The programmed cell death-1 (PD-1)/programmed cell death ligand-1 pathway has been shown to limit cell-mediated effector functions during chronic viral infections impeding clearance of pathogens. As a strategy to reverse this exhaustion and increase T cell polyfunctionality, PD-1 ligands were blocked in vivo using a recombinant macaque PD-1 fused to a macaque Ig-Fc (rPD-1-Fc) in SIVmac239-infected rhesus macaques during the early chronic phase of infection, either alone or in combination with antiretroviral therapy. In vitro blockade showed improvement of Ag-specific CD4(+) and CD8(+) T cells from monkeys chronically infected with SIV. Of note, a prolonged 5-d blockade in culture was beneficial for both gag-specific CD4(+) and CD8(+) T cells based on proliferation and dual cytokine production. Although the in vivo administration of rPD-1-Fc induced enhanced SIV-specific CD4 and CD8 T cell proliferation both in the blood and gut, it failed to alter plasma viremia. However, rPD-1-Fc administration in the context of antiretroviral therapy interruption induced a significant delay of viral load rebound. In addition, rPD-1-Fc administration in MamuA*001(+) monkeys led to both an increase in the frequencies and Ki67 expression of GagCM9(+) CD8(+) T cells in the blood and rectal mucosa and polyfunctionality of GagCM9(+) CD8(+) T cells in blood. In conclusion, however, our data suggest that PD-1/programmed cell death ligand-1 blockade using soluble rPD-1-Fc instead of anti-PD-1 mAb, although effective in rescuing the effector function of SIV-specific CD4(+) and CD8(+) T cells during the early chronic phase of infection, has limited clinical benefit.