E1K, a disease-modifying drug candidate for knee osteoarthritis, alleviates pain and regenerates cartilage simultaneously by inhibiting TGF-β1-mediated SMAD1/5/9 signaling in osteoarthritis models.

Abstract

OBJECTIVE: Transforming growth factor-β1 (TGF-β1) signaling is critical in joint homeostasis and osteoarthritis (OA) pathogenesis. Engedi 1000 (E1K), a novel short peptide, is a selective modulator of TGF-β1-induced SMAD1/5/9 signaling. We evaluated the analgesic and structure-modifying effects of E1K in OA models. DESIGN: The binding of E1K and TGF-β1 were investigated using molecular docking and surface plasmon resonance (SPR). The molecular effects of E1K were examined in human chondrocytes and bovine fibroblast-like synoviocytes (FLS). Analgesia was assessed in a monosodium iodoacetate (MIA)-induced OA rat model using the von Frey test. Structure modification were evaluated in an anterior cruciate ligament transection (ACLT)-induced OA rabbit model by OARSI scoring and immunohistochemistry (type II collagen, MMP13, ALK1, and ALK5). RESULTS: Molecular docking predicted, and SPR confirmed E1K binding to TGF-β1. In human chondrocytes, E1K selectively inhibited TGF-β1-induced SMAD1/5/9 signaling while preserving SMAD2/3 phosphorylation. It downregulated catabolic markers (MMP13, COL10A1), while COL2A1 remained unchanged. In bovine FLS, E1K significantly inhibited TGF-β1-induced NGF expression. In the MIA model, E1K significantly reduced mechanical allodynia at highest dose. In the ACLT model, E1K improved the cartilage integrity, reduced the OARSI score, and decreased the ALK1/ALK5 expression ratio. CONCLUSIONS: E1K promoted TGF-β1 signaling toward an anabolic profile by inhibiting catabolic signaling and regenerating cartilage, suggesting its potential as a disease-modifying osteoarthritis drug candidate with dual efficacy in pain relief and structural protection.