290 Sigma-2 Receptor Agonists Inhibit HIV Infection of Lymphocytes by Reducing Membrane Sphingomyelin Content KW Crawford*1,2, Z Liao3, JE Hildreth3, WD Bowen2 1Howard Univ Sch of Pharmacy, Washington, DC; 2Natl Inst of Diabetes, Digestive and Kidney Diseases, NIH, Bethesda, MD; and 3Johns Hopkins Univ Sch of Med, Baltimore, MD
Background: HIV infection of CD4+ lymphocytes and release of virions occurs in lipid rafts; cholesterol- and sphingolipid-rich microdomains of the plasma membrane. We have shown that reducing membrane cholesterol content also reduces HIV infectivity of lymphocytes and diminishes virulence of the virions released. Reducing membrane sphingomyelin may produce similar effects. Sigma-2 receptors are enigmatic receptors expressed in various tissues that recognize diverse psychoactive compounds. We have shown that sigma-2 receptor-activation reduces levels of membrane sphingomyelin in breast tumor cells. We also found that sigma-2 receptors are localized to lipid rafts. In addition, sigma-2 receptor activation may inhibit P-I-3’ kinase signaling, an effect that should inhibit HIV infection of lymphocytes and macrophages.
Methods: Membrane sphingomyelin was metabolically labeled in H9 lymphocytes by incubating the cells with [3H] palmitic acid in supplemented RPMI media for 48 h. Labeled media was removed and cells were incubated in the presence or absence of various concentrations of the sigma-2 receptor agonist CB-184 for 24 hrs. Total lipids were extracted and the dried organic layer was chromatagraphed on Silica G-60 plates. Sphingomyelin spots were scraped and quantified by liquid scintillation counting. CEMX174 cells transfected with a luciferase gene driven by the Simian Immunodeficiency Virus (SIV) LTR were used in infectivity assays. Cells were then treated with sigma-2 receptor agonists at various concentrations followed by incubation in the presence or absence (control) of HIV. Luciferase activity quantified HIV replication.
Results: Treatment of H9 cells with the sigma-2 receptor agonist CB-184 produced reductions of about 39% and 45% of control levels of membrane sphingomyelin at doses of 0.3 muM and 1muM, respectively. We observed dose-dependent inhibition of HIV replication by CB-184. This HIV-inhibitory effect was most pronounced at 0.5muM (< 10% of control), a dose within the range for producing reductions in sphingomyelin, yet non-toxic to the cells.
Conclusions: Sigma-2 receptor agonists, at non-toxic concentrations, may prevent HIV infection of lymphocytes, presumably by reducing membrane sphingomyelin. This host-targeted strategy should be further explored for potential clinical applications in the treatment of HIV-infection.
