Modulating CD226 and PD-(L)1 pathways improves CMV-specific CD8＋T cell responses in the absence of IL-2

Abstract

Glioblastoma (GBM) frequently expresses cytomegalovirus (CMV)antigens, making CMV-specific CD8＋T cells attractive candidatesfor adoptive immunotherapy due to their longevity andinherent tumor reactivity. However, these T cells encountersignificant immunosuppressive challenges within the GBMmicroenvironment, including cytokine scarcity and checkpointmediatedinhibition, which limit their proliferation and function. Here, we assessed strategies to overcome these limitationsby modulating immune checkpoint pathways. Antigenstimulation combined with IL-2 robustly expanded high-avidity(tetramer-high) CMV-specific T cells with significant enrichmentof CD62L＋ central memory (TCM) cells. In contrast, antigenstimulation alone modestly expanded tetramer-high cells withlimited TCM enrichment. PD-L1 blockade in the absence of IL-2favored expansion of tetramer-high CMV-specific CD8＋T cells,preserved CD62L expression, and enhanced CD226 expression. Furthermore, combining anti-PD-L1 blockade with ananti-CD226 agonist markedly enhanced proliferation, IFN-？production, and TCM enrichment in both tetramer-high andtetramer-low populations, reaching levels comparable to IL-2-supported conditions. Together, these findings highlight thatsimultaneous modulation of PD-L1 and CD226 pathways canrestore CMV-specific T cell function, offering a promising strategyto boost TCR-T efficacy in cytokine-deprived environments.