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Conjugate vaccines against encapsulated pathogens like Streptococcus pneumoniae face many challenges, including the existence of multiple serotypes with a diverse global distribution that constantly requires new formulations and higher coverage. Multivalency is usually achieved by combining capsular polysaccharide-protein conjugates from invasive serotypes, and for S. pneumoniae, this has evolved from 7- up to 20-valent vaccines. These glycoconjugate formulations often contain high concentrations of carrier proteins, which may negatively affect glycoconjugate immune response. This work broadens the scope of an efficient multicomponent strategy, leading to multivalent pneumococcal glycoconjugates assembled in a single synthetic operation. The bioconjugation method, based on the Ugi four-component reaction, enables the one-pot incorporation of two different polysaccharide antigens to a tetanus toxoid carrier, thus representing the fastest approach to achieve multivalency. The reported glycoconjugates incorporate three combinations of capsular polysaccharides 1, 6B, 14, and 18C from S. pneumoniae. The glycoconjugates were able to elicit functional specific antibodies against pneumococcal strains comparable to those shown by mixtures of the two monovalent glycoconjugates.

Peptide stapling is traditionally used to lock peptide conformations into α-helical structures using a variety of macrocyclization chemistries. In an endeavor to add a diversity-generating tool to this repertoire, we introduce a multicomponent stapling approach enabling the simultaneous stabilization of helical secondary structures and the exocyclic N-functionalization of the side chain-tethering lactam bridge. This is accomplished by means of a novel solid-phase methodology comprising, for the first time, the on-resin Ugi reaction-based macrocyclization of peptide side chains bearing amino and carboxylic acid groups. The exocyclic diversity elements arise from the isocyanide component used in the Ugi multicomponent stapling protocol, which allows for the incorporation of relevant fragments such as lipids, sugars, polyethylene glycol, fluorescent labels, and reactive handles. We prove the utility of such exocyclic reactive groups in the bioconjugation of a maleimide-armed lactam-bridged peptide to a carrier protein. The on-resin multicomponent stapling proved efficient for the installation of not only one, but also two consecutive lactam bridges having either identical or dissimilar N-functionalities. The easy access to helical peptides with a diverse set of exocyclic functionalities shows prospect for applications in peptide drug discovery and chemical biology.