{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"institution":[{"name":"bioRxiv"}],"indexed":{"date-parts":[[2026,9,2]],"date-time":"2026-09-02T16:16:27Z","timestamp":1788365787490,"version":"build-2803163510"},"posted":{"date-parts":[[2026,1,13]]},"group-title":"Cancer Biology","reference-count":52,"publisher":"openRxiv","license":[{"start":{"date-parts":[[2026,1,13]],"date-time":"2026-01-13T00:00:00Z","timestamp":1768262400000},"content-version":"vor","delay-in-days":0,"URL":"http:\/\/creativecommons.org\/licenses\/by\/4.0\/"}],"funder":[{"award":["CA259384"],"award-info":[{"award-number":["CA259384"]}],"id":[{"id":"https:\/\/ror.org\/01cwqze88","id-type":"ROR","asserted-by":"publisher"}]},{"award":["CA280623"],"award-info":[{"award-number":["CA280623"]}],"id":[{"id":"https:\/\/ror.org\/01cwqze88","id-type":"ROR","asserted-by":"publisher"}]}],"content-domain":{"domain":[],"crossmark-restriction":false},"short-container-title":[],"accepted":{"date-parts":[[2026,8,17]]},"abstract":"<jats:title>ABSTRACT<\/jats:title>\n                <jats:p>\n                  Li-Fraumeni Syndrome (LFS) is caused by germline pathogenic variants in\n                  <jats:italic>TP53<\/jats:italic>\n                  which predispose carriers to early onset cancer across multiple tissues. While genomically profiling those cancers has revealed factors contributing to their formation, little is understood about how LFS impacts clonal evolution in healthy tissues preceding cancer. Here, we use ultra-deep duplex sequencing (mean \u223c15,000\u00d7 depth) to investigate somatic mutation and selection in a family carrying the germline\n                  <jats:italic>TP53<\/jats:italic>\n                  p.R181H pathogenic variant and a cohort of non-carrier controls. In blood samples, the germline variant is associated with more mutations in a panel designed to capture genomewide mutagenesis, and with reduced positive selection on somatic\n                  <jats:italic>TP53<\/jats:italic>\n                  mutations, despite confounding by chemotherapy treatment in one individual.\n                  <jats:italic>DNMT3A<\/jats:italic>\n                  and\n                  <jats:italic>TET2<\/jats:italic>\n                  mutations appear positively selected and\n                  <jats:italic>GATA2<\/jats:italic>\n                  mutations negatively selected across the cohort, independent of the p.R181H status. Extensive multi-tissue sampling of 22 non-cancerous and 6 cancerous samples was also performed at autopsy in one individual with LFS who succumbed to esophageal cancer. Cross-tissue analysis reveals excess mutations in sun-exposed skin, esophagus and chronically-inflamed stomach tissue, and concordant mutations in the p.R248 hotspot of\n                  <jats:italic>TP53<\/jats:italic>\n                  across most (18\/28) tissue samples. Most somatic\n                  <jats:italic>TP53<\/jats:italic>\n                  mutations in LFS that can be assessed for phase arose on the chromosomal copy lacking the p.R181H variant. Our study reveals how the germline p.R181H variant reshapes baseline somatic mutation and selection in normal tissues and highlights the importance of understanding early somatic evolution in LFS prior to cancer development and treatment.\n                <\/jats:p>","DOI":"10.64898\/2026.01.12.699071","type":"posted-content","created":{"date-parts":[[2026,1,13]],"date-time":"2026-01-13T21:50:16Z","timestamp":1768341016000},"source":"Crossref","is-referenced-by-count":1,"title":["Ultra-deep duplex sequencing reveals unique features of somatic evolution in the normal tissues of a family with Li-Fraumeni syndrome"],"prefix":"10.64898","author":[{"ORCID":"https:\/\/orcid.org\/0000-0002-6660-8460","authenticated-orcid":false,"given":"Hunter L.","family":"Colegrove","sequence":"first","affiliation":[{"name":"Department of Genome Sciences, University of Washington; Seattle, WA, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Marianne E","family":"Dubard-Gault","sequence":"additional","affiliation":[{"name":"Fred