Polyclonal and monoclonal antibodies to liposomes having various phospholipid compositions have been produced. Binding of the anti-lipid bilayer antibodies is influenced both by the chemical composition and the physical state of the liposomal lipids. The antibodies to liposomes have a 'subsite' in the binding site that recognizes small soluble phosphorylated haptens such as nucleotides (e.g., ATP). The capacity of anti-liposome antibodies to bind to phosphate is also shared by antibodies to numerous other substances, including lipid A from Gram-negative bacterial lipopolysaccharide, cardiolipin, DNA, polynucleotides, and lipoteichoic acids from Gram-positive bacteria. Because of similarities of chemical structures between all of these molecules widespread immunological cross-reactivities are observed.

Animals↗

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Murine IgG subclass antibodies to antigens incorporated in liposomes containing lipid A.

The IgG subclass responses to antigens incorporated in liposomes containing lipid A were investigated using a synthetic malarial antigen (SPf66) and cholera toxin (CT). The antigen-specific IgG subclass response was determined in BALB/c mice immunized with either: (a) SPf66 encapsulated in liposomes containing lipid A, (b) CT bound to the surface of liposomes containing lipid A, or (c) both encapsulated SPf66 and surface-bound CT in the same liposomes. In each case the antibodies to SPf66, CT and lipid A demonstrated an IgG2a predominance. Liposomes containing lipid A not only increased the magnitude of the antibody response to liposomal antigens but elicited predominantly IgG2a subclass antibodies as well.

Animals↗

PubMedDate not supplied

Immunogenicity of Plasmodium falciparum circumsporozoite protein multiple antigen peptide vaccine formulated with different adjuvants.

Only low antibody levels were obtained from vaccinating human volunteers with single-chain peptide from the Plasmodium falciparum circumsporozoite protein (PfCSP). This resulted in modest protection against sporozoite challenge. In addition, HLA restriction limits the probability of synthesis of a vaccine effective for a diverse population. We report immunization studies with a multiple antigen peptide (MAP) system consisting of multiple copies of a B-cell epitope from the central repeat region of the PfCSP in combination with a universal T-cell epitope, the P2P30 portion of tetanus toxin. This MAP4(NANP)6P2P30 vaccine was highly immunogenic in four different strains of mice when used with various safe and nontoxic adjuvants. When this MAP vaccine was encapsulated in liposomes with lipid A and adsorbed to aluminium hydroxide and given three times at 4-week intervals, the resultant antibody prevented 100% of sporozoites from invading and developing into liver stage infection. This high degree of immunogenicity of MAP4(NANP)6P2P30 vaccine formulated in liposomes, lipid A and aluminum hydroxide provides the foundation for consideration of human trials with this formulation.

Adjuvants, Immunologic↗

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Role of complement activation in hypersensitivity reactions to doxil and hynic PEG liposomes: experimental and clinical studies.

Pegylated liposomal doxorubicin (Doxil) and 99mTc-HYNIC PEG liposomes (HPL) were reported earlier to cause hypersensitivity reactions (HSRs) in a substantial percentage of patients treated i.v. with these formulations. Here we report that (1) Doxil, HPL, pegylated phosphatidylethanolamine (PEG-PE)-containing empty liposomes matched with Doxil and HPL in size and lipid composition, and phosphatidylglycerol (PG)-containing negatively charged vesicles were potent C activators in human serum in vitro, whereas small neutral liposomes caused no C activation. (2) Doxil and other size-matched PEG-PE and/or PG-containing liposomes also caused massive cardiopulmonary distress with anaphylactoid shock in pigs via C activation, whereas equivalent neutral liposomes caused no hemodynamic changes. (3) A clinical study showed more frequent and greater C activation in patients displaying HSR than in non-reactive patients. These data suggest that liposome-induced HSRs in susceptible individuals may be due to C activation, which, in turn, is due to the presence of negatively charged PEG-PE in these vesicles.

Animals↗

PubMedDate not supplied

Heterogeneity of lipid A.

