Chimeric antigen receptors (CARs) are artificially engineered receptors that confer a desired specificity to immune effector T cells. As an HIV-1-specific CAR, CD4 zeta CAR has been extensively tested in vitro as well as in clinical trials. T cells modified with this CAR mediated highly potent anti-HIV-1 activities in vitro and were well-tolerated in vivo, but exerted limited effects on viral load and reservoir size due to poor survival and/or functionality of the transduced cells in patients. We hypothesize that ectopic expression of CD4 zeta on CD8(+) T cells renders them susceptible to HIV-1 infection, resulting in poor survival of those cells. To test this possibility, highly purified CD8(+) T cells were genetically modified with a CD4 zeta-encoding lentiviral vector and infected with HIV-1. CD8+ T cells were vulnerable to HIV-1 infection upon expression of CD4 zeta as evidenced by elevated levels of p24(Gag) in cells and culture supernatants. Concurrently, the number of CD4 zeta-modified CD8(+) T cells was reduced relative to control cells upon HIV-1 infection. To protect these cells from HIV-1 infection, we co-expressed two anti-HIV-1 shRNAs previously developed by our group together with CD4 zeta. This combination vector was able to suppress HIV-1 infection without impairing HIV-1-dependent effector activities of CD4 zeta. In addition, the number of CD4 zeta-modified CD8(+) T cells maintained similar levels to that of the control even under HIV-1 infection. These results suggest that protecting CD4 zeta-modified CD8(+) T cells from HIV-1 infection is required for prolonged HIV-1-specific immune surveillance. (C) 2015 Elsevier Inc. All rights reserved.