To develop gene-modified T-cell-based antileukemia adoptive immunotherapy, concomitant administration of CD4+ and CD8+ T cells that have been gene modified using identical HLA class I-restricted leukemia antigen-specific T-cell receptor (TCR) gene transfer has not yet been fully investigated. Here, using CD4+ and CD8+ T cells that had been gene modified with a retroviral vector expressing HLA-A*24:02-restricted and Wilms’ tumor 1 (WT1)-specific TCR-α/β genes and siRNAs for endogenous TCRs (WT1-siTCR/CD4+ T cells and WT1-siTCR/CD8+ T cells), we examined the utility of this strategy. WT1-siTCR/CD4+ T cells sufficiently recognized leukemia cells in an HLA class I-restricted manner and provided target-specific Th1 help for WT1-siTCR/CD8+ T cells. By using a xenografted mouse model, we found that WT1-siTCR/CD4+ T cells migrated to leukemia sites and subsequently attracted WT1-siTCR/CD8+ T cells via chemotaxis. Therapy-oriented experiments revealed effective enhancement of leukemia suppression mediated by concomitant administration of WT1-siTCR/CD4+ T cells and WT1-siTCR/CD8+ T cells. Importantly, this augmented efficacy in the presence of WT1-siTCR/CD4+ T cells was correlated with longer survival and enhanced formation of memory T cells by WT1-siTCR/CD8+ T cells. Collectively, our experimental findings strongly suggest that this strategy would be clinically advantageous for the treatment of human leukemia.