The development and potent antitumor efficacy of CD44/CD133 dual-targeting IL7Rα-armored CAR-T cells against glioblastoma

Tumor heterogeneity and an immunosuppressive microenvironment pose significant challenges for immunotherapy against solid tumors, particularly glioblastoma multiforme (GBM). Recent studies have highlighted the crucial role of glioma stem cells (GSCs) in tumor recurrence and therapeutic resistance. In this context, we developed a tandem chimeric antigen receptor (CAR)-T cell targeting CD44 and CD133 (PROM1), containing a truncated IL-7 receptor alpha intracellular domain (Delta 7R) between the CD28 costimulatory receptor and the CD3 zeta signaling chain (Tan zeta-T28-Delta 7R). Our target identification and validation were carried out using GSCs, samples from GBM patients, and the corresponding sequencing data. The antitumor efficacy of CAR-T cells was evaluated in patient-derived GSCs, intracranial xenograft models, patient-derived xenograft models, and glioblastoma organoids (GBOs). Single-cell RNA sequencing and mass cytometry were used to determine the immune phenotypes of CAR-T cells. We showed that locoregionally administered Tan zeta-T28-Delta 7R CAR-T cells induced longterm tumor regression with the desired safety outcomes. Patient-derived autologous Tan zeta-T28-Delta 7R CAR-T cells showed robust antitumor activity against GBOs. Our pre-clinical data has demonstrated the translational potential of Tan zeta-T28-Delta 7R CAR-T cell against GBM.