Fabry disease is caused by α-galactosidase A deficiency leading to accumulation of globotriaosylceramide (Gb3) in tissues. Clinical manifestations do not appear to correlate with total Gb3 levels. Studies examining tissue distribution of specific acyl chain species of Gb3 and upstream glycosphingolipids are lacking.

Our previous genetic, pharmacological and analogue protection studies identified the glycosphingolipid, Gb(3) (globotriaosylceramide, Pk blood group antigen) as a natural resistance factor for HIV infection. Gb(3) is a B cell marker (CD77), but a fraction of activated peripheral blood mononuclear cells (PBMCs) can also express Gb(3). Activated PBMCs predominantly comprise CD4+ T-cells, the primary HIV infection target. Gb(3) is the sole receptor for Escherichia coli verotoxins (VTs, Shiga toxins). VT1 contains a ribosome inactivating A subunit (VT1A) non-covalently associated with five smaller receptor-binding B subunits. The effect of VT on PHA/IL2-activated PBMC HIV susceptibility was determined. Following VT1 (or VT2) PBMC treatment during IL2/PHA activation, the small Gb(3)+/CD4+ T-cell subset was eliminated but, surprisingly, remaining CD4+ T-cell HIV-1(IIIB) (and HIV-1(Ba-L)) susceptibility was significantly reduced. The Gb(3)-Jurkat T-cell line was similarly protected by brief VT exposure prior to HIV-1(IIIB) infection. The efficacy of the VT1A subunit alone confirmed receptor independent protection. VT1 showed no binding or obvious Jurkat cell/PBMC effect. Protective VT1 concentrations reduced PBMC (but not Jurkat cell) proliferation by 50%. This may relate to the mechanism of action since HIV replication requires primary T-cell proliferation. Microarray analysis of VT1A-treated PBMCs indicated up regulation of 30 genes. Three of the top four were histone genes, suggesting HIV protection via reduced gene activation. VT blocked HDAC inhibitor enhancement of HIV infection, consistent with a histone-mediated mechanism. We speculate that VT1A may provide a benign approach to reduction of (X4 or R5) HIV cell susceptibility.

Many germ line diseases stem from a relatively minor disturbance in mutant protein endoplasmic reticulum (ER) 3D assembly. Chaperones are recruited which, on failure to correct folding, sort the mutant for retrotranslocation and cytosolic proteasomal degradation (ER-associated degradation-ERAD), to initiate/exacerbate deficiency-disease symptoms. Several bacterial (and plant) subunit toxins, retrograde transport to the ER after initial cell surface receptor binding/internalization. The A subunit has evolved to mimic a misfolded protein and hijack the ERAD membrane translocon (dislocon), to effect cytosolic access and cytopathology. We show such toxins compete for ERAD to rescue endogenous misfolded proteins. Cholera toxin or verotoxin (Shiga toxin) containing genetically inactivated (± an N-terminal polyleucine tail) A subunit can, within 2-4 hrs, temporarily increase F508delCFTR protein, the major cystic fibrosis (CF) mutant (5-10x), F508delCFTR Golgi maturation (<10x), cell surface expression (20x) and chloride transport (2x) in F508del CFTR transfected cells and patient-derived F508delCFTR bronchiolar epithelia, without apparent cytopathology. These toxoids also increase glucocerobrosidase (GCC) in N370SGCC Gaucher Disease fibroblasts (3x), another ERAD-exacerbated misfiling disease. We identify a new, potentially benign approach to the treatment of certain genetic protein misfolding diseases.

There continues to be no approved drugs for the treatment of Ebola virus disease (EVD). Despite a number of candidate drugs showing limited efficacy in vitro and/or in non-human primate studies, EVD continues to plaque certain areas of Africa without any efficacious treatments yet available. Recently, we have been exploring the potential for anti-malarial drugs to inhibit an in vitro model of Ebola Zaire replication using a transcription-competent virus-like particle (trVLP) assay. We examined the efficacy of chloroquine, amodiaquine and 36 novel anti-parasite quinoline derivatives at inhibiting Ebola virus replication. Drug efficacy was tested by trVLP assay and toxicity by MTT assay. Both chloroquine and amodiaquine were effective for inhibition of Ebola virus replication without significant toxicity. The half-maximal inhibitory concentration (IC 50) of chloroquine and amodiaquine to inhibit Ebola virus replication were IC 50, Chl = 3.95 µM and IC 50, Amo = 1.45 µM, respectively. Additionally, three novel quinoline derivatives were identified as having inhibitory activity and low toxicity for Ebola trVLP replication, with 2NH2Q being the most promising derivative, with an IC 50 of 4.66 µM. Quinoline compounds offer many advantages for disease treatment in tropical climates as they are cheap to produce, easy to synthesize and chemically stable. In this report, we have demonstrated the potential of anti-parasite quinolines for further investigation for use in EVD.

The influenza virus (IFV) acquires its envelope by budding from host cell plasma membranes. Using quantitative shotgun mass spectrometry, we determined the lipidomes of the host Madin-Darby canine kidney cell, its apical membrane, and the IFV budding from it. We found the apical membrane to be enriched in sphingolipids (SPs) and cholesterol, whereas glycerophospholipids were reduced, and storage lipids were depleted compared with the whole-cell membranes. The virus membrane exhibited a further enrichment of SPs and cholesterol compared with the donor membrane at the expense of phosphatidylcholines. Our data are consistent with and extend existing models of membrane raft-based biogenesis of the apical membrane and IFV envelope.