cancer target. Fluorescence-based screening assays using optimized USP substrates including various ubiquitin derivatives (ubiquitin precursor, branched ubiquitin chains) as well as specific, physiological substrates were developed. High-throughput screening performed on our chemically diverse library followed by different optimization programs resulted in the discovery of several series as novel USP7 inhibitors. Our progress made towards the specificity issue will be presented here with the identification of the first USP7-specific series. These will help further validate this novel class of molecular targets and may provide a structural basis for the development of new anticancer drugs. Note: This abstract was not presented at the AACR 101st Annual Meeting 2010 because the presenter was unable to attend. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 2642.

Abstract B193: Identification of selective inhibitors of ubiquitin specific proteases

Regulated protein turnover is primarily controlled by the ubiquitin‐proteasome system. The therapeutic efficacy of the proteasome inhibitor Bortezomib establishes this system as a valid anticancer therapeutic field. A promising alternative to targeting the proteasome itself lies upstream, at the level of the ubiquitin conjugation/deconjugation step to generate more specific anticancer agents. Ubiquitin‐specific proteases (USP) are involved in the deubiquitination of specific target substrates regulating their stability, subcellular localization and/or activation status. USP represent a druggable target class due to their thiol‐protease catalytic core which is amenable to pharmacological inhibition by small molecules. A genome‐wide RNAi screen of the catalytically active human USPs in cancer‐relevant cellular models and phenotypic assays allowed us to identify USP7/HAUSP and USP8/UBPY as promising cancer targets. Fluorescence‐based screening assays using optimized USP substrates including various ubiquitin derivatives (ubiquitin precursor, branched ubiquitin chains) as well as specific, physiological substrates were developed. High‐throughput screening performed on our chemically diverse library followed by different optimization programs resulted in the discovery of several series as novel USP inhibitors. Our progress made towards the specificity issue will be presented here with the identification of the first USP8‐ and USP7‐specific series. Citation Information: Mol Cancer Ther 2009;8(12 Suppl):B193.