A synthetic globotriaosylceramide analogue inhibits HIV-1 infection <Emphasis Type="Italic">in vitro</Emphasis> by two mechanisms

Previously, it was shown that the cell-membrane-expressed glycosphingolipid, globotriaosylceramide (Gb₃/P^k/CD77), protects against HIV-1 infection and may be a newly described natural resistance factor against HIV infection. We have now investigated the potential of a novel, water soluble, non-toxic and completely synthetic analogue of Gb₃/P^k (FSL-Gb₃) to inhibit HIV-1 infection in vitro. A uniquely designed analogue, FSL-Gb₃, of the natural Gb₃/P^k molecule was synthesized. HIV-1_IIIB (X4 virus) and HIV-1_Ba-L (R5 virus) infection of PHA/interleukin-2-activated, peripheral blood mononuclear cells (PBMCs) and Jurkat T cells in vitro was assessed, as well as infection of U87.CD4.CCR5 by various clinical R5 tropic viruses after treatment with FSL-Gb₃. We monitored Gb₃, CD4 and CXCR4 expression by fluorescent antibody cell sorting and viral replication by p24 ^gag ELISA. Total cellular Gb₃ was examined by glycosphingolipid extraction and thin layer chromatography. In vivo toxicity was monitored in mice by histological assessment of vital organs and lymphoid tissue. FSL-Gb₃ blocked X4 and R5 of both lab and clinical viral strains in activated PBMCs or the U87.CD4.CCR5 cell line with a 50% inhibitory concentration (IC₅₀) of approximately 200–250 μM. FACS and TLC overlay showed that FSL-Gb₃ can insert itself into cellular plasma membranes and that cellular membrane-absorbed FSL-Gb₃ is able to inhibit subsequent HIV-1 infection. There was no effect of FSL-Gb₃ on cell surface levels of CD4 or CXCR4. Thus, FSL-Gb₃ can inhibit HIV-1 by two mechanisms: direct inhibition of virus and inhibition of viral entry. Infusion of FSL-Gb₃ into laboratory mice at doses well in excess of theoretical therapeutic doses was tolerated with no untoward reactions. Our results demonstrate the potential utility of using a completely synthetic, water soluble globotriaosylceramide analogue, FSL-Gb₃, having low toxicity, for possible future use as a novel therapeutic approach for the systemic treatment of HIV/AIDS.