Showing papers by "Richard M. Epand published in 1994"

Cationic liposomes for direct gene transfer in therapy of cancer and other diseases.

Abstract: Cationic liposomes can mediate efficient delivery of DNA and DNA/protein complex to mammalian cells in vitro and in vivo. Cationic cholesterol derivatives mixed with phosphatidylethanolamine and sonicated to form small unilamellar vesicles can complex with DNA and mediate the entry into the cytosol from the endosome compartment. One of the liposome formulations, DC-Chol liposomes, is used in a gene therapy clinical trial for melanoma. Recently, we exploited these cationic liposomes for the delivery of trans-activating protein factors to regulate and control the expression of delivered transgenes in a protein dose-dependent manner. Bacteriophage T7 RNA polymerase was co-delivered with a reporter gene under the control of T7 promoter to allow cytoplasmic expression of the gene. Human immunodeficiency virus-1 transactivating protein was also codelivered with a reporter gene under the control of HIV-1 long terminal repeat. Finally, human tumor cells selected for cis-platin resistance or isolated from patients who have failed cis-platin therapy are highly transfectable with cationic liposomes. These results suggest a serial therapy protocol with cis-platin and gene therapy for malignancy.

HDL and apolipoprotein A-I protect erythrocytes against the generation of procoagulant activity.

Abstract: The appearance of anionic lipids on the extracellular surface of cells is required for the formation of the procoagulant complex that leads to the activation of prothrombin. Procoagulant activity would be expected to be inhibited by substances that stabilize the membrane structure and hence inhibit the transbilayer diffusion of phosphatidylserine from the cytoplasmic to the extracellular surface of the plasma membrane. The generation of procoagulant activity in human erythrocytes by A23187 and Ca2+ is inhibited by apolipoprotein A-I, its amphipathic peptide analogues, and high-density lipoprotein (HDL). These agents do not inhibit the Ca2+ loading of erythrocytes by A23187, nor do they inhibit the activation of prothrombin once the cells have been incubated at 37 degrees C with A23187 and Ca2+. Transbilayer diffusion of fluorescently labeled phosphatidylserine is inhibited by apolipoprotein A-I. These findings indicate that class A amphipathic helixes as well as lipoprotein particles and liposomes inhibit the transbilayer diffusion of phospholipids and procoagulant activity. This activity may contribute to the protective role of HDL against arteriosclerosis and thrombosis.

Mixing rates can markedly affect the kinetics of peptide-induced leakage from liposomes.

Abstract: Assays on vesicle aqueous content leakage are widely used in the study of peptide-lipid interactions. We found this assay to be affected by the mode of mixing vesicle and peptide solutions. This effect can lead to artifactual conclusions regarding the lytic activity of peptides. We demonstrate that the source of this artifact is that fast (millisecond range) peptide-membrane association creates a nonhomogeneous distribution which exists sufficiently long after peptide addition to markedly alter the course of leakage. Mixing problems can be overcome by using a stopped flow apparatus. It can also be diminished by switching to injecting a small volume of vesicle suspension into a large volume of peptide solution. Mixing rates are rarely considered in literature reports of peptide effects on liposomes. The same artifacts can also take place in a number of other assays of the activity of membrane active peptides on liposomes and thus their consideration is of general importance.

Fusion of influenza to liposomes is not inhibited by aliphatic primary alcohols.

Abstract: Reports of the antiviral activity of aliphatic alcohols led us to investigate the effects of aliphatic alcohols, from 10 to 20 carbons in length, on the phase transition behaviour of model phospholipids and on the fusion of influenza to liposomes. Contrary to the effects of many other antiviral agents, we find that alcohols are potent promoters of the inverted hexagonal phase. However, we also find that aliphatic alcohols have little effect on influenza fusion to liposomes. Eicosanol is the only aliphatic alcohol tested which substantially increases in fusion of influenza virus. We also find that long chain alcohols display multi-component bilayer to hexagonal phase transitions at higher mole fractions. This suggests that eicosanol may be facilitating fusion by creating defects between alcohol-rich and alcohol-poor regions of the lipid bilayer.

The Binding of Peptides and Proteins to Membranes Containing Anionic Lipid

Abstract: The interaction of proteins with lipids is one of the central problems of membrane biophysics. Of particular interest is that some proteins require a distinct category of lipids for their function. For example, several functionally related groups of proteins that interact preferentially with acidic lipids were discovered in recent years. They include vitamin K-dependent blood coagulation proteins (Jackson and Nemerson, 1980), annexins (Geisow et al., 1987), calpactins (Glenney, 1986), synexins (Creutz et al., 1983), the protein kinase C family (Nishizuka, 1989), and actin-severing or actin-cap-ping proteins (Stossel, 1989). Many other proteins, particularly enzymes, also require acidic lipids for activation (Cornell, 1991; Enyedi et al., 1987; Moritz et al., 1992). There are excellent reviews available on many of the particular groups of proteins that interact with acidic lipids (Geisow et al., 1987; Glenney, 1986; Jackson and Nemerson, 1980; Stossel, 1989). Therefore, we decided to focus this chapter on the roles played by acidic lipids in lipid-protein interactions. For the sake of clarity, we used a limited number of examples to describe the four distinct functions performed by acidic lipids. We also review recent models of the mechanism of acidic lipid-protein interaction.