Construction of an Anti-IL-1β scfv and TNFRI Fusion Protein and Its Therapeutic Effect on RA Mice Model

Abstract

IL-1β and TNF-α play key roles in the inflammatory response. Their abnormal expression may cause the occurrence of various diseases, such as RA. Recently, medicines of target TNF-α and IL-1β have become popular in the clinical practice. Although these biological agents can get mostly good results, they are not effective in all patients. The reason for this result may be that these biological agents could not fully inhibit a variety of inflammatory cytokines in the inflammatory response. In the present study, a fusion protein gene which encoded human interleukin-1β scfv and soluble TNF receptor I (sTNFRI) was cloned. A number of in vitro assays demonstrated that anti-IL-1β scfv/TNFRI simultaneously bound to both targets. The bioactivity assay showed that the fusion protein could inhibit both the cytotoxicity of hTNF-α on L929 cells and hIL-1β-induced proliferation of L929 cells, indicating that the fusion protein has the ability to neutralize both hTNF-α and hIL-1β. In this study, we established the chicken type II collagen-induced rheumatoid arthritis model in Kunming mice, and evaluated the pharmacological effect of the fusion protein in vivo. Model mice were randomly divided into 8 groups (n=8): CIA model control group, DEX treatment group (1 mg/kg), intraperitoneal treatment group (highdose: 5 mg/kg; medium-dose: 2 mg/kg; low-dose: 0.8 mg/kg), subcutaneous treatment group (high-dose: 5 mg/kg; medium- dose: 2 mg/kg; low-dose: 0.8 mg/kg), and healthy mice as control. The control group received the same volume of saline. The mice were administrated once every 2 days. Arthritis index, anti-CII antibody titers, cytokine levels, histopathological changes were examined. The results showed that anti-IL-1β scfv/TNFRI fusion protein could reduce the degree of joint swelling, inflammatory cell infiltration, synovial cell proliferation and the level of CII antibody in the sera. The Real-time PCR analysis showed that anti-IL-1β scfv/TNFRI had the ability to reduce the expression of IL-1β, TNF-α, IL-17A, MMP-3, IL-6 and improve the expression of IL-10 in a dose-dependent manner, suggesting that the fusion protein is the mediator for IL-1β and TNF-α involved in the RA process. Compared with DEX positive medicine control, anti-IL-1β scfv/TNFRI appeared more beneficial in treatment of CIA mice. The therapeutic effect of the anti-IL-1β scfv/TNFRI at 5mg/kg was significantly better than that of DEX treatment. So the anti-IL-1β scfv/TNFRI can become a candidate for treatment of RA.

Hierarchical, imbalanced pro-inflammatory cytokine networks govern the pathogenesis of chronic arthropathies

“…Indeed, simultaneous blocking of TNFa and IL-17 revealed a synergistic effect in reducing joint inflammation and protection from SB and cartilage damage in a mouse collageninduced arthritis (CIA) model 40,42 . Similar immune status rebalancing and therapeutic effects were reported upon targeting the other pairs of fragile nodes, such as IL-1b/TNFR1 43 , TNFa/IL-20 44 , or IL-6/21 45 . Significant associations between the HIPICN rebalancing and the beneficial clinical effect upon the combinatorial targeting of two fragile nodes, observed in the aforementioned studies, strongly suggest their causative role in RA pathogenesis.…”

Anti-arthritic activity of ethanol extract of Claoxylon indicum on Freund’s complete adjuvant-induced arthritis in mice

“…IL-1β and TNF-α are reported to play critical roles in the pathogenesis of RA [21, 22]. Therefore, we investigated whether the therapeutic effects of CIE treatment were associated with changes in the frequencies of cells producing IL-1β or TNF-α in the arthritic joints by immunohistochemical staining.…”

Case report: Recombinant human type II tumour necrosis factor receptor-antibody fusion protein induced occult hepatitis B virus reactivation leading to liver failure

“…Research has shown that rh TNFR:Fc has biological effects on several animal models of inflammation, including collagen-induced arthritis in mice. 27 The excessive immune cascade mediated by TNF-α may lead to hepatocyte apoptosis and damage, leading to acute liver failure. 28 The exact mechanisms of rh TNFR:Fc and HBVr are unknown; therefore, care should be taken when rh TNFR:Fc is administered to patients who have been confirmed as HBV carriers.…”