Exquisitely Specific anti-KRAS Biodegraders Inform on the Cellular Prevalence of Nucleotide-Loaded States

Mutations to RAS proteins (H-, N-, and K-RAS) are among the most common oncogenic drivers, and tumors harboring these lesions are some of the most difficult to treat. Although covalent small molecules against KRAS(G12C) have shown promising efficacy against lung cancers, traditional barriers remain for drugging the more prevalent KRAS(G12D) and KRAS(G12V) mutants. Targeted degradation has emerged as an attractive alternative approach, but for KRAS, identification of the required high-affinity ligands continues to be a challenge. Another significant hurdle is the discovery of a hybrid molecule that appends an E3 ligase-recruiting moiety in a manner that satisfies the precise geometries required for productive polyubiquitin transfer while maintaining favorable druglike properties. To gain insights into the advantages and feasibility of KRAS targeted degradation, we applied a protein-based degrader (biodegrader) approach. This workflow centers on the intracellular expression of a chimeric protein consisting of a high-affinity target-binding domain fused to an engineered E3 ligase adapter. A series of anti-RAS biodegraders spanning different RAS isoform/nucleotide-state specificities and leveraging different E3 ligases provided definitive evidence for RAS degradability. Further, these established that the functional consequences of KRAS degradation are context dependent. Of broader significance, using the exquisite degradation specificity that biodegraders can possess, we demonstrated how this technology can be applied to answer questions that other approaches cannot. Specifically, application of the GDP-state specific degrader uncovered the relative prevalence of the off-state of WT and various KRAS mutants in the cellular context. Finally, if delivery challenges can be addressed, anti-RAS biodegraders will be exciting candidates for clinical development.

Automated summary

Mutations in RAS proteins are common drivers of cancer, with some mutations being particularly difficult to treat. Targeted degradation has emerged as a promising alternative approach, but challenges remain in finding high-affinity ligands for KRAS and in discovering hybrid molecules that can effectively degrade the protein.