63P Single-cell RNA-seq combined with bulk RNA-seq revealed the involvement of pancreatic cancer tissue-resident macrophages in tumour progression and the immunotherapy response

Background: Personalized immunotherapy has proven to be crucial in the treatment of tumors with high genetic heterogeneity.Neoantigens derived from tumor mutations allow for precise immune activation.This study addresses the use of exosomes derived from dendritic cells pulsed with neoantigens in co-culture to enhance immune response.The combination of neoantigens and pulsing in dendritic cells optimizes immune activation, laying new therapeutic groundwork in cancer treatment.Methods: Exosomes were generated from dendritic cells subjected to co-culture with pulsing using tumor-specific neoantigens.Exosomes were characterized by Nanosight, with an average size of 104.7 AE 10.3 nm and concentration of 7.15 x 10 9 particles/ mL.The presence of the exosomal markers CD63, CD81 and Alix was confirmed by Western blot.To evaluate the efficacy of coculture and dendritic cell pulsing, Native T cell activation was measured by flow cytometry (CD69 and CD25), proinflammatory cytokine production (IFN-g, IL-6, TNF-a) by ELISA and Cytometric Bead Array, and apoptosis induction in tumor cells by lactate dehydrogenase (LDH) release.Results: Native T cell activation showed an increase of 50.7% (95% CI: 48.2-53.1%) in cultures treated with pulsed dendritic cell-derived exosomes.IFN-g production increased by 55.4% (95% CI: 53.6-57.2%),IL-6 by 49.8% (95% CI: 47.9-51.7%)and TNFa by 30.5% (95% CI: 28.8-32.1%).The apoptosis rate in treated tumor cells increased by 34.7% (95% CI: 32.4-36.8%)and LDH release increased by 31.9% (95% CI: 29.7-33.5%)versus controls.The results indicate an effective immune activation.Conclusions: This study demonstrates that exosomes derived from neoantigen-pulsed dendritic cells in coculture have great potential to activate specific immune responses and modify the tumor microenvironment.The observed increases in T cell activation, cytokine production and apoptosis suggest that this strategy can be successfully applied in personalized cancer immunotherapy, particularly in tumors resistant to conventional therapies.Further clinical studies are recommended to validate these results.