Pediatric cancers, including Ewing sarcoma (ES), are only weakly immunogenic and the tumor-patients' immune system often is devoid of effector T cells for tumor elimination. Based on expression profiling technology, targetable tumor-associated antigens (TAA) are identified and exploited for engineered T-cell therapy. Here, the specific recognition and lytic potential of transgenic allo-restricted CD8(+) T cells, directed against the ES-associated antigen 6-transmembrane epithelial antigen of the prostate 1 (STEAP1), was examined. Following repetitive STEAP1(130) peptide-driven stimulations with HLA-A*02:01(+) dendritic cells (DC), allo-restricted HLA-A*02:01(-) CD8(+) T cells were sorted with HLA-A*02:01/peptide multimers and expanded by limiting dilution. After functional analysis of suitable T cell clones via ELISpot, flow cytometry and xCELLigence assay, T cell receptors' (TCR) α- and β-chains were identified, cloned into retroviral vectors, codon optimized, transfected into HLA-A*02:01(-) primary T cell populations and tested again for specificity and lytic capacity in vitro and in a Rag2(-/-)γc(-/-) mouse model. Initially generated transgenic T cells specifically recognized STEAP1(130)-pulsed or transfected cells in the context of HLA-A*02:01 with minimal cross-reactivity as determined by specific interferon-γ (IFNγ) release, lysed cells and inhibited growth of HLA-A*02:01(+) ES lines more effectively than HLA-A*02:01(-) ES lines. In vivo tumor growth was inhibited more effectively with transgenic STEAP1(130)-specific T cells than with unspecific T cells. Our results identify TCRs capable of recognizing and inhibiting growth of STEAP1-expressing HLA-A*02:01(+) ES cells in vitro and in vivo in a highly restricted manner. As STEAP1 is overexpressed in a wide variety of cancers, we anticipate these STEAP1-specific TCRs to be potentially useful for immunotherapy of other STEAP1-expressing tumors.