Synthetic lethality results when mutant KRAS and EGFR proteins are co-expressed in human lung adenocarcinoma (LUAD) cells, revealing the biological basis for mutual exclusivity of KRAS and EGFR mutations. We have now defined the biochemical events responsible for the toxic effects by combining pharmacological and genetic approaches and to show that signaling through extracellular signal-regulated kinases (ERK1/2) mediates the toxicity. These findings imply that tumors with mutant oncogenes in the RAS pathway must restrain the activity of ERK1/2 to avoid toxicities and enable tumor growth. A dual specificity phosphatase, DUSP6, that negatively regulates phosphorylation of (P)-ERK is up-regulated in EGFR- or KRAS-mutant LUAD, potentially protecting cells with mutations in the RAS signaling pathway, a proposal supported by experiments with DUSP6-specific siRNA and an inhibitory drug. Targeting DUSP6 or other negative regulators might offer a treatment strategy for certain cancers by inducing the toxic effects of RAS-mediated signaling.

Cancer models based on cells derived from human embryonic stem cells (hESCs) may reveal why certain constellations of genetic changes drive carcinogenesis in specialized lineages. Here we demonstrate that inhibition of NOTCH signaling induces up to 10% of lung progenitor cells to form pulmonary neuroendocrine cells (PNECs), putative precursors to small cell lung cancers (SCLCs), and we can increase PNECs by reducing levels of retinoblastoma (RB) proteins with inhibitory RNA. Reducing levels of TP53 protein or expressing mutant KRAS or EGFR genes did not induce or expand PNECs, but tumors resembling early-stage SCLC grew in immunodeficient mice after subcutaneous injection of PNEC-containing cultures in which expression of both RB and TP53 was blocked. Single-cell RNA profiles of PNECs are heterogeneous; when RB levels are reduced, the profiles resemble those from early-stage SCLC; and when both RB and TP53 levels are reduced, the transcriptome is enriched with cell cycle–specific RNAs. Our findings suggest that genetic manipulation of hESC-derived pulmonary cells will enable studies of this recalcitrant cancer.

MEK inhibitors (MEKi) have limited efficacy in KRAS mutant lung adenocarcinoma (LUAD) patients, and this is attributed to both intrinsic and adaptive mechanisms of drug resistance. While many studies have focused on the former, there remains a dearth of data regarding acquired resistance to MEKi in LUAD. We established trametinib-resistant KRAS mutant LUAD cells through dose escalation and performed targeted MSK-IMPACT sequencing to identify drivers of MEKi resistance. Comparing resistant cells to their sensitive counterparts revealed alteration of genes associated with trametinib response. We describe a state of “drug addiction” in resistant cases where cells are dependent on continuous culture in trametinib for survival. We show that dependence on ERK2 suppression underlies this phenomenon and that trametinib removal hyperactivates ERK, resulting in ER stress and apoptosis. Amplification of KRAS^G12C occurs in drug-addicted cells and blocking mutant-specific activity with AMG 510 rescues the lethality associated with trametinib withdrawal. Furthermore, we show that increased KRAS^G12C expression is lethal to other KRAS mutant LUAD cells, consequential to ERK hyperactivation. Our study determines the drug-addicted phenotype in lung cancer is associated with KRAS amplification and demonstrates that toxic acquired genetic changes can develop de novo in the background of MAPK suppression with MEK inhibitors. We suggest that the presence of mutant KRAS amplification in patients may identify those that may benefit from a “drug holiday” to circumvent drug resistance. These findings demonstrate the toxic potential of hyperactive ERK signaling and highlight potential therapeutic opportunities in patients bearing KRAS mutations.

Large-scale analyses of cancer genomes are revealing patterns of mutations that suggest biologically significant ideas about many aspects of cancer, including carcinogenesis, classification, and preventive and therapeutic strategies. Among those patterns is “mutual exclusivity”, a phenomenon observed when two or more mutations that are commonly observed in samples of a type of cancer are not found combined in individual tumors. We have been studying a striking example of mutual exclusivity: the absence of co-existing mutations in the KRAS and EGFR proto-oncogenes in human lung adenocarcinomas, despite the high individual frequencies of such mutations in this common type of cancer. Multiple lines of evidence suggest that toxic effects of the joint expression of KRAS and EGFR mutant oncogenes, rather than loss of any selective advantages conferred by a second oncogene that operates through the same signaling pathway, are responsible for the observed mutational pattern. We discuss the potential for understanding the physiological basis of such toxicity, for exploiting it therapeutically, and for extending the studies to other examples of mutual exclusivity.

We recently described the synthetic lethality that results when mutant KRAS and mutant EGFR are coexpressed in human lung adenocarcinoma (LUAD) cells, revealing the biological basis for the mutual exclusivity of KRAS and EGFR mutations in lung cancers. We have now further defined the biochemical events responsible for the toxic effects of signaling through the RAS pathway. By combining pharmacological and genetic approaches, we have developed multiple lines of evidence that signaling through extracellular signal-regulated kinases (ERK1/2) mediates the toxicity. These findings imply that tumors with mutant oncogenes that drive signaling through the RAS pathway must restrain the activity of ERK1/2 to avoid cell toxicities and enable tumor growth. In particular, a dual specificity phosphatase, DUSP6, regulates phosphorylated (P)-ERK levels in lung adenocarcinoma cells, providing negative feedback to the RAS signaling pathway. Accordingly, inhibition of DUSP6 is cytotoxic in LUAD cells driven by either mutant KRAS or mutant EGFR, phenocopying the effects of co-expression of mutant KRAS and EGFR. Together, these data suggest that targeting DUSP6 or other feedback regulators of the EGFR-KRAS-ERK pathway may offer a strategy for treating certain cancers by exceeding an upper threshold of RAS-mediated signaling.