Fluoropolymer‐Mediated Delivery of a Dual TSHR/IGF1R‐Targeting CRISPR‐Cas9 System for Localized Therapy in Thyroid‐Associated Ophthalmopathy

Thyroid-associated ophthalmopathy (TAO), a vision-threatening and disfiguring autoimmune orbital disorder, remains a therapeutic challenge due to the lack of therapies with orbital specificity, sustained efficacy, and minimal side effects. Herein, we present G4F7-CRISPR, a fluoropolymer-based CRISPR-Cas9 delivery platform engineered for localized and efficient disruption of thyroid-stimulating hormone receptor (TSHR) and insulin-like growth factor 1 receptor (IGF1R), two key mediators of TAO pathogenesis. G4F7-CRISPR achieved high insertion/deletion frequencies in primary orbital fibroblasts (Tshr: 37.2%; Igf1r: 42.8%) and mature adipocytes (Tshr: 22.4%; Igf1r: 24.3%), and maintained robust editing efficiency in orbital adipose tissue of TAO mouse models (Tshr: 30.7%; Igf1r: 32.4%). In both TAO mouse models and 3D human orbital organoids, dual-gene editing of Tshr and Igf1r via G4F7-CRISPR significantly suppressed orbital adipogenesis, inflammation, and fibrosis, demonstrating superior therapeutic efficacy over either single-gene approaches. Comprehensive off-target analyses in both TAO mouse models and orbital organoids revealed minimal off-target activity. Furthermore, G4F7-CRISPR exhibited excellent short- and long-term ocular and systemic safety in TAO mouse models. Notably, it outperformed teprotumumab-the FDA-approved therapy for TAO-in both therapeutic efficacy and safety, highlighting its potential clinical advantages. Collectively, these findings highlight the translational promise of G4F7-CRISPR as a safe, precise, and clinically viable gene therapy for TAO.