Abstract
IntroductionHER2-positive breast cancer, accounting for 15%-20% of cases, remains challenging due to therapy resistance and immunosuppressive "cold tumor" microenvironments. Current strategies combining immunotherapy with chemotherapy or radiotherapy often face toxicity limitations. To address this, we developed HER2-targeted nanoliposomes co-delivering viral peptides and the STING agonist diABZI, aiming to convert cold tumors into immunologically active "hot tumors" by enhancing antigen spreading and immune recognition.MethodsViral peptides with high Human Leukocyte Antigen-A2 affinity were selected using NetMHCpan-4.1/4.0 and incorporated into nanoliposomes via thin-film dispersion. Trastuzumab F(ab')(2) fragments were conjugated for HER2-specific targeting. Nanoliposomes were characterized for size, stability, encapsulation efficiency (HPLC), and in vitro release. Immune efficacy was assessed via ELISPOT, flow cytometry (CD3+/CD8+/NK cells), and TCR β sequencing in HER2(+) SK-BR-3 and HER2(-) MCF-7 cells. Cytotoxicity and cellular uptake were evaluated using CCK-8 assays and fluorescence imaging.ResultsThe nanoliposomes exhibited uniform size (∼70 nm), stability (5% size variation over 25 days), and high encapsulation efficiency (75.5% for peptides). Targeted delivery to SK-BR-3 cells peaked at 60 µL (P < .05), with sustained release of peptides (52% at 48 h) and diABZI (46.2%). In HER2(+) cells, nanoliposomes synergistically enhanced IFN-γ (2.5-fold, P < .01) and granzyme B (3-fold, P < .05) secretion, overcoming antagonism seen with free agents. Flow cytometry revealed dominant CD8(+) T-cell infiltration (50.9% vs 0.67% in controls) and expanded NK/NKT populations. TCR β sequencing showed increased clonotype diversity (60,915 vs 57 574 clones) and reduced clonal dominance, indicating broadened antigen recognition.ConclusionOur HER2-targeted nanoliposomes effectively reprogrammed cold tumors by dual activation of innate (STING pathway) and adaptive (viral peptide-driven TCR diversity) immunity. The platform demonstrated robust targeting, safety, and immune activation, offering a promising strategy to overcome immunotherapy resistance. Future studies will validate in vivo efficacy and explore adaptations for other cold tumors via alternative targeting ligands.