<p dir="ltr"> Bispecific chimeric antigen receptor (CAR) T cell therapy has demonstrated significant success in B cell-related hematologic cancers. However, the application of CAR-T therapy in treating T-cell leukemia/lymphoma poses challenges due to the risk of T-cell fratricide resulting from shared antigens between normal T cells and malignant cells. To address this issue and to enhance the efficacy of CAR-T therapy, we first developed a novel approach involving the selection of target antigen-negative T cells through one-step magnetic selection and subsequent transduction with bispecific CAR gene. We then investigated the expansion, in vitro cytotoxicity, and in vivo anti-tumor activity of this novel bispecific CARs in T-cell malignancies.</p>