Chitosan microparticles and nanoparticles as biocompatible delivery vehicles for peptide and protein-based immunocontraceptive vaccines.

摘要

It has become increasingly recognized that polymer particle size can have a profound effect on the interactions of particle-based vaccines with antigen presenting cells (APCs) thereby influencing and modulating ensuing immune responses. With the aim of developing chitosan particle-based immunocontraceptive vaccines, we have compared the use of chitosan-based nanoparticles and chitosan-based microparticles as vaccine delivery vehicles for vaccine candidates based on luteinizing hormone-releasing hormone (LHRH). Particles, functionalized with chloroacetyl groups, which allows the covalent attachment of thiol-containing antigens, were able to adsorb ~60-70% of their weight of peptide-based antigen and 10-20% of their weight of protein-based antigen. Quantitation by amino acid analysis of antigen associated with particles demonstrated a correlation between associated antigen and the degree of chloracetylation of particles. Visualization of fluorescently labeled antigen-loaded particles by confocal microscopy indicated that the majority of antigen was localized at the particle surface with a smaller amount located in the interior. We also found that uptake of both fluoresceinated nanoparticles and microparticles by dendritic cells occurred in a manner dependent on particle concentration. Nanoparticles trafficked from the injection site to draining lymph nodes faster than microparticles; high numbers of nanoparticle-bearing cells appeared in draining lymph nodes on day 3 and microparticles on day 4. This difference in trafficking rate did not, however, appear to have any significant impact on the ensuing immune response because inoculation with both peptide-conjugated and protein-conjugated particles induced high levels of LHRH-specific antibodies. In the case of protein-conjugated particles, the levels of antibodies elicited were similar to those elicited following inoculation with antigen emulsified with complete Freund's adjuvant. The approach to vaccine design that we have described here could represent another useful method for inducing immune responses against microbial, viral and tumorigenic protein antigens.