Targeted Delivery of CD34 Aptamer-Coupled Tocilizumab Microspheres for Effective Treatment of Thyroid-Associated Ophthalmopathy

Thyroid-associated ophthalmopathy (TAO) is a debilitating autoimmune disorder linked to Graves' disease (GD) that is characterized by inflammation and remodeling of orbital tissues. This study focuses on the use of poly(lactic-co-glycolic) acid (PLGA)-based microspheres (MS) coupled with CD34-specific aptamers to enhance the targeted delivery of Tocilizumab (Toc), an IL-6 receptor monoclonal antibody, to CD34+ orbital fibroblasts, which is a critical cell type implicated in TAO.

The flow cytometry and aptamer-mediated pull-down assays were applied to detect the affinity of CD34 aptamers (Apts) for CD34+ orbital fibroblasts. The CD34 Apt-modified Toc-loaded MS (Toc-MS-CD34 Apt) were characterized by scanning electron microscopy (SEM), transmission electron microscopy (TEM), Fourier transform infrared spectroscopy (FT-IR), and agarose gel electrophoresis. The CCK-8 assay kit was used to examine cell viability. The EdU assay was used to assess cell proliferation. The scratch wound healing assay was applied to detect cell migration.

The affinity of CD34 aptamers for CD34+ orbital fibroblasts was confirmed, demonstrating high specificity and binding strength. The Toc-MS-CD34 Apt exhibited size uniformity and successful aptamer conjugation. In vitro, studies showed that Toc-MS-CD34 Apt effectively inhibited the viability, proliferation and cell activation, extracellular matrix (ECM) protein expression, and migration of TGF-β1-stimulated CD34+ orbital fibroblasts. In vivo, a TAO mouse model treated with anti-mouse IL-6R microspheres (anti-mmu-IL-6R-MS) and CD34 aptamer-modified anti-mmu-IL-6R microspheres (anti-mmu-IL-6R-MS-CD34 Apt) demonstrated significant reductions in tissue inflammation, fibrosis, and ECM protein levels, with notable inhibition of the STAT3 signaling pathway.

These findings highlight the potential of CD34 aptamer-coupled Toc microspheres as a targeted therapy for TAO, providing a comprehensive strategy that addresses local manifestations of the disease.