title: Antineoplastic agent busulfan regulates a network of genes related to coagulation and fibrinolysis.
abstract: Purpose Hepatic veno-occlusive disease (HVOD) is one of the major complications following hematopoietic stem cell transplantation (HSCT). Although high-dose busulfan is associated with the development of HVOD, the underlying molecular mechanisms are still unknown.Methods Transcriptional gene regulation by busulfan was profiled using Affymetrix GeneChip® Human Genome U133 Plus 2.0 arrays. Messenger RNA (mRNA) expression of regulated genes was assessed by TaqMan real-time polymerase chain reaction (PCR), and protein expression and secretion was determined by enzyme-linked immunosorbent assay (ELISA)in cell supernatants, lysates, and patient plasma.Results Plasma levels of plasminogen activator inhibitor(PAI)-1 significantly increased 48 h after starting busulfan treatment IV in children preconditioned for HSCT. In vitro,busulfan significantly induced plasminogen activator inhibitor-1 (PAI-1) expression in endothelium-like ECV304 cells in a concentration- and time-dependent manner. Comparative transcriptional profiling of busulfan-treated and control ECV304 cells identified differential expression of genes related to coagulation and fibrinolysis, including tissue factor, tissue factor pathway inhibitor-1, protein S, thrombospondin-1, urokinase receptor, and PAI-1, as well as activin A and transforming growth factor beta 1 (TGF-β1). Ingenuity pathway analysis (IPA) suggested TGF-β1 as a central modulator of gene regulation by busulfan. Consequently, expression of tissue factor, urokinase receptor, and PAI-1 mRNA and PAI-1 protein secretion induced by busulfan were significantly reduced by the activin A/TGF-β1 inhibitor SB 431542 in ECV304 and primary endothelial cells.Conclusions This is the first report that directly relates busulfan exposure to antifibrinolytic activity by PAI-1 and hypercoagulation possibly mediated by members of the TGF-β1 family. This suggests further research to evaluate activin A and TGF-β1 as potential targets for HVOD treatment.
authors.affiliation: Research Center of Pharmacology and Experimental Therapeutics,Department of Pharmacology, Ernst-Moritz-Arndt-Universityof Greifswald,Greifswald, Germany.
authors.forename: Janka
authors.initials: J
authors.lastname: Reimer
authors.forename: Sandra
authors.initials: S
authors.lastname: Bien
authors.forename: Sabine
authors.initials: S
authors.lastname: Ameling
authors.forename: Elke
authors.initials: E
authors.lastname: Hammer
authors.forename: Uwe
authors.initials: U
authors.lastname: Völker
authors.forename: Georg
authors.initials: G
authors.lastname: Hempel
authors.forename: Joachim
authors.initials: J
authors.lastname: Boos
authors.forename: Heyo K
authors.initials: HK
authors.lastname: Kroemer
authors.forename: Christoph A
authors.initials: CA
authors.lastname: Ritter
mesh_terms: D028341:Activins / Q000037:antagonists & inhibitors / Q000494:pharmacology; D018906:Antineoplastic Agents, Alkylating / Q000494:pharmacology*; D001549:Benzamides / Q000494:pharmacology; D001777:Blood Coagulation / Q000235:genetics*; D002066:Busulfan / Q000494:pharmacology*; D045744:Cell Line, Tumor; D002648:Child; D004149:Dioxoles / Q000494:pharmacology; D005342:Fibrinolysis / Q000235:genetics*; D020869:Gene Expression Profiling; D005786:Gene Expression Regulation / Q000187:drug effects*; D015972:Gene Expression Regulation, Neoplastic / Q000187:drug effects*; D053263:Gene Regulatory Networks / Q000187:drug effects*; D018380:Hematopoietic Stem Cell Transplantation / Q000009:adverse effects / Q000379:methods; D006504:Hepatic Veno-Occlusive Disease / Q000097:blood / Q000209:etiology / Q000235:genetics; D061307:Human Umbilical Vein Endothelial Cells / Q000187:drug effects; D006801:Humans; D017395:Plasminogen Activator Inhibitor 1 / Q000096:biosynthesis / Q000097:blood / Q000235:genetics; D012333:RNA, Messenger / Q000096:biosynthesis / Q000235:genetics; D053773:Transforming Growth Factor beta1 / Q000037:antagonists & inhibitors / Q000235:genetics / Q000494:pharmacology; D014409:Tumor Necrosis Factor-alpha / Q000494:pharmacology; D001749:Urinary Bladder Neoplasms / Q000097:blood / Q000188:drug therapy / Q000235:genetics
chemical_list: C459179:4-(5-benzo(1,3)dioxol-5-yl-4-pyridin-2-yl-1H-imidazol-2-yl)benzamide; D018906:Antineoplastic Agents, Alkylating; D001549:Benzamides; D004149:Dioxoles; D017395:Plasminogen Activator Inhibitor 1; D012333:RNA, Messenger; C051321:SERPINE1 protein, human; D053773:Transforming Growth Factor beta1; D014409:Tumor Necrosis Factor-alpha; C422405:activin A; D028341:Activins; D002066:Busulfan