NKT Cell-Driven Enhancement of Antitumor Immunity Induced by Clec9a-Targeted Tailorable Nanoemulsion

Invariant natural killer T (iNKT) cells are a subset of lymphocytes with immune regulatory activity. Their ability to bridge the innate and adaptive immune systems has been studied using the glycolipid ligand α-galactosylceramide (αGC). To better harness the immune adjuvant properties of iNKT cells to enhance priming of antigen-specific CD8⁺ T cells, we encapsulated both αGC and antigen in a Clec9a-targeted nanoemulsion (TNE) to deliver these molecules to cross-presenting CD8⁺ dendritic cells (DC). We demonstrate that, even in the absence of exogenous glycolipid, iNKT cells supported the maturation of CD8α⁺ DCs to drive efficient cross-priming of antigen-specific CD8⁺ T cells upon delivery of Clec9a/OVA-TNE. The addition of αGC to the TNE (Clec9a/OVA/αGC) further enhanced activation of iNKT cells, NK cells, CD8α⁺ DCs, and polyfunctional CD8⁺ T cells. When tested therapeutically against HPVE7-expressing TC-1 tumors, long-term tumor suppression was achieved with a single administration of Clec9a/E7 peptide/αGC TNE. Antitumor activity was correlated with the recruitment of mature DCs, NK cells, and tumor-specific effector CD8⁺ T cells to the tumor-draining lymph node and tumor tissue. Thus, Clec9a-TNE codelivery of CD8⁺ T-cell epitopes with αGC induces alternative helper signals from activated iNKT cells, elicits innate (iNKT, NK) immunity, and enhances antitumor CD8⁺ T-cell responses for control of solid tumors.