lipid-a and Malaria

Lipopolysaccharide (LPS) is a well-defined agonist of Toll-like receptor (TLR) 4 that activates innate immune responses and influences the development of the adaptive response during infection with Gram-negative bacteria. Many years ago, Dr. Edgar Ribi separated the adjuvant activity of LPS from its toxic effects, an effort that led to the development of monophosphoryl lipid A (MPL). MPL, derived from Salmonella minnesota R595, has progressed through clinical development and is now used in various product-enabling formulations to support the generation of antigen-specific responses in several commercial and preclinical vaccines. We have generated several synthetic lipid A molecules, foremost glucopyranosyl lipid adjuvant (GLA) and second-generation lipid adjuvant (SLA), and have advanced these to clinical trial for various indications. In this review we summarize the potential and current positioning of TLR4-based adjuvant formulations in approved and emerging vaccines.

Topics: Adjuvants, Immunologic; Alum Compounds; Animals; Glucosides; HIV Infections; Humans; Immunity, Cellular; Immunity, Humoral; Immunogenicity, Vaccine; Leishmaniasis; Leprosy; Lipid A; Liposomes; Malaria; Mice; Schistosomiasis; T-Lymphocytes, Helper-Inducer; Toll-Like Receptor 4; Tuberculosis; Vaccines

RTS,S is the world's most advanced malaria vaccine candidate and is intended to protect infants and young children living in malaria endemic areas of sub-Saharan Africa against clinical disease caused by Plasmodium falciparum. Recently, a pivotal Phase III efficacy trial of RTS,S began in Africa. The goal of the programme has been to develop a vaccine that will be safe and effective when administered via the Expanded Program for Immunization (EPI) and significantly reduce the risk of clinically important malaria disease during the first years of life. If a similar reduction in the risk of severe malaria and other important co-morbidities associated with malaria infection can be achieved, then the vaccine could become a major new tool for reducing the burden of malaria in sub-Saharan Africa. Encouraging data from the ongoing phase II programme suggest that these goals may indeed be achievable. This review discusses some of the unique challenges that were faced during the development of this vaccine, highlights the complexity of developing new vaccine technologies and illustrates the power of partnerships in the ongoing fight against this killer disease.

Topics: Adjuvants, Immunologic; Africa; Clinical Trials, Phase II as Topic; Clinical Trials, Phase III as Topic; Endemic Diseases; Epitopes; History, 20th Century; History, 21st Century; Humans; Lipid A; Malaria; Malaria Vaccines; Protozoan Proteins; Saponins

The need for potentiating immune responses to recombinant or subunit antigens has prompted GlaxoSmithKline (GSK) Biologicals to develop various Adjuvant Systems for the design of prophylactic and therapeutic vaccines. Adjuvant Systems are formulations of classical adjuvants mixed with immunomodulators, specifically adapted to the antigen and the target population. They can activate the appropriate innate immune system and subsequently impact on adaptive immune responses. AS04 is an Adjuvant System that has demonstrated significant achievements in several vaccines against viral diseases. AS02, another Adjuvant System, is being evaluated in various contexts, where a strong T-cell response is needed to afford protection. Likewise, AS01 has been developed for vaccines where the induction of a yet stronger T-cell-mediated immune response is required. Altogether, the promising clinical results strongly support the concept of Adjuvant Systems and allow for further development of new vaccines, best adapted to the target population and the immune mechanisms of protection.

Topics: Adjuvants, Immunologic; Adjuvants, Pharmaceutic; Aluminum Hydroxide; Animals; Drug Combinations; Drug Industry; Humans; Lipid A; Malaria; Saponins; Vaccines; Virus Diseases

Endemic malaria and helminth infections in sub-Saharan Africa can act as immunological modulators and impact responses to standard immunizations. We conducted a cohort study to measure the influence of malaria and helminth infections on the immunogenicity of the bivalent HPV-16/18 vaccine.. We evaluated the association between malaria and helminth infections, and HPV-16/18 antibody responses among 298 Tanzanian females aged 10-25 years enrolled in a randomized controlled trial of the HPV-16/18 vaccine. Malaria parasitaemia was diagnosed by examination of blood smears, and helminth infections were diagnosed by examination of urine and stool samples, respectively. Geometric mean antibody titres (GMT) against HPV-16/18 antibodies were measured by enzyme-linked immunosorbent assay.. Parasitic infections were common; one-third (30.4%) of participants had a helminth infection and 10.2% had malaria parasitaemia. Overall, the vaccine induced high HPV-16/18 GMTs, and there was no evidence of a reduction in HPV-16 or HPV-18 GMT at Month 7 or Month 12 follow-up visits among participants with helminths or malaria. There was some evidence that participants with malaria had increased GMTs compared to those without malaria.. The data show high HPV immunogenicity regardless of the presence of malaria and helminth infections. The mechanism and significance for the increase in GMT in those with malaria is unknown.

