Identification and Characterization of a Novel Nanobody Against Human CTGF to Reveal Its Antifibrotic Effect in an in vitro Model of Liver Fibrosis

Abstract

Rong Liu,1,* Min Zhu,2,* Jiaojiao Chen,3 Junwei Gai,2 Jing Huang,2 Yingqun Zhou,1 Yakun Wan,2 Chuantao Tu4 1Department of Gastroenterology, Shanghai Tenth People’s Hospital, Tongji University School of Medicine, Shanghai, 200072, People’s Republic of China; 2Shanghai Novamab Biopharmaceuticals Co., Ltd, Shanghai, 201318, People’s Republic of China; 3Department of Gastroenterology, Shanghai Fourth People’s Hospital, Tongji University School of Medicine, Shanghai, 200434, People’s Republic of China; 4Department of Gastroenterology, Shanghai Public Health Clinical Center, Fudan University, Shanghai, 201508, People’s Republic of China*These authors contributed equally to this workCorrespondence: Chuantao Tu; Yakun Wan, Email [email protected]; [email protected]: No agents are currently available for the treatment or reversal of liver fibrosis. Novel antifibrotic therapies for chronic liver diseases are thus urgently needed. Connective tissue growth factor (CTGF) has been shown to contributes profoundly to liver fibrogenesis, which makes CTGF as a promising target for developing antifibrotic agents.Methods: In this study, we identified a novel nanobody (Nb) against human CTGF (anti-CTGF Nb) by phage display using an immunized camel, which showed high affinity and specificity in vitro. LX-2 cells, the immortalized human hepatic stellate cells, were induced by transforming growth factor beta1 (TGFβ 1) as an in vitro model of liver fibrosis to verify the antifibrotic activity of the anti-CTGF Nb.Results: Our data demonstrated that anti-CTGF Nb effectively alleviated TGFβ 1-induced LX-2 cell proliferation, activation, and migration, and promoted the apoptosis of activated LX-2 cells in response to TGFβ 1. Moreover, the anti-CTGF Nb remarkably reduced the levels of TGFβ 1, Smad2, and Smad3 expression in LX-2 stellate cells stimulated by TGFβ 1.Conclusion: Taken together, we successfully identified a novel Nb against human CTGF, which exhibited antifibrotic effects in vitro by regulating the biological functions of human stellate cells LX-2.Graphical Abstract: Keywords: nanobody, connective tissue growth factor, liver fibrosis, hepatic stellate cells

Keywords

• nanobody
• connective tissue growth factor
• liver fibrosis
• hepatic stellate cells.