Abstract 4560: Use of ctDNA fragmentomics as a predictive biomarker in locally advanced non-small cell lung cancer

Abstract Introduction: Locally advanced non-small cell lung cancer (NSCLC) is a serious condition with a poor 5-year survival rate of approximately 30%. Standard treatment involves chemoradiotherapy (CRT) followed by immunotherapy (IO). However, predicting benefit prior to CRT and monitoring response post treatment are areas of unmet need. Cancers, like many tissues, release fragments of DNA into the bloodstream, which can be detected using whole genome sequencing (WGS). Research has shown that cancer cells tend to release shorter DNA fragments, typically between 90-150 base pairs, at higher rates than normal tissues. This study aims to analyze cell-free DNA fragment sizes (fragmentomics) as a predictive tool for assessing treatment response in patients with LA NSCLC. This tumor-uninformed approach is cost-effective and can be implemented using routine blood samples. Methods: Low-pass WGS at 0.1x coverage was performed on 40 pre-treatment plasma samples from patients LA NSCLC obtained from the UCL biobank. The median follow-up for the cohort was 33 months. To investigate the cancer origin of short DNA fragments (90-150 bp), mutation calling was conducted using Mutect2. Mutation density was defined as the number of mutations per million bases (MPM). Tumor volumes were obtained using CT imaging, and the percentage quantity of short DNA fragments was plotted against tumor volume. Enrichment of short fragments (SFE) <20% was assessed and correlated with clinical outcomes, including relapse and overall survival. Results: Short DNA fragments (90-150 bp) had a significantly higher mutation density (973.8 MPM) compared to unselected fragments (381.7 MPM; p<0.0001). This elevated mutational burden supports the hypothesis that shorter fragments are of cancer origin. Linear regression analysis showed no significant relationship between tumor volume and the percentage of short DNA fragments for the entire cohort (R2 = 0.035, p = 0.35). However, in lung squamous cell carcinoma (LUSC), a strong positive correlation was observed (R2 = 0.6787, p = 0.003). Suggesting a potential quantitative link between tumor burden and ctDNA quantity in this subtype. SFE was observed in 31 of 40 patients (78%) prior to treatment. SFE was found in similar proportions in LUSC (78%; 15/19) and lung adenocarcinoma (75%; 15/20). Mortality occurred in 45% (14/31) of patients with SFE, compared to 11.1% (1/9) of those without enrichment. Among patients for whom consolidation IO was indicated, relapse rates were higher in those with SFE (44%; 7/16) than in those without enrichment (25%; 1/4). Findings of the research were limited by sample size; however further centers have been recruited to address this issue. These findings suggest that fragmentomic patterns represent a cost-effective, non-invasive prognostic biomarker for LA NSCLC, with particular potential in LUSC. Ongoing research is exploring patterns following treatment. Citation Format: Kishen R. Patel, Anisha Mistry, Robert Monfries, Florent Mouliere, John R. Conibear, James Wilson, Karim Keshwani, Wing K. Liu, Crispin Hiley. Use of ctDNA fragmentomics as a predictive biomarker in locally advanced non-small cell lung cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2025; Part 1 (Regular Abstracts); 2025 Apr 25-30; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2025;85(8_Suppl_1):Abstract nr 4560.