Mutational order and epistasis determine the consequences of FBXW7 mutations during colorectal cancer evolution

Somatic driver mutations, in genes such as FBXW7, have been discovered in phenotypically normal colonic tissue, however their role in cancer initiation remains elusive. Using normal and gene-edited patient-derived human colon organoids as models of early tumour evolution we observed that FBXW7-/- mutations exerted an epistatic effect on subsequent transcription depending on the mutational background of the cell. Specifically, the timing of acquiring an FBXW7-/- mutation respective to an APC mutation, led to profound phenotypic and transcriptomic differences. When FBXW7 was mutated before APC, a near-normal cell state was maintained alongside repression of the APC transcriptional response. However, when APC was mutated before FBXW7, cells acquired classical cancer-stem cell features. Single-cell RNA sequencing revealed that mutation of FBXW7 in normal tissue subtly switched cells from adult to a foetal/regenerative stem cell state. Further analysis using transposase-accessible chromatin sequencing identified this cellular plasticity was driven by changes in chromatin accessibility of transcriptional start site regions associated with TEAD, SNAI1 and AP-1 motifs, which in turn activate the foetal-like state. Taken together, we demonstrate a critical role of FBXW7 mutations in preventing colorectal cancer initiation and provide exemplar evidence for the importance of epistasis and mutational order in cancer biology.