Anti-angiogenic activity of human endostatin is HIF-1-independent in vitro and sensitive to timing of treatment in a human saphenous vein assay.
Endostatin is a 20-kDa endogenous angiogenesis inhibitor that has recently been shown to inhibit the expression of vascular endothelial growth factor (VEGF), an angiogenic growth factor that is up-regulated by hypoxia via the HIF-1 transcription factor complex. To determine if the anti-angiogenic activity of endostatin involves a modulation of the HIF-1/VEGF pathway in cancer cells, experiments were conducted to establish what effect endostatin has on HIF-1 activity, HIF-1alpha protein production, and cellular localization in prostate cancer cells and endothelial cells. Endothelial cell tube formation was inhibited by endostatin purchased from Calbiochem (San Diego, CA) but not endostatin obtained from EntreMed (Rockville, MD). Subsequent experiments using Calbiochem endostatin showed that it did not alter HIF-1alpha protein production or cellular localization in any of the cell lines tested, nor did it alter HIF-1 transactivational activity in hypoxia. Whether or not this is also true in vivo remains to be determined. Nevertheless, these data suggest that the anti-angiogenic activity of endostatin is independent of the HIF-1/VEGF pathway. Immunocytochemical staining results do not indicate a decreased production of VEGF in Calbiochem endostatin-treated LNCaP or human umbilical vein endothelial cells (HUVEC). Treatment of rat aortic cross sections with human endostatin from Calbiochem resulted in a dose-dependent inhibition of microvessel outgrowth. Importantly, inhibition of vessel outgrowth by Calbiochem endostatin in a human saphenous vein angiogenesis assay required early treatment. In view of this in vitro data, we suggest that clinical trials involving endostatin treatment of late-stage disease may not adequately represent the efficacy of this drug in early-stage cancer.
Macpherson
Gordon R
GR
Molecular Pharmacology Laboratory, Laboratory of Receptor Biology and Gene Expression, and Cell and Cancer Biology Branch and Vascular Biology Faculty, National Cancer Institute, Bethesda, MD 20892, USA.
Ng
Sylvia S W
SS
Forbes
Siiri L
SL
Melillo
Giovanni
G
Karpova
Tatiana
T
McNally
James
J
Conrads
Thomas P
TP
Veenstra
Timothy D
TD
Martinez
Alfredo
A
Cuttitta
Frank
F
Price
Douglas K
DK
Figg
William D
WD
D020533:Angiogenesis Inhibitors / Q000627:therapeutic use*; D000818:Animals; D001013:Aorta, Thoracic / Q000187:drug effects; D002851:Chromatography, High Pressure Liquid; D043169:Endostatins / Q000627:therapeutic use*; D004730:Endothelium, Vascular / Q000187:drug effects; D015972:Gene Expression Regulation, Neoplastic; D006801:Humans; D051795:Hypoxia-Inducible Factor 1, alpha Subunit; D015151:Immunoblotting; D007124:Immunoenzyme Techniques; D008156:Luciferases / Q000378:metabolism; D008297:Male; D013058:Mass Spectrometry; D009389:Neovascularization, Pathologic / Q000188:drug therapy* / Q000378:metabolism; D011471:Prostatic Neoplasms / Q000098:blood supply*; D051381:Rats; D017207:Rats, Sprague-Dawley; D012501:Saphenous Vein / Q000187:drug effects; D013997:Time Factors; D014157:Transcription Factors / Q000378:metabolism*; D015533:Transcriptional Activation; D014407:Tumor Cells, Cultured; D042461:Vascular Endothelial Growth Factor A / Q000378:metabolism
D020533:Angiogenesis Inhibitors; D043169:Endostatins; C497442:HIF1A protein, human; D051795:Hypoxia-Inducible Factor 1, alpha Subunit; D014157:Transcription Factors; D042461:Vascular Endothelial Growth Factor A; D008156:Luciferases