How theKRASoncogene drives cancer growth remains poorly understood. Therefore, we established a systemwide portrait of KRAS- and extracellular signal–regulated kinase (ERK)–dependent gene transcription in KRAS-mutant cancer to delineate the molecular mechanisms of growth and of inhibitor resistance. Unexpectedly, our KRAS-dependent gene signature diverges substantially from the frequently cited Hallmark KRAS signaling gene signature, is driven predominantly through the ERK mitogen-activated protein kinase (MAPK) cascade, and accurately reflects KRAS- and ERK-regulated gene transcription in KRAS-mutant cancer patients. Integration with our ERK-regulated phospho- and total proteome highlights ERK deregulation of the anaphase promoting complex/cyclosome (APC/C) and other components of the cell cycle machinery as key processes that drive pancreatic ductal adenocarcinoma (PDAC) growth. Our findings elucidate mechanistically the critical role of ERK in driving KRAS-mutant tumor growth and in resistance to KRAS-ERK MAPK targeted therapies.

Primary/intrinsic and treatment-induced acquired resistance limit the initial response rate to and long-term efficacy of direct inhibitors of the KRASG12C mutant in cancer. To identify potential mechanisms of resistance, we applied a CRISPR/Cas9 loss-of-function screen and observed loss of multiple components of the Hippo tumor suppressor pathway, which acts to suppress YAP1/TAZ-regulated gene transcription. YAP1/TAZ activation impaired the antiproliferative and proapoptotic effects of KRASG12C inhibitor (G12Ci) treatment in KRASG12C-mutant cancer cell lines. Conversely, genetic suppression of YAP1/WWTR1 (TAZ) enhanced G12Ci sensitivity. YAP1/TAZ activity overcame KRAS dependency through two distinct TEAD transcription factor–dependent mechanisms, which phenocopy KRAS effector signaling. First, TEAD stimulated ERK-independent transcription of genes normally regulated by ERK (BIRC5, CDC20, ECT2, FOSL1, and MYC) to promote progression through the cell cycle. Second, TEAD caused activation of PI3K–AKT–mTOR signaling to overcome apoptosis. G12Ci treatment-induced acquired resistance was also caused by YAP1/TAZ-TEAD activation. Accordingly, concurrent treatment with pharmacologic inhibitors of TEAD synergistically enhanced KRASG12C inhibitor antitumor activity in vitro and prolonged tumor suppression in vivo. In summary, these observations reveal YAP1/TAZ-TEAD signaling as a crucial driver of primary and acquired resistance to KRAS inhibition and support the use of TEAD inhibitors to enhance the antitumor efficacy of KRAS-targeted therapies. Significance: YAP1/TAZ-TEAD activation compensates for loss of KRAS effector signaling, establishing a mechanistic basis for concurrent inhibition of TEAD to enhance the efficacy of KRASG12C-selective inhibitor treatment of KRASG12C-mutant cancers. See related commentary by Johnson and Haigis, p. 4005

To delineate the mechanisms by which the ERK1 and ERK2 mitogen-activated protein kinases support mutant KRAS–driven cancer growth, we determined the ERK-dependent phosphoproteome in KRAS-mutant pancreatic cancer. We determined that ERK1 and ERK2 share near-identical signaling and transforming outputs and that the KRAS-regulated phosphoproteome is driven nearly completely by ERK. We identified 4666 ERK-dependent phosphosites on 2123 proteins, of which 79 and 66%, respectively, were not previously associated with ERK, substantially expanding the depth and breadth of ERK-dependent phosphorylation events and revealing a considerably more complex function for ERK in cancer. We established that ERK controls a highly dynamic and complex phosphoproteome that converges on cyclin-dependent kinase regulation and RAS homolog guanosine triphosphatase function (RHO GTPase). Our findings establish the most comprehensive molecular portrait and mechanisms by which ERK drives KRAS-dependent pancreatic cancer growth.

Primary/intrinsic and treatment-induced acquired resistance limit the initial response rate to and long-term efficacy of direct inhibitors of the KRASG12C mutant in cancer. To identify potential mechanisms of resistance, we applied a CRISPR/Cas9 loss-of-function screen and observed loss of multiple components of the Hippo tumor suppressor pathway, which acts to suppress YAP1/TAZ-regulated gene transcription. YAP1/TAZ activation impaired the antiproliferative and proapoptotic effects of KRASG12C inhibitor (G12Ci) treatment in KRASG12C-mutant cancer cell lines. Conversely, genetic suppression of YAP1/WWTR1 (TAZ) enhanced G12Ci sensitivity. YAP1/TAZ activity overcame KRAS dependency through two distinct TEAD transcription factor–dependent mechanisms, which phenocopy KRAS effector signaling. First, TEAD stimulated ERK-independent transcription of genes normally regulated by ERK (BIRC5, CDC20, ECT2, FOSL1, and MYC) to promote progression through the cell cycle. Second, TEAD caused activation of PI3K–AKT–mTOR signaling to overcome apoptosis. G12Ci treatment-induced acquired resistance was also caused by YAP1/TAZ-TEAD activation. Accordingly, concurrent treatment with pharmacologic inhibitors of TEAD synergistically enhanced KRASG12C inhibitor antitumor activity in vitro and prolonged tumor suppression in vivo. In summary, these observations reveal YAP1/TAZ-TEAD signaling as a crucial driver of primary and acquired resistance to KRAS inhibition and support the use of TEAD inhibitors to enhance the antitumor efficacy of KRAS-targeted therapies.Significance:YAP1/TAZ-TEAD activation compensates for loss of KRAS effector signaling, establishing a mechanistic basis for concurrent inhibition of TEAD to enhance the efficacy of KRASG12C-selective inhibitor treatment of KRASG12C-mutant cancers.See related commentary by Johnson and Haigis, p. 4005

Data from TEAD Inhibition Overcomes YAP1/TAZ-Driven Primary and Acquired Resistance to KRAS^G12C Inhibitors