Figure 1

Protease Inhibitors effectively block cell-to-cell spread of HIV-1. A) Quantification of cell-to-cell spread of HIV-1 over time in presence of C_max LPV (14 μM) and B) C_max of DRV (12 μM). HIV-1infected Jurkat cells (donors) incubated with PI for 24 h and mixed with uninfected Jurkat cells (targets) with or without PIs. After DNA extraction, qPCR performed to measure appearance of HIV pol DNA. Data normalized to albumin housekeeping gene and expressed as the fold-increase in HIV DNA copy number over time relative to the baseline value at t = 0 h. Data show mean of triplicates, error bars represent the standard deviation of the mean (SD), **** P < 0.0001, ***P<0.001, ns: not significant, two-way ANOVA plus Bonferroni post-test. C) Reduced detection of 2-LTR circles following cell-to-cell spread of HIV-1 in the presence of LPV (14 μM). After 24 h co-culture of donor cells and target cells with or without LPV, 2-LTR circles were detected by qPCR. ** P < 0.05, Unpaired Student T-test. D) Confirmation of PI Gag maturation defect in HIV-1. HIV-1+ donor cells incubated with PIs, RTIs or left untreated for 24 h. Virus-containing supernatants harvested, purified and equal volumes of virus analyzed by SDS-PAGE and western blotting for HIV-1 Gag. E) LPV and F) DRV similarly inhibit cell-to-cell and cell-free HIV-1 spread. HIV-1 infected cells incubated with serial dilutions of PI for 24 h, mixed with target cells and HIV-1 DNA measured as described for A. For cell-free infections, virus-containing supernatant harvested from infected donor cells incubated with PI for 24 h was used to infect target cells. 24 h post-infection, DNA was extracted from pelleted target cells and qPCR performed. Error bars represent the SD of the mean of triplicates G) Increasing donor: target cell ratio in co-culture reduces the efficacy of LPV in blocking cell-to-cell spread of HIV-1. ****P < 0.0001, ***P < 0.001, ns: not significant, Two-way ANOVA with Bonferroni post-test.