ChIP-seq of Urinary Cell-Free Chromatin Infers Tissue Origin and Detects Tumor-Derived Cell-Free DNA in Bladder Cancer

Abstract

AbstractUrinary Cell-free DNA (cfDNA) holds valuable molecular insights into the processes occurring within the urinary system. The analysis of epigenetic markers such as histone post-translational modifications in both cells and in blood circulation offers detailed information regarding cell identity and underlying state. However it remains unclear whether urine contains cell-free chromatin with intact epigenetic marks. Here, we extend the use of cell-free chromatin immunoprecipitation followed by sequencing (cfChIP-seq) to human urine. We demonstrate that cell-free nucleosomes captured in urine preserve multiple histone post-translational modifications indicative of activation and repression. By analyzing the H3K4me3 promoter mark, we identified the primary tissues contributing to cfDNA in healthy individuals. Notably, we observe distinct populations of circulating nucleosomes in urine and plasma samples with a contribution from the kidney in healthy donors that are not detected in matched urine exfoliated cells or matching plasma samples. Additionally, we show that urine cf-nucleosomes can be used to detect pathologically driven changes in the urine of bladder cancer patients, reflecting tumor-associated transcriptional programs and immune responses. The cfChIP-seq signal from cancer and immune origins can be used to accurately identify cancer patient samples, achieving an area under the curve (AUC) of 0.97 in a validation set. Our findings highlight the potential of urine cell-free nucleosomes as accessible, noninvasive biomarkers for both basic research in renal physiology and monitoring urinary pathologies.Key FindingsUrine cell-free nucleosomes retain multiple histone marks informative of gene promoters and enhancers and can be successfully immunoprecipitated.Urine cfChIP-seq identifies bladder, kidney, and immune cells as the major contributing organs to the pool of urine cell-free nucleosomes.The populations of cell-free nucleosomes in urine and blood are distinct and mostly disjoint. Suggesting that there is little if any transport of nucleosomes across the blood-urine interface.We can detect pathologically driven changes in cell-free nucleosomes from the tumors and immune cells responding to the tumor.Urine cell-free nucleosomes as potential noninvasive biomarkers for detections of non-muscle invasive bladder cancer