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Thromb Haemost 2009; 102(05): 816-822
DOI: 10.1160/TH09-01-0061
Theme Issue Article
Schattauer GmbH

Binding between heparin and the integrin VLA-4

Martin Schlesinger
1   Department of Pharmacy, Rheinische Friedrich Wilhelms University, Bonn, Germany
,
Dirk Simonis
1   Department of Pharmacy, Rheinische Friedrich Wilhelms University, Bonn, Germany
,
Patrick Schmitz
1   Department of Pharmacy, Rheinische Friedrich Wilhelms University, Bonn, Germany
,
Juliane Fritzsche
1   Department of Pharmacy, Rheinische Friedrich Wilhelms University, Bonn, Germany
,
Gerd Bendas
1   Department of Pharmacy, Rheinische Friedrich Wilhelms University, Bonn, Germany
› Author AffiliationsFinancial support: This work was in part supported by grants from Deutsche Forschungsgemeinschaft (GRK 677 to JF, DS, and MS).
Further Information

Publication History

Received: 26 January 2009

Accepted after major revision: 28 June 2009

Publication Date:
27 November 2017 (online)

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Summary

Heparin possesses antimetastatic effects that were related to various molecular mechanisms beyond anticoagulant activities. The ability of heparin to interfere with the function of adhesion receptors in the metastatic course appears as a promising therapeutic approach. This refers to numerous findings that heparin attenuates metastasis in a selectin-dependent manner. We recently demonstrated that heparin interferes with the integrin VLA-4 on murine melanoma cells binding to VCAM-1. To confirm this activity and to obtain further insight into molecular recognition of heparin by VLA-4, we investigated the inhibition of VLA-4 mediated binding of human melanoma MV3 cells to immobilized VCAM-1 by different heparins. The size of heparin has an important impact on inhibition. Unfractionated heparin (UFH) and tinzaparin, a low-molecular-weight heparin (LMWH) representing a mean of about 18– 20 monomers, displayed high inhibitory activity. Fractionating tinzaparin to 14– 18 monomers reduced inhibition slightly, while the pentasaccharide fondaparinux was without effects. To confirm molecular recognition of tinzaparin by VLA-4,a surface acoustic wave-biosensor was applied. A VLA-4 containing membrane preparation of MV3 cells was immobilised at the sensors to allow for detection of kinetic binding constants of tinzaparin compared to VCAM-1. Tinzaparin binds to VLA-4 with affinity in the low micromolar range (4.61× 10−6 M), which clearly indicates specific molecular recognition. Furthermore, tinzaparin displays a nearly identical koff compared toVCAM-1 (5.13× 10−3 s-1 versus 3.44× 10–3 s-1) which is evident for interference with the ligand binding. The data provide evidence for a direct confirmation of heparin binding to VLA-4 and thus, contribute to understand the antimetastatic activity of heparin.