Hutchinson Cancer Center; Seattle, WA, USA"},{"name":"Department of Medicine, Division of Medical Genetics, University of Washington; Seattle, WA, USA"},{"name":"Providence Swedish Cancer Institute; Seattle, WA, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Henry","family":"Marshall","sequence":"additional","affiliation":[{"name":"Wellcome Sanger Institute; Hinxton, UK"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0001-9948-2131","authenticated-orcid":false,"given":"Brendan F.","family":"Kohrn","sequence":"additional","affiliation":[{"name":"Department of Laboratory Medicine and Pathology, University of Washington; Seattle, WA, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Thomas H.","family":"Smith","sequence":"additional","affiliation":[{"name":"TwinStrand Biosciences; Seattle, WA, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Zachary K.","family":"Norgaard","sequence":"additional","affiliation":[{"name":"TwinStrand Biosciences; Seattle, WA, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Fang Yin","family":"Lo","sequence":"additional","affiliation":[{"name":"TwinStrand Biosciences; Seattle, WA, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Elizabeth K.","family":"Schmidt","sequence":"additional","affiliation":[{"name":"TwinStrand Biosciences; Seattle, WA, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Jacob E.","family":"Higgins","sequence":"additional","affiliation":[{"name":"TwinStrand Biosciences; Seattle, WA, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"suffix":"III","given":"Charles C.","family":"Valentine","sequence":"additional","affiliation":[{"name":"TwinStrand Biosciences; Seattle, WA, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Desiree A.","family":"Marshall","sequence":"additional","affiliation":[{"name":"Department of Laboratory Medicine and Pathology, University of Washington; Seattle, WA, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"John I.","family":"Clark","sequence":"additional","affiliation":[{"name":"Department of Neurobiology & Biophysics, University of Washington; Seattle, WA, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Eric Q.","family":"Konnick","sequence":"additional","affiliation":[{"name":"Department of Laboratory Medicine and Pathology, University of Washington; Seattle, WA, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Jesse J.","family":"Salk","sequence":"additional","affiliation":[{"name":"TwinStrand Biosciences; Seattle, WA, USA"},{"name":"Department of Medicine, Division of Hematology and Oncology, University of Washington; Seattle, WA, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-1683-1680","authenticated-orcid":false,"given":"Marshall S.","family":"Horwitz","sequence":"additional","affiliation":[{"name":"Department of Laboratory Medicine and Pathology, University of Washington; Seattle, WA, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Raheleh","family":"Rahbari","sequence":"additional","affiliation":[{"name":"Wellcome Sanger Institute; Hinxton, UK"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0003-2915-089X","authenticated-orcid":false,"given":"Alison F.","family":"Feder","sequence":"additional","affiliation":[{"name":"Department of Genome Sciences, University of Washington; Seattle, WA, USA"},{"name":"Herbold Computational Biology Program, Fred Hutch Cancer Center; Seattle, WA, USA"},{"name":"Howard Hughes Medical Institute; Seattle, WA, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-0291-1858","authenticated-orcid":false,"given":"Rosa Ana","family":"Risques","sequence":"additional","affiliation":[{"name":"Department of Laboratory Medicine and Pathology, University of Washington; Seattle, WA, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]}],"member":"54368","reference":[{"key":"2026082009301274000_2026.01.12.699071v2.1","doi-asserted-by":"crossref","first-page":"4239","DOI":"10.1038\/s41467-023-39957-6","article-title":"Mutational signature dynamics shaping the evolution of oesophageal adenocarcinoma","volume":"14","year":"2023","journal-title":"Nat