NKG2D-based CAR-T cells for synovial sarcoma: A preclinical proof-of-concept study.

The prognosis for advanced synovial sarcoma remains poor, with limited therapeutic options. Recently, cloned T cell receptor (TCR)-engineered T cell therapy has shown promising results in patients with advanced synovial sarcoma; however, the number of patients eligible for this therapy is limited owing to human leukocyte antigen (HLA) restriction. To solve this problem, we focused on chimeric antigen receptor (CAR)-T cells that target ligands of NKG2D, an activating receptor of natural killer cells. Because NKG2D recognizes a family of eight ligands, we hypothesized that NKG2D-based CARs could function as an intrinsic multi-antigen targeting platform, potentially mitigating antigen heterogeneity and antigen escape, which are key barriers in CAR-T therapy for solid tumors. We first evaluated the surface expression of NKG2D ligands (NKG2DLs) in synovial sarcoma cell lines and examined their transcript levels in public bulk RNA-seq datasets from synovial sarcoma tissues. We then constructed NKG2D-based, 4-1BB-co-stimulated CAR-T cells. These CAR-T cells showed effector responses and antitumor effects against synovial sarcoma cells in vitro, as demonstrated by the results of the intracellular cytokine production, cytokine secretion, CD107a degranulation assay, WST-8 assay, and real-time cell analysis, and in vivo in an NSG mouse xenograft model of synovial sarcoma. Although further mechanistic, translational, and safety validation studies are required, these findings provide a preliminary disease-specific preclinical proof of concept for HLA-independent NKG2D-based CAR-T cell therapy in synovial sarcoma, particularly in patients who are ineligible for or have not responded to TCR-T therapy.