chemoDetect

Title
Chemotherapy and APOBEC mutational signatures in tumor and healthy cells revealed by cell-free DNA monitoring

Authors
Gustav Alexander Poulsgaard¹, Mathilde Diekema¹, Amanda Frydendahl¹, Ludvig Renbo Olsen¹, Mads Heilskov Rasmussen¹, Søren Besenbacher^1,2, Britt Elmedal Laursen^1,3, Deirdre Cronin-Fenton^1,4, Radhakrishnan Sabarinathan⁵, Claus Lindbjerg Andersen¹, Jakob Skou Pedersen^1,2

Author affiliations

1. Department of Molecular Medicine, Department of Clinical Medicine, Aarhus University & Aarhus University Hospital, Denmark
2. Bioinformatics Research Centre, Aarhus University, Denmark
3. Department of Biomedicine, Aarhus University, Denmark
4. Department of Clinical Epidemiology, Department of Clinical Medicine, Aarhus University & Aarhus University Hospital, Denmark
5. National Centre for Biological Research, Tata Institute of Fundamental Research, Bengaluru, India

Abstract
Genotoxic chemotherapies such as 5-fluorouracil (5-FU) and oxaliplatin exert their cytotoxic effects through DNA damage. Yet their mutational consequences in healthy human tissues remain poorly understood. To track therapy-associated mutational processes in vivo, we analyzed read-level mismatches from 1,230 whole-genome-sequenced plasma cell-free DNA (cfDNA) samples collected longitudinally from 144 colorectal cancer patients, of which 88% received adjuvant chemotherapy. In cancer patients, cfDNA comprises both circulating tumor DNA (ctDNA) and DNA released from healthy cells. Paired-read consensus calling and stringent filtering yielded high-confidence cfDNA-fragment mismatch profiles, enabling detection of mutational signatures. Mixed-effects models revealed that chemotherapy exposure was associated with enrichment of the 5-FU mutational signature in ctDNA and platinum-associated signatures in both ctDNA and healthy cfDNA, alongside elevated APOBEC-related signatures independent of ctDNA. Time-series analyses showed persistent platinum and APOBEC signatures in healthy cfDNA more than one year after therapy. Together, these findings demonstrate that chemotherapy leaves durable mutational imprints in both cancer and healthy cells.