Commun"},{"key":"2026082009301274000_2026.01.12.699071v2.2","doi-asserted-by":"crossref","first-page":"23134","DOI":"10.1038\/s41598-024-73587-2","article-title":"Frequency and spectrum of mutations in human sperm measured using duplex sequencing correlate with trio-based de novo mutation analyses","volume":"14","year":"2024","journal-title":"Sci Rep"},{"key":"2026082009301274000_2026.01.12.699071v2.3","doi-asserted-by":"crossref","first-page":"15499","DOI":"10.3390\/ijms232415499","article-title":"Unraveling the Structural Changes in the DNA-Binding Region of Tumor Protein p53 (TP53) upon Hotspot Mutation p53 Arg248 by Comparative Computational Approach","volume":"23","year":"2022","journal-title":"Int J Mol Sci"},{"key":"2026082009301274000_2026.01.12.699071v2.4","doi-asserted-by":"publisher","DOI":"10.1126\/science.aax3649"},{"key":"2026082009301274000_2026.01.12.699071v2.5","doi-asserted-by":"publisher","DOI":"10.1038\/s41588-020-00710-0"},{"key":"2026082009301274000_2026.01.12.699071v2.6","unstructured":"Boruah N , Hoyos D , Moses R , Hausler R , Desai H , Le AN , Good M , Kelly G , Raghavakaimal A , Tayeb M , et al. 2024. Distinct genomic and immunologic tumor evolution in germline TP53-driven breast cancers. 2024.04.03.588009. https:\/\/www.biorxiv.org\/content\/10.1101\/2024.04.03.588009v1 (Accessed December 16, 2025)."},{"key":"2026082009301274000_2026.01.12.699071v2.7","doi-asserted-by":"publisher","DOI":"10.1200\/JCO.2014.59.5728"},{"key":"2026082009301274000_2026.01.12.699071v2.8","doi-asserted-by":"crossref","unstructured":"Calvet F , Blanco Martinez-Illescas R , Mui\u00f1os F , Tretiakova M , Latorre-Esteves ES , Fredrickson J , Andrianova M , Pellegrini S , Huber AR , Ramis-Zaldivar JE , et al. 2025. Sex and smoking bias in the selection of somatic mutations in human bladder. Nature 1\u20139.","DOI":"10.1038\/s41586-025-09521-x"},{"key":"2026082009301274000_2026.01.12.699071v2.9","doi-asserted-by":"crossref","unstructured":"Cheng J , Novati G , Pan J , Bycroft C , \u017demgulyt\u0117 A , Applebaum T , Pritzel A , Wong LH , Zielinski M , Sargeant T , et al. 2023. Accurate proteome-wide missense variant effect prediction with AlphaMissense. Science 381: eadg7492.","DOI":"10.1126\/science.adg7492"},{"key":"2026082009301274000_2026.01.12.699071v2.10","doi-asserted-by":"publisher","DOI":"10.1016\/j.stem.2017.07.010"},{"key":"2026082009301274000_2026.01.12.699071v2.11","doi-asserted-by":"publisher","DOI":"10.1016\/S1470-2045(21)00580-5"},{"key":"2026082009301274000_2026.01.12.699071v2.12","doi-asserted-by":"crossref","first-page":"100242","DOI":"10.1016\/j.xhgg.2023.100242","article-title":"Genome-first approach of the prevalence and cancer phenotypes of pathogenic or likely pathogenic germline TP53 variants","volume":"5","year":"2024","journal-title":"Hum Genet Genomics Adv"},{"key":"2026082009301274000_2026.01.12.699071v2.13","doi-asserted-by":"publisher","DOI":"10.1016\/j.celrep.2019.07.001"},{"key":"2026082009301274000_2026.01.12.699071v2.14","doi-asserted-by":"publisher","DOI":"10.1007\/S10689-024-00419-7\/TABLES\/1"},{"key":"2026082009301274000_2026.01.12.699071v2.15","doi-asserted-by":"publisher","DOI":"10.1056\/NEJMoa1409405"},{"key":"2026082009301274000_2026.01.12.699071v2.16","unstructured":"Hart SFM , Alcala N , Feder AF , Harris K . 2026. A signature-agnostic test for differences between tumor mutation spectra reveals carcinogen and ancestry effects. Commun Biol. https:\/\/www.nature.com\/articles\/s42003-026-09652-5 (Accessed March 13, 2026)."