Remote immune processes revealed by immune-derived circulating cell-free DNA

Blood cell counts often fail to report on immune processes occurring in remote tissues. Here we use 25 immune cell type-specific methylation patterns in circulating cell-free DNA (cfDNA) for studying 26 human immune cell dynamics. We characterized cfDNA released from specific immune cell types in 27 healthy individuals (N=242), cross sectionally and longitudinally. Immune cfDNA levels had no 28 individual steady state as opposed to blood cell counts, suggesting that cfDNA concentration reflects 29 adjustment of cell survival to maintain homeostatic cell numbers. We also observed selective elevation 30 of immune-derived cfDNA upon perturbations of immune homeostasis. Following influenza 31 vaccination (N=92), B-cell-derived cfDNA levels increased prior to elevated B-cell counts and 32 predicted efficacy of antibody production. Patients with Eosinophilic Esophagitis (N=21) and B-cell 33 lymphoma (N=27) showed selective elevation of eosinophil and B-cell cfDNA respectively, which 34 were undetectable by cell counts in blood. Immune-derived cfDNA provides a novel biomarker for 35 monitoring immune responses to physiological and pathological processes that are not accessible using 36 conventional methods. To further examine the relative presence of immune cfDNA in plasma and whole blood, we employed an independent set of samples and an independent technology to measure and interpret methylation markers. We performed deconvolution of methylomes obtained by whole genome bisulfite sequencing (85x coverage), on genomic DNA from whole blood, and matched plasma cfDNA from 23 healthy donors. *** Raw data is unavailable due to patient privacy concerns ***