Physical activity specifically evokes release of cell-free DNA from granulocytes thereby affecting liquid biopsy

Abstract

Cell-free DNA (cfDNA) methylation-based diagnostics is a promising approach in oncology and hematooncology. Exercise impacts immune homeostasis and leads to a rapid and marked increase of cfDNA levels in blood. Since the origin of cfDNA during exercise remains elusive, the implications for liquid biopsy are unknown. In this study, we identified the source of cfDNA in 10 healthy untrained individuals before, immediately after, and 30 min after exercise, and in 6 patients with myeloid neoplasms or acute leukemia under resting conditions. A pyrosequencing assay was used to analyze the methylation levels of four CpGs, representing DNA from granulocytes, lymphocytes, monocytes, and non-hematopoietic cells. After exercise, cfDNA was almost exclusively released from granulocytes, with cell type specific proportions increasing significantly from 54.1% to 90.2%. Exercise did not trigger the release of cfDNA from lymphocytes or other analyzed cell types, whereas a small amount of cfDNA was released from monocytes. Compared to healthy people, patients with hematological malignancies show significantly higher cfDNA levels at rest with 48.1 (19.1; 78) vs. 8.5 (8.2; 9.5) ng/ml, data expressed as median (25th; 75th percentiles), and considerably higher levels of lymphocyte specific hypomethylated cg17587997 (P<.001). Hence, exercise-induced cfDNA elevations can compromise diagnostic accuracy.

How to Translate DNA Methylation Biomarkers Into Clinical Practice

“…Tissue composition can then be reliably estimated using deconvolution algorithms, as each cell only has two copies of chromosomal DNA ( Houseman et al, 2012 ; Schmidt et al, 2020 ). With such assays it is for instance possible to investigate the origin of cell-free DNA in blood plasma as a liquid biopsy of potentially affected tissues ( Moss et al, 2018 ; Neuberger et al, 2022 ).…”

“…For example, blood samples might be taken from venous blood or capillary blood by finger pricks, shipped as dried blood spot or at liquid state, frozen or at room temperature ( Sontag et al, 2022 ). For liquid biopsies, it may also be relevant if DNA is extracted from serum or plasma ( Constancio et al, 2020 ), or if the donor performed physical activity before sample collection ( Neuberger et al, 2022 ). Similar considerations are also required for specimen from urine, stool, airway, or other tissues ( Locke et al, 2019 ).…”