Characteristics, origin, and potential for cancer diagnostics of ultrashort plasma cell-free DNA
- Irena Hudecova1,
- Christopher G. Smith1,
- Robert Hänsel-Hertsch2,
- Chandra Chilamakuri1,
- James A. Morris1,
- Aadhitthya Vijayaraghavan1,
- Katrin Heider1,
- Dineika Chandrananda3,
- Wendy N. Cooper1,
- Davina Gale1,
- Javier Garcia-Corbacho4,
- Simon C. Pacey5,
- Richard Baird5,
- Nitzan Rosenfeld1 and
- Florent Mouliere6,7
Abstract
Current evidence suggests that plasma cell-free DNA (cfDNA) is fragmented around a mode of 166 bp. Data supporting this view has been mainly acquired through the analysis of double-stranded cfDNA. The characteristics and diagnostic potential of single-stranded and damaged double-stranded cfDNA in healthy individuals and cancer patients remain unclear. Here, through a combination of high-affinity magnetic bead-based DNA extraction and single-stranded DNA sequencing library preparation (MB-ssDNA), we report the discovery of a large proportion of cfDNA fragments centred at ~50 bp. We show that these 'ultrashort' cfDNA fragments have a greater relative abundance in plasma of healthy individuals (median = 19.1% of all sequenced cfDNA fragments, n = 28) than in plasma of patients with cancer (median = 14.2%, n = 21, P < 0.0001). The ultrashort cfDNA fragments map to accessible chromatin regions of blood cells, particularly in promoter regions with the potential to adopt G-quadruplex (G4) DNA secondary structures. G4-positive promoter chromatin accessibility is significantly enriched in ultrashort plasma cfDNA fragments from healthy individuals relative to patients with cancers (P < 0.0001), in whom G4-cfDNA enrichment is inversely associated with copy number aberration-inferred tumor fractions. Our findings redraw the landscape of cfDNA fragmentation by identifying and characterizing a novel population of ultrashort plasma cfDNA fragments. Sequencing of MB-ssDNA libraries could facilitate the characterization of gene regulatory regions and DNA secondary structures via liquid biopsy. Our data underline the diagnostic potential of ultrashort cfDNA through classification for cancer patients.
- Received May 3, 2021.
- Accepted December 16, 2021.
- Published by Cold Spring Harbor Laboratory Press
This manuscript is Open Access.
This article, published in Genome Research, is available under a Creative Commons License (Attribution-NonCommercial 4.0 International license), as described at http://creativecommons.org/licenses/by-nc/4.0/.
