Abstract B063: Differential pathway engagement and the biological consequences of KRAS variants in cancer

Abstract KRAS is a proto-oncogene found mutated in >90% of pancreatic ductal adenocarcinomas (PDAC) and encodes for a GTPase that mediates signaling transduction through the mitogen-activated protein kinase (MAPK) and PI3K/AKT pathways. Mutations in amino acids G12, G13, and Q61 prevent its inactivation by regulatory GTPase-activating proteins (GAPs), resulting in its constitutive activation. Previous work indicates that KRAS mutants possess unique biochemical differences that could result in divergent allele-specific signaling networks, underscoring unique vulnerabilities and response to treatment. Furthermore, preclinical and clinical data reveal allele-specific differences in tumor initiation capacity and patient prognosis in PDAC. Therefore, dissecting the biochemical and biological differences among KRAS variants would allow the identification of new allele-specific strategies for therapy. We hypothesize that the biochemical differences between KRAS variants result in differential signaling pathway engagement, affecting cellular behavior and tumorigenic properties. To study the biology of KRAS mutant variants in PDAC, we used CRISPR-Cas9 to generate KRAS-deficient PDAC cell lines which survived KRAS ablation through adaptive and reversible signaling rewiring. Our initial observations suggested that, at comparable expression levels, KRAS mutants diverge in their baseline signaling through the canonical MAPK and PI3K pathways. Therefore, we leveraged this unique isogenic system to uncover allele-specific differences between KRAS variants in global gene expression (by RNA-seq) and phosphoproteomic signaling networks (by data-independent acquisition mass spectroscopy; DIA-MS). We further employed a multiplexed competitive fitness approach using a sequence-based readout to assess changes in cellular fitness in response to drug perturbations (KRASi, PI3Ki, SHP2i, ERKi, RTKi and MEKi). Our findings suggest that KRAS mutants cluster into groups that diverge in downstream pathway engagement and gene expression, leading to cluster-specific differences in drug sensitivity. Our findings suggest that elucidating the signaling differences between KRAS alleles and their biological functions could translate to novel therapeutic strategies in PDAC and other KRAS mutant cancers. Citation Format: Yanixa Quiñones Avilés, Jaffarguriqbal Singh, Wenxue Li, Yong Kong, Yansheng Liu, Mandar Deepak Muzumdar. Differential pathway engagement and the biological consequences of KRAS variants in cancer [abstract]. In: Proceedings of the AACR Special Conference on Pancreatic Cancer; 2022 Sep 13-16; Boston, MA. Philadelphia (PA): AACR; Cancer Res 2022;82(22 Suppl):Abstract nr B063.