RMAD1, a Novel Cell-Penetrating Peptide Derived from ADARB2: Preclinical Insights into Antigen Uptake and T Cell Activation.

The development of effective cancer vaccines remains a major challenge in oncology, largely due to limited antigen delivery and suboptimal T cell priming. Here, we report RMAD1, a novel human-derived cell-penetrating peptide (CPP) originating from the ADARB2 (Adenosine Deaminase RNA Specific B2) protein, identified through an intra-dermal delivery technology (IDDT) platform, and evaluate its potential as a vaccine delivery enhancer. RMAD1 exhibited superior intracellular delivery compared with conventional CPPs and preferential uptake by antigen-presenting cells (APCs), including dendritic cells and macrophages. RMAD1 conjugated vaccines showed enhanced accumulation in draining lymph nodes and facilitated efficient antigen cross-presentation through the MHC class I pathway. In murine E.G7-OVA and TC-1 tumor models, RMAD1 conjugated vaccines induced robust antigen-specific CD8⁺ T cell responses across peripheral blood, lymphoid organs, and tumor tissues. Functional analyses revealed increased IFN-γ and TNF-α production by both CD8⁺ and CD4⁺ T cells, accompanied by a reduction in Foxp3⁺CD25⁺ regulatory T cells. In addition, RMAD1 conjugation promoted epitope spreading and established durable immunological memory, resulting in protection against tumor rechallenge. Therapeutic efficacy was further demonstrated in a TC-1 lung metastasis model, where RMAD1-based vaccination significantly reduced metastatic burden. Importantly, RMAD1-vaccinated mice exhibited therapeutic efficacy comparable to cisplatin treatment, while demonstrating a favorable safety profile. Together, these findings position RMAD1 as a next-generation CPP platform that outperforms existing peptides in enhancing antigen delivery and anti-tumor immunity, offering a promising strategy for advancing cancer vaccine development.