T cell malignancies are a group of hematologic cancers with high recurrence and mortality rates. CD5 is highly expressed in X85% of T cell malignancies, although normal expression of CD5 is restricted to thymocytes, T cells, and B1 cells. However, CD5 expression on chimeric antigen receptor (CAR)-T cells leads to CAR-T cell fratricide. Once this limitation is overcome, CD5-targeting CAR-T therapy could be an attractive strategy to treat T cell malignancies. Here, we report the selection of novel CD5-targeting fully human heavy-chain variable (FHVH) domains for the development of a biepitopic CAR, termed FHV(H)3/V(H)1, containing FHV(H)1 and FHV(H)3, which were validated to bind different epitopes of the CD5 antigen. To prevent fratricide in CD5 CAR-T cells, we optimized the manufacturing procedures of a CRISPR-Cas9-based CD5 knockout (CD5KO) and lentiviral transduction of anti-CD5 CAR. In vitro and in vivo functional comparisons demonstrated that biepitopic CD5KO FHV(H)3/V(H)1 CAR-T cells exhibited enhanced and longer lasting efficacy; produced moderate levels of cytokine secretion; showed similar specificity profiles as either FHV(H)1, FHV(H)3, or the clinically tested H65; and is therefore suitable for further development.