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Manni Luthra-Guptasarma, Maryada Sharma, Shweta Sharma, Anil Tiwari, Vishali Gupta, Amod Gupta; Immunospecific targeting of fibronectin towards an anti-fibrotic therapy, with special emphasis on proliferative vitreoretinopathy (PVR). Invest. Ophthalmol. Vis. Sci. 2013;54(15):6261.
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Proliferative vitreoretinopathy (PVR) involves fibrotic pathology. Fibrosis is characterized by excessive accumulation of scar tissue due to exaggerated deposition of extracellular matrix components (ECM). Contractile membranes formed during PVR contain various cell types, including retinal pigment epithelial cells (RPE). The ECM surrounding RPE cells mainly consists of the protein, fibronectin. Together with the vitreous, fibronectin creates microenvironments in which RPE cells proliferate. Our purpose was to target fibronectin, using phage display-selected and re-engineered single chain variable fragment (scFv) antibodies, to prevent ECM deposition.
Phage display-based human scFv antibody library screening was done against the N-terminal 30 kDa region of fibronectin to identify scFv antibodies capable of binding to this region. One chosen antibody (Fn52) was engineered to introduce an N-terminal RGDS tag, to create the Fn52RGDS antibody. Both antibodies were used in cultures of D407 RPE cells grown in the presence of patient-derived vitreous and sub-retinal fluid. A variety of cell-based assays were used to examine the effect of the antibodies on cell viability, proliferation, adhesion, migration, collagen gel contraction, and fibronectin deposition levels.
We have successfully developed fully human, fibronectin-specific single chain variable fragment antibodies that act in one, or both, of the following ways: i) they bind to cryptic sites in fibronectin, preventing its self polymerization/fibrillogenesis, and ii) they interact with cell surface receptors (integrins) through an ‘RGD’ sequence tag, blocking downstream signaling. Fn52RGDS effectively inhibits hallmark features of fibrosis, such as proliferation, migration, adhesion, fibronectin polymerization, matrix metalloprotease (MMP) expression, as well as reduced collagen gel contraction (a model of fibrotic tissue remodeling).
Fn52RGDS is a novel anti-fibrotic candidate therapeutic agent that may be used to reduce fibrosis in PVR and also in pathological situations involving tumors and thrombosis.
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