IMMU-12. Locoregional delivery of GPC2 CAR T cells is safe and effective in preclinical models of pediatric central nervous system tumors

Pediatric brain tumors are the leading cause of cancer death in children and new targeted therapies are urgently needed. Glypican 2 (GPC2) is highly expressed on multiple pediatric central nervous system (CNS) tumors, including embryonal tumors and high-grade gliomas. Following initial proof-of-concept using mRNA-based CAR T cells, we optimized a lentiviral-based GPC2 CAR using a D3 GPC2 antibody with CD28 transmembrane and 4-1BB/CD3zeta co-stimulatory domains (D3-GPC2-BBz CAR) which is being tested in neuroblastoma (NCT05650749). Here we tested D3-GPC2-BBz CARs in GPC2-expressing pediatric CNS tumors. D3-GPC2-BBz CAR T cells were selectively cytotoxic to group 3 medulloblastoma cell lines in vitro: 7316-4509 showed >75% reduction in cell viability at 1:5 E:T ratio, p<0.01, and 7316-10374 showed >75% reduction in cell viability at 1:1 E:T ratio, p<0.05. Similarly, GPC2 CAR T cells showed robust antitumor activity against a H3G34 high-grade glioma cell line KNS42: 69% reduction in cell viability at 5:1 E:T ratio, p<0.001. In a 7316-4509 group 3 orthotopic xenograft, lateral ventricle delivery of a single 1e6 dose of D3-GPC2-BBz CAR T cells resulted in durable tumor regression (mean radiance at 95 days 1.40e6 vs. 1.09e9 for control CD19 CARs, p<0.0001) with substantially prolonged overall survival (OS; median survival >200 days vs. 147 days for CD19 CARs, p<0.0001). In a thalamic H3K27M DMG orthotopic xenograft model (7316-3058), lateral ventricle delivery of a single 1e6 dose of GPC2 CAR T cells also significantly extended OS (87 vs. 69 days for CD19 CARs, p<0.0001). No toxicity was observed in GPC2 CAR-treated animals, including no evidence of tumor inflammation-associated neurotoxicity. Overall, ventricular delivery of D3-GPC2-BBz CAR T cells showed potent antitumor efficacy in preclinical models of pediatric CNS tumors without toxicity, supporting the clinical testing of GPC2 CAR T cells in children with GPC2-expressing CNS tumors.