Anti-EMP2 diabody blocks epithelial membrane protein 2 (EMP2) and FAK mediated collagen gel contraction in ARPE-19 cells

Abstract

Epithelial membrane protein 2 (EMP2) regulates collagen gel contraction by the retinal pigment epithelium cell line ARPE-19 by modulating FAK activation. Collagen gel contraction is one in vitro model for an aberrant wound healing response, proliferative vitreoretinopathy (PVR), which occurs as a complication of severe ocular trauma. The purpose of this study is to investigate whether EMP2 specific recombinant diabody decreases activation of FAK and collagen gel contraction in ARPE-19. Anti-EMP2 diabody was recombinantly constructed from a human phage library-derived clone selected for reactivity against an extracellular domain of human EMP2. ARPE-19 cells were exposed to an anti-EMP2 or control diabody, and toxicity, adhesion, and migration were assessed respectively through toluidine blue exclusion, binding to collagen type 1, and a migration assay. Collagen gel contraction was assessed using an in vitro assay. FAK activation was evaluated using Western blot. Exposure to anti-EMP2 diabody resulted in a 75% reduction in EMP2 protein levels at 4 hours. No significant toxicity was observed with anti-EMP2 diabody at levels that maximally reduced EMP2. Anti-EMP2 diabody, but not control diabody significantly reduced collagen gel contraction (p<0.001), and was not secondary to changes in adhesion or migration. Concordantly, anti-EMP2 diabody as compared to a control diabody reduced collagen stimulated FAK activation (p=0.01). Anti-EMP2 diabody decreases EMP2 protein levels, FAK activation, and collagen gel contraction by ARPE-19 cells without an adverse effect on cell survival. Modulation of EMP2 using anti-EMP2 diabody could be a new approach for targeting EMP2 and pathologic consequences associated with EMP2.

Epithelial Membrane Protein 2 Is a Prognostic Indictor for Patients with Urothelial Carcinoma of the Upper Urinary Tract

“…42 Expression of EMP2 was shown to drive the cellular response toward a FAK-dependent pathway. 43 In addition, activation of the Src and FAK family members and downstream signaling proteins are associated with a good prognosis in UC of the bladder. 44 As a result, we speculate that reciprocal regulation of EMP2 and caveolin-1 levels in cancer cells may reshape the cellular surface-receptor repertoire and its attendant responsiveness to the local environment.…”

Proliferative vitreoretinopathy: A new concept of disease pathogenesis and practical consequences

“…Thus it may be possible to use antagonists to endogenous integrin mediators or reagents designed to inhibit integrin activity to prevent or treat PVR. Recent preclinical studies have shown that decreasing the expression of epithelial membrane protein 2, a RPE cell integrin, or inhibition of it by directed antibodies reduces the onset of PVR formation (Morales et al, 2012;Telander et al, 2011). The applicability of these in vitro results to human disease is not yet known.…”

High expression of EMP1 predicts a poor prognosis and correlates with immune infiltrates in bladder urothelial carcinoma

“…A number of previous studies have stated that members of the EMP family affect the integrin heterodimer repertoire on the plasma membrane, and modulation of the expression or localization of EMP proteins may alter the surface repertoire of molecules ( 54 – 56 ). The surface molecular repertoire, including major histocompatibility complex 1 proteins, integrins and other immunoglobulin superfamily members such as CD54 and glycosylphosphatidyl-inositol-linked proteins may be altered as the expression of EMP2 changes ( 57 ).…”