Abstract 3444: Spatial profiling of recurrent glioblastoma in a Phase I clinical trial reveals favorable immune remodeling induced by intracerebroventricular CAR T therapy

Abstract Recurrent glioblastoma (rGBM) is an aggressive brain tumor with median survival under one year after standard chemoradiation. Antigen heterogeneity, immune exclusion, and a suppressive tumor microenvironment (TME) limit responses to immunotherapy. A first-in-human phase 1 trial of intracerebroventricular EGFR/IL13Rα2 CAR T cells (CART-EGFR-IL13Rα2) in EGFR-amplified rGBM was feasible, produced manageable neurotoxicity, and induced radiographic tumor regressions in a subset of patients (NCT05168423).To understand how this therapy reshapes the local TME, we analyzed paired tumor resections from 6 patients enrolled in the phase 1 trial, with specimens obtained from the primary intracranial disease site at trial enrollment (pre-treatment) and at radiographic progression after CART-EGFR-IL13Rα2 infusion. Multimodal spatial profiling included regional transcriptomic and protein mapping (GeoMx), single-cell whole-transcriptome imaging (CosMx), and high-resolution spatial transcriptomics (Visium HD). We annotated tumor, myeloid, lymphoid, and stromal compartments and derived composite scores for stemness, invasion, cell death, and immune regulation. Neighborhood- and interaction-based analyses were used to compare cellular states and cell-cell communication.Across patients, post-treatment samples showed reduced expression of CAR target antigen and a shift in tumor-intrinsic programs toward less stem-like, less migratory, and more apoptotic states, despite radiographic progression. The post-treatment TME was remodeled, with fewer suppressive myeloid- and B-cell-rich niches and increases in interferon-responsive and T cell-associated activation programs. Spatial interaction analyses indicated that pre-treatment rGBM contained dense networks of myeloid-tumor and myeloid-T-cell contacts consistent with impaired antigen presentation and effector function. Post-treatment specimens, in contrast, showed partial disruption of these suppressive circuits and the emergence of microenvironments more permissive to T-cell infiltration and activity.In the parent phase 1 trial, CART-EGFR-IL13Rα2 was feasible & induced radiographic tumor regressions in a subset of patients. This correlative spatial analysis suggests that prior EGFR/IL13Rα2 CAR T exposure can leave a less suppressive, more immunologically engaged TME at the primary site, even in resections obtained at radiographic progression. Together, these data support the idea that intracerebroventricular CAR T therapy may condition rGBM for subsequent immunotherapy. Myeloid and B-cell interactions are highlighted as candidate targets for armoring next-generation CAR T cells and for designing rational combination and sequencing strategies. Citation Format: Wesley V. Wilson, MacLean P. Nasrallah, Nakial Cross, Yael A. Day, Vanessa Gonzalez, Rachel M. Leskowitz, Amy Marshall, Julie K. Jadlowsky, Gabriela Plesa, Donald L. Siegel, Elizabeth O. Hexner, Joseph A. Fraietta, Carl H. June, Stephen J. Bagley, Donald O’Rourke, Zev Binder, Andrew J. Rech. Spatial profiling of recurrent glioblastoma in a Phase I clinical trial reveals favorable immune remodeling induced by intracerebroventricular CAR T therapy [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 3444.