Co-blockade of TGFβ and PD-1 Reinvigorates Glioblastoma-Infiltrating CD8+ T Cells That Characteristically Upregulate TGFβRI Expression

Abstract Purpose: Clinical trials have shown limited efficacy of anti–PD-1 treatment for glioblastoma (GBM). In this study, we examined the expression of TGFβ type I receptor (TGFβRI) in GBM-infiltrating CD8+ T cells and the characteristics of TGFβRI+CD8+ T cells. We examined the ex vivo effects of the co-blockade of PD-1 and TGFβ on the functions of GBM-infiltrating CD8+ T cells. Experimental Design: Using flow cytometry, we examined the phenotypes of tumor-infiltrating CD8+ T cells from newly diagnosed patients with GBM. We performed single-cell RNA/T-cell receptor sequencing to characterize the tumor-infiltrating TGFβRI+CD8+ T cells. We also examined the effects of co-blockade of PD-1 and TGFβ on the functions of tumor-infiltrating CD8+ T cells in ex vivo assays. Results: GBM-infiltrating CD8+ T cells expressed significantly increased levels of TGFβRI compared with peripheral blood CD8+ T cells. Among tumor-infiltrating CD8+ T cells, TGFβRI+CD8+ T cells exhibited increased expression of immune checkpoint inhibitory receptors, and tumor antigen–specific cells were enriched in TGFβRI+CD8+ T cells. Single-cell profiling revealed that tumor-infiltrating TGFBR1+CD8+ T cells demonstrated more clonal expansion and upregulation of T-cell receptor signaling genes compared with TGFBR1−CD8+ T cells. In vitro, anti-CD3 stimulation upregulated TGFβRI expression on CD8+ T cells. Patients with GBM with a high frequency of TGFβRI+CD8+ T cells presented with increased TGFβ signaling intensity. Importantly, combined blockade of PD-1 and TGFβ significantly enhanced the functions of tumor-infiltrating CD8+ T cells ex vivo. Conclusions: Our findings provide a basis for further investigation of the co-blockade of PD-1 and TGFβ for the treatment of patients with GBM.