Poly(hydrophobic amino acid) Conjugates for the Delivery of Multiepitope Vaccine against Group A Streptococcus.

Peptide-based vaccines are composed of small, defined, antigenic peptide epitopes. They are designed to induce well-controlled immune responses. Multiple epitopes are often employed in these vaccines to cover strain variability of a pathogen. However, peptide epitopes cannot stimulate adequate immune responses on their own and require an adjuvant (immune stimulant) and/or delivery system. Here, we designed and synthesized a multiepitope vaccine candidate against Group A Streptococcus (GAS) composed of several B-cell epitopes (J8, PL1, and 88/30) derived from GAS M-protein, universal PADRE T-helper cell epitope, and a polyleucine self-adjuvanting unit. The vaccine components were conjugated together (using mercapto-maleimide and azide-alkyne Huisgen cycloaddition reactions) or delivered as a mixture. The conjugated multiepitope vaccine candidate self-assembled into small nanoparticles and chain-like aggregated nanoparticles (CLANs) that were able to induce the production of J8-, PL1-, and 88/30-specific antibodies in mice. The multiepitope conjugate and the physical mixture of conjugates bearing the individual epitopes produced similar nanoparticles and induced comparable immune responses. Hence, simple physical mixing can replace complex chemical conjugation to produce multiepitope nanoparticles with equivalent morphology and immunological efficacy. This greatly simplifies vaccine production.

Plasmodium falciparum formins are essential for invasion and sexual stage development.

The malaria parasite uses actin-based mechanisms throughout its lifecycle to control a range of biological processes including intracellular trafficking, gene regulation, parasite motility and invasion. In this work we assign functions to the Plasmodium falciparum formins 1 and 2 (FRM1 and FRM2) proteins in asexual and sexual blood stage development. We show that FRM1 is essential for merozoite invasion and FRM2 is required for efficient cell division. We also observed divergent functions for FRM1 and FRM2 in gametocyte development. Conditional deletion of FRM1 leads to a delay in gametocyte stage progression. We show that FRM2 controls the actin and microtubule cytoskeletons in developing gametocytes, with premature removal of the protein resulting in a loss of transmissible stage V gametocytes. Lastly, we show that targeting formin proteins with the small molecule inhibitor of formin homology domain 2 (SMIFH2) leads to a multistage block in asexual and sexual stage parasite development.

Poly(hydrophobic amino acid)-Based Self-Adjuvanting Nanoparticles for Group A Streptococcus Vaccine Delivery.

Peptide antigens have been widely used in the development of vaccines, especially for those against autoimmunity-inducing pathogens and cancers. However, peptide-based vaccines require adjuvant and/or a delivery system to stimulate desired immune responses. Here, we explored the potential of self-adjuvanting poly(hydrophobic amino acids) (pHAAs) to deliver peptide-based vaccine against Group A Streptococcus (GAS). We designed and synthesized self-assembled nanoparticles with a variety of conjugates bearing a peptide antigen (J8-PADRE) and polymerized hydrophobic amino acids to evaluate the effects of structural arrangement and pHAAs properties on a system's ability to induce humoral immune responses. Immunogenicity of the developed conjugates was also compared to commercially available human adjuvants. We found that a linear conjugate bearing J8-PADRE and 15 copies of leucine induced equally effective, or greater, immune responses than commercial adjuvants. Our fully defined, adjuvant-free, single molecule-based vaccine induced the production of antibodies capable of killing GAS bacteria.

Poly(amino acids) as a potent self-adjuvanting delivery system for peptide-based nanovaccines.

To be optimally effective, peptide-based vaccines need to be administered with adjuvants. Many currently available adjuvants are toxic, not biodegradable; they invariably invoke adverse reactions, including allergic responses and excessive inflammation. A nontoxic, biodegradable, biocompatible, self-adjuvanting vaccine delivery system is urgently needed. Herein, we report a potent vaccine delivery system fulfilling the above requirements. A peptide antigen was coupled with poly-hydrophobic amino acid sequences serving as self-adjuvanting moieties using solid-phase synthesis, to produce fully defined single molecular entities. Under aqueous conditions, these molecules self-assembled into distinct nanoparticles and chain-like aggregates. Following subcutaneous immunization in mice, these particles successfully induced opsonic epitope-specific antibodies without the need of external adjuvant. Mice immunized with entities bearing 15 leucine residues were able to clear bacterial load from target organs without triggering the release of soluble inflammatory mediators. Thus, we have developed a well-defined and effective self-adjuvanting delivery system for peptide antigens.