Investigation of KRAS Dependency Bypass and Functional Characterization of All Possible KRAS Missense Variants
Ly, Seav Huong
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AbstractThe importance of oncogenic KRAS in human cancers have prompted intense efforts to target KRAS and its effectors. To anticipate the development of resistance to these strategies, we previously performed a genome-scale expression screen to identify genes that bypass KRAS oncogenic dependency. Here we test thirty-seven genes that scored over five standard deviations and find that overexpression of LIM homeobox 9 (LHX9), a transcription factor involved in embryonic development, robustly rescues the suppression of KRAS in vitro and xenograft models. Furthermore, LHX9 substantially decreases cell sensitivity to KRASG12C and MEK1/2 inhibitors in KRAS-dependent cells. Based on RNA-seq and ChIP-seq analysis, we observe that LHX9 promotes both KRAS-associated and KRAS-unassociated expression profiles. Importantly, we show that LHX9-mediated upregulation of STAT3 and YAP1 expression is a major contributor to the rescue of KRAS suppression. Together these observations identify LHX9 as a transcription factor that regulates pathways that permit proliferation of KRAS-dependent cells following KRAS suppression.
KRAS mutation is the major genetic alteration responsible for its oncogenicity, Although the majority of KRAS mutations occur at codon 12, 13, and 61, mutations that alter other positions in KRAS are found frequently in human cancers, and these variants of unknown significance (VUS) complicate the interpretation of clinical cancer profiling. Here we performed saturation mutagenesis of KRAS by creating a library of all possible alleles of KRAS. We characterized the transforming ability of each variant by measuring its ability to confer anchorage-independent growth to an immortalized human cell line. The strength of the observed transformation phenotypes correlates strongly with the frequency of observed mutations in human cancers. In addition, we identified alleles that are rarely mutated but exhibit a strong transformation phenotype and other alleles that do not appear to exhibit transforming functions. These observations provide strong evidence that oncogenic KRAS alleles are selected during tumorigenesis and establish a comprehensive resource to infer the significance of KRAS alleles identified by molecular analysis of human tumors.
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