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Antineoplastic agent busulfan regulates a network of genes related to coagulation and fibrinolysis

  • Pharmacodynamics
  • Published:
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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.

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Abbreviations

HSCT:

Hematopoietic stem cell transplantation

HVOD:

Hepatic veno-occlusive disease

PAI-1:

Plasminogen activator inhibitor-1

TGF-β1:

Transforming growth factor beta-1

TFPI-1:

Tissue factor pathway inhibitor-1

THBS-1:

Thrombospondin-1

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Acknowledgements

This work was financially supported by the Wilhelm-Sander Foundation, Munich, Germany (Grant No. 2003.103.1, HKK)

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Authors and Affiliations

  1. Research Center of Pharmacology and Experimental Therapeutics, Department of Pharmacology, Ernst-Moritz-Arndt-University of Greifswald, Greifswald, Germany

    Janka Reimer, Sandra Bien, Heyo K. Kroemer & Christoph A. Ritter

  2. Department of Functional Genomics, Interfaculty Institute of Genetics and Functional Genomics, Ernst-Moritz-Arndt-University of Greifswald, Greifswald, Germany

    Sabine Ameling, Elke Hammer & Uwe Völker

  3. Institute of Pharmacy, Ernst-Moritz-Arndt-University of Greifswald, Friedrich-Ludwig-Jahn-Straße 17, 17487, Greifswald, Germany

    Christoph A. Ritter

  4. Klinik und Poliklinik für Kinder- und Jugendmedizin, Pädiatrische Hämatologie/Onkologie, Universitätsklinikum Münster, Münster, Germany

    Georg Hempel & Joachim Boos

  5. Institut für Pharmazeutische und Medizinische Chemie, Westfälische Wilhelms-Universität Münster, Münster, Germany

    Georg Hempel

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  1. Janka Reimer
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Reimer, J., Bien, S., Ameling, S. et al. Antineoplastic agent busulfan regulates a network of genes related to coagulation and fibrinolysis. Eur J Clin Pharmacol 68, 923–935 (2012). https://doi.org/10.1007/s00228-011-1209-y

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