biostudies-literatureLu WAmerican Chemical SocietyNational Cancer InstituteNCI NIH HHSWelch Foundation3356-3371https://www.ebi.ac.uk/biostudies/studies/S-EPMC11190964biostudies-literatureUnknown66(5)The c-Jun N-terminal kinases (JNKs) are members of the mitogen-activated protein kinase (MAPK) family, which includes JNK1-JNK3. Interestingly, JNK1 and JNK2 show opposing functions, with JNK2 activity favoring cell survival and JNK1 stimulating apoptosis. Isoform-selective small molecule inhibitors of JNK1 or JNK2 would be useful as pharmacological probes but have been difficult to develop due to the similarity of their ATP binding pockets. Here, we describe the discovery of a covalent inhibitor YL5084, the first such inhibitor that displays selectivity for JNK2 over JNK1. We demonstrated that YL5084 forms a covalent bond with Cys116 of JNK2, exhibits a 20-fold higher <i>K</i>_inact/<i>K</i>_I compared to that of JNK1, and engages JNK2 in cells. However, YL5084 exhibitJournal of medicinal chemistryDevelopment of a Covalent Inhibitor of c-Jun N-Terminal Protein Kinase (JNK) 2/3 with Selectivity over JNK1.PMC11190964I-1829P50 CA070907RSG-18-039-01-DMCP50CA07090720Geng QLu WSinha DTse JChe JMarto JALi LAnderson KCLiu YGurbani DYou IWen KZhang TGray NSMeyer CJHe ZJi WGao YFicarro SBYu TKim AYWestover KDfalseDevelopment of a Covalent Inhibitor of c-Jun N-Terminal Protein Kinase (JNK) 2/3 with Selectivity over JNK1.The c-Jun N-terminal kinases (JNKs) are members of the mitogen-activated protein kinase (MAPK) family, which includes JNK1-JNK3. Interestingly, JNK1 and JNK2 show opposing functions, with JNK2 activity favoring cell survival and JNK1 stimulating apoptosis. Isoform-selective small molecule inhibitors of JNK1 or JNK2 would be useful as pharmacological probes but have been difficult to develop due to the similarity of their ATP binding pockets. Here, we describe the discovery of a covalent inhibitor YL5084, the first such inhibitor that displays selectivity for JNK2 over JNK1. We demonstrated that YL5084 forms a covalent bond with Cys116 of JNK2, exhibits a 20-fold higher <i>K</i>_inact/<i>K</i>_I compared to that of JNK1, and engages JNK2 in cells. However, YL5084 exhibit2023-01-01T00:00:00Z2023 Mar2025-04-20T02:17:01.764Z2025-04-20T02:17:01.764ZS-EPMC111909643682683310.1021/acs.jmedchem.2c01834