Characterization of Somatic Mutations in the Normal Colon Using Duplex Sequencing to Evaluate Colorectal Cancer Risk

Abstract

The accumulation of somatic mutations in normal tissues has been established over the last decade. However, we lack an understanding of how these mutations might predispose individuals to cancer. In this study we expand on previous findings to demonstrate that duplex sequencing enables the detection of variants at extremely variant allele fractions (VAFs) in normal colon from patients without cancer or polyps, with polyps, and those with colorectal cancer (CRC). By applying advanced computational techniques, we characterize the potential pathogenicity of these mutations, developing a framework for assessing positive selection in the normal colon. Our data indicates that individuals with CRC have a higher frequency of mutations in key CRC driver genes in their normal colon, specifically in APC, FBXW7, and PIK3CA, than patients without cancer. Furthermore, patients with cancer exhibit a higher frequency of pathogenic large clones (pathogenic variants with >1 duplex read) in KRAS and TP53 in normal colon, indicating an increased prevalence of positive selection and clonal expansions driven by mutations in those genes. This selection is not random, with mutations clustering in colorectal cancer gene hotspots. These results reveal that the clonal landscape of CRC driver genes differs between the normal colon of individuals with cancer and without cancer, providing insight into how the somatic genome may predict a patient's risk for developing cancer.