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

Abstract

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.

Polyphenylalanine as a self-adjuvanting delivery system for peptide-based vaccines: the role of peptide conformation

“…A similar tendency to self-assemble was observed with polyleucine antigen conjugates. [19][20][21] Particles were highly polydisperse due to aggregates formation. Consequently, particle size distribution measured by dynamic light scattering (DLS) did not fully correspond with that observed through transmission electron microscopy (TEM) (Supplementary Fig.…”

“…The GAS B-cell epitope-poly(hydrophobic amino acid) conjugates induced significant humoral immune responses; however, only the compound bearing a 15-leucine unit induced the production of superior IgG titers. [19][20][21] The antibodies produced were opsonic against GAS clinical isolates. Interestingly, mice treated with the compound bearing 10 phenylalanines did not produce opsonic antibodies, in contrast to its 10-leucine analogue, despite both inducing the same IgG titers.…”

Development of Multilayer Nanoparticles for the Delivery of Peptide-Based Subunit Vaccine against Group A Streptococcus

“…Antibody opsonization assays were performed with GAS clinical isolate strains provided by the Princess Alexandra Hospital: strains GC203 (wound swab) and D3840 (nasopharynx swab) [ 34 ]. The two strains were streaked on Todd Hewitt broth (THB) agar plates with 5% yeast supplementation and incubated at 37 °C for 24 h. Single colonies from the plates were inoculated in 5 mL of THB + 5% yeast and incubated until bacterial replication reached approximately 4.6 × 10 6 colony forming units (CFU)/mL.…”

“…The mixtures were added to 96-well plates containing a 2-fold dilution of bacterial culture. The plates were incubated at 37 °C for 3 h. Then, 10 μL aliquots from the wells were streaked onto plates with THB agar + 5% yeast and 5% horse blood, and incubated for another 24 h [ 34 ]. Bacterial survival rates were calculated based on the number of colonies on the plate.…”

Liposomal Formulations of a Polyleucine–Antigen Conjugate as Therapeutic Vaccines against Cervical Cancer

“…The application of fully defined natural hydrophobic amino acids serving as monomers may overcome all disadvantages of classic polymer-based delivery systems. We have demonstrated that a pHAA sequence consisting of pLeu conjugated to the J8 B-cell epitope from group A Streptococcus self-assembled into nanoparticles stimulated the production of high titers of opsonic antibodies in mice [ 23 , 24 ]. Moreover, pLeu -based peptide vaccines were very effective in triggering humoral immunity against hookworm and in parasite clearance from infected animals [ 25 , 26 , 27 ].…”