Cell-free DNA methylome and fragmentome analysis for disease relapse monitoring in patients with Ewing Sarcoma

Liquid biopsies and cell-free DNA (cfDNA) offer minimally invasive methods for the diagnosis and monitoring of Ewing Sarcoma (EwS). EwS have a low tumour mutational burden and their detection with plasma cfDNA is challenging. We hypothesised that analysing the cfDNA methylome and fragmentome could enhance sensitivity for detecting EwS and identifying early disease recurrence. We conducted whole-genome and methylome sequencing of cfDNA from 68 serial samples of 15 patients with EwS and 3 patients with CIC-rearranged sarcoma (CIC). With EwingSign, a new machine learning model, we identified EwS or CIC in a test set for 10 out of 12 patients at diagnosis and 15 out of 18 clinically confirmed relapse events. 0 out of 29 non-cancer controls were detected positive with EwingSign. These findings indicate that cfDNA methylome and fragmentome analysis, if validated in a larger cohort, could improve disease detection, monitoring and relapse identification in patients with EwS.