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Summary Combinatorial inhibition of effector and feedback pathways is a promising treatment strategy for KRAS mutant cancers. However, the particular pathways that should be targeted to optimize therapeutic responses are unclear. Using CRISPR/Cas9, we systematically mapped the pathways whose inhibition cooperates with drugs targeting the KRAS effectors MEK, ERK, and PI3K. By performing 70 screens in models of KRAS mutant colorectal, lung, ovarian, and pancreas cancers, we uncovered universal and tissue-specific sensitizing combinations involving inhibitors of cell cycle, metabolism, growth signaling, chromatin regulation, and transcription. Furthermore, these screens revealed secondary genetic modifiers of sensitivity, yielding a SRC inhibitor-based combination therapy for KRAS / PIK3CA double-mutant colorectal cancers (CRCs) with clinical potential. Surprisingly, acquired resistance to combinations of growth signaling pathway inhibitors develops rapidly following treatment, but by targeting signaling feedback or apoptotic priming, it is possible to construct three-drug combinations that greatly delay its emergence. [ABSTRACT FROM AUTHOR]

Although KRAS has long been considered undruggable, direct KRASG12C inhibitors have shown promising initial clinical efficacy. However, the majority of patients still fail to respond. Adaptive feedback reactivation of RAS-mitogen-activated protein kinase (MAPK) signaling has been proposed by our group and others as a key mediator of resistance, but the exact mechanism driving reactivation and the therapeutic implications are unclear. We find that upstream feedback activation of wild-type RAS, as opposed to a shift in KRASG12C to its active guanosine triphosphate (GTP)-bound state, is sufficient to drive RAS-MAPK reactivation in a KRASG12C-independent manner. Moreover, multiple receptor tyrosine kinases (RTKs) can drive feedback reactivation, potentially necessitating targeting of convergent signaling nodes for more universal efficacy. Even in colorectal cancer, where feedback is thought to be primarily epidermal growth factor receptor (EGFR)-mediated, alternative RTKs drive pathway reactivation and limit efficacy, but convergent upstream or downstream signal blockade can enhance activity. Overall, these data provide important mechanistic insight to guide therapeutic strategies targeting KRAS. [Display omitted] • Adaptive RAS-MAPK feedback reactivation occurs following KRASG12C inhibition • RTK-mediated feedback activation of wild-type NRAS and HRAS bypasses KRASG12C • Inhibitor combinations abrogate RAS reactivation, boosting efficacy in KRASG12C CRC • Targeting convergent signaling nodes overcomes adaptive resistance by multiple RTKs Ryan et al. provide evidence that feedback reactivation of the RAS-MAPK pathway through wild-type NRAS and HRAS, as opposed to a shift in KRASG12C to its active GTP-bound state, can drive adaptive resistance to KRASG12C inhibitors in a KRASG12C-independent manner. [ABSTRACT FROM AUTHOR]

We apply genetic screens to delineate modulators of KRAS mutant pancreatic ductal adenocarcinoma (PDAC) sensitivity to ERK inhibitor treatment, and we identify components of the ATR-CHK1 DNA damage repair (DDR) pathway. Pharmacologic inhibition of CHK1 alone causes apoptotic growth suppression of both PDAC cell lines and organoids, which correlates with loss of MYC expression. CHK1 inhibition also activates ERK and AMPK and increases autophagy, providing a mechanistic basis for increased efficacy of concurrent CHK1 and ERK inhibition and/or autophagy inhibition with chloroquine. To assess how CHK1 inhibition-induced ERK activation promotes PDAC survival, we perform a CRISPR-Cas9 loss-of-function screen targeting direct/indirect ERK substrates and identify RIF1. A key component of non-homologous end joining repair, RIF1 suppression sensitizes PDAC cells to CHK1 inhibition-mediated apoptotic growth suppression. Furthermore, ERK inhibition alone decreases RIF1 expression and phenocopies RIF1 depletion. We conclude that concurrent DDR suppression enhances the efficacy of ERK and/or autophagy inhibitors in KRAS mutant PDAC. [Display omitted] • Pancreatic cancer cells are dependent on CHEK1 and DNA damage response genes • CHK1 inhibition causes compensatory ERK and autophagy activation • Concurrent CHK1 and ERK inhibition causes synergistic loss of MYC • Concurrent CHK1, ERK, and autophagy inhibition synergistically suppresses growth Klomp et al. show that CHK1 is essential for KRAS mutant pancreatic cancer cell growth. CHK1 inhibition causes apoptotic growth suppression, MYC loss, and compensatory ERK and autophagy activation. Concurrent CHK1, ERK, and/or autophagy inhibition enhances apoptotic growth suppression. Additionally, genetic depletion of ERK-regulated DNA damage repair protein RIF1 phenocopies ERK inhibition. [ABSTRACT FROM AUTHOR]