title: Intracellular TGF-beta receptor blockade abrogates Smad-dependent fibroblast activation in vitro and in vivo.
abstract: Fibrosis, the hallmark of scleroderma, is characterized by excessive synthesis of collagen and extracellular matrix proteins and accumulation of myofibroblasts. Transforming growth factor-beta (TGF-beta), a potent inducer of collagen synthesis, cytokine production, and myofibroblast transdifferentiation, is implicated in fibrosis. Profibrotic TGF-beta responses are induced primarily via the type I activin-like receptor kinase 5 (ALK5) TGF-beta receptor coupled to Smad signal transducers. Here, we investigated the effect of blocking ALK5 function with SM305, a novel small-molecule kinase inhibitor, on fibrotic TGF-beta responses. In normal dermal fibroblasts, SM305 abrogated the ligand-induced phosphorylation, nuclear import, and DNA-binding activity of Smad2/3 and Smad4, and inhibited Smad2/3-dependent transcriptional responses. Furthermore, SM305 blocked TGF-beta-induced extracellular matrix gene expression, cytokine production, and myofibroblast transdifferentiation. In unstimulated scleroderma fibroblasts, SM305 caused a variable and modest reduction in type I collagen levels, and failed to abrogate constitutive nuclear accumulation of Smad2/3, or alter the proportion of smooth muscle actin stress fiber-positive fibroblasts. In vivo, SM305 prevented TGF-beta-induced Smad2/3 phosphorylation type I collagen (COL1)A2 promoter activation in dermal fibroblasts. Taken together, these results indicate that SM305 inhibits intracellular TGF-beta signaling through selective interference with ALK5-mediated Smad activation, resulting in marked suppression of profibrotic responses induced by TGF-beta in vivo and in vitro.
authors.affiliation: Division of Rheumatology, Northwestern University Feinberg School of Medicine, Chicago, Illinois 60611, USA.
authors.forename: Wataru
authors.initials: W
authors.lastname: Ishida
authors.forename: Yasuji
authors.initials: Y
authors.lastname: Mori
authors.forename: Gabriella
authors.initials: G
authors.lastname: Lakos
authors.forename: Lihong
authors.initials: L
authors.lastname: Sun
authors.forename: Feng
authors.initials: F
authors.lastname: Shan
authors.forename: Scott
authors.initials: S
authors.lastname: Bowes
authors.forename: Serene
authors.initials: S
authors.lastname: Josiah
authors.forename: Wen-Cherng
authors.initials: WC
authors.lastname: Lee
authors.forename: Juswinder
authors.initials: J
authors.lastname: Singh
authors.forename: Leona E
authors.initials: LE
authors.lastname: Ling
authors.forename: John
authors.initials: J
authors.lastname: Varga
mesh_terms: D000818:Animals; D001706:Biopsy; D002455:Cell Division / Q000187:drug effects; D002478:Cells, Cultured; D020405:Dermis / Q000166:cytology; D005347:Fibroblasts / Q000166:cytology / Q000378:metabolism; D006801:Humans; D066298:In Vitro Techniques; D051379:Mice; D008822:Mice, Transgenic; D047428:Protein Kinase Inhibitors / Q000494:pharmacology*; D011720:Pyrazoles / Q000494:pharmacology*; D011804:Quinolines / Q000494:pharmacology*; D018125:Receptors, Transforming Growth Factor beta / Q000037:antagonists & inhibitors* / Q000378:metabolism; D012595:Scleroderma, Systemic / Q000188:drug therapy* / Q000378:metabolism* / Q000473:pathology; D051785:Smad Proteins / Q000378:metabolism*; D051899:Smad2 Protein / Q000378:metabolism; D051900:Smad3 Protein / Q000378:metabolism; D051901:Smad4 Protein / Q000378:metabolism
chemical_list: C506615:HTS 466284; D047428:Protein Kinase Inhibitors; D011720:Pyrazoles; D011804:Quinolines; D018125:Receptors, Transforming Growth Factor beta; C497782:SMAD2 protein, human; C497790:SMAD3 protein, human; C497796:SMAD4 protein, human; D051785:Smad Proteins; D051899:Smad2 Protein; D051900:Smad3 Protein; D051901:Smad4 Protein