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In vitro characterisation of the anti-intravasative properties of the marine product heteronemin

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Abstract

Metastases destroy the function of infested organs and are the main reason of cancer-related mortality. Heteronemin, a natural product derived from a marine sponge, was tested in vitro regarding its properties to prevent tumour cell intravasation through the lymph-endothelial barrier. In three-dimensional (3D) cell cultures consisting of MCF-7 breast cancer cell spheroids that were placed on lymph-endothelial cell (LEC) monolayers, tumour cell spheroids induce “circular chemorepellent-induced defects” (CCIDs) in the LEC monolayer; 12(S)-Hydroxyeicosatetraenoic acid (12(S)-HETE) and NF-κB activity are major factors inducing CCIDs, which are entry gates for tumour emboli intravasating the vasculature. This 3D co-culture is a validated model for the investigation of intravasation mechanisms and of drugs preventing CCID formation and hence lymph node metastasis. Furthermore, Western blot analyses, NF-κB reporter, EROD, SELE, 12(S)-HETE, and adhesion assays were performed to investigate the properties of heteronemin. Five micromolar heteronemin inhibited the directional movement of LECs and, therefore, the formation of CCIDs, which were induced by MCF-7 spheroids. Furthermore, heteronemin reduced the adhesion of MCF-7 cells to LECs and suppressed 12(S)-HETE-induced expression of the EMT marker paxillin, which is a regulator of directional cell migration. The activity of CYP1A1, which contributed to CCID formation, was also inhibited by heteronemin. Hence, heteronemin inhibits important mechanisms contributing to tumour intravasation in vitro and should be tested in vivo.

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Abbreviations

ALOX:

Lipoxygenase A

CCIDs:

Circular chemorepellent-induced defects

CYP:

Cytochrome P450

EMT:

Epithelial to mesenchymal transition

EROD:

Ethoxyresorufin-O-deethylase

HUVEC:

Human umbilical vein endothelial cell

ICAM-1:

Intercellular adhesion molecule 1

LEC:

Lymph-endothelial cell

MCL2:

Myosin light chain 2

SELE:

Selectin E

12(S)-HETE 12(S):

Hydroxyeicosatetraenoic acid

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Acknowledgments

We wish to thank Toni Jäger for preparing the figures and to Prof Ulrichova for kindly providing HUVEC cells. Heteronemin was a kind gift of Professor Marcel Jaspars (University of Aberdeen) and was originally obtained from the NCI open repository. This work was supported by the Austrian Science Fund, FWF, grant number P20905-B13 (to W.M.), and grant numbers S10704-B03 and S10704-B13 (to V.M.D.), by the European Union, FP7 Health Research, project number HEALTH-F4-2008-202047 (to W.M.), further by a grant of the Fellinger foundation (to G.K.), grants of the Herzfelder family foundation (to T.S., H.D., P.S. and M.G.), a grant of the “Hochschuljubilaeumsstiftung der Stadt Wien” grant number H-2498/2011 (to P.S.), a scholarship from the Austrian exchange service OeAD (to K.J.). Grant Agency of the Czech Republic GACR (P505/11/1163) and ED0007/01/01 (to L.R. and M.S) from the Centre of the Region Haná for Biotechnological and Agricultural Research is also gratefully acknowledged.

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Authors and Affiliations

  1. Institute of Clinical Pathology, Medical University of Vienna, Waehringer Guertel 18-20, 1090, Vienna, Austria

    Sabine Kopf, Katharina Viola, Nicole Kretschy, Mathias Teichmann, Caroline Vonach, Benedikt Giessrigl, Nicole Huttary, Ingrid Raab, Sigurd Krieger & Georg Krupitza

  2. Department of Pharmacognosy, University of Vienna, Althanstrasse 14, 1090, Vienna, Austria

    Atanas G. Atanasov & Verena M. Dirsch

  3. Department for Clinical Pharmacy and Diagnostics, Faculty of Life Sciences, University of Vienna, Althanstrasse 14, 1090, Vienna, Austria

    Kanokwan Jarukamjorn & Walter Jäger

  4. Department of Pharmaceutical Chemistry, Faculty of Pharmaceutical Sciences, Khon Kaen University, Khon Kaen, 40002, Thailand

    Kanokwan Jarukamjorn

  5. Faculty of Science, Centre of the Region Haná for Biotechnological and Agricultural Research, Palacký University, Šlechtitelů 11, 783 71, Olomouc, Czech Republic

    Lucie Rarova & Miroslav Strnad

  6. Department of Medical and Chemical Laboratory Diagnostics, General Hospital of Vienna, Medical University of Vienna, Waehringer Guertel 18-20, 1090, Vienna, Austria

    Philipp Saiko & Thomas Szekeres

  7. Laboratoire de Biologie Moléculaire et Cellulaire du Cancer, Hôpital Kirchberg, 9 Rue Edward Streichen, 2540, Luxembourg, Luxembourg

    Marc Schumacher & Marc Diederich

  8. Department of Vascular Biology and Thrombosis Research, Medical University of Vienna, Lazarettgasse 19, 1090, Vienna, Austria

    Rainer de Martin

  9. Department of Medicine I, Institute of Cancer Research, Medical University of Vienna, Borschkegasse 8a, 1090, Vienna, Austria

    Wolfgang Mikulits & Michael Grusch

  10. Institute of Medical Genetics, Medical University of Vienna, Währinger Strasse 10, 1090, Vienna, Austria

    Helmut Dolznig

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  1. Sabine Kopf
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Correspondence to Georg Krupitza.

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Kopf, S., Viola, K., Atanasov, A.G. et al. In vitro characterisation of the anti-intravasative properties of the marine product heteronemin. Arch Toxicol 87, 1851–1861 (2013). https://doi.org/10.1007/s00204-013-1045-1

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