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Endothelin-1 induces the transactivation of vascular endothelial growth factor receptor-3 and modulates cell migration and vasculogenic mimicry in melanoma cells

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Abstract

Endothelin receptor B (ETBR) is a G-protein-coupled receptor overexpressed in melanoma, blood, and lymphatic endothelial cells. Given that aberrant signal transduction can be mediated through cross talk between receptors, here, we explore the functional relationship between ETBR and the vascular endothelial growth factor receptor (VEGFR)-3 system and how this cross talk might influence the aggressive behavior of melanoma cells. The expression of VEGFR-3 and its ligands, VEGF-C and VEGF-D, significantly increased after activating ETBR by ET-1 in primary and metastatic melanoma cell lines. These effects, similarly to those induced by hypoxia, were mediated by hypoxia-inducible factor (HIF)-1α and HIF-2α. ET-1 caused the phosphorylation of VEGFR-3, which was accompanied by the activation of the downstream signaling molecules, such as MAPK and AKT. Inhibition of c-Src activity or silencing of the scaffold protein β-arrestin-1 reduced ET-1-induced VEGFR-3 phosphorylation, demonstrating that, upon ET-1 stimulus, β-arrestin-1 is involved with c-Src in the ETBR-mediated VEGFR-3 transactivation. Moreover, ET-1 in combination with VEGF-C further increased VEGFR-3, MAPK, and AKT phosphorylation and markedly promoted cell migration and vasculogenic mimicry. Dual inhibition of ETBR and VEGFR-3 was required for the effective inhibition of these effects, as well as for VEGFR-3 phosphorylation, demonstrating that ETBR cross talk with VEGFR-3 enhances cell plasticity and motility. Finally, in melanoma xenografts, ETBR antagonist inhibited tumor growth and the activation of the VEGF-C/VEGFR-3 axis, indicating that targeting ETBR may improve melanoma treatment acting directly or indirectly by impairing ETBR cross talk with VEGFR-3.

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References

  1. Chudnovsky Y, Khavari PA, Adams AE (2005) Melanoma genetics and the development of rational therapeutics. J Clin Invest 115:813–824

    PubMed  CAS  Google Scholar 

  2. Maniotis AJ, Folberg R, Hess A, Seftor EA, Gardner LM, Pe’er J, Trent JM, Meltzer PS, Hendrix MJ (1999) Vascular channel formation by human melanoma cells in vivo and in vitro: vasculogenic mimicry. Am J Pathol 155:739–752

    Article  PubMed  CAS  Google Scholar 

  3. Kirschmann DA, Seftor EA, Hardy KM, Seftor RE, Hendrix MJ (2012) Molecular pathways: vasculogenic mimicry in tumor cells: diagnostic and therapeutic implications. Clin Cancer Res 18:2726–2732. doi:10.1158/1078-0434

    Article  PubMed  CAS  Google Scholar 

  4. Vartanian A, Stepanova E, Grigorieva I, Solomko E, Belkin V, Baryshnikov A, Lichinitser M (2011) Melanoma vasculogenic mimicry capillary-like structure formation depends on integrin and calcium signaling. Microcirculation 18:390–399

    Article  PubMed  CAS  Google Scholar 

  5. Garbe C, Eigentler TK, Keilholz U, Hauschild A, Kirkwood JM (2011) Systematic review of medical treatment in melanoma: current status and future prospects. Oncologist 16:5–24

    Article  PubMed  CAS  Google Scholar 

  6. Ferrara N, Gerber HP, LeCouter J (2003) The biology of VEGF and its receptors. Nat Med 9:669–676

    Article  PubMed  CAS  Google Scholar 

  7. Joukov V, Pajusola K, Kaipainen A, Chilov D, Lahtinen I, Kukk E, Saksela O, Kalkkinen N, Alitalo K (1996) A novel vascular endothelial growth factor, VEGF-C, is a ligand for the Flt4 (VEGFR-3) and KDR (VEGFR-2) receptor tyrosine kinases. EMBO J 15:1751

    PubMed  CAS  Google Scholar 

  8. Alitalo K (2011) The lymphatic vasculature in disease. Nat Med 17:1371–1380

    Article  PubMed  CAS  Google Scholar 

  9. Skobe M, Hamberg LM, Hawighorst T, Schirner M, Wolf GL, Alitalo K, Detmar M (2001) Concurrent induction of lymphangiogenesis, angiogenesis, and macrophage recruitment by vascular endothelial growth factor-C in melanoma. Am J Pathol 159:893–903

    Article  PubMed  CAS  Google Scholar 

  10. Su JL, Yen CJ, Chen PS, Chuang SE, Hong CC, Kuo IH, Chen HY, Hung MC, Kuo ML (2007) The role of the VEGF-C/VEGFR-3 axis in cancer progression. Br J Cancer 96:541–545

    Article  PubMed  CAS  Google Scholar 

  11. Clarijs R, Schalkwijk L, Hofmann UB, Ruiter DJ, de Waal RM (2002) Induction of vascular endothelial growth factor receptor-3 expression on tumor microvasculature as a new progression marker in human cutaneous melanoma. Cancer Res 62:7059–7065