In a previous study, lipid A from Shigella flexneri was separated into eight distinct fractions by thin-layer chromatography (TLC). In the present investigation, lipid A from different gram-negative bacteria was analyzed by thin-layer chromatography for its heterogeneity and compared with lipid A of S. flexneri. Purified lipid A preparations from six different enterobacterial species were heterogeneous, but based on Rf, comparable bands were found among all preparations. Changing the culture conditions by using two other culture media resulted in very minor changes in the TLC appearance. Different serotypes of Shigella grown under the same conditions also showed very minor differences. Two bands (5 and 8) in each preparation of lipid A comigrated with lysophosphatidylethanolamine and phosphatidylethanolamine, respectively, and both bands were stained with ninhydrin and molybdenum blue. Therefore, phosphatidylethanolamine, and probably lysophosphatidylethanolamine, are common constitutents of unfractioned lipid A preparations, and both phospholipids can occur even in highly purified unfractionated lipid A.

Chromatography, Thin Layer↗

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Mitogenic activities of lipid A and liposome-associated lipid A: effects of epitope density.

Among eight purified lipid A fractions, there was heterogeneity of mitogenic activity, but the relative activity could not be predicted by limulus amebocyte lysate assay. Analysis of mitogenic activity after the incorporation of lipid A into liposomes demonstrated that epitope density of lipid A was a crucial factor that strongly modulated expression of lipid A. The results suggest that lipid A hydrophobic groups are responsible for mitogenic activity and that the surface accessibility of lipid A hydrophobic groups in membranes may be influenced by epitope density. The data are compared with previous results derived from parallel studies on limulus amebocytes. Both approaches lead to the conclusion that the activities of purified lipid A fractions are heterogeneous and that epitope density is critically important for the expression of biologic activity of membrane-associated lipid A.

Animals↗

PubMedDate not supplied

Mitogenic response of lymphocytes from C3H/HeJ mice in the presence of lipid A and lipid A fractions.

Purified lipid A and eight fractions of lipid A were examined for the ability to elicit a mitogenic response in spleen lymphocytes of endotoxin-resistant C3H/HeJ mice. The pattern of results obtained with different lipid A fractions was similar to that observed with spleen cells from the endotoxin-responsive C57Bl/6J mouse. Although the absolute mitogenic response of C3H/HeJ lymphocytes to lipid A was diminished, the cells apparently were completely normal with respect to the pattern of relative mitogenic responses induced by purified fractions of lipid A.

Animals↗

PubMedDate not supplied

Antibodies to lipid A: occurrence in humans.

Lipid A, the toxic part of the bacterial endotoxin, is a common antigen for many gram-negative bacteria. Antibodies to lipid A occur naturally in humans; they have been found in 10%-34%, and even up to 73%, of individuals tested, as detected by indirect hemolysis and enzyme-linked immunosorbent assay (ELISA), respectively. Inflammatory bowel diseases (Crohn's disease or ulcerative colitis) cause changes in the level of antibodies to lipid A, as compared with that found in healthy control subjects. Increased levels of antibodies to lipid A are seen in both children and adults with infections due to gram-negative bacteria, such as urinary tract infections (UTI). The highest titers of IgG in serum, as detected by ELISA, have been recorded in patients with development or progression of renal scarring associated with UTI. Since lipid A may play a role in the pathogenesis of renal impairment, the determination of the level of antibodies to lipid A may help in the diagnosis of certain forms of UTI. Possible beneficial roles of antibodies to lipid A during septicemia caused by gram-negative bacteria in humans are still unclear.

Antibodies, Bacterial↗

PubMedDate not supplied

Enhancement by lipid A of mucosal immunogenicity of liposome-associated cholera toxin.

Two methods that might enhance the mucosal immunogenicity of a protein antigen, cholera toxin (CT), were studied in rats: association of CT with liposomes, and coadministration of CT with lipid A. Enteric priming by CT was not enhanced when the antigen was trapped within liposomes or bound to their surface via GM1 ganglioside, nor was it improved when CT was mixed with lipid A or with liposomes containing lipid A. However, lipid A did enhance priming by liposome-associated CT when the lipid A was incorporated into CT-bearing liposomes. It is concluded that lipid A can act as an adjuvant for a local IgA response to a mucosally applied antigen, at least when lipid A and the antigen are associated on a liposome carrier.

Animals↗