Project Summary/Abstract** Activating mutations in the RAS oncogenes are found across cancers, and contribute to a malignant phenotype through deregulation of normal proliferation and differentiation. As sequencing data from whole cancer genomes becomes available, it has become apparent that activating Ras mutations can be found at many different amino acids. Moreover, the distribution of specific Ras mutations and the isoform in which they occur vary highly with tissue type, suggesting that the mechanism for Ras-driven oncogenesis depends on tissue- specific genetic interactions. Among epithelial cancers, colorectal cancer (CRC) is unique in that it enriches for the rarer alleles of K-Ras, such as K-RasA146T and K-RasG13D. A key difference between CRC and other epithelial cancers (such as pancreatic or lung) is that loss of Apc, and therefore activation of Wnt signaling, is very frequent, and often required for tumorigenesis. Preliminary evidence from our lab shows that, in the colon, K-RasA146T, K- RasG13D, and K-RasG12D expression manifest in distinct phenotypes, with differing levels of Ras activation. However, in the background of Apc loss, and therefore in CRC, the weaker alleles of K-Ras (G13D and A146T) are now hyper-activated, and phenotypically indistinguishable from K-RasG12D. I hypothesize that the loss of Apc, and therefore activation of Wnt signaling, serves to activate the weak alleles of K-Ras, likely by altering expression of the GTPase activating proteins (GAPs) that regulate K-Ras. I propose to test this hypothesis in mouse colon organoids, using established methods to externally modulate Wnt levels, and to see how this affects K-Ras activation in the context of the various mutant alleles. Additionally, using existing conditional mouse models for these K-Ras mutant alleles, I will characterize their phenotypic and molecular properties in the pancreas, and determine if the absence of Wnt signaling in this tissue serves to enrich a certain subset of mutations. The findings generated in this study will help establish the importance of tissue-specific genetic interactions of K-Ras – an area of study important to use precision medicine to design therapeutic strategies targeting K-Ras mutant cancers.