EXTH-135. Intrathecally delivered T cells precomplexed with a Bispecific T-cell Engager target CNS tumors

Abstract BACKGROUND Despite recent advances, glioblastoma (GBM) and leptomeningeal disease (LMD) continue to have a dismal clinical trajectory. Immunotherapeutic strategies, particularly targeted T-cell–based therapeutics, offer promising new approaches. We investigated a novel targeted T-cell strategy where naïve T cells were pre-complexed with a bispecific T-cell engager (BTE) targeting the tumor-associated antigen IL13Rα2 and delivered intrathecally (pBTE). This approach is designed to circumvent key limitations of CNS immunotherapy, including poor penetration across the blood-tumor barrier (BTB) and blood–CSF barrier (BCB), as well as the need to overcome the immunologically cold tumor microenvironment (TME). METHODS The retention of pBTE on T cells was assessed by flow cytometry. In vitro, cytotoxicity of pBTE to glioma cells was measured using a Cr-51 release assay. pBTE distribution was assessed via confocal imaging of 100 µm frozen brain sections. Intracranial and cisterna magna injections established GBM and triple-negative breast cancer LMD (LMD-BC) PDX models, respectively. Survival was assessed using Kaplan-Meier analysis, and significance was assessed via log-rank and t-tests. RESULTS In vitro, 95% of T cells retained surface-bound BTE after 24 hours and demonstrated significant cytotoxicity against GBM6 glioma cells (P = 0.0002). Confocal microscopy confirmed the localization of fluorescently labeled pBTE within perivascular spaces and the tumor bed at 6 hours post-injection. In vivo, pBTE significantly improved median survival in GBM6 NSG-MHC I/II DKO PDX models (55.5 vs. 35.5 days; P = 0.0007). A larger pBTE-Fc fusion construct similarly prolonged survival (46 vs. 34 days; P = 0.0043). In an LMD-BC model using MDA-MB-231 cells, pBTE improved survival (32 vs. 26 days; P = 0.0694), with complete tumor regression and long-term survival observed in 3 of 9 mice. CONCLUSION The IL13Rα2-specific BTE, precomplexed with T cells, showed potent in vitro and in vivo activity, and represents a novel immunotherapeutic strategy for GBM and LMD.