The colorectal adenoma-carcinoma sequence has provided a paradigmatic framework for understanding the successive somatic genetic changes and consequent clonal expansions leading to cancer. As for most cancer types, however, understanding of the earliest phases of colorectal neoplastic change, which may occur in morphologically normal tissue, is comparatively limited. Here, we whole genome sequenced hundreds of normal crypts from 42 individuals. Signatures of multiple mutational processes were revealed, some ubiquitous and continuous, others only found in some individuals, in some crypts or during certain periods of life. Likely driver mutations were present in ~1% of normal colorectal crypts in middle-aged individuals, indicating that adenomas and carcinomas are rare outcomes of a pervasive process of neoplastic change across morphologically normal colorectal epithelium. Colorectal cancers exhibit substantially elevated mutation burdens relative to normal cells. Sequencing normal colorectal cells provides quantitative insights into the genomic and clonal evolution of cancerdriver mutations, which conceivably are morphologically indistinguishable from normal cells, are similarly unclear. In large part, these deficiencies are due to the technical challenge of identifying somatic mutations in normal tissues, which are composed of myriad microscopic cell clones. Several different approaches have been adopted to address this 4-14 , revealing signatures of common somatic mutational processes in normal cells of the small and large intestine, liver, blood, skin, and nervous system. Thus far, however, studies have not been of sufficient scale to characterise variation in signature activity or detect less frequent processes 4-14. Remarkably high proportions of normal skin, oesophageal, and endometrial epithelial cells have been shown to be members of clones already carrying driver mutations 10,11,15,16 , and large mutant clones have been detected in blood 17-20. The extent of this phenomenon in the colon, an organ with a high cancer incidence, has not been investigated. Colonic epithelium is a contiguous cell sheet organised into ~15,000,000 crypts each composed of ~2,000 cells 21. Towards the base of each crypt resides a small number of stem cells ancestral to the maturing and differentiated cells in the crypt 22. These stem cells stochastically replace one another through a process of neutral drift 23,24 such that all stem cells, and thus all cells, in a crypt derive from a single ancestor stem cell that existed in recent years 25-27. The somatic mutations that were present in this ancestor are thus found in all ~2,000 descendant cells and can be revealed by DNA sequencing of an individual crypt. These stem cells are thought to be the cells of origin of colorectal cancers 28. To characterise the earliest stages of colorectal carcinogenesis, somatic mutation burdens, mutational signatures, clonal dynamics, and the frequency of driver mutations in normal colorectal epithelium were explored by sequencing individual colorect...
