Targeting KRASG13C: Exploring Warhead Orientation with Cyclic Linker-Based Inhibitors

The small GTPase KRAS is a key driver of carcinogenesis when mutated, and significant progress has been made in targeting KRAS ^G12C and other oncogenic variants. Building on our previous work demonstrating the potential of nucleotide-based inhibitors with an acrylamide warhead to target KRAS ^G13C , we designed and synthesized a library of nucleotide-based compounds with cyclic linkers to explore the effect of warhead orientation on reactivity toward Cys13. Using mass spectrometry, kinetic studies, and protein X-ray crystallography, we validated the binding and reactivity of these modulators. In addition, computational predictions of the conformational space of the linkers and warheads provided insights into their reactivity, which agreed well with the experimental data. These findings advance our understanding of the structure-reactivity relationship in these nucleotide-based KRAS inhibitors and will be the basis for further optimization.