Plasma Cell-Free DNA-Based Next-Generation Sequencing for Molecular Profiling of Diffuse Gliomas

Abstract

To evaluate the utility of plasma-derived cell-free DNA (cfDNA) using targeted next-generation sequencing (NGS) for molecular profiling of diffuse gliomas (DG) by assessing concordance with matched tumor tissue DNA, mutation profiles, variant allele frequencies, therapeutically actionable alterations, and survival associations. A total of 30 DG cases were included and 10 healthy control were included. Paired plasma cfDNA and formalin-fixed paraffin-embedded tumor tissue DNA were analyzed using targeted next-generation sequencing (NGS). Plasma cfDNA concentrations were higher in patients with DG (n = 30) than in healthy controls (P = .0002), though this comparison was limited by the small, unmatched control cohort (n = 10). Targeted sequencing predominantly identified single nucleotide variants (81%). Gene-level concordance between plasma cfDNA and tumor tissue DNA was 90.0%, whereas ≥1 matched variant was identified in 76.7% of cases. Plasma cfDNA detectability and concordance varied across diffuse glioma subtypes, and multiple therapeutically actionable alterations involving receptor tyrosine kinase, MAPK, PI3K/AKT/mTOR, and mismatch repair pathways were identified. Higher plasma cfDNA concentrations were associated with shorter survival, whereas cases with matched plasma and tissue variants demonstrated longer survival; however, these associations were not statistically significant. Plasma cfDNA-based liquid biopsy may serve as a minimally invasive approach for molecular profiling of diffuse gliomas, demonstrating high gene-level concordance with tumor tissue DNA and detection of clinically relevant, potentially actionable genomic alterations. Although variant-level concordance and detectability varied across glioma subtypes, these findings support a complementary role for plasma cfDNA analysis in diffuse gliomas. Further prospective studies, including serial sampling and matched leukocyte controls, are needed to improve analytical sensitivity and define its clinical utility.