},{"key":"2026082009301274000_2026.01.12.699071v2.17","doi-asserted-by":"crossref","first-page":"100179","DOI":"10.1016\/j.xgen.2022.100179","article-title":"Uncovering novel mutational signatures by de novo extraction with SigProfilerExtractor","volume":"2","year":"2022","journal-title":"Cell Genomics"},{"key":"2026082009301274000_2026.01.12.699071v2.18","doi-asserted-by":"publisher","DOI":"10.1038\/s41568-021-00335-3"},{"key":"2026082009301274000_2026.01.12.699071v2.19","doi-asserted-by":"publisher","DOI":"10.1038\/nprot.2014.170"},{"key":"2026082009301274000_2026.01.12.699071v2.20","doi-asserted-by":"publisher","DOI":"10.1038\/s41586-022-05448-9"},{"key":"2026082009301274000_2026.01.12.699071v2.21","doi-asserted-by":"crossref","first-page":"1978","DOI":"10.1158\/0008-5472.CAN-24-1136","article-title":"The Prolonged Half-Life of the p53 Missense Variant R248Q Promotes Accumulation and Heterotetramer Formation with Wild-Type p53 to Exert the Dominant-Negative Effect","volume":"85","year":"2025","journal-title":"Cancer Res"},{"key":"2026082009301274000_2026.01.12.699071v2.22","doi-asserted-by":"publisher","DOI":"10.1038\/s41388-021-02141-5"},{"key":"2026082009301274000_2026.01.12.699071v2.23","doi-asserted-by":"crossref","first-page":"1800","DOI":"10.1001\/jamaoncol.2021.4398","article-title":"Analysis of the Li-Fraumeni Spectrum Based on an International Germline TP53 Variant Data Set: An International Agency for Research on Cancer TP53 Database Analysis","volume":"7","year":"2021","journal-title":"JAMA Oncol"},{"key":"2026082009301274000_2026.01.12.699071v2.24","doi-asserted-by":"publisher","DOI":"10.1093\/nar\/gkw227"},{"key":"2026082009301274000_2026.01.12.699071v2.25","doi-asserted-by":"crossref","first-page":"411","DOI":"10.1038\/s41586-025-09584-w","article-title":"Somatic mutation and selection at population scale","volume":"647","year":"2025","journal-title":"Nature"},{"key":"2026082009301274000_2026.01.12.699071v2.26","doi-asserted-by":"publisher","DOI":"10.1038\/s41467-022-35727-y"},{"key":"2026082009301274000_2026.01.12.699071v2.27","doi-asserted-by":"crossref","unstructured":"Liu M , Wu Y , Jiang N , Boot A , Rozen SG. 2023. mSigHdp: hierarchical Dirichlet process mixture modeling for mutational signature discovery. NAR Genomics Bioinforma 5: lqad005.","DOI":"10.1093\/nargab\/lqad005"},{"key":"2026082009301274000_2026.01.12.699071v2.28","doi-asserted-by":"publisher","DOI":"10.1038\/s41586-022-05072-7"},{"key":"2026082009301274000_2026.01.12.699071v2.29","doi-asserted-by":"publisher","DOI":"10.1002\/cncr.30248"},{"key":"2026082009301274000_2026.01.12.699071v2.30","doi-asserted-by":"publisher","DOI":"10.1016\/j.yjmcc.2021.07.004"},{"key":"2026082009301274000_2026.01.12.699071v2.31","doi-asserted-by":"publisher","DOI":"10.1038\/S41568-020-0290-X"},{"key":"2026082009301274000_2026.01.12.699071v2.32","doi-asserted-by":"crossref","first-page":"1492","DOI":"10.1158\/0008-5472.CAN-21-3607","article-title":"Colorectal Cancer Is Associated with the Presence of Cancer Driver Mutations in Normal Colon","volume":"82","year":"2022","journal-title":"Cancer Res"},{"key":"2026082009301274000_2026.01.12.699071v2.33","doi-asserted-by":"publisher","DOI":"10.1038\/nrclinonc.2014.41"},{"key":"2026082009301274000_2026.01.12.699071v2.34","doi-asserted-by":"crossref","first-page":"291","DOI":"10.1016\/j.stemcr.2019.07.005","article-title":"Gata2 as a Crucial Regulator of Stem Cells in Adult Hematopoiesis and Acute Myeloid Leukemia","volume":"13","year":"2019","journal-title":"Stem Cell Rep"},{"key":"2026082009301274000_2026.01.12.699071v2.35","doi-asserted-by":"crossref","first-page":"1684","DOI":"10.1038\/s41588-025-02234-x","article-title":"The long-term effects of chemotherapy on normal blood cells","volume":"57","year":"2025","journal-title":"Nat Genet"},{"key":"2026082009301274000_2026.01.12.699071v2.36","doi-asserted-by":"publisher","DOI":"10.1158\/1078-0432.CCR-06-2545"},{"key":"2026082009301274000_2026.01.12.699071v2.37","doi-asserted-by":"publisher","DOI":"10.1038\/sj.bjc.6601302"},{"key":"2026082009301274000_2026.01.12.699071v2.38","doi-asserted-by":"crossref","unstructured":"Moses R , Indeglia A , Levine AS , Hausler R , Kelly G , Miller SA , Anez I , Heller M , Delgado R , Orr C , et al. 2025. Variation at the R181 residue of p53 confers loss of p53 DNA binding cooperativity with the retention of mitochondrial-associated apoptosis. 