Grant: Project Grant

In the past decade, we have witnessed a burst of knowledge about cellular processes that are deregulated in human cancer cells. One such biological process is the regulation of protein stability by ubiquitin, a small protein that is assembled to other proteins (so-called "ubiquitination") and signals for their destruction or functional changes. Among several protein families involved in this process, ubiquitin ligases (~600 in human) have attracted increasing attention as anti-cancer targets, as they catalyze the attachment of ubiquitin and therefore determine the specificity of ubiquitination. In particular, the HECT family members (28 in human) are deemed as promising targets due to the availability of an active site and other biochemical properties. However, existing inhibitory molecules of HECT ligases fall short in terms of potency, selectivity, and generally have had limited utility in probing unknown HECT mechanisms. To tackle this problem, we have used protein design and engineering approaches to systematically develop protein modulators for HECT ligases. We showed that the synthetic molecules could bind HECT ligases tightly on places other than the active site and were able to compromise or enhance their biological function. In this proposal, we will generate potent and specific non-active site inhibitors of selected oncogenic HECT ligases and validate their function in human cells. Additional biochemical and cellular analyses will provide information on molecular mechanisms by which HECT ligases function and further aid the design of chemical inhibitors as drug leads. In summary, this work will generate unique probes to understand and manipulate HECT ligases in normal and cancerous cells and will provide novel routes for cancer therapy.