(556f) A Self-Assembled pH-Responsive Multi-Component Platform for Oral Vaccination

Next-generation vaccine research must achieve safe, cost-effective and easily administered vaccines with 1) needle-free delivery mechanism, and 2) immunization with subunit vaccines. Oral vaccines improve ease of administration and confer protection at mucosal surfaces for more robust immune responses. Subunit vaccines (e.g. protein antigens) are desirable to overcome safety issues associated with current attenuated or inactivated oral vaccines, but are poorly immunogenic and absorbed, requiring the use of adjuvants. We have previously demonstrated success using anionic hydrogel carriers to deliver protein therapeutics, such as insulin and calcitonin, in a pH-dependent manner for targeted intestinal delivery. Successful vaccination strategies require direct antigen internalization and elicitation of immunogenic responses.

We have developed a system that combines two biomaterial platforms. In its core, polyanhydride nanoparticles (PNPs) based on 1,6-bis-(p-carboxyphenoxy)hexane (CPH) and sebacic acid (SA) serve simultaneously as adjuvant and delivery vehicle of subunit antigens. These are protected by pH-responsive polymers based on poly(ethylene glycol) (PEG) and poly(methacrylic acid) (PMAA), shielding a payload of PNPs through the harsh conditions of the gastrointestinal tract for selective release in the small intestine, proximal to antigen-sampling M cells. This platform is referred to as Polyanhdyride-Releasing MicroParticle Technology, or PROMPT.

PROMPT formulations were synthesized by pH-mediated anti-solvent precipitation to encapsulate PNPs. Briefly, the pH-responsive copolymer, denoted P(MAA-g-EG), was synthesized by UV initiated free radical polymerization of MAA and polyethylene glycol methacrylate and characterized by ¹³C NMR, FT-IR, and GPC. PNP formulations were synthesized by anti-solvent nanoprecipitation to encapsulate model protein antigen, ovalbumin. P(MAA-g-EG) dispersions were flocculated in the presence of PNPs by desolvating the polymer through pH reduction. PROMPT particle morphology and PNP encapsulation were evaluated by scanning electron microscopy and confocal microscopy, respectively. In vitro release kinetics were evaluated at physiological conditions and quantified using the micro bicinchoninic acid (microBCA) assay. Enzyme-linked immunosorbent assay (ELISA) was used to evaluate that antigenicity of ovalbumin released from PROMPT formulations. Cytocompatibility of self-assembled PROMPT formulations was evaluated in two cell lines: Caco-2 intestinal epithelial cells and RAW264.7 macrophages. RAW264.7 cells were also used to evaluate PNP internalization. Results indicated that PROMPT formulations can achieve targeted release of PNPs in intestinal conditions. Formulations preserve protein antigenicity, and nanoparticles released from PROMPT are readily internalized by macrophages, indicating PROMPT has potential as an adaptable platform for oral subunit vaccine administration.

Acknowledgements: This work was supported by a grant from the National Institutes of Health (5-R01-EB-000246-20) and a NSF GRFP. We thank the research group of Prof. Balaji Narasimhan from Iowa State University for their collaboration on this project.