Download PDF
Thromb Haemost 2008; 100(06): 1166-1175
DOI: 10.1160/TH08-05-0332
Cellular Proteolysis and Oncology
Schattauer GmbH

Melanoma cell adhesion can be blocked by heparin in vitro: Suggestion of VLA-4 as a novel target for antimetastatic approaches

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

Publication History

Received: 29 May 2008

Accepted after major revision: 12 September 2008

Publication Date:
23 November 2017 (online)

Also available at
    Permissions and Reprints

Summary

The clinical benefit of heparin in cancer patients to prolong survival can be attributed to non-anticoagulant mechanisms. Since adhesion molecules are crucially involved in tumour cell metastasis, their inhibition offers an attractive approach for interfering with the metastatic cascade. Heparin is known to attenuate metastasis in a selectin-dependent manner and possessesa variety of additional effects that are thought to influence tumour cell dissemination, proliferation, and angiogenesis. We investigated the adhesion behaviour of B16F10 melanoma cells in vitro regarding selectin- and VLA-4/VCAM-1-mediated binding to get an insight into underlying mechanisms of melanoma cell metastasis. We show that B16F10 cells display binding ability to P- and L-selectin as well as to isolated platelets. In contrast, B16F10 cells did not adhere to immobilized P-selectin under flow. This contributes to recent findings that elucidate a major role of platelet P-selectin for microemboli formation and thus, facilitating metastasis. In contrast, B16F10 cells adhered to endothelial cells under flow, which could partly be inhibited by a function-blocking anti-VCAM-1 mAb. To emphasizeVCAM-1 function, we analyzed cell adhesion at immobilizedVCAM-1 and observed an integrin dependency. Inhibition experiments reveal that heparin influences VLA-4-mediated binding pathways. By a combination of different techniques we prove that the site of heparin action is ratherVLA-4 thanVCAM-1.To our knowledge, this is the first time that heparin is shown to interfere with theVLA-4/VCAM-1 interaction leading to the suggestion of a novel heparin target. Our results may contribute to the understanding of how heparin exerts its anti-metastatic activity.

