Glycolipid–peptide conjugates designed to release vaccine components within target cells ensuring potent CD1d dependent T cell responses.

An important goal of vaccination against viruses and virus-driven cancers is to elicit cytotoxic CD8+ T cells specific for virus-derived peptides. CD8+ T cell responses can be enhanced by engaging help from natural killer T (NKT) cells. We have produced synthetic vaccines that induce strong peptide-specific CD8+ T cell responses in vivo by incorporating an NKT cell-activating glycolipid. Here we examine the effect of a glycolipid-peptide conjugate vaccine incorporating an NKT cell-activating glycolipid linked to an MHC class I-restricted peptide from a viral antigen in human peripheral blood mononuclear cells. The vaccine induces CD1d-dependent activation of human NKT cells following enzymatic cleavage, activates human dendritic cells in an NKT-cell dependent manner, and generates a pool of activated antigen-specific CD8+ T cells with cytotoxic potential. Compared to unconjugated peptide, the vaccine upregulates expression of genes encoding interferon-γ, CD137 and granzyme B. A similar vaccine incorporating a peptide from the clinically-relevant human papilloma virus (HPV) 16 E7 oncoprotein induces cytotoxicity against peptide-expressing targets in vivo, and elicits a better antitumor response in a model of E7-expressing lung cancer than its unconjugated components. Glycolipid-peptide conjugate vaccines may prove useful for the prevention or treatment of viral infections and tumors that express viral antigens.

Prostate cancer is the second most common cancer in men worldwide. Despite an abundance of prostate-specific antigens, immunotherapies have yet to become a standard of care, potentially limited by T-cell dysfunction. Up to 10% of human circulating T-cells, and a significant fraction in the urogenital tract, are mucosal-associated invariant T (MAIT) cells. MAIT cells express stereotyped T-cell receptors that recognize riboflavin metabolites derived from microbes presented by MR-1. We evaluated the number, phenotype and function of circulating MAIT cells, alongside two other innate-like T (ILT) -cell subsets, in men with prostate cancer and age- and sex-matched controls. MAIT cells in men with prostate cancer circulated at similar frequencies to controls, but their cytokine production and proliferation was impaired. In contrast, the function of two other ILT-cell populations (natural killer T-cells and Vγ9Vδ2 T-cells) was not impaired. In both patients and controls, MAIT cells expressed high levels of the immune checkpoint molecule PD-1 at rest, while upregulation of PD-1 in response to the MR-1 ligand 5-amino-6D-ribitylaminouracil (5-A-RU) was greater in patients. 5-A-RU also induced upregulation of PD-L1 and -L2 RNA in primary mononuclear cells. We confirmed that circulating MAIT cell number and function were preserved before and during anti-PD1 therapy with pembrolizumab in a cohort of patients with melanoma. In vitro, 5-A-RU enhanced mononuclear cell cytotoxicity against the PD-L1 positive prostate cancer cell line PC3 in an MR-1-dependent manner. Addition of pembrolizumab enhanced this cytotoxicity, and was associated with increased MAIT cell expression of CD107a and IFN-γ. We conclude that prostate cancer is associated with MAIT-cell dysfunction, and that this might be overcome through the application of potent MR-1 ligands with PD-1 blockade. These findings may have implications for the development of cancer immunotherapies that exploit MAIT cells.

Through chemical synthesis we have designed out the toxicity associated with the powerful vaccine adjuvant α-GalCer.