The most effective bispecific antibody (bsAb) to date has been the bispecific T-cell engager (BiTE) blinatumomab against CD19 and CD3. Nevertheless, a sizeable portion of patients did not respond to treatments or eventually relapsed after an initial response, and the recurrence rate increased significantly due to escape or downregulation of the CD19 antigen. 

For a study, researchers created a unique method to create a CD19/CD22/CD3 trispecific antibody (tsAb) by site-specifically fusing anti-CD19 scFv (FMC63) and anti-CD22 nanobody (Nb25) to the designated locations of the CD3 antigen-binding fragment (Fab, SP34) to increase anticancer activity and prevent immune escape. Through the use of the technique, immunological synapses between target cells and T cells—mediated by CD19/CD22/CD3—can be formed as effectively as possible. 

Optimized tsAb showed significantly improved antitumor efficacy and the capacity to override immune escape compared with the corresponding bsAbs alone or in combination, as well as with blinatumomab, and can be superior for inducing T-cell-specific cytotoxicity and cytokine production against CD19+ and/or CD22+ tumor cells. Additionally, tsAb therapy can considerably extend survival and result in the long-term eradication of main B-ALL patient samples in the PDX model. Site-specific recombination was used in the innovative technique, which offered a unique insight into the structural optimization of T-cell-redirected multispecific antibodies. 

It may have broad applicability to solid tumors and diverse and resistant tumor populations.