Inflammatory Type 2 Conventional Dendritic Cells Induced by Vaccine Adjuvant Promote Antigen Cross-Presentation through Autophagy Inhibition 2305812

Abstract Introduction Cross-presentation (XPT) is pivotal for inducing antigen (Ag)-specific CD8+ T cell responses, which is linked to better therapeutic outcomes. However, in the context of therapeutic vaccination, how to promote T cell cross-priming in vivo remains unclear. This motivates elucidating the mechanisms underlying XPT by specific cross-presenting subsets under physiological conditions. Methods Ag-specific cytotoxic responses in EG7-OVA tumor model upon adjuvanted vaccination with Th1-stimulating adjuvant was assessed for the mechanism underlying cross-priming. The key cross-presenting subsets under adjuvanted vaccination were analyzed by flow cytometry. The potential pathways related to induced XPT were discovered by RNA sequencing (RNA-seq) and pathway analysis and the results were validated by immunoblotting and OT-I XPT assay. Unbiased single-cell RNA-seq was employed to define the counterpart in human immune system (HIS) mice. Results Adjuvanted vaccination elicited a robust Ag-specific anti-tumor response and expanded inflammatory cDC2 (inf-cDC2) in tumor model. In knockout mice studies, inf-cDC2 emerges as the dominant cross-presenting subset during vaccination. RNA-seq analysis highlighted autophagy-related pathways, next confirming that autophagy inhibition drives XPT. Mechanistically, impeding the fusion of autophagosomes and lysosomes in DCs could prime T cells by prolonged XPT. In HIS mice, DC3 was identified as the phenotypic counterpart of inf-cDC2 under vaccination. Adjuvant-induced pathways were highly enriched in autophagy and autophagy inhibition exclusively within DC3 promotes XPT. Conclusion We identified murine inf-cDC2 and human DC3–not cDC1–are the key player in XPT in the context of adjuvanted vaccination and this marked function strongly depends on autophagy blockade. Prioritizing autophagy inhibition, particularly in these DCs, could effectively augment Ag-specific anti-tumor immunity. This bridges the gap between preclinical and clinical investigations in cancer vaccinology. Funding Source n/a Topic Categories Classical and Non-Classical Antigen Presenting Cells (APC)