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Biodegradable microspheres may represent a potential tool for the delivery of combination vaccines. We demonstrate strong immunogenicity of five co-encapsulated antigens after a single subcutaneous inoculation in guinea pigs. Tetanus- and diphtheria-specific antibodies were not significantly affected by the presence of either antigen or by the presence of pertussis or Haemophilus influenzae type b (Hib) antigens. Microsphere formulations gave better protection against diphtheria toxin than did two injections of a licensed tetravalent vaccine. Finally, a synthetic malaria peptide antigen (PfCS) also encapsulated in PLGA microspheres increased diphtheria and tetanus-specific immunity and improved protection against diphtheria. These findings demonstrate the potential of microspheres as an alternative and promising strategy for combination vaccines with a further aptitude in reducing the number of inoculations required to gain functional immunity. [Copyright &y& Elsevier]

Previous studies demonstrated in vitro phagocytosis of poly(lactide-co-glycolide) (PLGA) microspheres (MS) by macrophages and dendritic cells and the biodistribution of fluorescent PLGA particles following oral or intranasal administration. In this study, we report the uptake and biodistribution of sub-cutaneously administered fluorescent labelled PLGA MS loaded with diphtheria toxoid (DT). The cell type and percentage of fluorescent positive cells were determined by flow cytometry and confirmed by fluorescent microscopy. Fluorescent particles were detected inside cells of the peritoneal flush as early as 10 min post-inoculation, predominantly in cells of macrophage morphology. In vivo trafficking of PLGA particles following a sub-cutaneous immunisation of mice appeared to be governed by macrophages. However, in the first week after inoculation, dendritic cells played a significant role in the uptake and digestion of the microspheres, thereby triggering the immune response against the antigen. Fluorescent PLGA MS were also observed in cells of lymphoid tissues such as mesenteric lymph nodes (MLN) and spleen (S). However, microsphere fluorescence in lymphoid tissues decreased rapidly, as they were degraded inside the cells, thereby enabling the presentation of the antigen to specific cells of the immune system. To our knowledge, this is the first time the fate of immunogenic PLGA microspheres was studied in vivo following a sub-cutaneous injection route. [Copyright &y& Elsevier]

Abstract: There is increasing concern over the efficacy of existing vaccines for diphtheria and there is interest in the development of a mucosally active formulation which might improve local protection. Lipophilic immune stimulating complexes (ISCOMS) containing Quil A are active by both parenteral and mucosal routes and here we have established methods for incorporating palmitified diphtheria toxoid (DT) into ISCOMS. The resulting formulation was immunogenic by the subcutaneous, oral and intranasal routes, with very low doses of DT inducing systemic humoral immune responses, as well as cell mediated immunity including both γIFN and IL5 production. Intranasal immunisation with DT in ISCOMS also stimulated significant local antibody production in tracheal washes, as well as cellular immunity in draining lymphoid tissues and serum neutralising antibodies. Finally, subcutaneous immunisation of guinea pigs with DT in ISCOMS primed protective immunity against challenge with diphtheria holotoxin more efficiently than the equivalent doses of DT in the conventional alum vaccine. ISCOMS based vaccines may provide a novel strategy for mucosal and systemic immunisation against diphtheria. [Copyright &y& Elsevier]