Keywords

Heparins - cell-cell interactions - integrins - cancer - melanoma
 

  • References

  • 1 Borsig L, Wong R, Feramisco J. et al. Heparin and cancer revisited: mechanistic connections involving platelets, P-selectin, carcinoma mucins, and tumour metastasis. Proc Natl Acad Sci USA 2001; 98: 3352-3357.
    CrossrefPubMedSearch in Google Scholar
  • 2 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
  • 3 Läubli H, Stevenson JL, Varki A. et al. L-selectin facilitation of metastasis involves temporal induction of Fut7-dependent ligands at sites of tumour cell arrest. Cancer Res 2006; 66: 1536-1542.
    CrossrefPubMedSearch in Google Scholar
  • 4 Nieswandt B, Hafner M, Echtenacher B. et al. Lysis of tumour cells by natural killer cells is in mice is impeded by platelets. Cancer Res 1999; 59: 1295-1300.
    PubMedSearch in Google Scholar
  • 5 Borsig L, Wong R, Hynes RO. et al. Synergistic effects of L- and P-selectin in facilitating tumour metastasis can involve non-mucin ligands and implicate leukocytes as enhancers of metastasis. Proc Natl Acad Sci USA 2002; 99: 2193-2198.
    CrossrefPubMedSearch in Google Scholar
  • 6 Krause T, Turner GA. Are selectins involved in metastasis?. Clinical & Experimental Metastasis 1999; 17: 183-192.
    CrossrefPubMedSearch in Google Scholar
  • 7 Elices MJ, Osborn L, Takada Y. et al. VCAM-1 on activated endothelium interacts with the leukocyte integrin VLA-4 ata site distinct from the VLA-4/fibronectin binding site. Cell 1990; 60: 577-584.
    CrossrefPubMedSearch in Google Scholar
  • 8 Mattila P, Majuri ML, Renkonen R. VLA-4 integrin on sarcoma cell lines recognizes endothelial VCAM-1. Differential regulation of the VLA-4 avidity on various sarcoma cell lines. Int J Cancer 2006; 52: 918-923.
    PubMedSearch in Google Scholar
  • 9 Klemke M, Weschenfelder T, Konstandin MH. et al. High affinity interaction of integrin a4b1 (VLA-4) and vascular cell adhesion molecule 1 (VCAM-1) enhances migration of human melanoma cells across activated endothelial cell layers. J Cell Physiol 2007; 212: 368-374.
    CrossrefPubMedSearch in Google Scholar
  • 10 Okahara H, Yagita H, Miyake K. et al. Involvement of very late activation antigen 4 (VLA-4) and vascular cell adhesion molecule1 (VCAM-1) in tumour necrosis factor alpha enhancement of experimental metastasis. Cancer Res 1994; 54: 3233-3236.
    PubMedSearch in Google Scholar
  • 11 Garofalo A, Chirivi RG, Foglieni C. et al. Involvement of the very late antigen 4 integrin on melanoma in interleukin 1-augmented experimental metastases. Cancer Res 1995; 55: 414-419.
    PubMedSearch in Google Scholar
  • 12 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
  • 13 Kakkar AK, Levine MN, Kadziola Z. et al. Low molecular weight heparin, therapy with dalteparin, and survival in advanced cancer: the fragmin advanced malignancy outcome study (FAMOUS). J Clin Oncol 2004; 22: 1944-1948.
    CrossrefPubMedSearch in Google Scholar
  • 14 Altinbas M, Coskun HS, Er O. et al. A randomized clinical trial of combination chemotherapy with and without low-molecular-weight heparin in small cell lung cancer. J Thromb Haemost 2004; 02: 1266-1271.
    CrossrefPubMedSearch in Google Scholar
  • 15 Klerk CP, Smorenburg SM, Otten HM. et al. The effect of low molecular weight heparin on survival in patients with advanced malignancy. J Clin Oncol 2005; 23: 2130-2135.
    CrossrefPubMedSearch in Google Scholar
  • 16 Lee AY, Rickles FR, Julian JA. et al. Randomized comparison of low molecular weight heparin and coumarin derivatives on the survival of patients with cancer and venous thromboembolism. J Clin Oncol 2005; 23: 2123-2129.
    CrossrefPubMedSearch in Google Scholar
  • 17 Niers TM, Klerk CP, DiNisio M. et al. Mechanisms of heparin induced anti-cancer activity in experimental cancer models. Crit Rev Oncol Hematol 2007; 61: 195-207.
    CrossrefPubMedSearch in Google Scholar
  • 18 Yoshitomi Y, Nakanishi H, Kusano Y. et al. Inhibition of experimental lung metastases of Lewis lung carcinoma cells by chemically modified heparin with reduced anticoagulant activity. Cancer Lett 2004; 207: 165-174.
    CrossrefPubMedSearch in Google Scholar
  • 19 Kragh M, Binderup L, Vig Hjarnaa PJ. et al. Nonanti-coagulant heparin inhibits metastasis but not primary tumour growth. Oncol Rep 2005; 14: 99-104.
    PubMedSearch in Google Scholar
  • 20 Hostettler N, Naggi A, Torri G. et al. P-selectin and heparanase-dependent antimetastatic activity of nonanticoagulant heparins. Faseb J 2007; 21: 3562-3572.
    CrossrefPubMedSearch in Google Scholar
