Abstract In human HER2+ breast cancer, neoadjuvant intratumoral delivery of conventional type 1 dendritic cells loaded with HER2 peptides (HER2 DCs) reprograms the immune milieu, induces potent antitumor regression, and, unexpectedly, recruits natural killer T (NKT)-like cells, which can surveil lipids on CD1D-expressing antigen-presenting cells. Given the high infiltration of immunosuppressive M2-like macrophages and myeloid-derived suppressor cells (MDSCs), we hypothesized that this DC immunotherapy could leverage invariant NKT (iNKT) cells to target these myeloid cells via CD1D. CD1D knockout in murine bone marrow-derived HER3 DCs abrogated the effectiveness of immunotherapy in the EO771 breast cancer and B16 melanoma models, yet did not induce phenotypic deficiencies in antigen presentation or apoptosis rates. Conversely, loading DCs with the prototypical iNKT lipid antigen, alpha-galactosylceramide (α-GalCer DCs), further enhanced tumor regression across multiple tumor models. Additionally, in the HER2+ TUBO breast cancer model, intratumoral adoptive cell transfer of iNKT cells and α-GalCer + HER2 DCs simultaneously further enhanced the immunotherapy and tumor control. Through flow cytometric assays of iNKT cells cocultured with bone marrow-derived macrophages or MDSCs, iNKT cells demonstrated enhanced apoptosis of α-GalCer-loaded bone marrow-derived MDSCs that was contingent on CD1D interaction. However, intratumoral injections of free α-GalCer abrogated iNKT and DC immunotherapy in the same TUBO model. These findings indicate that iNKT cells require context- and cell-specific activation to participate in successful reprogramming of the tumor immune microenvironment. Citation Format: Vincent Lok, Saurabh Garg, Colin Snyder, Amy Aldrich, Doris Wiener, Brian J. Czerniecki. Intratumoral dendritic cell immunotherapy rewires the tumor microenvironment through contextualized invariant natural killer T-cell interactions abstract. In: Proceedings of the AACR Immuno-Oncology Conference (AACR IO): Discovery and Innovation in Cancer Immunology: Revolutionizing Treatment through Immunotherapy; 2026 Feb 18-21; Los Angeles, CA. Philadelphia (PA): AACR; Cancer Immunol Res 2026;14(2 Suppl):Abstract nr C017.
Lok et al. (Wed,) studied this question.