Cell-free DNA-derived immune cell ratios uncover cancer-associated systemic changes

Abstract Genome-wide coverage patterns of plasma cell-free DNA (cfDNA) fragments reflect nucleosome positioning in the cells of origin, enabling non-invasive inference of cell-type contributions and transcriptional activity. While the majority of cfDNA originates from hematopoietic cells, the diagnostic and biological relevance of this fraction remains underexplored. Here, we performed cfDNA-based deconvolution of blood cell types by integrating transcription start site (TSS) coverage profiles from plasma whole-genome sequencing with single-cell transcriptomic reference data. By correlating cfDNA TSS coverage with gene expression across 457 blood cell types, we ranked their relative contributions to the cfDNA pool. We analyzed 788 pre-treatment and longitudinal plasma samples from patients with localized colorectal cancer (CRC), muscle-invasive bladder cancer (MIBC), as well as 30 samples from healthy controls. In healthy individuals, cfDNA TSS coverage profiles reflected blood gene expression, and the inferred cell type contributions recapitulated the known hematopoietic composition. In cancer patients, we observed a significant increase in cfDNA contributions from lymphocytes, including T cells and plasma cells, and decreased contributions from monocytes and granulocytes. These immune-derived signatures distinguished CRC (AUC=0.793) and MIBC (AUC=0.745) patients from healthy controls. Longitudinal analysis of immune cell-type contributions revealed treatment-associated changes in the relative abundance of classical monocytes and plasma cells, although these temporal dynamics were not predictive of relapse or outcome. Together, these findings suggest that cfDNA-derived immune signatures may capture aspects of systemic immune remodeling in cancer, potentially providing a complementary non-invasive biomarker in liquid biopsies beyond tumor-derived signals.