Nanodiamond-Mediated Targeted Delivery of Nanobodies and Immunostimulatory RNA for Breast Cancer Therapy.

Triple-negative breast cancer (TNBC) is an aggressive subtype lacking defined molecular targets and characterized by high rates of recurrence and metastasis. Aberrant activation of the epidermal growth factor receptor (EGFR) contributes to tumor progression and immune evasion in TNBC. Although EGFR inhibitors can temporarily suppress tumor growth, compensatory signaling and therapeutic resistance limit their effectiveness. Therapeutic strategies that modulate multiple pathways while enhancing antitumor immunity are needed, and selective nanoparticle-based delivery offers a means to improve potency while reducing nonspecific toxicity. We developed ND-dsRNA-VHH, a biocompatible carbon-based nanomaterial platform that codelivers EGFR-specific nanobodies (VHHs) and immunostimulatory double-stranded RNA, polyinosinic-polycytidylic acid (poly(I:C)). Optimized ND surface chemistry supported efficient dsRNA payload and stable VHH conjugation, yielding nanoparticles with EGFR-binding specificity and serum stability. Subsequent studies demonstrated that ND-dsRNA-VHH induced apoptosis, oxidative stress, and immunogenic cell death, leading to dendritic cell activation. Additional assessments indicated that treatment with ND-dsRNA-VHH reduced tumor growth, extended survival, increased T-cell infiltration, and shifted the tumor microenvironment toward a more proinflammatory, immunologically active state. The modular nature of this platform supports ligand exchange for broader applicability across EGFR-driven malignancies such as glioblastoma, underscoring its potential to enhance immunotherapy through combined ICD induction and immune priming.