Keywords

Heparin - melanoma cell adhesion - VLA-4 - antimetastatic activity
 

  • References

  • 1 Schmidmaier R, Baumann P. ANTI-ADHESION evolves to a promising therapeutic concept in oncology. Curr Med Chem 2008; 15: 978-990.
    CrossrefPubMedSearch in Google Scholar
  • 2 Borsig L, Wong R, Hynes RO. et al. Synergistic effects of L-and P-selectin in facilitating tumor metastasis can involve non-mucin ligands and implicate leukocytes as enhancers of metastasis. Proc Natl Acad Sci U S A 2002; 99: 2193-2198.
    CrossrefPubMedSearch in Google Scholar
  • 3 Ludwig RJ, Boehme B, Podda M. et al. Endothelial P-selectin as a target of heparin action in experimental melanoma lung metastasis. Cancer Res 2004; 64: 2743-2750.
    CrossrefPubMedSearch in Google Scholar
  • 4 Okahara H, Yagita H, Miyake K. et al. Involvement of very late activation antigen 4 (VLA-4) and vascular cell adhesion molecule 1 (VCAM-1) in tumor necrosis factor alpha enhancement of experimental metastasis. Cancer Res 1994; 54: 3233-3236.
    PubMedSearch in Google Scholar
  • 5 Schadendorf D, Heidel J, Gawlik C. et al. Association With Clinical Outcome of Expression of VLA-4 in Primary Cutaneous Malignant Melanoma as Well as P-selectin and E-selectin on Intratumoral Vessels. J Natl Cancer Inst 1995; 87: 366-371.
    CrossrefPubMedSearch in Google Scholar
  • 6 Fritzsche J, Simonis D, Bendas G. Melanoma cell adhesion can be blocked by heparin in vitro: Suggestion of VLA-4 as a novel target for antimetastatic approaches. Thromb Haemost 2008; 100: 1166-1175.
    Thieme ConnectPubMedSearch in Google Scholar
  • 7 Liang S, Dong C. Integrin VLA-4 enhances sialyl-Lewisx/a-negative melanoma adhesion to and extravasation through the endothelium under low flow conditions. Am J Physiol Cell Physiol 2008; 295: C701-707.
    CrossrefPubMedSearch in Google Scholar
  • 8 Akl EA, van Doormaal FF, Barba M. et al. Parenteral anticoagulation may prolong the survival of patients with limited small cell lung cancer: a Cochrane systematic review. J Exp Clin Cancer Res 2008; 27: 4.
    CrossrefPubMedSearch in Google Scholar
  • 9 Borsig L. Selectins facilitate carcinoma metastasis and heparin can prevent them. News Physiol Sci 2004; 19: 16-21.
    PubMedSearch in Google Scholar
  • 10 Mousa SA, Mohamed S. Inhibition of endothelial cell tube formation by the low molecular weight heparin, tinzaparin, is mediated by tissue factor pathway inhibitor. Thromb Haemost 2004; 92: 627-633.
    Thieme ConnectPubMedSearch in Google Scholar
  • 11 Jayson GC, Gallagher JT. Heparin oligosaccharides: inhibition of the biological activity of bFGF on Caco-2 cells. Br J Cancer 1997; 75: 9-16.
    CrossrefPubMedSearch in Google Scholar
  • 12 Vlodavsky I, Abboud-Jarrous G, Elkin M. et al. The impact of heparanase and heparin on cancer metastasis and angiogenesis. Pathophysiol Haemost Thromb 2006; 35: 116-127.
    CrossrefPubMedSearch in Google Scholar
  • 13 Sobel M, Fish WR, Toma N. et al. Heparin modulates integrin function in human platelets. J Vasc Surg 2001; 33: 587-594.
    CrossrefPubMedSearch in Google Scholar
  • 14 Diamond MS, Alon R, Parkos CA. et al. Heparin is an adhesive ligand for the leukocyte integrin Mac-1 (CD11b/CD18). J Cell Biol 1995; 130: 1473-1482.
    CrossrefPubMedSearch in Google Scholar
  • 15 Vorup-Jensen T, Chi L, Gjelstrup LC. et al. Binding between the integrin α Xβ 2 (CD11c/CD18) and Heparin. J Biol Chem 2007; 282: 30869-30877.
    CrossrefPubMedSearch in Google Scholar
  • 16 Da Silva MS, Horton JA, Wijelath JM. et al. Heparin modulates integrin-mediated cellular adhesion: specificity of interactions with alpha and beta integrin subunits. Cell Commun Adhes 2003; 10: 59-67.
    CrossrefPubMedSearch in Google Scholar
  • 17 Perpeet M, Glass S, Gronewold TM. et al. SAW Sensor System for Marker-Free Molecular Interaction Analysis. Anal Lett 2006; 39: 1747-1757.
    CrossrefPubMedSearch in Google Scholar
  • 18 Gronewold TM, Baumgartner A, Quandt E. et al. Discrimination of single mutations in cancer-related gene fragments with a surface acoustic wave sensor. Anal Chem 2006; 78: 4865-4871.
