Immune checkpoint inhibitors, such as PD-1 and TIGIT, are crucial dysregulators of CD8 T cell function during chronic HIV-1/SIV infections. Importantly, these checkpoint inhibitors are highly expressed on the surface of CD4 T cells that harbor latent HIV. We have previously demonstrated that an anti-HIV/anti-CD3 bispecific T-cell engager (BiTE) can be used to redirect functionally compromised follicular CD8 (fCD8) T cells to kill HIV infected cells in vitro. Here, we hypothesize that adding an extra specificity to target PD-1 and TIGIT to the BiTE will further enhance the functional activities of CD8 T cells and simultaneously lower the threshold for reactivation of latently infected cells in HIV infection.
We generated trifunctional anti-HIV/anti-CD3/anti-PD-1 and anti-TIGIT T-cell engagers by linking the scFvs from either anti-TIGIT and anti-PD-1 antibodies to the BiTE molecule. HIV-infected cell lines and primary cells isolated from lymph nodes (LN) of chronically SHIV-infected rhesus macaques were used as target cells in the in vitro and ex vivo killing assays to test whether trifunctional T-cell engagers could enhance the cytolytic activities of functionally compromised CD8 T cells. We also examined whether trifunctional T-cell engagers could stimulate CD8 T-cell lysis of HIV-infected CD4 T cells using a primary HIV latency model. Furthermore, multiparameter flow cytometry was used to investigate the effects of trifunctional T-cell engagers on CD8 and CD4 T cell polyfunctionality.
We found that trifunctional anti-HIV/anti-CD3/anti-TIGIT and anti-PD1 increased the killing capability of CD8 T cells compared to the bifunctional anti-HIV/anti-CD3 BiTE in the in vitro and ex vivo killing assays. We also demonstrated that trifunctional anti-HIV-1/anti-CD3/anti-TIGIT and anti-PD1 enhanced the CD8 T cell lysis of latently infected cells. Furthermore, trifunctional anti-HIV-1/anti-CD3/anti-TIGIT and anti-PD1 were shown to increase antigen-specific CD107a degranulation, levels of granzyme B, cytokine and chemokine release by CD8 T cells, which could potentially underlie the observed increase in the killing capability of CD8 T cells.
Our results indicate that the use of trifunctional T cell engagers targeting immune checkpoints, PD1 and TIGIT, may serve as novel immunotherapeutic strategies to eliminate infected cells in HIV infected individuals.