Ruibin Liu, Zhi Yue, Cheng-Chieh Tsai, and Jana Shen
J. Am. Chem. Soc., 2019
DOI: 10.1021/jacs.8b13248
Targeted covalent inhibitor design is gaining increasing interest and acceptance. A typical covalent kinase inhibitor design targets a reactive cysteine; however, this strategy is limited due to the low abundance of cysteine and acquired drug resistance from point mutations. Inspired by the recent development of lysine-targeted chemical probes, we asked if nucleophilic (reactive) catalytic lysines are common based on the published crystal structures of the human kinome. Using a newly developed pKa prediction tool based on continuous constant pH molecular dynamics, the catalytic lysines of 8 unique kinases from various human kinase groups were retro- and prospectively predicted to be nucleophilic, when kinase is in the rare DFG-in/aC-out type of conformation. Importantly, other reactive lysines as well as cysteines at various locations were also identified. Based on the finding, we proposed a new strategy based on modification of selective type II reversible kinase inhibitors to discover highly selective, lysine-targeted covalent inhibitors. Traditional covalent drugs were discovered serendipitously; the presented tool, which can assess the reactivities of any potentially targetable residues, may assist and accelerate the rational discovery of new covalent inhibitors. Another significant finding of the work is that lysines and cysteines in kinases may adopt neutral and charged states at physiological pH, respectively. This finding may shift the current paradigm of computational studies of kinases, which assume standard protonation states.
Linking of fragments in neighboring binding sites is one of the optimization strategies in fragment-based drug discovery, where additive or even more substantial bioactivity improvements can be realized. However, such efforts present a considerable challenge when one fragment binds covalently to the target protein, as small modifications can influence the correct positioning of the covalent warhead toward the targeted nucleophilic residue. Here, we present a case study of fragment linking that yielded single-digit micromolar, covalent inhibitors of the SARS-CoV-2 main protease, starting from fragments that were inactive in the biochemical assay. Using structural information from a recent, high-throughput crystallographic fragment screen, we show that the success of fragment linking in the design of targeted covalent inhibitors is heavily impacted by several factors, including the warhead type, the labeling chemistry, and even subtle changes in the designed linker. Notably, we observe that induced fit effects might override the original fragment orientations in the linked molecule, highlighting the need for reliable structure verification, especially in consecutive rounds of fragment elaboration.
Levente Kollár, Levente M. Mihalovits, Dávid Bajusz, DamijanKnez, József Simon, Blake H. Balcomb, Daren Fearon, Stanislav Gobec, György M. K...
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Design, synthesis and biological evaluation of the activity-based probes for FGFR covalent inhibitorDandan Zhu, Zijian Zheng, Huixin Huang, Xiaojuan Chen, Shuhong Zhang, Zhuchu Chen, Ting Liu, Guangyu Xu, Ying Fu, Yongheng Chen, European Jo...
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Yoav Shamir, Nir London bioRxiv 2025.03.19.642201 doi: https://doi.org/10.1101/2025.03.19.642201 Recent years have seen an explosion in the...
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DOI Ansgar Oberheide, Maxime van den Oetelaar, Jakob Scheele, Jan Borggräfe, Semmy Engelen, Michael Sattler, Christian Ottmann, ...
