Here, we report the fragment-based discovery of BI-9321, a potent, selective and cellular active antagonist of the NSD3-PWWP1 domain. The human NSD3 protein is encoded by the WHSC1L1 gene located in the 8p11-p12 amplicon, frequently amplified in breast and squamous lung cancer. Recently, it was demonstrated that the PWWP1 domain of NSD3 is required for the viability of acute myeloid leukemia cells. To further elucidate the relevance of NSD3 in cancer biology, we developed a chemical probe, BI-9321, targeting the methyl-lysine binding site of the PWWP1 domain with sub-micromolar in vitro activity and cellular target engagement at 1 µM. As a single agent, BI-9321 downregulates Myc messenger RNA expression and reduces proliferation in MOLM-13 cells. This first-in-class chemical probe BI-9321, together with the negative control BI-9466, will greatly facilitate the elucidation of the underexplored biological function of PWWP domains.
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Evaluating the use of absolute binding free energy in the fragment optimisation process
Communications Chemistry Open Access 05 September 2022
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The authors declare that the data supporting the findings of this study are available within the publication and its Supplementary Information files or have been deposited in the RCSB Protein Data Bank (PDB, http://www.rcsb.org) with the following accession numbers: selenomethionine labeled (6G3P); unlabeled (6G3T); X-ray structure of NSD3-PWWP1 in complex with compound 3 (6G24); X-ray structure of NSD3-PWWP1 in complex with compound 4 (6G25); X-ray structure of NSD3-PWWP1 in complex with compound 5 (6G27); X-ray structure of NSD3-PWWP1 in complex with compound 6 (6G29); X-ray structure of NSD3-PWWP1 in complex with compound 8 (6G2B); X-ray structure of NSD3-PWWP1 in complex with compound 9 (6G2C); X-ray structure of NSD3-PWWP1 in complex with compound 13 (6G2E); X-ray structure of NSD3-PWWP1 in complex with compound 16 (6G2F); X-ray structure of NSD3-PWWP1 in complex with compound BI-9321 (6G2O).
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The Structural Genomics Consortium is a registered charity (no. 1097737) that receives funds from AbbVie; Bayer Pharma AG; Boehringer Ingelheim; Canada Foundation for Innovation; Eshelman Institute for Innovation; Genome Canada; Innovative Medicines Initiative (EU/EFPIA) (ULTRA-DD grant no. 115766); Janssen; Merck & Co.; Novartis Pharma AG; Ontario Ministry of Economic Development and Innovation; Pfizer; São Paulo Research Foundation-FAPESP; Takeda and the Wellcome Trust. We thank the Expose team for data collection at the Swiss Light Source beamlines X06SA and X06DA. We thank D. Daniels, M. Robers and C. Corona from Promega for advising on the NanoBRET and target engagement assays, and acknowledge the Natural Sciences and Engineering Research Council of Canada (NSERC) for a postdoctoral fellowship awarded to D.D.
J.B., U.R., R.A.N., M. Petronczki, M.Z., N.M., K.R., A.Z., M.M., T.W., S.Z., H.A., H.B., C.M.R., A.H., T.K., M.C., B.S., S.W., D.H., X.-L.C., J.E.F., B.M., A.W.-P., T.G., G.B., M. Pearson and D.B.M. are employees of Boehringer Ingelheim.
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Böttcher, J., Dilworth, D., Reiser, U. et al. Fragment-based discovery of a chemical probe for the PWWP1 domain of NSD3. Nat Chem Biol 15, 822–829 (2019). https://doi.org/10.1038/s41589-019-0310-x
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