title: Endoglin requirement for BMP9 signaling in endothelial cells reveals new mechanism of action for selective anti-endoglin antibodies.
abstract: Endoglin (ENG), a co-receptor for several TGFβ-family cytokines, is expressed in dividing endothelial cells alongside ALK1, the ACVRL1 gene product. ENG and ACVRL1 are both required for angiogenesis and mutations in either gene are associated with Hereditary Hemorrhagic Telangectasia, a rare genetic vascular disorder. ENG and ALK1 function in the same genetic pathway but the relative contribution of TGFβ and BMP9 to SMAD1/5/8 activation and the requirement of ENG as a co-mediator of SMAD phosphorylation in endothelial cells remain debated. Here, we show that BMP9 and TGFβ1 induce distinct SMAD phosphorylation responses in primary human endothelial cells and that, unlike BMP9, TGFβ only induces SMAD1/5/8 phosphorylation in a subset of immortalized mouse endothelial cell lines, but not in primary human endothelial cells. We also demonstrate, using siRNA depletion of ENG and novel anti-ENG antibodies, that ENG is required for BMP9/pSMAD1 signaling in all human and mouse endothelial cells tested. Finally, anti-ENG antibodies that interfere with BMP9/pSMAD1 signaling, but not with TGFβ1/pSMAD3 signaling, also decrease in vitro HUVEC endothelial tube formation and inhibit BMP9 binding to recombinant ENG in vitro. Our data demonstrate that BMP9 signaling inhibition is a key and previously unreported mechanism of action of TRC105, an anti-angiogenic anti-Endoglin antibody currently evaluated in clinical trials.
authors.affiliation: Oncology Research, Amgen, South San Francisco, CA, USA. oliviern@amgen.com
authors.forename: Olivier
authors.initials: O
authors.lastname: Nolan-Stevaux
authors.forename: Wendy
authors.initials: W
authors.lastname: Zhong
authors.forename: Stacey
authors.initials: S
authors.lastname: Culp
authors.forename: Kathy
authors.initials: K
authors.lastname: Shaffer
authors.forename: Joseph
authors.initials: J
authors.lastname: Hoover
authors.forename: Dineli
authors.initials: D
authors.lastname: Wickramasinghe
authors.forename: Astrid
authors.initials: A
authors.lastname: Ruefli-Brasse
mesh_terms: D000818:Animals; D000888:Antibodies, Anti-Idiotypic / Q000494:pharmacology*; D015703:Antigens, CD / Q000235:genetics / Q000378:metabolism*; D017209:Apoptosis; D015153:Blotting, Western; D002465:Cell Movement; D049109:Cell Proliferation; D002478:Cells, Cultured; D000071063:Endoglin; D004730:Endothelium, Vascular / Q000166:cytology / Q000378:metabolism*; D005434:Flow Cytometry; D055427:Growth Differentiation Factor 2; D055412:Growth Differentiation Factors / Q000235:genetics / Q000378:metabolism*; D006801:Humans; D051379:Mice; D010766:Phosphorylation; D011485:Protein Binding; D012333:RNA, Messenger / Q000235:genetics; D034741:RNA, Small Interfering / Q000235:genetics; D051381:Rats; D060888:Real-Time Polymerase Chain Reaction; D011956:Receptors, Cell Surface / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism*; D020133:Reverse Transcriptase Polymerase Chain Reaction; D051785:Smad Proteins / Q000235:genetics / Q000378:metabolism*; D016212:Transforming Growth Factor beta / Q000235:genetics / Q000378:metabolism*
chemical_list: D000888:Antibodies, Anti-Idiotypic; D015703:Antigens, CD; C064206:ENG protein, human; D000071063:Endoglin; C525887:GDF2 protein, human; D055427:Growth Differentiation Factor 2; D055412:Growth Differentiation Factors; D012333:RNA, Messenger; D034741:RNA, Small Interfering; D011956:Receptors, Cell Surface; D051785:Smad Proteins; D016212:Transforming Growth Factor beta