Bispecific T-cell engager constructs and future directions. The ‘classical’ T-cell engager structure, consisting of two antigen-binding variable fragments, devoid of the Fc domain of their parent antibody, linked by a short flexible linker region. It tethers the epsilon subunit of the T cell to EGFRvIII expressing tumor cells, activating the T cell, which then releases granzymes and perforins, resulting in tumor cell death. This approach is currently being evaluated in phase I clinical trials (NCT04903795 and NCT03296696) (B) CiTEs include the extracellular domain of a checkpoint inhibitor (such as PD-1) fused to a traditional T-cell engager scaffold. This allows for synergistic checkpoint blockade alongside T-cell tethering and activation and has demonstrated enhanced efficacy in vitro and in vivo in acute myeloid leukemia (AML).129 (C) SMiTEs are constructed from two separate classical T-cell engagers which target separate antigens. These can offer costimulation of tethered T cells and provide regional checkpoint blockade while also offering the traditional lytic effects formed by the tumor–T cell synapse. (D) TRiTEs can tether T cells to the tumor while also delivering costimulatory signals (eg, via interaction with CD28). (E) CAR T cells which can be engineered to secrete T-cell engagers have been described by Choi et al, who developed a bicistronic construct that resulted in expression of a CAR specific for EGFRvIII, which could also secrete T-cell engagers specific for EGFR wild type, which would only have effect in the local tumor environment. These demonstrated promising efficacy against heterogeneous mouse models of glioblastoma.60 (F) DART proteins are a novel take on the bispecific construct, where two variable fragment chains are linked by disulfide bonds and non-covalent forces, which may result in greater stability and enhanced cytotoxicity.142 CAR, chimeric antigen receptor; CiTE, checkpoint inhibitory T-cell engager; DART, dual-affinity retargeting; EGFR, epidermal growth factor receptor; SMiTE, simultaneous multiple interaction T-cell engager; TCR, T-cell receptor; TRiTE, trispecific T-cell engager.