title: Biological activity of a novel nonpeptide antagonist to the interleukin-6 receptor 20S,21-epoxy-resibufogenin-3-formate.
abstract: Interleukin (IL)-6 is a key mediator in the regulation and coordination of the immune response and participates in pathogenesis of cancer cachexia, autoimmune disease, and postmenopausal osteoporosis. In the course of a screening program aimed at IL-6 inhibitor from natural products, we isolated 20S,21-epoxy-resibufogenin-3-formate (ERBF) from bufadienolide and examined the effect of ERBF on activities of various cytokines. ERBF dose dependently suppressed IL-6 activity and caused a parallel rightward shift of dose-response curves to IL-6 at concentrations of 0.03 to 10 ng/ml. Analysis of data yields a pA(2) of 5.12 and a slope of 0.99. Selectivity of ERBF on activity of cytokines was examined using cytokine-dependent cell lines. ERBF did not affect IL-2-dependent growth of CTLL-2 cells, IL-3-dependent growth of Baf3 cells, or tumor necrosis factor (TNF)alpha-induced growth suppression in TNFalpha-sensitive L929 cells. ERBF also did not affect IL-4-stimulated expression of FcepsilonR II receptor (CD23) in U-937 cells, the IL-8-induced chemotaxis of human neutrophils, or nerve growth factor-stimulated neuronal differentiation in PC-12 cells. In contrast, ERBF dose dependently suppressed IL-6-induced neuronal differentiation in PC-12 cells. Furthermore, ERBF suppressed only IL-6-induced osteoclast formation without affecting osteoclast formation induced by IL-11, leukemia inhibitory factor, and 1alpha,25-dihydroxyvitamin D(3). In receptor binding assay, unbound (free) IL-6 was increased in a dose-dependent manner by pretreatment with ERBF on IL-6 receptor (IL-6R), suggesting that ERBF suppresses binding of IL-6 to IL-6R. These results clearly indicate that ERBF is a novel specific small molecule to show IL-6 receptor antagonist activity.
authors.affiliation: The Kitasato Institute, Tokyo, Japan. mhayashi@lisci.kitasato-u.ac.jp
authors.forename: Masahiko
authors.initials: M
authors.lastname: Hayashi
authors.forename: Mun-Chual
authors.initials: MC
authors.lastname: Rho
authors.forename: Akiko
authors.initials: A
authors.lastname: Fukami
authors.forename: Akiko
authors.initials: A
authors.lastname: Enomoto
authors.forename: Shinobu
authors.initials: S
authors.lastname: Nonaka
authors.forename: Yoshino
authors.initials: Y
authors.lastname: Sekiguchi
authors.forename: Tadashi
authors.initials: T
authors.lastname: Yanagisawa
authors.forename: Ayano
authors.initials: A
authors.lastname: Yamashita
authors.forename: Toshihiko
authors.initials: T
authors.lastname: Nogawa
authors.forename: Yoshiaki
authors.initials: Y
authors.lastname: Kamano
authors.forename: Kanki
authors.initials: K
authors.lastname: Komiyama
mesh_terms: D000818:Animals; D001854:Bone Marrow Cells / Q000166:cytology / Q000187:drug effects; D002018:Bufanolides / Q000494:pharmacology*; D002454:Cell Differentiation; D002455:Cell Division / Q000187:drug effects; D002460:Cell Line; D002634:Chemotaxis, Leukocyte / Q000187:drug effects*; D018920:Coculture Techniques; D016207:Cytokines / Q000494:pharmacology; D006801:Humans; D051379:Mice; D015394:Molecular Structure; D009474:Neurons / Q000166:cytology / Q000187:drug effects; D010010:Osteoclasts / Q000166:cytology / Q000187:drug effects; D016716:PC12 Cells; D010673:Pheochromocytoma; D051381:Rats; D019947:Receptors, Interleukin-6 / Q000037:antagonists & inhibitors*; D011994:Recombinant Proteins / Q000037:antagonists & inhibitors; D020298:U937 Cells
chemical_list: C466795:20,21-epoxyresibufogenin-3-formate; D002018:Bufanolides; D016207:Cytokines; D019947:Receptors, Interleukin-6; D011994:Recombinant Proteins