Abstract 2689: Single-cell multiome sequencing reveals distinct transcriptional programs associated with ecDNA amplifications in medulloblastoma.

Abstract Extrachromosomal DNA (ecDNA) is a key driver of intratumoral heterogeneity and has been linked to therapeutic resistance and poor patient outcomes. However, technical limitations in identifying, isolating, and analyzing ecDNA at the single-cell level have hindered our understanding of its role in tumor development and treatment response. We have previously developed a single-cell multiome(RNA + ATAC) sequencing approach that enables the simultaneous analyses of ecDNA and its gene expression profiles. Initial application of this method in a medulloblastoma tumor of the sonic hedgehog subgroup revealed distinct transcriptional signatures in ecDNA-positive compared to ecDNA-negative and non-malignant cells. Building on these findings, we performed multiome single-cell sequencing in an additional cohort of medulloblastoma specimens harboring ecDNA amplifications. Initial analyses revealed distinct functional characteristics between ecDNA-positive and ecDNA-negative tumor cells. For example, ecDNA-positive tumor cells show up-regulation of DNA replication, recombination, and damage repair pathways, while ecDNA-negative cells demonstrated increased expression of synaptic activity, cell signaling, and morphogenesis pathways. Our ongoing analysis focuses on further defining the transcriptional programs that distinguish ecDNA-positive from ecDNA-negative tumor cells, ultimately informing future functional genetic and pharmacological studies targeting transcriptional dependencies in ecDNA-driven medulloblastoma tumors. Citation Format: Hui Hui, Yan Yuen Lo, Jessica Wang, Rishaan Kenkre, Jon D. Larson, Sunita Sridhar, Owen Chapman, Lukas Chavez. Single-cell multiome sequencing reveals distinct transcriptional programs associated with ecDNA amplifications in medulloblastoma [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2026; Part 1 (Regular Abstracts); 2026 Apr 17-22; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2026;86(7 Suppl):Abstract nr 2689.