    CrossrefPubMedSearch in Google Scholar
  • 19 Gronewold TM, Glass S, Quandt E. et al. Monitoring complex formation in the blood-coagulation cascade using aptamer-coated SAW sensors. Biosens Bioelectron 2005; 20: 2044-2052.
    CrossrefPubMedSearch in Google Scholar
  • 20 Andra J, Bohling A, Gronewold TM. et al. Surface acoustic wave biosensor as a tool to study the interaction of antimicrobial peptides with phospholipid and lipopolysaccharide model membranes. Langmuir 2008; 24: 9148-9153.
    CrossrefPubMedSearch in Google Scholar
  • 21 Deisingh AK, Thompson M. Biosensors for the detection of bacteria. Can J Microbiol 2004; 50: 69-77.
    CrossrefPubMedSearch in Google Scholar
  • 22 Casalinuovo IA, Pierro D, Bruno E. et al. Experimental use of a new surface acoustic wave sensor for the rapid identification of bacteria and yeasts. Lett Appl Microbiol 2006; 42: 24-29.
    CrossrefPubMedSearch in Google Scholar
  • 23 Tamarin O, Comeau S, Dejous C. et al. Real time device for biosensing: design of a bacteriophage model using love acoustic waves. Biosens Bioelectron 2003; 18: 755-763.
    CrossrefPubMedSearch in Google Scholar
  • 24 Cavic BA, Hayward GL, Thompson M. Acoustic waves and the study of biochemical macromolecules and cells at the sensor-liquid interface. Analyst 1999; 124: 1405-1420.
    CrossrefPubMedSearch in Google Scholar
  • 25 Hong S, Ergezen E, Barbee K. et al. BAEC adhesion analysis using Thickness Shear Mode sensor. Conf Proc IEEE Eng Med Biol Soc 2005; 01: 1047-1050.
    PubMedSearch in Google Scholar
  • 26 Fritzsche J, Alban S, Ludwig RJ. et al. The influence of various structural parameters of semisynthetic sulfated polysaccharides on the P-selectin inhibitory capacity. Biochem Pharmacol 2006; 72: 474-485.
    CrossrefPubMedSearch in Google Scholar
  • 27 Lever JE. Active amino acid transport in plasma membrane vesicles from Simian virus 40-transformed mouse fibroblasts. Characteristics of electrochemical Na+ gradient-stimulated uptake. J Biol Chem 1977; 252: 1990-1997.
    PubMedSearch in Google Scholar
  • 28 Kaplan RS, Pedersen PL. Determination of microgram quantities of protein in the presence of milligram levels of lipid with amido black 10B. Anal Biochem 1985; 150: 97-104.
    CrossrefPubMedSearch in Google Scholar
  • 29 Swerlick RA, Lee KH, Li LJ. et al. Regulation of vascular cell adhesion molecule 1 on human dermal microvascular endothelial cells. J Immunol 1992; 149: 698-705.
    PubMedSearch in Google Scholar
  • 30 Stevenson JL, Choi SH, Varki A. Differential metastasis inhibition by clinically relevant levels of heparins--correlation with selectin inhibition, not antithrombotic activity. Clin Cancer Res 2005; 11: 7003-7011.
    CrossrefPubMedSearch in Google Scholar
  • 31 Sotnikov I, Hershkoviz R, Grabovsky V. et al. Enzymatically quiescent heparanase augments T cell interactions with VCAM-1 and extracellular matrix components under versatile dynamic contexts. J Immunol 2004; 172: 5185-5193.
    CrossrefPubMedSearch in Google Scholar
  • 32 Levy-Adam F, Feld S, Suss-Toby E. et al. Heparanase facilitates cell adhesion and spreading by clustering of cell surface heparan sulfate proteoglycans. PLoS ONE 2008; 03: e2319.
    CrossrefPubMedSearch in Google Scholar
  • 33 Clark K, Newham P, Burrows L. et al. Production of recombinant soluble human integrin alpha4beta1. FEBS Lett 2000; 471: 182-186.
    CrossrefPubMedSearch in Google Scholar
  • 34 McEver RP. Selectin-carbohydrate interactions during inflammation and metastasis. Glycoconj J 1997; 14: 585-591.
    CrossrefPubMedSearch in Google Scholar
  • 35 Chigaev A, Zwartz G, Graves SW. et al. Alpha4beta1 integrin affinity changes govern cell adhesion. J Biol Chem 2003; 278: 38174-38182.
    CrossrefPubMedSearch in Google Scholar
  • 36 Simonis D, Fritzsche J, Alban S. et al. Kinetic analysis of heparin and glucan sulfates binding to P-selectin and its impact on the general understanding of selectin inhibition. Biochemistry 2007; 46: 6156-6164.
    CrossrefPubMedSearch in Google Scholar