PULMONARY DELIVERY OF PNEUMOCCOCAL VACCINE USING NANOCOMPOSITE MICROPARTICLE CARRIERS VIA DRY POWDER INHALATION

Abstract

S. pneumoniae is one of the most significant human pathogens, causing high morbidity and mortality rates globally. Although there are vaccine available such as PPV 23, PCV7, PCV10, and PCV13, they are ineffective in some situations due to the differing epidemiology of various serotypes depending on the site of infection and the geographical location. Furthermore, they are expensive to produce and distribute. Universal research is presently concentrated on establishing other pneumococcalvaccine approaches such as using pneumococcal surface protein A (PspA) which relate to pathogenesis and are common to all serotypes. In this study polymeric nanoparticles (NPs) encapsulating PspA4Pro were incorporated into microcarriers using L-leucine and spray dried to produce nanocomposite micro#particles (NCMPs) dry powder for inhalation. Parameters for the preparation of protein-loaded polyester poly (Glycerol Adipate-co-I‰-Pentadecalactone), (PGA-co-PDL) NCMPs were optimised using Taguchi design and BSA as a model protein, by the double emulsion solvent evaporation method followed by spray drying. Particle size was mainly affected by the polymer mass and small particle size ≤ 500nm was achieved. The most important factor for obtaining a high BSA loading was BSA concentration. The spray drying process was optimised to produce NCMPs with a porous corrugated surface, 50% yield, MMAD of 1.71±0.10 I¼m and FPF% of 78.57±0.1%. Adsorption of chitosan hydrochloride (CHL) onto PGA-co-PDL NPs can be used assuccessful strategies to produce cationic NPs. Cationic NPs were prepared with similarparticle size to anionic NPs ≤ 500nm. The In vitro aerosolisation performance ofcationic NPs/NCMPs showed FPF% of 46.79±11.21% and MMAD of 1.49±0.29 I¼m. Further cell viability studies on A549 cell line showed a good profile with a cell viability of 79±4.7% for anionic NPs/NCMPs and 78.85±9.96% for cationic xviii NPs/NCMPs at 2.5 mg/ml concentration after 24 h exposure. The previous results introduced a successful method for preparing anionic and cationic NPs/NCMPs for delivering PspA4Pro as dry powder via inhalation. The particle size of PspAPro4 loaded anionic NPs and cationic NPs were 310±25.3 nm and 409.7±49.5 nm, respectively, to be effectively taken up by dendritic cells (DCs). The PspA4Pro loading in anionic NPs was 65.73±5.6 I¼g/mg and in cationic NPs was 9.84±1.4 I¼g/mg. The PspA4Pro released from anionic and cationic NPs/NCMPs preserved its primary and secondary structure as evaluated by SDS-PAGE and circular dichroism. In vitro release studies showed that the anionic NPs/NCMPs formulations achieved a cumulative release of 21.01±1.5% while the cationic NPs/NCMPs formulation released 83.13 ±0.84% after 48 h. DCs uptake studies provide evidence of particles uptake by DCs upon incubation for 1 h as visualized by confocal microscopy. These results indicate the use of optimised methods for developing polymeric based NCMPs for vaccine delivery via inhalation against pneumococcal diseases.