title: HSulf-1 inhibits angiogenesis and tumorigenesis in vivo.
abstract: We previously identified HSulf-1 as a down-regulated gene in several tumor types including ovarian, breast, and hepatocellular carcinomas. Loss of HSulf-1, which selectively removes 6-O-sulfate from heparan sulfate, up-regulates heparin-binding growth factor signaling and confers resistance to chemotherapy-induced apoptosis. Here we report that HSulf-1 expression in MDA-MB-468 breast carcinoma clonal lines leads to reduced proliferation in vitro and reduced tumor burden in athymic nude mice in vivo. Additionally, xenografts derived from HSulf-1-expressing stable clones of carcinoma cells showed reduced vessel density, marked necrosis, and apoptosis, indicative of inhibition of angiogenesis. Consistent with this observation, HSulf-1-expressing clonal lines showed reduced staining with the endothelial marker CD31 in Matrigel plug assay, indicating that HSulf-1 expression inhibits angiogenesis. More importantly, HSulf-1 expression in the xenografts was associated with a reduced ability of vascular endothelial cell heparan sulfate to participate in a complex with fibroblast growth factor 2 (FGF-2) and its receptor tyrosine kinase FGF receptor 1c. In vitro, short hairpin RNA-mediated down-regulation of HSulf-1 in human umbilical vein endothelial cells (HUVEC) resulted in an increased proliferation mediated by heparan sulfate-dependent FGF-2, hepatocyte growth factor, and vascular endothelial growth factor 165 (VEGF165) but not by heparan sulfate-independent VEGF121. HSulf-1 down-regulation also enhanced downstream signaling through the extracellular signal-regulated kinase pathway compared with untreated cells. Consistent with the role of heparan sulfate glycosaminoglycan sulfation in VEGF-mediated signaling, treatment of HUVEC cells with chlorate, which inhibits heparan sulfate glycosaminoglycan sulfation and therefore mimics HSulf-1 overexpression, led to an attenuated VEGF-mediated signaling. Collectively, these observations provide the first evidence of a novel mechanism by which HSulf-1 modulates the function of heparan sulfate binding VEGF165 in proliferation and angiogenesis.
authors.affiliation: Department of Experimental Pathology, Mayo Clinic Cancer Center, Rochester, Minnesota 55905, USA.
authors.forename: Keishi
authors.initials: K
authors.lastname: Narita
authors.forename: Julie
authors.initials: J
authors.lastname: Staub
authors.forename: Jeremy
authors.initials: J
authors.lastname: Chien
authors.forename: Kristy
authors.initials: K
authors.lastname: Meyer
authors.forename: Maret
authors.initials: M
authors.lastname: Bauer
authors.forename: Andreas
authors.initials: A
authors.lastname: Friedl
authors.forename: Sundaram
authors.initials: S
authors.lastname: Ramakrishnan
authors.forename: Viji
authors.initials: V
authors.lastname: Shridhar
mesh_terms: D000818:Animals; D001943:Breast Neoplasms / Q000098:blood supply* / Q000235:genetics / Q000473:pathology / Q000628:therapy*; D048708:Cell Growth Processes / Q000502:physiology; D045744:Cell Line, Tumor; D015536:Down-Regulation; D042783:Endothelial Cells / Q000166:cytology / Q000378:metabolism / Q000502:physiology; D005260:Female; D016222:Fibroblast Growth Factor 2 / Q000378:metabolism; D006497:Heparan Sulfate / Q000378:metabolism; D006801:Humans; D051379:Mice; D008819:Mice, Nude; D009389:Neovascularization, Pathologic / Q000235:genetics / Q000378:metabolism / Q000473:pathology / Q000628:therapy; D010051:Ovarian Neoplasms / Q000098:blood supply* / Q000235:genetics / Q000378:metabolism / Q000628:therapy*; D034741:RNA, Small Interfering / Q000235:genetics; D051496:Receptor, Fibroblast Growth Factor, Type 1 / Q000378:metabolism; D015238:Sulfotransferases / Q000096:biosynthesis / Q000235:genetics / Q000502:physiology*; D014162:Transfection; D042461:Vascular Endothelial Growth Factor A / Q000378:metabolism; D023041:Xenograft Model Antitumor Assays
chemical_list: D016222:Fibroblast Growth Factor 2; D006497:Heparan Sulfate; D034741:RNA, Small Interfering; D051496:Receptor, Fibroblast Growth Factor, Type 1; D015238:Sulfotransferases; D042461:Vascular Endothelial Growth Factor A; C467484:VEGFA protein, human; C470791:SULF1 protein, human