Cell-free DNA from germline TP53 mutation carriers reflects cancer-like fragmentation patterns

Sep 21, 2026

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Germline pathogenic TP53 variants drive the high lifetime cancer risk characteristic of Li-Fraumeni syndrome (LFS). A fragmentomic analysis was performed on 199 cell-free DNA (cfDNA) samples from 82 TP53 mutation carriers and 30 TP53-wildtype controls to clarify functional drivers of cfDNA fragmentation in LFS. LFS shows increased prevalence of A/T nucleotides at fragment ends, dysregulated nucleosome positioning at p53 binding sites, and locus-specific shifts in chromatin accessibility. Machine learning distinguished TP53 mutant versus wildtype cfDNA (AUC 0.710–1.000), and longitudinal fragmentation signals supported early cancer detection.

LanguageEnglish

CategoryResearch & Report

FormatPDF

Pages16

Reading Time40 min

AccessFree to download

Abstract and background

Study cohort and objectives

Fragmentomic features and mechanisms

Classification and early detection approach

Results: patient cohort

Article [https://doi.org/10.1038/s41467-024-51529-w](https://doi.org/10.1038/s41467-024-51529-w) Cell-free DNA from germline TP53 mutation carriers reﬂect cancer-like fragmentation patterns Received: 11 September 2023 | Accepted: 7 August 2024 | | --- | | | | Check for updates | Derek Wong1, Maha Tageldein 1,2, Ping Luo 1, Erik Ensminger 1,2, Jeffrey Bruce1, Leslie Oldﬁeld1, Haifan Gong3, Nicholas William Fischer 3, Brianne Laverty2,3, Vallijah Subasri 2,3,4, Scott Davidson5,6, Reem Khan5,6, Anita Villani5,7, Adam Shlien 5,6, Raymond H. Kim 1,3,8,9 , David Malkin 2,6,8 & Trevor J. Pugh 1,2,9 Germline pathogenic TP53 variants predispose individuals to a high lifetime risk ofdeveloping multiple cancersand are the hallmark feature of Li-Fraumeni syndrome (LFS). Our group has previously shown that LFS patients harbor shorter plasma cell-free DNA fragmentation; independent of cancer status. To understand the functional underpinning of cfDNA fragmentation in LFS, we conducted a fragmentomic analysis of 199 cfDNA samples from 82 TP53 mutation carriers and 30 healthy TP53-wildtype controls. We ﬁnd that LFS individuals exhibit an increased prevalence of A/T nucleotides at fragment ends, dysregulated nucleosome positioning at p53 binding sites, and locispeciﬁc changes in chromatin accessibility at development-associated transcription factor binding sites and at cancer-associated open chromatin regions. Machine learning classiﬁcation resulted in robust differentiation between TP53 mutant versus wildtype cfDNA samples (AUC ROC = 0.710–1.000) and intra-patient longitudinal analysis of ctDNA fragmentation signal enabled early cancer detection. These results suggest that cfDNA fragmentation may be a useful diagnostic tool in LFS patients and provides an important baseline for cancer early detection. Somatic mutations in the tumor suppressor gene TP53 are the most common genetic alterations across all human cancers1 and germline TP53 variants are the hallmark feature of Li-Fraumeni Syndrome (LFS; OMIM \#151623), a highly penetrant hereditary cancer syndrome (HCS)2. Individuals with TP53 mutations (TP53m-carrier) have an estimated life-time risk of ~75% in males and ~100% in females for developing, often multiple, cancers3,4, most commonly soft tissue sarcoma, osteosarcoma, brain cancers, breast cancer, and adrenocortical carcinoma5,6. Cell-free DNA (cfDNA) are fragments of DNA released, mainly by apoptotic cells, into bodily ﬂuids such as lymphocytes into the blood plasma7. Sequencing of blood cfDNA has become an attractive method to explore for early cancer detection and monitoring in individuals with HCS, such as LFS8,9. One emerging “omic”, fragmentomics, is the 1Princess Margaret Cancer Center, University Health Network, Toronto, Ontario, Canada. 2Department of Medical Biophysics, University of Toronto, Toronto, Ontario, Canada. 3The Hospital for Sick Children, University of Toronto, Toronto, Ontario, Canada. 4Vector Institute, Toronto, Ontario, Canada. 5Genetics and Genome Biology, The Hospital for Sick Children Research Institute, Toronto, Ontario, Canada. 6Department of Pediatrics, University of Toronto, Torotno, Ontario, Canada. 7Division of Hematology/Oncology, The Hospital for Sick Children, Toroton, Ontario, Canada. 8Toronto General Hospital Research Institute, Toronto, Ontario, Canada. 9Ontario Institute of Cancer Research, Toronto, Ontario, Canada. e-mail: [Raymond.Kim@uhn.ca](Raymond.Kim@uhn.ca); [david.malkin@sickkids.ca](david.malkin@sickkids.ca); [trevor.pugh@utoronto.ca](trevor.pugh@utoronto.ca) study of cfDNA fragmentation and is based on ﬁndings that fragment preservation in circulation is related to nucleosome protection and highly correlated to the chromatin landscape of the cell-of-origin10. This is evidenced by a characteristic peak at 167 bp, corresponding to the length of DNA occupied by one nucleosome10. Studies have shown that TP53m-carriers, even prior to cancer onset, exhibit unique

FAQs

What was the main goal of analyzing cfDNA in Li-Fraumeni syndrome?

To understand the functional basis of cfDNA fragmentation patterns in LFS and assess whether cfDNA fragmentation can support cancer detection.

How were the samples collected and grouped in the study?

Plasma samples were obtained from TP53 mutation carriers and TP53-wildtype controls, with TP53 mutation carriers grouped by clinical cancer history at collection into LFS-Healthy, LFS-Past Cancer, and LFS-Active Cancer.

Which cfDNA fragmentation signals helped differentiate TP53 mutation carriers from wildtype controls?

LFS cfDNA showed increased A/T nucleotides at fragment ends, altered nucleosome positioning at p53 binding sites, and locus-specific changes in chromatin accessibility, enabling robust classification between mutant and wildtype samples.

Sep 21, 2026

Uploaded by Ezra