Figure 1.
Gnidimacrin activated HIV-1 production in chronically infected cells at picomolar concentrations.
(a) Chemical structures of gnidimacrin and prostratin. (b) Activation of HIV-1 production by gnidimacrin and prostratin. ACH-2 or U1 cells were treated with gnidimacrin or prostratin at various concentrations for two days. Each data point in the figure represents the mean +/− standard deviation of three independent experiments.
Figure 2.
Gnidimacrin inhibited HIV-1 NL4-3 infection at picomolar concentrations.
(a) MT4 cells were infected with NL4-3 in the presence of gnidimacrin, AZT, or prostratin for 4 days. The virus replication in the absence of antivirals is defined as 100% (control) virus production. Each data point in the figure represents the mean +/− standard deviation of three independent experiments. (b) Down regulation of CD4 and CXCR4 by gnidimacrin. MT4 cells were treated with gnidimacrin at various concentrations for one day before FACS analysis. The anti-CD4 monoclonal antibody OKT4 and the anti-CXCR4 monoclonal antibody 12G5 were used for the FACS analysis. The color assignment to each assay condition is the same for both FACS panels. A histogram of the FACS data on % receptor down regulation expressed as ratio of mean fluorescence intensity (MFI) is available as additional file 1 (Figure S1).
Figure 3.
Gnidimacrin exhibited highly selective cytotoxicity against HIV-1 chronically infected cells.
ACH-2, U1, and U937 cells were treated with the compounds for three days before the cytotoxicity was determined by using a Promega cell viability assay kit. MT4 and PBMCs were treated with the compounds parallel to the antiviral assays for 4 days and 7 days, respectively. The viability of the cells cultured in the absence of the tested compounds is defined as 100% control. Each data point in the figure represents the average of two independent experiments.
Figure 4.
Gnidimacrin inhibited R5 virus infection at picomolar concentrations.
(a) HIV-1 BaL and NL4-3 were used to infect CD8-depleted PHA activated PBMCs. The virus replication in the absence of antivirals is defined as 100% control virus production. Each data point in the figure represents the mean +/− standard deviation of three independent experiments. (b) Down regulation of HIV-1 receptors on PBMCs. CD8-depleted PHA activated PBMCs were treated with 1 nM of gnidimacrin or 1 uM of prostratin for 24 hr or 48 hr. Background binding was performed with the same protocol as other conditions, except that primary antibodies were not included in the assay. The color assignment to each assay condition is the same for all six FACS panels. A histogram of the FACS data on % receptor down regulation expressed as ratio of mean fluorescence intensity (MFI) is available as additional file 2 (Figure S2).
Table 1.
Gnidimacrin inhibited R5 viruses at low picomolar concentrations.
Figure 5.
Enzastaurin abrogated the anti-HIV-1 activity of gnidimacrin and prostratin.
CD8-depleted PHA activated PBMCs were treated with enzastaurin at indicated concentrations for 2 hours. The enzastaurin treated PBMCs were then infected with HIV-1 BaL in the presence of various concentrations of enzastaurin and 0.1 nM of gnidimacrin or 0.5 uM of prostratin. The culture supernatants were assayed for p24 on day 7 post infection. The virus replication (p24) in the presence of enzastaurin without gnidimacrin or prostratin is defined as 100% control. Each data point in the figure represents the mean +/− standard deviation of three independent experiments.