Bispecific antibodies cross-link and redirect CD8 ⁺ T cells to kill cytomegalovirus-infected cells

Adoptive transfer studies of ex vivo-expanded autologous cytomegalovirus (CMV)-specific CD8⁺ T lymphocytes (CTLs) in immunocompromised hosts demonstrate a central role for viral control, but this therapeutic approach is not generally feasible. We present T cell-redirecting bispecific antibodies (TRBAs) that cross-link CD3ε on CTLs to gH protein on CMV-infected cells, including versions where a single-chain antibody is appended to the carboxyl terminus of the heavy chain of a regular antibody, or where both light and heavy chains have two tandem variable regions in a crossover ordered design. Both versions bind both antigens and mediate the specific clearance of infected cells with the release of interferon-γ when added to CTLs with CMV-infected cells. TRBA-mediated clearance is visualized with physical clustering of CTLs with CMV-infected cells. These results illustrate the mechanism and utility of these TRBAs as potential therapeutic candidates for disseminated CMV infection, with potential advantages compared to two prior CMV-specific TRBAs.