Summary Whole-genome-sequencing (WGS) of human tumors has revealed distinct mutation patterns that hint at the causative origins of cancer. We examined mutational signatures in 324 WGS human-induced pluripotent stem cells exposed to 79 known or suspected environmental carcinogens. Forty-one yielded characteristic substitution mutational signatures. Some were similar to signatures found in human tumors. Additionally, six agents produced double-substitution signatures and eight produced indel signatures. Investigating mutation asymmetries across genome topography revealed fully functional mismatch and transcription-coupled repair pathways. DNA damage induced by environmental mutagens can be resolved by disparate repair and/or replicative pathways, resulting in an assortment of signature outcomes even for a single agent. This compendium of experimentally induced mutational signatures permits further exploration of roles of environmental agents in cancer etiology and underscores how human stem cell DNA is directly vulnerable to environmental agents. Video Abstract

Mutational signatures are patterns of mutations that arise during tumorigenesis. We present an enhanced, practical framework for mutational signature analyses. Applying these methods on 3,107 whole genome sequenced (WGS) primary cancers of 21 organs reveals known signatures and nine previously undescribed rearrangement signatures. We highlight inter-organ variability of

Mutational signatures are imprints of pathophysiological processes arising through tumorigenesis. We generated isogenic CRISPR-Cas9 knockouts (Δ) of 43 genes in human induced pluripotent stem cells, cultured them in the absence of added DNA damage, and performed whole-genome sequencing of 173 subclones. Δ OGG1, Δ UNG, Δ EXO1, Δ RNF168, Δ MLH1, Δ MSH2, Δ MSH6, Δ PMS1, and Δ PMS2 produced marked mutational signatures indicative of being critical mitigators of endogenous DNA modifications. Detailed analyses revealed mutational mechanistic insights, including how 8-oxo-dG elimination is sequence-context-specific while uracil clearance is sequence-context-independent. Mismatch repair (MMR) deficiency signatures are engendered by oxidative damage (C>A transversions), differential misincorporation by replicative polymerases (T>C and C>T transitions), and we propose a ‘reverse template slippage’ model for T>A transversions. Δ MLH1, Δ MSH6, and Δ MSH2 signatures were similar to each other but distinct from Δ PMS2 . Finally, we developed a classifier, MMRDetect, where application to 7,695 WGS cancers showed enhanced detection of MMR-deficient tumors, with implications for responsiveness to immunotherapies.

Whole-genome sequencing (WGS) permits comprehensive cancer genome analyses, revealing mutational signatures, imprints of DNA damage, and repair processes that have arisen in each patient’s cancer. We performed mutational signature analyses on 12,222 whole-genome–sequenced tumor-normal matched pairs from patients recruited via the UK National Health Service (NHS). We contrasted our results with two independent cancer WGS datasets—from the International Cancer Genome Consortium (ICGC) and the Hartwig Medical Foundation (HMF)—involving 18,640 whole-genome–sequenced cancers in total. Our analyses add 40 single and 18 double substitution signatures to the current mutational signature tally. We show for each organ that cancers have a limited number of common signatures and a long tail of rare signatures, and we provide a practical solution for applying this concept of common versus rare signatures to future analyses.