Fragmentation-Induced Coverage Biases in Cell-Free DNA Sequencing Affect the Clinical Sensitivity of Liquid Biopsies

Area of Science:

• Genomics
• Molecular Biology
• Cancer Research

Background:

• Cell-free DNA (cfDNA) in blood exhibits epigenetic-dependent fragmentation, differing from stable genomic DNA.
• Extensive cfDNA fragmentation can cause uneven sequencing coverage and loss of genetic information.
• The impact of cfDNA fragmentation on detecting clinically relevant variants non-invasively is not fully understood.

Purpose of the Study:

• To investigate the effects of cfDNA fragmentation on variant detection sensitivity in liquid biopsies.
• To analyze locus-specific and allele-specific coverage biases in cfDNA sequencing.
• To assess the implications of these biases for cancer variant detection, disease tracking, and clonal hematopoiesis assessment.

Main Methods:

• Analysis of clinical genomic data from 67,129 samples across 99 cancer cohorts.
• Examination of 0.4 million variants, focusing on known oncogenic hotspots in genes like TP53, EGFR, and NRAS.
• Longitudinal analysis of cfDNA samples to evaluate disease tracking reliability.

Main Results:

• cfDNA fragmentation introduces locus-specific and allele-specific coverage biases in liquid biopsies.
• These biases affect the sensitivity of detecting somatic variants in clinically actionable genes.
• Biases can compromise longitudinal disease tracking, potentially leading to false negatives for minimal residual disease and clonal extinction.

Conclusions:

• cfDNA fragmentation significantly impacts the reliability of variant detection and disease monitoring in liquid biopsies.
• Caution is advised when interpreting cfDNA sequencing data, especially for assessing clonal hematopoiesis.
• Fragmentation-driven biases necessitate careful consideration for accurate non-invasive cancer diagnostics and monitoring.