Tailored multiepitope PEI-PLGA nanovaccines functionalized with poly(I:C) promote neoantigen-specific T-cell responses in melanoma patients

Melanoma remains a major global health burden, with more than 510,000 new cases and 96,000 deaths projected worldwide by 2040. Over the past decade, neoantigen-based cancer vaccines have emerged as a promising immunotherapeutic strategy for the treatment of melanoma. Nevertheless, beyond the challenges associated with antigen delivery, an important unanswered question is whether combining multiple neoantigens within a single vaccine construct maintains neoantigen-specific T-cell recognition of individual neoepitopes. To address this issue, we developed a nanovaccine platform based on polyethyleneimine–poly(lactic-co-glycolic acid) (PEI-PLGA) nanoparticles (NPs) encapsulating patient-specific multiepitope neoantigen peptides and coated with poly(I:C), enabling the delivery of multiepitope neoantigen peptides and adjuvant within a single formulation. Following physicochemical characterization, their immunogenic potential was assessed ex vivo using peripheral blood mononuclear cells (PBMCs) isolated from patients with melanoma. Activation-induced marker (AIM) assays and IL-2 cytokine secretion were used to assess the effects of delivering multiepitope neoantigens and poly(I:C), which significantly enhanced T-cell activation in two of the three tested patients. Importantly, nanovaccine stimulation elicited immune responses against the 15mer-epitope melanoma-associated antigen using ELISPOT. Analysis using the individual three individual peptide epitopes identified pre-existing immune recognition of specific regions contained preferential immune recognition of specific regions within the multiepitope. These findings demonstrate immune recognition of the multiepitope antigen delivered by the nanovaccine and identify the peptide region(s) most frequently recognized within this sequence, but do not establish induction or maintenance of peptide-specific responses. Collectively, these findings demonstrate that multiepitope PEI-PLGA nanovaccines allow neoantigen-specific T-cell responses while evaluating baseline T-cell reactivity against individual neoantigens, supporting their development as a personalized immunotherapeutic strategy for melanoma.