Abstract 1253: Single-cell analysis reveals intricate subclonal heterogeneity and evolution in response to standard of care treatments in glioblastoma

Abstract Glioblastoma multiforme (GBM) is the most common and lethal brain cancer. Standard of care treatments (SOC) are limited to surgery, radiation and chemotherapies, providing an average overall survival of only ∼16 months. Tumor heterogeneity and cellular plasticity are major contributions to treatment resistance in this disease. We constructed an integrated patient-level and cohort-level framework and analyzed single-nucleotide RNA sequencing data in matched primary and recurrence specimens to understand tumor heterogeneity, evolution, and drug resistance in SOC treated GBM patients (N=23). Patient-level tumor analysis revealed distinct tumor cell transcriptional subclones representing widespread intra-patient heterogeneity. Tumor evolution analysis from primary to recurrence revealed diverse treatment responses representing sensitive, intrinsic resistant, and acquired resistant subclones. Further characterization of tumor subclones and clustering analyses indicated that tumor heterogeneity under SOC treatments was linked to diverse biological mechanisms including neural development and oncogenic pathways, contributing to varying levels of subclonal resistance. Our analysis also highlighted intrinsic resistance was prominent as primary tumors often harbored substantial proportion of resistant tumor cells. This insight on resistant status at cell level then allowed us to compare resistant to sensitive cells and identify both established and potentially novel mechanisms of SOC resistance that were not always obvious in the comparison between primary and recurrence samples. These included activation of PI3K/AKT, MAPK, TGF-β, EMT, and WNT signaling pathways. Taken together, our novel analytical strategy revealed complex tumor heterogeneity and evolution landscape in SOC treated GBM, offering insights on potential mechanisms driving these phenomena and providing a resource for further exploration and development of therapeutic strategies. Citation Format: Yueshan Zhao, Xiaoyu Lu, Verah Nyarige, Junfei Zhao, Gonzalo Lopez, Ann Forslund, Maria Ortiz-Estevez, Jorge Benitez-Hernandez, Wei Zhang, Celia Fontanillo, Kai Wang, Ha Dang. Single-cell analysis reveals intricate subclonal heterogeneity and evolution in response to standard of care treatments in glioblastoma [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 1253.