Bulk RNA-Seq from CRISPR-Cas9 Screening Identifies Resistance Mechanisms to KRAS Inhibition in Pancreatic Cancer

KRAS inhibitors (KRASi) targeting various KRAS mutations have entered clinical trials for pancreatic cancer. Despite promising preliminary clinical responses, most patients relapse due to intrinsic or acquired resistance. Thus, combination treatments are essential to extend the efficacy of KRAS-targeted therapies. To further determine genetic mechanisms of KRASi resistance, we performed KRASi-anchored CRISPR-Cas9 loss-of-function screens in KRASG12D, KRASG12C, KRASG12R, and KRASQ61H mutant PDAC cell lines, using six KRASi, to identify genes that modulate sensitivity to KRAS inhibition. Several hits from the screens, including EGFR, CK2, p110α and p110γ, and YAP were validated by combining targeted inhibitors with KRASi. KRASQ61H-mutant PDAC cell lines were intrinsically less dependent on KRAS for survival than other KRAS mutational subtypes. Further, EGFR inhibitor erlotinib synergized with the RAS(ON) multi-selective inhibitor RMC-7977 in KRASQ61H-mutant PDAC cell lines and in cell lines with highly active EGFR by mitigating ERK rebound activity. KRASi-resistant cell lines featured sustained ERK/MAPK dependence despite decreased ERK activity. Together, these findings enhance the understanding of intrinsic and acquired resistance to KRASi and identify therapeutic vulnerabilities that can potentially be exploited for KRASi combination therapies in pancreatic cancer patients.