Topics: Adjuvants, Immunologic; Adolescent; Adult; Aluminum Hydroxide; Antibodies, Viral; Antibody Formation; Child; Double-Blind Method; Female; Helminthiasis; Human papillomavirus 16; Human papillomavirus 18; Humans; Lipid A; Malaria; Papillomavirus Infections; Papillomavirus Vaccines; Tanzania; Young Adult

We conducted a phase 1 trial of candidate malaria vaccine RTS,S/AS02A in western Kenya to determine its safety and immunogenicity in healthy adults in an area hyperendemic for malaria. Twenty adults were enrolled and received RTS,S/AS02A (50 microg of RTS,S in 0.5 mL of AS02A) by intramuscular injection on a 0-, 28-, and 178-day schedule. All 60 scheduled immunizations were given, and 18 of 20 volunteers completed the last study visit on day 210. The vaccine was safe and well-tolerated. There were no vaccine-related severe adverse events. The most common solicited adverse events associated with immunization were injection site pain and headache. The geometric mean concentration of antibodies to circumsporozoite protein was 1.9 microg/mL at baseline and it increased 2-4 weeks after each dose to 16, 17.8, and 36.6 microg/mL, respectively. These safety and immunogenicity data from adults in hyperendemic Kenya are comparable to data reported earlier from two trials in west African adults in hypo-endemic and meso-endemic areas of The Gambia. We conclude that in this small study, RTS,S/AS02A is safe and similarly immunogenic in malaria-exposed African adults of different ethnicity in different transmission settings.

Topics: Adolescent; Adult; Animals; Antibodies, Protozoan; Antibodies, Viral; Antibody Formation; Drug Combinations; Endemic Diseases; Epitopes, T-Lymphocyte; Female; Hepatitis B Surface Antigens; Humans; Kenya; Lipid A; Malaria; Malaria Vaccines; Male; Protozoan Proteins; Saponins; Time Factors; Vaccines, DNA

In the present study, the combined adjuvant effect of 7DW8-5, a potent α-GalCer-analog, and monophosphoryl lipid A (MPLA), a TLR4 agonist, on the induction of vaccine-induced CD8(+) T-cell responses and protective immunity was evaluated. Mice were immunized with peptides corresponding to the CD8(+) T-cell epitopes of a malaria antigen, a circumsporozoite protein of Plasmodium yoelii, and a tumor antigen, a Wilms Tumor antigen-1 (WT-1), together with 7DW8-5 and MPLA, as an adjuvant. These immunization regimens were able to induce higher levels of CD8(+) T-cell responses and, ultimately, enhanced levels of protection against malaria and tumor challenges compared to the levels induced by immunization with peptides mixed with 7DW8-5 or MPLA alone. Co-administration of 7DW8-5 and MPLA induces activation of memory-like effector natural killer T (NKT) cells, i.e. CD44(+)CD62L(-)NKT cells. Our study indicates that 7DW8-5 greatly enhances important synergistic pathways associated to memory immune responses when co-administered with MPLA, thus rendering this combination of adjuvants a novel vaccine adjuvant formulation.

Topics: Amino Acid Sequence; Animals; CD8-Positive T-Lymphocytes; Cell Line, Tumor; Drug Synergism; Epitopes, T-Lymphocyte; Galactosylceramides; HLA-A2 Antigen; Humans; Immunization; Immunologic Memory; Interferon-gamma; Killer Cells, Natural; Lipid A; Malaria; Mice, Inbred BALB C; Mice, Inbred C57BL; Mice, Transgenic; Models, Immunological; Peptides; Plasmodium yoelii; Protective Agents; Protozoan Proteins; Toll-Like Receptor 4; WT1 Proteins

Nanosuspensions are an important class of delivery system for vaccine adjuvants and drugs. Previously, we developed a nanosuspension consisting of the synthetic TLR4 ligand glucopyranosyl lipid adjuvant (GLA) and dipalmitoyl phosphatidylcholine (DPPC). This nanosuspension is a clinical vaccine adjuvant known as GLA-AF. We examined the effects of DPPC supplier, buffer composition, and manufacturing process on GLA-AF physicochemical and biological activity characteristics.. DPPC from different suppliers had minimal influence on physicochemical and biological effects. In general, buffered compositions resulted in less particle size stability compared to unbuffered GLA-AF. Microfluidization resulted in rapid particle size reduction after only a few passes, and 20,000 or 30,000 psi processing pressures were more effective at reducing particle size and recovering the active component than 10,000 psi. Sonicated and microfluidized batches maintained good particle size and chemical stability over 6 months, without significantly altering in vitro or in vivo bioactivity of GLA-AF when combined with a recombinant malaria vaccine antigen.. Microfluidization, compared to water bath sonication, may be an effective manufacturing process to improve the scalability and reproducibility of GLA-AF as it advances further in the clinical development pathway. Various sources of DPPC are suitable to manufacture GLA-AF, but buffered compositions of GLA-AF do not appear to offer stability advantages over the unbuffered composition.