  • 21 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
  • 22 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
  • 23 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
  • 24 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
  • 25 Kim YJ, Borsig L, Varki NM. et al. P-selectin deficiency attenuates tumour growth and metastasis. Proc Natl Acad Sci USA 1998; 95: 9325-9330.
    CrossrefPubMedSearch 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 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
  • 28 Robert C, Fuhlbrigge RG, Kieffer JD. et al. Interaction of dendritic cells with skin endothelium: a new perspective on immunosurveillance. J Exp Med 1999; 189: 627-636.
    CrossrefPubMedSearch in Google Scholar
  • 29 Ma YQ, Geng JG. Heparan sulfate-like proteoglycans mediate adhesion of human malignant melanoma A375 cells to P-selectin under flow. J Immunol 2000; 165: 558-565.
    CrossrefPubMedSearch in Google Scholar
  • 30 Hanley W, McCarty O, Jadhav S. et al. Single molecule characterization of P-selectin/ligand binding. J Biol Chem 2003; 278: 10556-10561.
    CrossrefPubMedSearch in Google Scholar
  • 31 Aigner S, Sthoeger ZM, Fogel M. et al. CD24, a mucin-type glycoprotein, is a ligand for P-selectin on human tumour cells. Blood 1997; 89: 3385-3395.
    PubMedSearch in Google Scholar
  • 32 Garcia J, Callewaert N, Borsig L. P-selectin mediates metastatic progression through binding to sulfatides on tumour cells. Glycobiology 2007; 17: 185-196.
    CrossrefPubMedSearch in Google Scholar
  • 33 Dardik R, Savion N, Kaufmann Y. et al. Thrombin promotes platelet-mediated melanoma cell adhesion to endothelial cells under flow conditions: role of platelet glycoproteins P-selectin and GPIIb-IIIA. Br J Cancer 1998; 77: 2069-2075.
    CrossrefPubMedSearch in Google Scholar
  • 34 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
  • 35 Kim YJ, Borsig L, Han HL. et al. Distinct selectin ligands on colon carcinoma mucins can mediate pathological interactions among platelets, leukocytes, and endothelium. Am J Pathol 1999; 155: 461-472.
    CrossrefPubMedSearch in Google Scholar
  • 36 Ohyama C, Tsuboi S, Fukuda M. Dual roles of sialyl Lewis X oligosaccharides in tumour metastasis and rejection by natural killer cells. EMBO J 1999; 18: 1516-1525.
    CrossrefPubMedSearch in Google Scholar
  • 37 Sikorski EE, Hallmann R, Berg EL. et al. The Peyer’s patch high endothelial receptor for lymphocytes, the mucosal vascular addressin, is induced on a murine endothelial cell line by tumour necrosis factoralpha and IL-1. J Immunol 1993; 151: 5239-5250.
    PubMedSearch in Google Scholar
  • 38 Gosk S, Moos T, Gottstein C. et al. VCAM-1 directed immunoliposomes selectively target tumour vasculature in vivo. Biochim Biophys Acta 2008; 1778: 854-863.
    CrossrefPubMedSearch in Google Scholar
  • 39 Robledo MM, Bartolomé RA, Longo N. et al. Expression of functional chemokine receptors CXCR3 and CXCR4 on human melanoma cells. J Biol Chem 2001; 276: 45098-45105.
    CrossrefPubMedSearch in Google Scholar
  • 40 Cardones AR, Murakami T, Hwang ST. CXCR4 enhances adhesion of B16 tumour cells to endothelial cells in vitro and in vivo via β1 integrin. Cancer Res 2003; 63: 6751-6757.
    PubMedSearch in Google Scholar
  • 41 Luo BH, Carman CV, Springer TA. Structural basis of integrin regulation and signaling. Annu Rev Immunol 2007; 25: 619-647.
    CrossrefPubMedSearch in Google Scholar
  • 42 Vonderheide RH, Springer TA. Lymphocyte adhesion through very late activation antigen 4: Evidence for a novel binding site in the alternatively spliced domain of vascular cell adhesion molecule1 and an additional α4 integrin counter-receptor on stimulated endothelium. J Exp Med 1992; 175: 1433-1442.
    CrossrefPubMedSearch in Google Scholar
  • 43 Saini A, Seller Z, Davies D. et al. Activation status and function of the VLA-4 (a4b1) integrin expressed on human melanoma cell lines. Int J Cancer 1997; 73: 264-270.
    CrossrefPubMedSearch in Google Scholar
  • 44 Nakayama J, Guan XC, Nakashima M. et al. In vitro comparison between mouse B16 and human melanoma cell lines of the expression of ICAM-1 induced by cytokines and/or hyperthermia. J Dermatol 1997; 24: 351-360.
    CrossrefPubMedSearch in Google Scholar
  • 45 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
  • 46 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
  • 47 Vorup-Jensen T, Chi L, Gjelstrup LC. et al. Binding between the integrin αXβ2 (CD11c/CD18) and heparin. J Biol Chem 2007; 282: 30869-30877.
    CrossrefPubMedSearch in Google Scholar