June 12-17, 2016
Pneumonia represents important infection in young children (under five years old). Even with efficient vaccines, this disease was responsible for 992,000 deaths in 2015 and it was globally equivalent to 15% of all deaths in children. Bio-Manguinhos/Oswaldo Cruz Foundation (Brazil), supplies in their National Immunization Program the 10-vallent pneumococcal vaccine. This vaccine contains 10 of the 92 serotypes of this pathogen, and it´s obtained through fermentations and purifications of the capsular polysaccharides of each serotype, followed by chemical couplings to specific carrier proteins. This work presents a proposal for industrial process of only one recombinant protein antigen, the PsaA (pneumococcal surface adhesion A with 37 kDa), potentially capable to act against most of all prevalent serotypes of streptococcus pneumoniae. Based on protein expression in prokaryotic system and purification, this process reached high purity levels and yield superior to conventional fermentation systems. Biomass was obtained in bench scale reactor and studies of expression times verified that it could be reduced in 60% from presented previously. Moreover, in purification steps was observed good resolution using anion exchange (DEAE Sepharose FF®). For desalting, it was observed that gel filtration technique could be replaced for tangential cross flow filtration, better to scale-up, and using 10 kDa membranes. The developed process resulted in overall yield of 35-40%, based in BCA analysis. Electrophoretic analysis (SDS-PAGE) of the purified sample showed a band with molecular weight of 37 kDa corresponding to 80% of identified proteins. As formulation proposal, chitosan in different concentrations were used to prepare capsules of the antigen resulting in encapsulation efficiency between 26 and 36% that can be evaluated as an intranasal delivery system. This data show that the methodology developed is promising for use in industrial process aimed at obtain a recombinant vaccine.
Ana Maria P. Santos, Mariana Miguez, Ana Paula Argondizzo, and Maria Helena Rocha-Leão, "Development of a production process for a recombinant protein pneumococcal vaccine" in "Vaccine Technology VI", Laura Palomares, UNAM, Mexico Manon Cox, Protein Sciences Corporation, USA Tarit Mukhopadhyay, University College London, UK Nathalie Garçon, BIOASTER Technology Research Institute, FR Eds, ECI Symposium Series, (2016). http://dc.engconfintl.org/vaccine_vi/100