Topics: 1,2-Dipalmitoylphosphatidylcholine; Adjuvants, Immunologic; Animals; Antigens, Protozoan; Buffers; Cytokines; Drug Stability; Female; Lipid A; Lymphocytes; Macrophages; Malaria; Malaria Vaccines; Mice; Mice, Inbred C57BL; Nanostructures; Particle Size; Plasmodium berghei; Protozoan Proteins; Recombinant Proteins; Sonication; Suspensions; Toll-Like Receptor 4

Topics: Adjuvants, Immunologic; Africa, Western; Clinical Trials, Phase II as Topic; Clinical Trials, Phase III as Topic; Drug Combinations; Humans; Lipid A; Malaria; Malaria Vaccines; Saponins

The goal of this project was the evaluation of a novel immunomodulatory adjuvant for human use, OM-174, which is a soluble adjuvant derived from Escherichia coli lipid A. For this study, we used a synthetic peptide, known for its safety and reproducibility and the murine model of BALB/c mice. The long peptide (PbCS 242-310) used corresponds to the C-terminal region of the circumsporozoite protein (CSP) that is the major protein on the surface of Plasmodium sporozoites. Subcutaneous injections of PbCS 242-310 in combination with soluble adjuvant OM-174 induced long lasting peptide-specific antibody titres comparable to those obtained by immunization with incomplete Freund's adjuvant (IFA). The ex vivo evaluation of the CD8(+) T cell response by IFN-gamma ELISPOT assay revealed that the injection of polypeptide with OM-174 adjuvant induced, compared to IFA, a similar and an eight-fold increased frequency of peptide-specific lymphocytes in the draining lymph-nodes and in the spleen, respectively. The CD8(+) T-cells are specific for the sequence PbCS 245-253, a well-known H-2K(d)-restricted CTL epitope, and are cytotoxic as shown in a chromium release assay. Immunization of BALB/c mice with this polypeptide in combination with adjuvant OM-174 conferred a protection after challenge with live Plasmodium berghei sporozoites.The strong antibody and CTL responses observed to a synthetic peptide in mice, the safety profile of the adjuvant and its extensive physico-chemical characterization suggest that OM-174 has a potential use in vaccine formulations for humans.

Topics: Adjuvants, Immunologic; Animals; Antibodies, Protozoan; Antigens, Protozoan; Disease Models, Animal; Enzyme-Linked Immunosorbent Assay; Humans; Lipid A; Lipopolysaccharides; Malaria; Malaria Vaccines; Mice; Mice, Inbred BALB C; Peptide Fragments; Plasmodium berghei; Protozoan Proteins

Most malariologists believe that optimal malaria vaccines will induce protective immune responses against different stages of the parasite's life cycle. A multiple antigen peptide (MAP) vaccine based on the Plasmodium yoelii circumsporozoite protein (PyCSP) protects mice against sporozoite challenge by inducing antibodies that prevent sporozoites from invading hepatocytes. A purified recombinant protein vaccine based on the P. yoelii merozoite surface protein-1 (PyMSP-1) protects mice against challenge with infected erythrocytes, presumably by inducing antibodies against the erythrocytic stage of the parasite. We now report studies designed to determine if the PyMSP-1 vaccine protects against challenge with sporozoites, the stage encountered in the field, and if immunization with a combination of the PyCSP and PyMSP-1 vaccines provides additive or synergistic protection against sporozoite challenge. In two experiments, using TiterMax or Ribi R-700 as adjuvant, 3 of 19 mice immunized with the PyMSP-1 vaccine were completely protected against sporozoite challenge. The remaining mice had significantly delayed onset and lower levels of peak parasitemia than did control mice (11.1 +/- 2.8% vs. 36.7 +/- 1.6% in experiment #2, P < 0.01). Immunization with the combination vaccine reduced by approximately 50% the level of antibodies induced to PyCSP and PyMSP-1, as compared to that induced by the individual components. However, in two experiments, there was evidence of additive protection. Six of 19 (31.6%) immunized with the PyCSP vaccine, 3 of 19 (15.8%) immunized with the PyMSP-1 vaccine, and 10 of 19 (52.6%) immunized with the combination were completely protected against sporozoit challenge. This modest increase in protection in the combination group may be a reflection of additive anti-PyCSP and anti-PyMSP-1 immunity, since mice in the combination group had diminished levels of antibodies to each components. These studies indicate that considerable work may be required to optimize the construction, delivery, and assessment of multi-stage malaria vaccines.

