1S. M. Rowe, A. Price, D. J. Murphy, J. Lin, E. N. Nartey, A. Chaikuad, K. Wong, J. E. Cottom, N. O. Concha, R. A. Reid, E. R. Dickinson, M. Jundt, K. Kammerer, M. Steidel, T. Mathieson, T. Werner, E. K. Grant, C. K. Stanborough, M. Rouah, J. Wojno-Picon, P. Pogány, J. Pettinger, D. J. Norman, H. Wilders, F. Rianjongdee, G. Valdes-Garcia, N. Nevins, R. Shenje, R. K. Thalji, C. Chung, H. C. Eberl, G. Neubauer, D. House, Y. Rao, M. P. Martino and J. T. Bush,
ChemRxiv, 2025
https://doi.org/10.26434/chemrxiv-2025-tvdzn
We report a ‘direct-to-biology’ (D2B) approach for optimising covalent acrylamide binders of protein targets and apply this to the identification of a selective and cell-active inhibitor of Werner (WRN) helicase. Inhibition of WRN helicase activity exhibits a synthetic lethal relationship with cancers displaying high microsatellite instability (MSI-H) and is being pursued as a therapeutic strategy in the clinic. Using intact-protein liquid chromatography-mass spectrometry (LC-MS) screening, we identified acrylamide fragment binders of the WRN helicase domain and then used covalent D2B chemistry to optimise these initial hits. Our efforts ultimately afforded a potent covalent inhibitor of WRN-mediated DNA unwinding, which displays selective, concentration-dependent cellular engagement of WRN, and demonstrates synthetic lethality in an MSI-H setting. Furthermore, our inhibitor targets a distinct conformation of WRN helicase compared to the current clinical covalent inhibitor, presenting a complementary approach for covalent inhibition of WRN helicase. This work demonstrates how D2B chemistry platforms can be used to explore structure-activity relationships in a modular fashion, while reducing investment of human and material resources.