2025.08.12.669925. https:\/\/www.biorxiv.org\/content\/10.1101\/2025.08.12.669925v1 (Accessed December 5, 2025).","DOI":"10.1101\/2025.08.12.669925"},{"key":"2026082009301274000_2026.01.12.699071v2.39","doi-asserted-by":"crossref","first-page":"5215","DOI":"10.1182\/bloodadvances.2024012929","article-title":"Impact of cancer therapy on clonal hematopoiesis mutations and subsequent clinical outcomes","volume":"8","year":"2024","journal-title":"Blood Adv"},{"key":"2026082009301274000_2026.01.12.699071v2.40","doi-asserted-by":"publisher","DOI":"10.1101\/cshperspect.a001008"},{"key":"2026082009301274000_2026.01.12.699071v2.41","doi-asserted-by":"crossref","unstructured":"Oshima MU , Higgins J , Jenkins I , Randolph T , Smith T , Valentine C , Salk J , Yeung C , Beppu L , Campbell J , et al. 2024. Characterization of clonal dynamics using duplex sequencing in donor-recipient pairs decades after hematopoietic cell transplantation. Sci Transl Med 16: eado5108.","DOI":"10.1126\/scitranslmed.ado5108"},{"key":"2026082009301274000_2026.01.12.699071v2.42","doi-asserted-by":"publisher","DOI":"10.1038\/s41588-019-0525-5"},{"key":"2026082009301274000_2026.01.12.699071v2.43","doi-asserted-by":"publisher","DOI":"10.1053\/j.gastro.2021.10.038"},{"key":"2026082009301274000_2026.01.12.699071v2.44","first-page":"1","article-title":"TP53 somatic evolution in the normal endometrium of Black and White individuals","volume":"197","year":"2025","journal-title":"Gynecol Oncol"},{"key":"2026082009301274000_2026.01.12.699071v2.45","doi-asserted-by":"publisher","DOI":"10.1016\/j.celrep.2019.05.109"},{"key":"2026082009301274000_2026.01.12.699071v2.46","doi-asserted-by":"publisher","DOI":"10.1053\/j.gastro.2014.04.036"},{"key":"2026082009301274000_2026.01.12.699071v2.47","doi-asserted-by":"publisher","DOI":"10.1053\/j.gastro.2018.03.047"},{"key":"2026082009301274000_2026.01.12.699071v2.48","doi-asserted-by":"crossref","unstructured":"Sun Zhang A , Omran M , Arthur C , Malmgren H , Barbosa-Matos R , Seixas S , Oliveira C , Tham E , Bajalica-Lagercrantz S . 2025. TP53 p.R181H is enriched in the Swedish cohort (SWEP53) and associated with a distinct breast and prostate phenotype. Sci Rep 15: 35033.","DOI":"10.1038\/s41598-025-22407-2"},{"key":"2026082009301274000_2026.01.12.699071v2.49","doi-asserted-by":"crossref","unstructured":"Takahashi K , Nakada D , Goodell M . 2024. Distinct landscape and clinical implications of therapy-related clonal hematopoiesis. J Clin Invest 134. https:\/\/www.jci.org\/articles\/view\/180069 (Accessed December 15, 2025).","DOI":"10.1172\/JCI180069"},{"key":"2026082009301274000_2026.01.12.699071v2.50","doi-asserted-by":"crossref","first-page":"2422","DOI":"10.3390\/cancers13102422","article-title":"Rely on Each Other: DNA Binding Cooperativity Shapes p53 Functions in Tumor Suppression and Cancer Therapy","volume":"13","year":"2021","journal-title":"Cancers"},{"key":"2026082009301274000_2026.01.12.699071v2.51","doi-asserted-by":"crossref","first-page":"1695","DOI":"10.1038\/s41588-025-02235-w","article-title":"Clonal evolution of hematopoietic stem cells after autologous stem cell transplantation","volume":"57","year":"2025","journal-title":"Nat Genet"},{"key":"2026082009301274000_2026.01.12.699071v2.52","doi-asserted-by":"crossref","unstructured":"Yokoyama A , Watanabe K , Inoue Y , Hirano T , Tamaoki M , Hirohashi K , Kawaguchi S , Ishida Y , Takeuchi Y , Kishimoto Y , et al. 2025. Somatic mosaicism in the buccal mucosa reflects lifestyle and germline risk factors for esophageal squamous cell carcinoma. Sci Transl Med 17: eadq6740.","DOI":"10.1126\/scitranslmed.adq6740"}],"container-title":[],"original-title":[],"link":[{"URL":"https:\/\/syndication.highwire.org\/content\/doi\/10.64898\/2026.01.12.699071","content-type":"unspecified","content-version":"vor","intended-application":"similarity-checking"}],"deposited":{"date-parts":[[2026,8,20]],"date-time":"2026-08-20T16:30:28Z","timestamp":1787243428000},"score":1,"resource":{"primary":{"URL":"http:\/\/biorxiv.org\/lookup\/doi\/10.64898\/2026.01.12.699071"}},"subtitle":[],"short-title":[],"issued":{"date-parts":[[2026,1,13]]},"references-count":52,"URL":"https:\/\/doi.org\/10.64898\/2026.01.12.699071","relation":{},"subject":[],"published":{"date-parts":[[2026,1,13]]},"subtype":"preprint"}}