Abstract B025: The RAF/MEK clamp avutometinib (VS-6766) enhances antitumor efficacy of KRAS G12C and G12D inhibitors through vertical inhibition of RAS, RAF and MEK. &lt;/title&gt;
Authors: Coma Silvia , Musteanu Monica , Caffarra Cristina , Mira Alessia , Patrucco Enrico , Dilly Julien , Aguirre Andrew J. , Barbacid Mariano , Ambrogio Chiara , Pachter Jonathan A. 
Published: May 2023

Abstract
KRAS is the most frequently mutated RAS family member. Within non-small cell lung cancer (NSCLC), G12C is the most prevalent KRAS mutation (~13%) and the KRAS G12C inhibitors (G12Ci) sotorasib and adagrasib are now FDA approved. However, it has been shown that simultaneous targeting of multiple nodes in the MAPK pathway may be optimal for deeper and more durable responses. Furthermore, acquired mutations in the MAPK pathway occur clinically upon progression on sotorasib or adagrasib, altogether supporting the need for clinical combinations with G12Ci. Based on the clinical success of G12Ci, several KRAS G12D inhibitors (G12Di) are now being developed as G12D is the most prevalent KRAS mutation across human cancers including pancreatic (~28%) and colorectal (~11%) cancers. Avutometinib (VS-6766; avuto) is a RAF/MEK clamp that potently inhibits MEK kinase activity and induces a dominant negative RAF-MEK complex preventing phosphorylation of MEK by ARAF, BRAF and CRAF. This unique mechanism allows avuto to block MEK signaling without the compensatory re-activation of MEK that appears to limit the efficacy of MEK-only inhibitors. Here, we tested whether addition of avuto to G12Ci or G12Di could improve MAPK pathway blockade and anti-tumor efficacy. Avuto was synergistic with sotorasib and adagrasib in reducing viability of a panel of KRAS G12C cancer cell lines in 3D culture, and the combination of avuto + G12Ci showed improved depth and duration of MAPK pathway inhibition, stronger inhibition of cell cycle/proliferation markers and stronger activation of pro-apoptotic markers relative to G12Ci alone. In mouse models, avuto enhanced efficacy of G12Ci in KRAS G12C NSCLC xenograft models. Furthermore, avuto potentiated G12Ci efficacy in a KRAS(+/G12C);p53null NSCLC GEMM model with avuto + G12Ci combination inducing complete response in 25% of the mice. To evaluate efficacy of avuto against KRAS mutations that have been observed clinically upon acquired resistance to G12Ci, proliferation of Ba/F3 cells expressing these mutations was assessed. While avuto potently inhibited (IC50 &lt; 15 nM) proliferation of cells bearing KRAS G12C/R68S, G12C/H95D or G12C/Y96C mutations, adagrasib showed diminished potency (IC50 ≥ 85 nM) against all 3 of these mutations and sotorasib showed diminished potency (IC50 ≥ 150 nM) against G12C/R68S and G12C/Y96C. Accordingly, avuto strongly inhibited tumor growth of an in vivo model expressing KRAS G12C/Y96D, while sotorasib was ineffective. Similarly, avuto was found to be synergistic with the G12Di MRTX1133 in reducing viability of a panel of KRAS G12D cancer cell lines, and avuto enhanced anti-tumor efficacy of MRTX1133 in KRAS G12D colorectal (CR3300) and pancreatic (PA1252) cancer PDX models. These results support ongoing clinical studies of avutometinib in combination with sotorasib (NCT05074810) and adagrasib (NCT05375994) for patients with KRAS G12C NSCLC. Additionally, these preclinical data support combination of avutometinib with a G12D inhibitor for patients with KRAS G12D colorectal or pancreatic cancers.
Citation Format: Silvia Coma, Monica Musteanu, Cristina Caffarra, Alessia Mira, Enrico Patrucco, Julien Dilly, Andrew J. Aguirre, Mariano Barbacid, Chiara Ambrogio, Jonathan A. Pachter. The RAF/MEK clamp avutometinib (VS-6766) enhances antitumor efficacy of KRAS G12C and G12D inhibitors through vertical inhibition of RAS, RAF and MEK. &lt;/title&gt; [abstract]. In: Proceedings of the AACR Special Conference: Targeting RAS; 2023 Mar 5-8; Philadelphia, PA. Philadelphia (PA): AACR; Mol Cancer Res 2023;21(5_Suppl):Abstract nr B025.