A comparative analysis of CD70-directed CAR-T cells for glioblastoma treatment demonstrates a superior efficacy of the ligand-based construct.

CD70, a member of the tumor necrosis factor receptor superfamily, is expressed in glioblastoma (GB), where it promotes tumor growth, migration, and immunosuppression. Accordingly, it has emerged as a target for chimeric antigen receptor (CAR)-T cell therapy. Despite the influence of CAR structure on therapeutic efficacy, no comparative studies have evaluated different CD70-directed CAR designs in GB. Our study addressed this gap. We first validated CD70 expression in transcriptomic datasets, patient tissue, and GB cell lines. We then generated CD70-specific CAR-T cells featuring distinct target recognition and co-stimulatory domains (CD27z, LF28z, and LFBBz) and performed phenotypic characterization. Using co-culture systems and 3D cerebral organoids, we showed that all constructs eliminated target cells in a CD70-dependent manner, with CD27z secreting the highest levels of Th1 cytokines. This functional advantage translated into superior survival in an orthotopic GB mouse model. Based on these findings, we developed a panel of murine CD27-based constructs, all of which demonstrated potent antitumor activity in vitro and in immunocompetent GB mouse models, further underscoring the therapeutic promise of CD27 integration into the CAR design. Collectively, our comparative analysis highlights the superior efficacy of the ligand-based construct, supporting its incorporation into a clinical trial targeting CD70 in recurrent GB.