Immunogenicity of Self-Associated Aggregates and Chemically Cross-Linked Conjugates of the 42 kDa Plasmodium falciparum Merozoite Surface Protein-1

London School of Hygiene and Tropical Medicine, United Kingdom
PLoS ONE (Impact Factor: 3.23). 06/2012; 7(6):e36996. DOI: 10.1371/journal.pone.0036996
Source: PubMed


Self-associated protein aggregates or cross-linked protein conjugates are, in general, more immunogenic than oligomeric or monomeric forms. In particular, the immunogenicity in mice of a recombinant malaria transmission blocking vaccine candidate, the ookinete specific Plasmodium falciparum 25 kDa protein (Pfs25), was increased more than 1000-fold when evaluated as a chemical cross-linked protein-protein conjugate as compared to a formulated monomer. Whether alternative approaches using protein complexes improve the immunogenicity of other recombinant malaria vaccine candidates is worth assessing. In this work, the immunogenicity of the recombinant 42 kDa processed form of the P. falciparum merozoite surface protein 1 (MSP1(42)) was evaluated as a self-associated, non-covalent aggregate and as a chemical cross-linked protein-protein conjugate to ExoProtein A, which is a recombinant detoxified form of Pseudomonas aeruginosa exotoxin A. MSP1(42) conjugates were prepared and characterized biochemically and biophysically to determine their molar mass in solution and stoichiometry, when relevant. The immunogenicity of the MSP1(42) self-associated aggregates, cross-linked chemical conjugates and monomers were compared in BALB/c mice after adsorption to aluminum hydroxide adjuvant, and in one instance in association with the TLR9 agonist CPG7909 with an aluminum hydroxide formulation. Antibody titers were assessed by ELISA. Unlike observations made for Pfs25, no significant enhancement in MSP1(42) specific antibody titers was observed for any conjugate as compared to the formulated monomer or dimer, except for the addition of the TLR9 agonist CPG7909. Clearly, enhancing the immunogenicity of a recombinant protein vaccine candidate by the formation of protein complexes must be established on an empirical basis.

Download full-text


Available from: Richard L Shimp,
1 Follower
27 Reads
  • Source
    • "Thus, observations made in vaccine development may help to understand the role of aggregation in the unwanted immunogenicity of biotherapeutic products. In animal models oligomeric and multimeric forms of protein antigens have been shown to be more immunogenic than the monomeric forms (Denis et al., 2007; Qian et al., 2012; Rudra et al., 2010). For example, multimerization of papaya mosaic virus capsid protein as a carrier protein for hepatitis C virus is critical for immunogenicity of the vaccine in mice, which is absent with the monomeric form (Denis et al., 2007). "
    [Show abstract] [Hide abstract]
    ABSTRACT: Abstract The elicitation of anti-drug antibodies (ADA) against biotherapeutics can have detrimental effects on drug safety, efficacy, and pharmacokinetics. The immunogenicity of biotherapeutics is, therefore, an important issue. There is evidence that protein aggregation can result in enhanced immunogenicity; however, the precise immunological and biochemical mechanisms responsible are poorly defined. In the context of biotherapeutic drug development and safety assessment, understanding the mechanisms underlying aggregate immunogenicity is of considerable interest. This review provides an overview of the phenomenon of protein aggregation, the production of unwanted aggregates during bioprocessing, and how the immune response to aggregated protein differs from that provoked by non-aggregated protein. Of particular interest is the nature of the interaction of aggregates with the immune system and how subsequent ADA responses are induced. Pathways considered here include 'classical' activation of the immune system involving antigen presenting cells and, alternatively, the breakdown of B-cell tolerance. Additionally, methods available to screen for aggregation and immunogenicity will be described. With an increased understanding of aggregation-enhanced immune responses, it may be possible to develop improved manufacturing and screening processes to avoid, or at least reduce, the problems associated with ADA.
    Journal of Immunotoxicology 08/2013; 11(2). DOI:10.3109/1547691X.2013.821564 · 2.05 Impact Factor
  • [Show abstract] [Hide abstract]
    ABSTRACT: Background: Acquired immunity to malaria develops with increasing age and repeated infections. Understanding immune correlates of protection from malaria would facilitate vaccine development and identification of biomarkers that reflect changes in susceptibility resulting from ongoing malaria control efforts. Methods: The relationship between immunoglobulin G (IgG) antibody and both interferon γ (IFN-γ) and interleukin 10 (IL-10) responses to the 42-kD C-terminal fragment of Plasmodium falciparum merozoite surface protein 1 (MSP142) and the risk of (re)infection were examined following drug-mediated clearance of parasitemia in 94 adults and 95 children in an area of holoendemicity of western Kenya. Results: Positive IFN-γ enzyme-linked immunosorbent assay (ELISA) and enzyme-linked immunosorbent spot assay (ELISPOT) responses to MSP142 3D7 were associated with delayed time to (re)infection, whereas high-titer IgG antibodies to MSP142 3D7 or FVO alleles were not independently predictive of the risk of (re)infection. When IFN-γ and IL-10 responses were both present, the protective effect of IFN-γ was abrogated. A Cox proportional hazard model including IFN-γ, IL-10, MSP142 3D7 IgG antibody responses, hemoglobin S genotype, age, and infection status at baseline showed that the time to blood-stage infection correlated positively with IFN-γ responses and negatively with IL-10 responses, younger age, and asymptomatic parasitemia. Conclusions: Evaluating combined allele-specific cellular and humoral immunity elicited by malaria provides a more informative measure of protection relative to evaluation of either measure alone.
    The Journal of Infectious Diseases 03/2013; 208(1). DOI:10.1093/infdis/jit134 · 6.00 Impact Factor
  • [Show abstract] [Hide abstract]
    ABSTRACT: As an agonist to innate immune system, Salmonella flagellin has been proven to be a potent adjuvant either admixed or genetically fused with antigens and applied to a variety of vaccines against infectious diseases. However, relatively little is known about its carrier-adjuvant effect for conjugate vaccine. Conjugation is an effective approach often used to make haptens such as some peptides and polysaccharides immunogenic and in some cases used to make poor immunogens more immunogenic. In the current study, Salmonella flagellin was tested for its carrier-adjuvant effect in a peptide conjugation. The recombinant Salmonella flagellin (rFliC) purified from Escherichia coli was firstly modified by maleimide groups, then coupled with a synthetic peptide (EXP153:CDNNLVSGP) that is a B-cell epitope derived from Plasmodium falciparum exported protein-1 to generate the conjugate of EXP153-rFliC. Bioactivity assay showed that both chemical modification and conjugation did not apparently impair the TLR5-ligand activity of rFliC. EXP153-rFliC was used to immunize BALB/c mice via subcutaneous route, and the sera obtained from immunized mice were examined by ELISA and IFA. While no detectable antibody responses were induced by the peptide admixed with rFliC, the robust peptide-specific antibody responses were observed in mice immunized with the peptide conjugated to rFliC in the absence of any additional adjuvant. The immune sera induced by the conjugate recognized the native protein of malaria parasite. The data obtained from this study demonstrate the carrier-adjuvant activity of Salmonella flagellin in peptide conjugate immunization and indicate its promising application for conjugate vaccine research and development. Copyright © 2015. Published by Elsevier Ltd.
    Vaccine 03/2015; 33(17). DOI:10.1016/j.vaccine.2015.03.006 · 3.62 Impact Factor