Discovery of potent degraders of pan-KRAS based on a novel KRAS binder.

Abstract

3113 Background: As the most frequently mutated oncogene, KRAS alteration occurring in approximately 25% of all malignancies. Despite extensive efforts, targeting KRAS has proven to be challenging due to its structure and complex function. Recently, direct KRAS G12C inhibitors, such as Sotorasib and Adagrasib have occurred first successes. However, therapeutic approaches targeting other variants beyond G12C are under significant unmet needs. Recent developments in protein degradation technologies, such as Proteolysis-Targeting Chimeras (PROTACs), bring new hope for targeting pan KRAS. Based on our novel warheads, potent pan-KRAS degraders were designed and synthesized. Methods: The ability of compounds to degrade KRAS protein was evaluated using western blotting. The anti-tumor efficacy was assessed in vitro through different mutant cell lines. As a proof of concept study in vivo , an experiment in subcutaneous xenograft mouse model was performed. Pharmacokinetic studies were conducted in mice, with serial blood samples analyzed by (LC−MS)/MS. Results: A novel series of warheads exhibiting exceptional enzymatic and cellular activity against pan-KRAS has been successfully obtained. Based on these warheads, more than 100 degraders were meticulously designed and synthesized incorporating a diverse array of linkers and E3 ligands. Through a comprehensive evaluation, two series of compounds have demonstrated a remarkable KRAS degradation property and downstream inhibition at concentrations below 10 nM in SW620 G12V and GP2D G12D cells. Cell proliferation assay demonstrated that the IC 50 values of these degraders are ranging from 0.01 to 30 nM in the MIA PaCa-2 G12C , GP2D G12D SW620 G12V and LOVO G13D cell lines, without affecting the viability of KRAS-independent cell lines (selectivity > 500-fold). These compounds also possess favorable PK properties in mice (clearance < 10 mL/min/kg; IV, 2 mpk, AUC > 5000 ng·hr/mL) and good safety profile (hERG IC 50 > 30 µM). Moreover, these compounds showed strong antitumor activity in xenograft mouse model in vivo. Conclusions: The innovative linker elongation and branching, coupled with modifications of KRAS binder portion significantly contributed to potent pan-KRAS degraders, which demonstrate excellent pharmacokinetics and exhibit remarkable efficacy both in vitro and in vivo. The IND-enabling studies are being conducted and the regulatory IND filing will be completed in 2025.