Limits...
A small-molecule inhibitor directed against the chemokine receptor CXCR4 prevents its use as an HIV-1 coreceptor.

Doranz BJ, Grovit-Ferbas K, Sharron MP, Mao SH, Goetz MB, Daar ES, Doms RW, O'Brien WA - J. Exp. Med. (1997)

Bottom Line: ALX40-4C inhibited HIV-1 use of the coreceptor CXCR4 by T- and dual-tropic HIV-1 strains, whereas use of CCR5 by M- and dual-tropic strains was not inhibited.ALX40-4C blocked stromal-derived factor (SDF)-1alpha-mediated activation of CXCR4 and binding of the monoclonal antibody 12G5 to cells expressing CXCR4.Overlap of the ALX40-4C binding site with that of 12G5 and SDF implicates direct blocking of Env interactions, rather than downregulation of receptor, as the mechanism of inhibition.

View Article: PubMed Central - PubMed

Affiliation: Department of Pathology and Laboratory Medicine, University of Pennsylvania, Philadelphia 19104, USA.

ABSTRACT
The chemokine receptor CXCR4 is the major coreceptor used for cellular entry by T cell- tropic human immunodeficiency virus (HIV)-1 strains, whereas CCR5 is used by macrophage (M)-tropic strains. Here we show that a small-molecule inhibitor, ALX40-4C, inhibits HIV-1 envelope (Env)-mediated membrane fusion and viral entry directly at the level of coreceptor use. ALX40-4C inhibited HIV-1 use of the coreceptor CXCR4 by T- and dual-tropic HIV-1 strains, whereas use of CCR5 by M- and dual-tropic strains was not inhibited. Dual-tropic viruses capable of using both CXCR4 and CCR5 were inhibited by ALX40-4C only when cells expressed CXCR4 alone. ALX40-4C blocked stromal-derived factor (SDF)-1alpha-mediated activation of CXCR4 and binding of the monoclonal antibody 12G5 to cells expressing CXCR4. Overlap of the ALX40-4C binding site with that of 12G5 and SDF implicates direct blocking of Env interactions, rather than downregulation of receptor, as the mechanism of inhibition. Thus, ALX40-4C represents a small-molecule inhibitor of HIV-1 infection that acts directly against a chemokine receptor at the level of Env-mediated membrane fusion.

Show MeSH

Related in: MedlinePlus

Inhibition of cell–cell fusion. 293T cells expressing 89.6, JR-FL, or BH8 Envs and T7 polymerase were mixed with PA317-T4 cells  expressing CD4, the coreceptor indicated, and a luciferase reporter gene  under control of a T7 promoter in the presence or absence of 10 μM  ALX40-4C. Fusion of the cell populations results in quantitative luciferase expression. Results are presented as in Fig. 1 with 89.6 normalized to 100% for CXCR4.
© Copyright Policy
Related In: Results  -  Collection


getmorefigures.php?uid=PMC2199097&req=5

Figure 3: Inhibition of cell–cell fusion. 293T cells expressing 89.6, JR-FL, or BH8 Envs and T7 polymerase were mixed with PA317-T4 cells expressing CD4, the coreceptor indicated, and a luciferase reporter gene under control of a T7 promoter in the presence or absence of 10 μM ALX40-4C. Fusion of the cell populations results in quantitative luciferase expression. Results are presented as in Fig. 1 with 89.6 normalized to 100% for CXCR4.

Mentions: To confirm that the inhibitory effects we observed were due to Env-mediated fusion events, we used a cell–cell fusion assay that depends only on Env-mediated membrane fusion for signal activity (7, 23). In this assay, murine PA317-T4 cells which express human CD4, the coreceptor of interest, and contain luciferase under control of the T7 promoter, are mixed with 293T cells that express the Env protein of interest and T7 polymerase. Fusion of the two cell populations results in cytoplasmic mixing and expression of luciferase. Use of the cell–cell fusion assay confirmed our infection results by demonstrating ALX40-4C inhibition of fusion between cells expressing dual (89.6) and T-tropic (BH8) Env proteins and cells expressing CD4 and CXCR4 (Fig. 3). Fusion between cells expressing dual- and M-tropic (JR-FL) Env proteins and cells expressing CD4 and CCR5 was not significantly affected.


