Liver resident-memory CD8+ T cells (TRM cells) can kill liver-stage Plasmodium-infected cells and prevent malaria, but simple vaccines for generating this important immune population are lacking. Here, we report the development of a fully synthetic self-adjuvanting glycolipid-peptide conjugate vaccine designed to efficiently induce liver TRM cells. Upon cleavage in vivo, the glycolipid-peptide conjugate vaccine releases an MHC I–restricted peptide epitope (to stimulate Plasmodium-specific CD8+ T cells) and an adjuvant component, the NKT cell agonist α-galactosylceramide (α-GalCer). A single dose of this vaccine in mice induced substantial numbers of intrahepatic malaria-specific CD8+ T cells expressing canonical markers of liver TRM cells (CD69, CXCR6, and CD101), and these cells could be further increased in number upon vaccine boosting. We show that modifications to the peptide, such as addition of proteasomal-cleavage sequences or epitope-flanking sequences, or the use of alternative conjugation methods to link the peptide to the glycolipid improved liver TRM cell generation and led to the development of a vaccine able to induce sterile protection in C57BL/6 mice against Plasmodium berghei sporozoite challenge after a single dose. Furthermore, this vaccine induced endogenous liver TRM cells that were long-lived (half-life of ~425 days) and were able to maintain >90% sterile protection to day 200. Our findings describe an ideal synthetic vaccine platform for generating large numbers of liver TRM cells for effective control of liver-stage malaria and, potentially, a variety of other hepatotropic infections.

Highlights d The Plasmodium ribosomal protein RPL6 is expressed during liver-stage infection d RPL6 can be targeted by specific liver T RM cells for efficient parasite elimination d Prime-and-trap vaccination targeting RPL6 induces effective protection against malaria d RPL6 is highly conserved across global P. falciparum clinical isolates

Self-adjuvanting vaccines consisting of peptide epitopes conjugated to immune adjuvants are a powerful way of generating antigen-specific immune responses. We previously showed that a Plasmodium-derived peptide conjugated to a rearranged...

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Liver resident-memory CD8+ T cells (TRM cells) can kill liver-stage Plasmodium-infected cells and prevent malaria, but simple vaccines for generating this important immune population are lacking. Here, we report the development of a fully synthetic self-adjuvanting glycolipid-peptide conjugate vaccine designed to efficiently induce liver TRM cells. Upon cleavage in vivo, the glycolipid-peptide conjugate vaccine releases an MHC I-restricted peptide epitope (to stimulate Plasmodium-specific CD8+ T cells) and an adjuvant component, the NKT cell agonist a-galactosylceramide (a-GalCer). Following transfer of a transgenic CD8+ T cell population, a single dose of this vaccine induced substantial numbers of intrahepatic CD8+ T cells expressing canonical markers of liver TRM cells (CD69, CXCR6 and CD101), and these cells could be further increased in number upon vaccine boosting. We show that modifications to the peptide, such as addition of proteosomal-cleavage sequences or epitope-flanking sequences, or the use of alternative conjugation methods to link the peptide to the glycolipid, improved liver TRM cell generation and led to the development of a vaccine able to induce sterile protection in C57BL/6 mice against P. berghei sporozoite challenge after a single dose. Incorporation of a cognate malaria antigen into the vaccine resulted in the generation of large numbers of long-lived liver TRM cells derived from the endogenous T cell population that were capable of providing sterile immunity. Our findings describe an ideal synthetic vaccine platform for generating large numbers of liver TRM cells for effective control of liver-stage malaria and, potentially, a variety of other hepatotropic infections

Liver resident-memory CD8+ T cells (TRM cells) provide important protection against liver-stage Plasmodium infections. Here, we show that a glycolipid-peptide conjugate vaccine designed to be cleaved in vivo to release the NKT cell agonist a-galactosylceramide (a-GalCer), together with a CD8+ T cell malarial epitope, efficiently induced intrahepatic malaria-specific T cells expressing canonical markers of liver TRM (CD69, CXCR6 and CD101). While sterile protection was achieved in a proportion of mice using a single vaccine dose, a combined prime-boost regimen induced higher numbers of liver TRM cells and more robust protection. Improved liver TRM cell generation and sterile immunity using a single dose of the vaccine were obtained in all mice by extending the C-and N-terminal residues of the minimal epitope, and by modifying the linker to enhance a-GalCer release. Our findings describe an ideal synthetic vaccine platform to generate large numbers of liver TRM cells that is potentially valuable for controlling a variety of hepatotropic infections.