Development of Low-Nanomolar Covalent Epoxide Inhibitors of Tubulin Detyrosinating Enzymes VASH1&2

The development of small molecule inhibitors targeting the set of microtubules' post-translational modifications, also known as the "tubulin code", remains an underexplored area in medicinal chemistry. The recent discovery of the VASH1 and VASH2 enzymes, which are responsible for tubulin detyrosination─a modification associated with neurodegeneration and cancer─prompted us to develop new molecules that inhibit their activity. In this study, we conducted the first QSAR analysis of VASH proteases. Through rational substrate-based design, we identified our lead compound, LV87, as a potent epoxide-based covalent inhibitor of tubulin detyrosination <i>in cellulo</i>. Specificity assessments against other cysteine proteases and incubations with nonspecific nucleophiles demonstrated that LV87 selectively targets VASH1/2. Additionally, safety data, serum, and microsome stability tests support the notion that LV87 is a selective and potent inhibitor of tubulin detyrosination, paving the way for further optimization and applications.