99P The differentially fragmented regions in cell-free DNA of colorectal cancer patients

Cell-free DNA analysis in cancer patients may reveal tumor-specific alterations, including mutations, aberrant methylation, and chromatin state changes. The latter are connected to shifts in transcription programs and may be considered as epigenetic markers of cancer. Several whole-genome sequencing studies have shown that the chromatin state of parental cells may be partially decoded from cfDNA fragmentation features. In this preliminary study, we aim to test an approach for identifying differentially fragmented regions in cell-free DNA of cancer patients. The study involved cfDNA samples from 24 patients with colon adenocarcinoma, stages I (n=3), II (n=6), III (n=7), and IV (n=8), and 13 healthy individuals. We designed a targeted NGS panel based on a single-primer extension protocol to analyze the lengths of cfDNA fragments. The panel comprised 81 primers covering COAD-specific open-chromatin regions extracted previously from TCGA data. We developed a novel approach for targeted cfDNA fragmentation analysis in multiply regions of interest. We have experimentally identified 10 genomic regions with significant (p<0.01) differences in cfDNA fragmentation in patients with stage IV colorectal cancer and healthy controls. Our evidence supports the concept that targeted analysis of cfDNA fragmentation may facilitate the detection of tumor presence. This strategy may reveal a novel class of cost-effective biomarkers that will serve as analytes themselves or might be incorporated into multidimensional liquid biopsy assays in combination with somatic mutations or aberrant methylation.