Abstract: The study is aimed to evaluate the anti-HIV-1 effect of chloroquine in combination with antihuman immunodeficiency virus (HIV) drugs, and inhibition of plasmacytoid dendritic cells (pDC) activation and type I interferon (IFN-I) production by Toll-like receptor 7 (TLR7) agonist stimulation. We investigated the anti-HIV-1_ⅢB, HIV-1_KM018 activity of chloroquine and chloroquine combined with rategrivir (RAL), enfuvirtide (T-20), indinavir (IDV) and efavirenz (EFV) in vitro by luciferase activity assay system and ELISA method for p24 antigen. We measured the effect of chloroquine on the activation of pDC in combination with RAL and IDV, respectively. Quantitative PCR was used to evaluate the activity of chloroquine in combination with RAL and IDV in the upregulation of interferon (IFN)-α and IFN-β. Chloroquine showed less cytotoxicity to C8166, TZM-bl and PBMC cells, and the 50% cytotoxic concentration values were 85.02 ±0.28, 73.67 ±5.10 and 91.84 ±4.10 μmol·L^-1, respectively. The anti-HIV-1_ⅢB activity of chloroquine combination with RAL, T-20, IDV and EFV were moderate in synergy, strong in synergy, additive and moderate antagonism, respectively. The anti-HIV-1_KM018 activity of chloroquine in combination with RAL, IDV were moderate synergy, minor synergy. There was no significant difference between the chloroquine monotherapy and chloroquine combined with RAL, IDV in the down-regulation of pDC activation and IFN-α, IFN-β expression levels. We have found that chloroquine combined with different anti-HIV drugs represent different degrees of synergism, antagonism or additive anti-HIV-1 effect. Chloroquine in combination with RAL and IDV did not have influence on the inhibitory effect of chloroquine on pDC activation and type I interferon secretion induced by TLR7 agonist. The results suggest that chloroquine may be used to enhance the therapeutic activities of anti-HIV medicines.