FIGURE 5.

CK548 inhibits HIV-1 infection and viral nuclear migration. A, an HIV Rev-dependent indicator (GFP) T cell, Rev-CEM, was pretreated with DMSO, CK548 or CK636 for 1 h, and then infected with HIV-1_NL4–3 for 2 h. Cells were washed, and the inhibitors were added back after infection. HIV-dependent GFP expression was measured at 72 h postinfection by flow cytometry. To exclude drug cytotoxicity, PI was added into the cell suspension prior to flow cytometry, and only viable cells (R1 gate) were used for measuring GFP expression. B, CEM-SS T cells were pre-treated with DMSO or CK548 for 1 h, and then infected with HIV-1_NL4–3 as above. Viral replication was measured by p24 release. C, drug cytotoxicity was meaured in CK548-treated and infected cells by PI staining and flow cytometry. Shown is the percentage of PI-positive cells. D–F, CEM-SS T cells were pretreated with DMSO or CK548 for 1 h, and then infected with a single-cycle HIV-1(Env) that was pseudotyped with HIV-1 gp160. Cells were infected for 2 h, washed, and cultured in the presence of the drug for 48 h. Total cellular DNA was extracted at 12 and 48 h postinfection and then PCR-amplified to quantify viral DNA (D) and viral 2-LTR-circles (E). The relative ratio of 2-LTR circles to viral total DNA was also plotted (F). ND, not detectable. G, effect of CK548 on viral budding and release was also measured. Cells were infected with HIV-1(Env) for 12 h, washed three times, and then incubated with CK548 at different dosages. Viral p24 release was measured.