In situ reprogramming of fibroblasts into antigen-presenting pseudo-dendritic cells via IFN-β-engineered protoplast-derived exosomes delivered by microneedle arrays to enhance adaptive immunity.

Rationale: Dendritic cells (DCs) play a crucial role in adaptive immune responses; however, ex vivo differentiation strategies face operational complexities and reduced cellular viability. In situ reprogramming of resident cells into antigen-presenting cells represents a promising alternative approach for enhancing local immune responses. Methods: We initially introduce the novel concept of pseudo-DCs, in situ transforming intradermal fibroblasts into DC-like cells using an engineered exosome-loaded microneedle (MN) array. Specifically, engineered nano-protoplasts expressing interferon-beta (IFN-β) and loaded with varicella-zoster virus glycoprotein E (VZV gE) were used to stimulate DCs and derive immunostimulatory exosomes. These exosomes were integrated into a microarray-based delivery system for intradermal application. Results: The engineered exosomes (IdE@E) induced resident fibroblasts to upregulate DC surface co-stimulatory markers (CD80/86) and effectively present the model antigen. Transcriptome analysis also revealed significant upregulation of genes associated with immune response and antigen presentation in IdE@E-treated cells. In vivo studies demonstrated that MN array-delivered IdE@E effectively induced the expression of DC and activation markers from fibroblasts in dermis. Furthermore, MN array-delivered IdE@E significantly elevated the population of IFN-γ+CD8+ T cells in both lymph nodes and spleen, indicating enhanced local and systemic immune responses. Conclusions: This novel in situ reprogramming method represents a paradigm shift in precision immunotherapies, leveraging exosome-mediated cellular mimicry to enhance adaptive immunity without complete cellular transformation. This scalable framework holds significant promise for immunotherapy and could revolutionize personalized immunotherapy.