Background/Purpose: Progranulin (PGRN) is a multifunctional growth factor. Recently, we reported that PGRN and its derived protein Atsttrin-α (referred to as “Atsttrin” in our previous publication) directly bound to TNF receptors (TNFR), blocked the binding of TNF to TNFR and inhibited TNF activity (Tang, W., et al, Science, 2011 Apr 22;332 (6028):478-484). It is well established that TNF family ligands bind to receptors in a heterohexameric 3:3 complex. Atsttrin-α was comprised of the three functional fragments of PGRN, thus we hypothesized that the three fragments of Atsttrin-α act independently for interacting with TNFR. If so, change of order of the three fragments would not affect binding activity to TNFRs. In this study, we created a novel engineered protein, Atsttrin-β which comprised the same fragments as Atsttrin-α but in a different order. The purpose of this project is to determine whether the novel protein Atsttrin-β is able to block the binding of TNF-α to TNFR, and has therapeutic effect in inflammatory arthritis, as does Atsttrin-α.

Methods: Yeast-two hybrid assay was used to compare the binding to TNFR between Atsttrin-α and Atsttrin-β; Solid-phase binding was performed to determine the Atsttrin-β inhibition of TNF-α binding to TNFRs; Collagen-induced arthritis(CIA) in DBA-IJ mice and TNF-α transgenic (hTNF-tg) mice that spontaneous inflammatory arthritis. Mice were divided into various groups and injected intraperitoneally with PBS, etanercept, Atsttrin-α and Atsttrin-β. Clinical assessment for arthritis scores, micro CT for bone erosion and histological analysis of joint sections were performed.

Results: Yeast-two hybrid assay showed that Atsttrin-β bound to TNFR1 and TNFR2, as did Atsttrin-α; Solid-phase binding assay revealed that Atsttrin-β inhibited TNF-α from binding to TNFRs in a dose-dependent manner. Clinical scores, Micro CT and histological analysis of joints from CIA model and hTNF-tg mice demonstrated that Atsttrin-β attenuated synovial proliferation, infiltration of inflammatory cells, cartilage destruction and bone erosion compared with vehicle-treated mice, and these effects were shown in dose-dependent manners.

Conclusion:  These findings suggest the three functional fragments of Atsttrin-α worked independently in binding to TNFRs and blocking TNF-α activity, and the novel engineered protein Atsttrin-β has anti-inflammatory effect and may represent a promising alternative in treatment of rheumatoid arthritis.

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ACR Meeting Abstracts - https://acrabstracts.org/abstract/atsttrin-a-an-engineered-protein-derived-from-progranulin-growth-factor-binds-to-tnf-receptors-and-exhibits-potent-anti-inflammatory-activity-in-mice/