Topics: Animals; Antibodies, Protozoan; Cell Wall Skeleton; Cord Factors; Enzyme-Linked Immunosorbent Assay; Female; Immunoblotting; Immunoglobulin Isotypes; Lipid A; Malaria; Merozoite Surface Protein 1; Mice; Mice, Inbred BALB C; Parasitemia; Plasmodium yoelii; Poloxalene; Protein Precursors; Protozoan Proteins; Vaccination; Vaccines, Combined; Vaccines, Synthetic

This study describes the safety and immunogenicity of a liposome-based vaccine injected into human subjects. Thirty healthy adult male volunteers were immunized with a liposome-encapsulated recombinant protein (R32NS181) containing epitopes from the repeat region of the circumsporozoite protein of Plasmodium falciparum. This antigen had previously been found to be poorly immunogenic in humans when it was adsorbed with Al(OH)3. In the present study, R32NS181 was encapsulated in liposomes containing monophosphoryl lipid A that were subsequently adsorbed to Al(OH)3. Increasing doses of liposomes containing antigen and monophosphoryl lipid A were used, but the liposomes were always adsorbed to the same dose of Al(OH)3. R32-specific serum IgG antibody responses to liposome-encapsulated R32NS181 were much higher than levels attained previously in humans with R32NS181 adsorbed to Al(OH)3. Geometric mean specific IgG levels after three doses ranged from 14 to 33 micrograms/ml. Sera from volunteers receiving the two highest doses inhibited P. falciparum sporozoite invasion of cultured hepatoma cells by an average of 92%, a result that was again superior to previously reported vaccines. Moderate but acceptable transient local reactogenicity was noted at high doses of the vaccine formulation, but little or no systemic toxicity was seen despite liposomal monophosphoryl lipid A doses up to 2200 micrograms. We conclude that encapsulation of poorly immunogenic circumsporozoite protein repeat peptides in monophosphoryl lipid A-containing liposomes is a successful adjuvant strategy in humans for inducing high levels of specific antibody production.

Topics: Adjuvants, Immunologic; Adult; Amino Acid Sequence; Antibodies, Protozoan; Cells, Cultured; Dose-Response Relationship, Immunologic; Humans; Immunoglobulin G; In Vitro Techniques; Lipid A; Liposomes; Malaria; Male; Molecular Sequence Data; Recombinant Fusion Proteins; Vaccines, Synthetic

A liposome-encapsulated cloned protein (R32tet32) containing sequences from the tetrapeptide repeat region of the circumsporozoite protein of Plasmodium falciparum sporozoites was examined for immunogenicity with rabbits and monkeys. Effects of adjuvants were tested by encapsulation of the antigen in liposomes either lacking or containing lipid A and adsorption with aluminum hydroxide (ALUM). When rabbits were immunized with R32tet32 alone, a primary antibody response was not seen and a secondary response did not appear until 32 to 36 weeks after boosting. Immunization with ALUM-adsorbed R32tet32 resulted in a minimal primary antibody response. A moderate secondary antibody response was detected within 2 weeks after boosting, but antibody levels decreased to preimmunization levels 8 weeks after boosting. When R32tet32 was encapsulated in liposomes containing lipid A, strong primary and secondary antibody responses were observed. Strong primary and secondary responses also were obtained when R32tet32 was encapsulated in liposomes either containing or lacking lipid A and the liposomes were adsorbed with ALUM. The strongest antibody response was obtained by immunization with ALUM-adsorbed liposomes containing lipid A and R32tet32, suggesting that the adjuvant effects of liposomes, lipid A, and ALUM were additive or synergistic.

Topics: Adjuvants, Immunologic; Aluminum Hydroxide; Animals; Antibodies, Protozoan; Antigens, Protozoan; Antigens, Surface; Lipid A; Liposomes; Macaca mulatta; Malaria; Plasmodium falciparum; Protozoan Proteins; Rabbits

To test for the presence of a lymphocyte mitogenic factor in malaria, sera were obtained from 10 patients with malaria (9 with falciparum and one with vivax), and 10 noninfected controls. The sera from the malarial patients caused an increased blastogenesis in mouse splenic lymphocyte cultures and inhibited hemagglutination between lipid-A-coated erythrocytes and lipid-A antibodies. None of the sera were positive using the limulus amebocyte lysate test. These results could be interpreted to demonstrate that patients with falciparum malaria have a circulating mitogen which cross-reacts with endotoxin. However, alternate explanations must be considered, including an hypothesis that antiglobulins and/or immune complexes in the sera of malarious patients both caused the blastogenesis of mouse spleen cells and inhibited hemagglutination to lipid-A antibodies.

Topics: Adult; Hemagglutination Inhibition Tests; Humans; Interleukin-2; Limulus Test; Lipid A; Lymphocyte Activation; Lymphokines; Malaria; Male; Plasmodium falciparum; Plasmodium vivax