Abstract
Despite dozens of tools to identify mutational signatures in cancer samples, there is not an established metric for quantifying whether signature exposures differ significantly between two heterogeneous groups of samples. We demonstrate that a signature-agnostic metric - the aggregate mutation spectrum distance permutation method (AMSD) - can rigorously determine whether mutational exposures differ between groups, a hypothesis that is not directly addressed by signature analysis. First, we reanalyze a study of carcinogen exposure in mice, determining that eleven of twenty tested carcinogens produce significant mutation spectrum shifts. Only three of these carcinogens were previously reported to induce distinct mutational signatures, suggesting that many carcinogens perturb mutagenesis by altering the composition of endogenous signatures. Next, we interrogate whether patient ancestry has a measurable impact on human tumor mutation spectra, finding significant ancestry-associated differences across ten cancer types. Some have been previously reported, such as elevated SBS4 in African lung adenocarcinomas, while some have not to our knowledge been reported, such as elevated SBS17a/b in European esophageal carcinomas. These examples suggest that AMSD is a robust tool for detecting differences among groups of tumors or other mutated samples, complementing descriptive signature deconvolution and enabling the discovery of environmental and genetic influences on mutagenesis.
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Data availability
All analyses in this study use publicly available datasets, and figures and results can be reproduced using the code available at https://github.com/sfhart33/AMSD_cancer_mutation_spectra. Preprocessed mutation spectra are included in the repository, while raw data can be accessed from:
• Mouse carcinogen exposure: https://github.com/team113sanger/mouse-mutatation-signatures/blob/master/starting_data/snvs.rds
• Asbestos exposure: https://github.com/IARCbioinfo/MESOMICS_data/tree/main/phenotypic_map/MESOMICS
• TCGA ancestry metadata: https://gdc.cancer.gov/about-data/publications/CCG-AIM-2020
• TCGA somatic mutations: https://gdc.cancer.gov/about-data/publications/mc3-2017
The original implementation of the AMSD as a method for identifying mutator alleles is also available on github: https://github.com/quinlan-lab/proj-mutator-mapping.
Code availability
The Aggregate Mutation Spectrum Distance permutation test is implemented as the R package “mutspecdist”, available at https://github.com/sfhart33/mutspecdist. All analyses in this study use publicly available datasets, and figures and results can be reproduced using the code available at https://github.com/sfhart33/AMSD_cancer_mutation_spectra.
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Acknowledgements
We thank Harris and Feder lab members for figure feedback, Sayre Coombs for graphic design feedback, and Tom Sasani for developing the original implementation of AMSD and manuscript feedback. This work was possible due to funding from NIH training grant T32-HG000035 supporting S.F.M.H., Worldwide Cancer Research grant 24-0106 to N.A., NIH grant 1DP2CA280623-01 to A.F.F., NIH NIGMS grant 2R35M133428-06 to K.H., and the Allen Discovery Center for Cell Lineage Tracing. Where authors are identified as personnel of the International Agency for Research on Cancer/World Health Organization, the authors alone are responsible for the views expressed in this article and they do not necessarily represent the decisions, policy or views of the International Agency for Research on Cancer/World Health Organization. The results published here are in part based on data generated by the the TCGA Research Network (https://www.cancer.gov/tcga) and by the Rare Cancers Genomics initiative (www.rarecancersgenomics.com).
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S.F.M.H., A.F.F., and K.H. contributed to study conceptualization and design. S.F.M.H. performed the data analysis. S.F.M.H., N.A., A.F.F., and K.H. interpreted the results. S.F.M.H. wrote the original draft of the manuscript. S.F.M.H., N.A., A.F.F., and K.H. contributed to review and editing of the manuscript.
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Hart, S.F.M., Alcala, N., Feder, A.F. et al. A signature-agnostic test for differences between tumor mutation spectra reveals carcinogen and ancestry effects. Commun Biol (2026). https://doi.org/10.1038/s42003-026-09652-5
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DOI: https://doi.org/10.1038/s42003-026-09652-5
