3335 - Genome-Wide Cell-Free DNA Fragmentation Analyses for Early, Non-Invasive Detection of Treatment Response to Standard of Care Adjuvant Radiation Treatment in High-Grade Gliomas

A. V. Annapragada¹, A. Bartolomucci¹, S. Short¹, K. Boyapati¹, A. Li², D. Roche², S. Patterson², D. M. Bruno², D. Kaplin², C. Jackson³, J. Weingart³, K. J. Redmond², V. Velculescu¹, R. Scharpf¹, J. Phallen¹, D. Mathios^4,5, and L. R. Kleinberg²; ¹The Sidney Kimmel Comprehensive Cancer Center, Johns Hopkins University School of Medicine, Baltimore, MD, ²Department of Radiation Oncology and Molecular Radiation Sciences, Johns Hopkins University School of Medicine, Baltimore, MD, ³Department of Neurosurgery, Johns Hopkins University School of Medicine, Baltimore, MD, ⁴Department of Neurological Surgery, Washington University School of Medicine, St. Louis, MO, ⁵The Brain Tumor Center, Siteman Cancer Center, Washington University School of Medicine, St. Louis, MO

Purpose/Objective(s): Biomarkers to assess response to standard chemoradiation following resection for high grade glioma (HGG) are imprecise. Assessments often require multiple MRIs months apart and fail to differentiate treatment response from progression, in a disease with median progression-free survival (PFS) around 7 months. We previously developed DELFI (DNA Evaluation of Fragments for Early Interception) to use cost-effective, low-coverage, whole-genome sequencing (WGS) and machine learning to evaluate millions of cfDNA fragments in the blood that can reflect cancer-related genomic and epigenomic changes. Here we analyze fragmentation in patients with HGG to identify noninvasive biomarkers of RT response and disease progression. Materials/

Methods: We enrolled 39 patients with primary HGG with 116 plasma liquid biopsies collected pre-surgery, pre-RT, 2, 4 and 6 weeks on-RT, and 1-month post-RT. We extracted cfDNA from 58 liquid biopsies (n=17 patients, with n=5, 16, 13, 8, 14 and 2 draws at the pre-surgery, pre-RT, 2, 4 and 6 weeks on-RT, and 1-month post-RT time points, respectively) and performed WGS of cfDNA fragments. We analyzed cfDNA fragmentation profiles by summarizing the ratio of short (100-150 bp) to long (151-220 bp) fragments in 5Mb bins genome-wide, and correlated profiles between timepoints for each patient. We estimated PFS as time between diagnosis and first indication of new treatment post-RT.
Results: Pre-surgery cfDNA concentrations trended higher than at pre-RT (mean 18.9 ng/mL vs. 9.7 ng/mL) and further decreased on-RT and post-RT (mean 5.9 and 5.6 ng/mL, respectively). Fragmentation profiles pre-RT were more correlated to on-RT profiles than to pre-surgery profiles (median correlation 0.94 vs. 0.88), possibly reflecting high pre-surgery tumor burden that falls after resection and adjuvant treatment. At four weeks on-RT, a higher correlation to the pre-RT fragmentation profile was associated with longer PFS (p=0.01, median PFS 15.6 vs. 4.8 months for patients with correlations above and below the mean, respectively), suggesting that patients with fragmentomes resembling their lowest tumor burden profile survive longer before progression.
Conclusion: We provide early evidence that genome-wide cfDNA fragmentation profiles reflect relative HGG tumor burden, and that changes to fragmentation may capture early molecular signs of progression. These changes are detectable on-RT and post-RT and are associated with PFS in as few as four weeks on-RT. This interim analysis demonstrates the utility of cfDNA fragmentomes for noninvasive monitoring of treatment response in patients with HGG. We are continuing patient accrual, sample collection and sequencing (goal n=100 patients enrolled) with the objective of identifying response biomarkers to support real time treatment modification.