Clonal biases dictate availability of colonic cancer driver mutations for transformation

Abstract Aged normal tissues harbour cancer mutations predisposing to transformation. However, previous studies of human colon have not allowed a comparison of the prevalence and behaviours of different pro-oncogenic events. Here, hotspot regions in colorectal cancer genes (APC, KRAS, TP53, FBXW7, CTNNB1) were analysed by targeted amplicon sequencing in normal tissue samples from 56 patients covering 76,800 colonic crypts. Cancer driver mutations were detected in all genes, including APC. Inference of clone dynamics revealed that FBXW7 R465C and KRAS G12 missense mutations have strong positive biases in clone fixation and expansion, respectively. Modelling of mutational co-occurrences and temporal ordering show that selection biases associated with early biallelic loss of APC progressively favours an ‘APC first’ acquisition of driver events leading to cancer with age. At younger ages KRAS activation is equally likely to be first due to its associated biases. Finally, spatial transcriptomics highlighted phenotypic heterogeneity in KRAS mutant clones of the same genotype, with mixed lineage presentation seen in a proportion of them, a state that has been linked with increased risk of transformation in other organs.