MHC class i-restricted TCR-transgenic CD4⁺ T cells against STEAP1 mediate local tumor control of ewing sarcoma in vivo

In this study we report the functional comparison of T cell receptor (TCR)-engineered major histocompatibility complex (MHC) class I-restricted CD4⁺ versus CD8⁺ T cells targeting a peptide from six transmembrane epithelial antigen of the prostate 1 (STEAP1) in the context of HLA-A*02:01. STEAP1 is a tumor-associated antigen, which is overexpressed in many cancers, including Ewing sarcoma (EwS). Based on previous observations, we postulated strong antitumor potential of tumor-redirected CD4⁺ T cells transduced with an HLA class I-restricted TCR against a STEAP1-derived peptide. We compared CD4⁺ T cell populations to their CD8⁺ counterparts in vitro using impedance-based xCELLigence and cytokine/granzyme release assays. We further compared antitumor activity of STEAP¹³⁰-TCR transgenic (tg) CD4⁺ versus CD8⁺ T cells in tumor-bearing xenografted Rag2^-/- γc^-/- mice. TCR tgCD4⁺ T cells showed increased cytotoxic features over time with similar functional avidity compared to tgCD8⁺ cells after 5–6 weeks of culture. In vivo, local tumor control was equal. Assessing metastatic organotropism of intraveniously (i.v.) injected tumors, only tgCD8⁺ cells were associated with reduced metastases. In this analysis, EwS-redirected tgCD4⁺ T cells contribute to local tumor control, but fail to control metastatic outgrowth in a model of xenografted EwS.