Inhibition of Toll-Like Receptor-Mediated Inflammation In Vitro and In Vivo by a Novel Benzoxaborole

Abstract

Pro-inflammatory cytokines play a critical role in the development of autoimmune and inflammatory diseases. Targeting the cytokine environment has proven efficient for averting inflammation. In this study, we reported that, a benzoxaborole analog, inhibited TLR2-, TLR3-, TLR4-, and TLR5-mediated TNF-a, IL-1b, and IL-6 release from human PBMCs and isolated monocytes with IC 50 values ranging from 18 to 580 nM, and the inhibition was mediated at the transcriptional level. Topical administration of AN3485 significantly reduced PMA-induced contact dermatitis and oxazolone-induced delayed-type hypersensitivity in mice, indicating its capability of penetrating skin and potential topical application in skin inflammation. Oral administration of AN3485 showed dose-dependent suppression of LPSinduced TNF-a and IL-6 production in mice with an ED 90 of 30 mg/kg. Oral AN3485, 35 mg/kg, twice a day, suppressed collagen-induced arthritis in mice over a 20-day period. The potent anti-inflammatory activity in in vitro and in vivo disease models makes AN3485 an attractive therapeutic lead for a variety of cutaneous and systemic inflammatory diseases.

Treatment of Skin Inflammation with Benzoxaborole Phosphodiesterase Inhibitors: Selectivity, Cellular Activity, and Effect on Cytokines Associated with Skin Inflammation and Skin Architecture Changes

“…To induce IL-22 production, PBMCs were stimulated for 24 hours with anti-CD2/CD3/CD28 beads (cell/bead 5 1:1) (Miltenyi Biotec, Bergisch Gladbach, Germany). Human monocytes were isolated as described by Dong et al (2013) with a purity of 85%-90% and were stimulated with 1 mg/ml LPS and 100 ng/ml IFN-g for 24 hours to induce IL-23 production. The cell culture supernatants were collected for cytokine analysis.…”

“…Phospho-CREB was detected by an antibody specific to phosphoserine (S133) (Santa Cruz Biotechnology, Santa Cruz, CA). To address the effect of the benzoxaborole PDE4 inhibitors on extracellular signal-regulated kinase (ERK) phosphorylation, human T cells were isolated as described by Dong et al (2013) with a purity of $ 90%. The T cells were incubated at 37°C for 2 hours and then either not treated or treated with compd3 or dBcAMP for 15 minutes prior to stimulation with human T-activator CD3/CD28 beads (Thermo Fisher Scientific, Grand Island, NY).…”

“…Ten female CD-1 mice (20-30 g) per group were used in this experiment, which was based on a protocol described by Kuchera et al (1993) with a few modifications as described by Dong et al (2013). Three milligrams of compd3 dissolved in acetone/ethanol [1:1 (v/v)] was applied 30 minutes before and 15 minutes after the phorbol myristate acetate (PMA) challenge.…”

Linking Phenotype to Kinase: Identification of a Novel Benzoxaborole Hinge-Binding Motif for Kinase Inhibition and Development of High-Potency Rho Kinase Inhibitors

“…Human peripheral blood mononuclear cells (PBMCs) were stimulated and treated with different compounds to measure the inhibitory effects on cytokine secretion, as described by Akama et al (2013) and Dong et al (2013). In brief, PBMCs were incubated with the following drugs and stimulators: lipopolysaccharide (LPS) for 24 hours to release tumor necrosis factora (TNF-a) and interleukin (IL)-6; phytohemagglutinin (PHA) for 24 hours to release IL-2 and interferon-g (IFN-g); and PHA for 48 hours to release IL-5 and IL-13.…”

“…The pharmacokinetics (PK) of compound 5 in rats was studied using methods similar to those by Dong et al (2013). Adult male Sprague-Dawley rats received 6-[4-…”