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Recently, targeting inflammatory cytokines in the pathogenic process of rheumatoid arthritis is now performed as a feasible biological method in therapy. However, treatments against single cytokine are often difficult to achieve the ideal therapeutic effect. Multi-target drugs permit more effective suppression of inflammation. In this study, we constructed an IgG-like bispecific antibody targeting IL-1β and IL-17A and expressed it in mammalian cells. The therapeutic efficacy was studied in CIA (collagen-induced arthritis) mice, which were administrated with either FL-BsAb1/17 (IgG-like bispecific antibody targeting IL-1β and IL-17A) or monovalent IL-1β Mab or IL-17A Mab (anti-IL-1β/IL-17A monoclonal antibody). We noticed that FL-BsAb1/17 had better effect on alleviating clinical symptom by significantly lowering arthritis score and relieving histological lesion on aspect of less damnification in synovial hyperplasia and cartilage destruction than monovalent Mab alone. In addition, FL-BsAb1/17 was more potent in inhibiting IL-1β, IL-17A, IL-6, TNF-α and anti-CCP antibody in the serum and in down-regulating the expression of IL-1β, IL-17A, IL-6, TNF-α, MMP-3 and RANKL in the spleen, compared to monovalent Mab alone. Further, the anti-inflammatory effect of FL-BsAb1/17 was demonstrated by significantly depressing Th17 cells expansion through decreasing phosphorylated STAT3 in the spleen of the CIA mice. FLS (fibroblast-like synoviocytes) from RA patients were used to examine the therapeutic efficacy of FL-BsAb1/17 in human pathological tissue. FL-BsAb1/17 could significantly decrease the production of IL-6 induced by IL-1β and/or IL-17A in FLS. In conclusion, FL-BsAb1/17 has the possibility to be a promising therapeutic agent for RA. [ABSTRACT FROM AUTHOR]

Rheumatoid arthritis (RA) is a chronic, systemic and autoimmune disease with overexpression inflammation cytokines. The biological therapy targeting these inflammatory cytokines has been applied for the clinic. Drugs aimed at a single target are ineffective in some patients with RA. However, double-target and multi-target drugs have huge advantages in therapy. Interleukin-1β (IL-1β) and Interleukin-17A (IL-17A) are the keys to inflammatory factors in RA. The bispecific antibody(BsAb)against both human IL-1β and human IL-17A was formed and expressed in E. coli. The binding specificity and efficiency of the BsAb was tested by enzyme-linked immunosorbent assay, Western blotting and several cells experiments including THP-1, 3T3-L1 and L929 in vitro. Different doses of BsAb (5, 2, 0.8 mg/kg) were compared in collagen-induced arthritis (CIA) mice, with Adalimumab and Dexamethasone as the positive control. The effects on mice were determined by the degree of arthritis severity, cytokines levels, the level of IgG against CII and rheumatoid factor level in serum, the transcription levels of relative cytokines in the spleen, and histological damage. Furthermore, the activation of NF-κB was analyzed by Western blotting. In conclusion, BsAb can bind with IL-1β and IL-17A to alleviate the severity of arthritis, to decrease the levels of inflammation cytokines in serum, to down-regulate the expression of IL-1β, IL-2, IL-6, IL-17A, TNF-α, IFN-γ, and MMP-3, to up-regulate the expression of IL-10, to relieve histological damage and to inhibit bone destruction. BsAb inhibited nuclear translocation of the p65 subunit and cytoplasm IκB-α degradation by blocking IL-1β and IL-17A, the upstream of NF-κB pathway. High doses of BsAb had a more beneficial effect on CIA mice than Adalimumab and Dexamethasone. Thus, these results indicate that BsAb can be regarded as an ideal candidate for RA therapy. [ABSTRACT FROM AUTHOR]

• Targeting IL-1 and IL-17 suppressed the immune cascade to reduce systemic sclerosis. • Systemic sclerosis oxidative stress level was regulated by a cytokine network. • Targeting IL-1 and IL-17 could inhibit fibrosis through the TGF-β/Smad2/3 pathway. Systemic sclerosis (SSc) is induced by variety of factors and eventually causes multiple organ damage. In recent years, biological agents targeting cytokines and cell surface molecules have gradually come to market. Here, the anti-inflammatory and antifibrotic effects of a novel bispecific antibody (FL-BsAb1/17) targeting interleukin-17A (IL-17A) and interleukin-1β (IL-1β) were detected. Bleomycin (BLM) was subcutaneously injected for 21 consecutive days to establish the SSc mouse model, and mice were subsequently treated with etanercept or different doses (1, 5, 10 mg/kg) of FL-BsAb1/17. The results showed that FL-BsAb1/17 treatment (10 mg/kg, 5 mg/kg) significantly attenuated BLM-induced SSc-like inflammation by inhibiting the expression of inflammatory factors (IL-17A, IL-1β, IL-8, IL-22, IL-23, IL-6) and fibrosis, with specific outcomes of dermis thickening and lung fibrosis, by inhibiting the expression of fibrotic factors (TGF-β, α-sma, Col-1, Col-3) in the serum, skin and lungs. In addition, FL-BsAb1/17 (10 mg/kg, 5 mg/kg) downregulated protein levels of TGF-β and phosphorylated Smad2/3 in the skin and lungs and reduced collagen 1 protein levels. This indicated that FL-BsAb1/17 can inhibit the development of fibrosis by inhibiting the TGF-β/Smad2/3 signaling pathway. FL-BsAb1/17 (10 mg/kg, 5 mg/kg) could also effectively reduce the content of MDA, increase the activity of SOD and CAT, and improve the total antioxidant capacity (T-AOC). In conclusion, FL-BsAb1/17 alleviated BLM-induced SSc by downregulating inflammatory cascades, relieving oxidative stress and inhibiting TGF-β/Smad2/3 signaling. These data suggest that FL-BsAb1/17 has potential as a novel therapeutic candidate for SSc. [ABSTRACT FROM AUTHOR]