A small-molecule inhibitor directed against the chemokine receptor CXCR4 prevents its use as an HIV-1 coreceptor.

Doranz BJ, Grovit-Ferbas K, Sharron MP, Mao SH, Goetz MB, Daar ES, Doms RW, O'Brien WA - J. Exp. Med. (1997)

Inhibition of cell–cell fusion. 293T cells expressing 89.6, JR-FL, or BH8 Envs and T7 polymerase were mixed with PA317-T4 cells  expressing CD4, the coreceptor indicated, and a luciferase reporter gene  under control of a T7 promoter in the presence or absence of 10 μM  ALX40-4C. Fusion of the cell populations results in quantitative luciferase expression. Results are presented as in Fig. 1 with 89.6 normalized to 100% for CXCR4.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC2199097&req=5

Figure 3: Inhibition of cell–cell fusion. 293T cells expressing 89.6, JR-FL, or BH8 Envs and T7 polymerase were mixed with PA317-T4 cells expressing CD4, the coreceptor indicated, and a luciferase reporter gene under control of a T7 promoter in the presence or absence of 10 μM ALX40-4C. Fusion of the cell populations results in quantitative luciferase expression. Results are presented as in Fig. 1 with 89.6 normalized to 100% for CXCR4.
Mentions: To confirm that the inhibitory effects we observed were due to Env-mediated fusion events, we used a cell–cell fusion assay that depends only on Env-mediated membrane fusion for signal activity (7, 23). In this assay, murine PA317-T4 cells which express human CD4, the coreceptor of interest, and contain luciferase under control of the T7 promoter, are mixed with 293T cells that express the Env protein of interest and T7 polymerase. Fusion of the two cell populations results in cytoplasmic mixing and expression of luciferase. Use of the cell–cell fusion assay confirmed our infection results by demonstrating ALX40-4C inhibition of fusion between cells expressing dual (89.6) and T-tropic (BH8) Env proteins and cells expressing CD4 and CXCR4 (Fig. 3). Fusion between cells expressing dual- and M-tropic (JR-FL) Env proteins and cells expressing CD4 and CCR5 was not significantly affected.

Bottom Line: ALX40-4C inhibited HIV-1 use of the coreceptor CXCR4 by T- and dual-tropic HIV-1 strains, whereas use of CCR5 by M- and dual-tropic strains was not inhibited.ALX40-4C blocked stromal-derived factor (SDF)-1alpha-mediated activation of CXCR4 and binding of the monoclonal antibody 12G5 to cells expressing CXCR4.Overlap of the ALX40-4C binding site with that of 12G5 and SDF implicates direct blocking of Env interactions, rather than downregulation of receptor, as the mechanism of inhibition.

View Article: PubMed Central - PubMed

Affiliation: Department of Pathology and Laboratory Medicine, University of Pennsylvania, Philadelphia 19104, USA.

ABSTRACT
The chemokine receptor CXCR4 is the major coreceptor used for cellular entry by T cell- tropic human immunodeficiency virus (HIV)-1 strains, whereas CCR5 is used by macrophage (M)-tropic strains. Here we show that a small-molecule inhibitor, ALX40-4C, inhibits HIV-1 envelope (Env)-mediated membrane fusion and viral entry directly at the level of coreceptor use. ALX40-4C inhibited HIV-1 use of the coreceptor CXCR4 by T- and dual-tropic HIV-1 strains, whereas use of CCR5 by M- and dual-tropic strains was not inhibited. Dual-tropic viruses capable of using both CXCR4 and CCR5 were inhibited by ALX40-4C only when cells expressed CXCR4 alone. ALX40-4C blocked stromal-derived factor (SDF)-1alpha-mediated activation of CXCR4 and binding of the monoclonal antibody 12G5 to cells expressing CXCR4. Overlap of the ALX40-4C binding site with that of 12G5 and SDF implicates direct blocking of Env interactions, rather than downregulation of receptor, as the mechanism of inhibition. Thus, ALX40-4C represents a small-molecule inhibitor of HIV-1 infection that acts directly against a chemokine receptor at the level of Env-mediated membrane fusion.

Show MeSH
Related in: MedlinePlus