Towards the Development of a Mucosally Active Vaccine against Group A Streptococcus (GAS)

Mehfuz Zaman (2011). Towards the Development of a Mucosally Active Vaccine against Group A Streptococcus (GAS) PhD Thesis, School of Chemistry & Molecular Bioscience, The University of Queensland.

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Author Mehfuz Zaman
Thesis Title Towards the Development of a Mucosally Active Vaccine against Group A Streptococcus (GAS)
School, Centre or Institute School of Chemistry & Molecular Bioscience
Institution The University of Queensland
Publication date 2011-09
Thesis type PhD Thesis
Supervisor Professor Istvan Toth
Dr. Micahel Batzloff
Total pages 142
Total colour pages 3
Total black and white pages 139
Language eng
Subjects 03 Chemical Sciences
Abstract/Summary Vaccination is the administration of material (a vaccine) that results in immunity to a disease or a pathogen. Vaccination in general is considered to be one of the most effective methods for prevention of infectious diseases. The evidence for the efficacy of many vaccines, such as the influenza vaccine, the human papilloma virus vaccine and the small pox and chicken pox vaccine among others highlight the contribution of vaccines to the betterment of public health. Streptococcus pyogenes or Group A Streptococcus (GAS) infection is a common cause of pharyngitis, scarlet fever and impetigo. GAS infection results in other more severe complications, namely acute rheumatic fever/rheumatic heart disease, streptococcal reactive arthritis, etc. It is the post-infectious, immune-mediated sequelae that cause severe morbidity and mortality in developing countries. The global prevalence of serious streptococcal infections and the continued endemicity of fatal post-streptococcal sequelae in developing countries have resulted in renewed interest in GAS, especially in developing a vaccine to prevent the associated diseases. Traditional vaccines (killed or live attenuated) have been effective against many bacterial and viral infectious diseases. However, application of these traditional techniques in many cases including GAS has not shown the same success. Risks associated with the use of live–attenuated pathogens for a GAS vaccine include the development of autoimmune diseases. Other vaccine strategy such as subunit vaccines (recombinant proteins or peptides) and carrier protein conjugated vaccine are hampered by the lack of suitable adjuvant, carriers and delivery systems. Mucosal surface of the upper respiratory tract is a primary site of GAS infections. As such an optimal GAS vaccine would induce both mucosal and systemic immune responses. This would prevent primary GAS infection and the development of GAS-associated diseases. The current thesis is focused on the design, synthesis and immunological evaluation of novel delivery systems against GAS. The first chapter reviews strategies utilized in the field of GAS vaccine development, basis of mucosal immunity and new findings in immunology and mechanism of action of particulate polymers and lipopeptides which represent the basis of our novel strategies for GAS vaccine development. In the chapters that follow, the immunological evaluation of a polyacrylate polymer based delivery system is described in chapter 2. This vaccine delivery system consisted of a target GAS B cell epitope (J14) covalently attached to a polyacrylic acid based polymer imparting adjuvanting activity. Chemical attachment of J14 was essential for immunogenicity and following systemic and intranasal administration of this nanoparticulate construct without additional adjuvant, J14-specific systemic IgG was induced. The antibodies elicited were also capable of in vitro opsonisation of a GAS strain found in endemic areas. The promising preliminary data shows the potential of this novel self-adjuvanting polyacrylate polymer based construct as a peptide vaccine delivery platform, which may afford promising opportunities for treating systemic GAS infection. It is also of relevance to other disease models where systemic immune responses are desired. A major aim of this thesis was to develop and optimize an intranasally administered, lipopeptide based mucosally active GAS vaccine candidate. The third chapter includes immunological assessment of a lipopeptide vaccine library composed of: (i) J14 and (ii) a lipid moiety based on lipoamino acids (LAA). Systemic J14-specific IgG antibodies were detected following intranasal immunization of inbred B10.Br (H-2k) and outbred Swiss mice without the need for an additional adjuvant. The effect of changing the position of J14, and lipid moiety attachment on antibody titer was assessed. The point of lipid moiety and J14 attachment had a significant influence on systemic J14-specific IgG antibody titer in outbred Swiss mice. Overall, the optimal lipopeptide GAS vaccine featured a C-terminal lipid moiety, conjugated through a central lysine residue and J14 on the lysine side-chain. The fourth chapter describes the design, synthesis and evaluation of a three component mucosally active GAS vaccine candidate, composed of: (i) a universal helper T cell epitope (P25), (ii) the target GAS B cell epitope (J14), and (iii) a LAA based lipid moiety. Structure activity relationship of the synthesized lipopeptides demonstrated that the choice of B and T helper epitope had a significant impact on mucosal IgA response. Replacement of P25 with another T helper epitope (TH2R) or replacing the B cell epitope with another GAS epitope (88/30) abolished IgA activity in B10.Br and Swiss mice. No significant differences were observed for the systemic IgG response. Our data would suggest that mucosal activity by the lipopeptide of interest is due to the sum of all of its three components rather than due to an individual component. Further investigation is needed to determine the contributing factors for mucosal immune response. Bactericidal assay showed the mucosally active lipopeptide was capable of in vitro opsonisation of a virulent GAS strain. IgG isotyping displayed the lipopeptide elicits a mixed T helper type 1(Th1)/Th2 response. Dynamic light scattering (DLS) measurements and transmission electron microscopy (TEM) images were used to analyse the particle size and the lipopeptide of interest formed particles in the nanometer range in aqueous solution. The fifth chapter identifies our lipopeptide vaccine strategy as a novel Toll-like receptor (TLR) 2 targeting synthetic ligand. A LAA based lipid moiety composed of two alkyl chains of 16 carbons was found to be optimal for TLR2 activation regardless of the position of lipid attachment.
Keyword mucosal Immunity
group A streptococcus
toll-like receptor 2
intranasal immunization
polyacrylic acid
Additional Notes color pages = pages 53, 77 and 102. Grayscale pages = page 103 landscape page = page 142

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