    PubMed  CAS  Google Scholar 

  12. Kilvaer TK, Valkov A, Sorbye S, Smeland E, Bremnes RM, Busund LT, Donnem T (2010) Profiling of VEGFs and VEGFRs as prognostic factors in soft tissue sarcoma: VEGFR-3 is an independent predictor of poor prognosis. PLoS One 5:e15368

    Article  PubMed  CAS  Google Scholar 

  13. Marchiò S, Primo L, Pagano M, Palestro G, Albini A, Veikkola T, Cascone I, Alitalo K, Bussolino F (1999) Vascular endothelial growth factor-C stimulates the migration and proliferation of Kaposi’s sarcoma cells. J Biol Chem 274:27617–27622

    Article  PubMed  Google Scholar 

  14. Kodama M, Kitadai Y, Tanaka M, Kuwai T, Tanaka S, Oue N, Yasui W, Chayama K (2008) Vascular endothelial growth factor C stimulates progression of human gastric cancer via both autocrine and paracrine mechanisms. Clin Cancer Res 14:7205–7214

    Article  PubMed  CAS  Google Scholar 

  15. Simiantonaki N, Jayasinghe C, Michel-Schmidt R, Peters K, Hermanns MI, Kirkpatrick CJ (2008) Hypoxia-induced epithelial VEGF-C/VEGFR-3 upregulation in carcinoma cell lines. Int J Oncol 32:585–592

    PubMed  CAS  Google Scholar 

  16. Semenza GL (1998) Hypoxia-inducible factor 1: master regulator of O2 homeostasis. Curr Opin Genet Dev 8:588–594

    Article  PubMed  CAS  Google Scholar 

  17. Liang X, Yang D, Hu J, Hao X, Gao J, Mao Z (2008) Hypoxia inducible factor-alpha expression correlates with vascular endothelial growth factor-C expression and lymphangiogenesis/angiogenesis in oral squamous cell carcinoma. Anticancer Res 28:1659–1666

    PubMed  Google Scholar 

  18. Schoppmann SF, Fenzl A, Schindl M, Bachleitner-Hofmann T, Nagy K, Gnant M, Horvat R, Jakesz R, Birner P (2006) Hypoxia inducible factor-1alpha correlates with VEGF-C expression and lymphangiogenesis in breast cancer. Breast Cancer Res Treat 99:135–141

    Article  PubMed  CAS  Google Scholar 

  19. Mouawad R, Spano JP, Comperat E, Capron F, Khayat D (2009) Tumoural expression and circulating level of VEGFR-3 (Flt-4) in metastatic melanoma patients: correlation with clinical parameters and outcome. Eur J Cancer 45:1407–1414

    Article  PubMed  CAS  Google Scholar 

  20. Mehnert JM, McCarthy MM, Jilaveanu L, Flaherty KT, Aziz S, Camp RL, Rimm DL, Kluger HM (2010) Quantitative expression of VEGF, VEGF-R1, VEGF-R2, and VEGF-R3 in melanoma tissue microarrays. Hum Pathol 41:375–384

    Article  PubMed  CAS  Google Scholar 

  21. Achen MG, Williams RA, Minekus MP, Thornton GE, Stenvers K, Rogers PA, Lederman F, Roufail S, Stacker SA (2001) Localization of vascular endothelial growth factor-D in malignant melanoma suggests a role in tumour angiogenesis. J Pathol 193:147–154

    Article  PubMed  CAS  Google Scholar 

  22. Little PJ, Burch ML, Al-aryahi S, Zheng W (2011) The paradigm of G protein receptor transactivation: a mechanistic definition and novel example. Sci World J 11:709–714

    Article  CAS  Google Scholar 

  23. Bittner M, Meltzer P, Chen Y, Jiang Y, Seftor E, Hendrix M, Radmacher M, Simon R, Yakhini Z, Ben-Dor A et al (2000) Molecular classification of cutaneous malignant melanoma by gene expression profiling. Nature 406:536–540

    Article  PubMed  CAS  Google Scholar 

  24. Bagnato A, Rosanò L, Spinella F, Di Castro V, Tecce R, Natali PG (2004) Endothelin B receptor blockade inhibits dynamics of cell interactions and communications in melanoma cell progression. Cancer Res 64:1436–1443

    Article  PubMed  CAS  Google Scholar 

  25. Spinella F, Garrafa E, Di Castro V, Rosanò L, Nicotra MR, Caruso A, Natali PG, Bagnato A (2009) Endothelin-1 stimulates lymphatic endothelial cells and lymphatic vessels to grow and invade. Cancer Res 69:2669–2676

    Article  PubMed  CAS  Google Scholar 

  26. Spinella F, Rosanò L, Del Duca M, Di Castro V, Nicotra MR, Natali PG, Bagnato A (2010) Endothelin-1 inhibits prolyl hydroxylase domain 2 to activate hypoxia-inducible factor-1alpha in melanoma cells. PLoS One 5:e11241

    Article  PubMed  Google Scholar 

  27. