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Group A streptococcus (GAS; Streptococcus pyogenes), known as the ‘flesh-eating bacterium’, is a human bacterial pathogen that normally causes benign infections (e.g. sore throat and pyoderma), but is also responsible for severe invasive infections (e.g. ‘flesh-eating’ disease and toxic shock syndrome), heart disease, and kidney failure. A safe commercial GAS vaccine is yet to be developed. Individual GAS antigens demonstrate potential universal expression across all GAS serotypes (>200 known), with dramatically reduced concern for autoimmune complications, and compelling efficacy in preclinical testing in mice. In this study, we developed a stepwise conjugation strategy, copper-catalysed alkyne–azide cycloaddition reaction (CuAAC), followed by mercapto–maleimide conjugation, to synthesise a multiantigenic, self-adjuvanting, peptide-based vaccine candidate against GAS. This multiantigenic vaccine includes two GAS antigens, J8 and NS1, a T-helper epitope, PADRE, and a self-adjuvanting moiety, dipalmitoyl serine.

Development of a synthetic vaccine against group A streptococcal infection is increasingly paramount due to the induction of autoimmunity by the main virulent factor – M protein. Peptide vaccines, however, are generally poorly immunogenic, necessitating administration with carriers and adjuvants. One of the promising approaches to deliver antigenic peptides is to assemble peptides on a suitable template which directs the attached peptides to form a well defined tertiary structure. For self-adjuvanting human vaccines, the conjugation of immunostimulatory lipids has been demonstrated as a potentially safe method. This study describes the design and optimized synthesis of two lipopeptide conjugated carbohydrate templates and the assembling of peptide antigens. These lipopeptide–carbohydrate assembled multivalent vaccine candidates were obtained in high yield and purity when native chemical ligation was applied. Circular dichroism studies indicated that the template-assembled peptides form four α-helix bundles. The developed technique extends the use of carbohydrate templates and lipopeptide conjugates for producing self-adjuvanting and topology-controlled vaccine candidates.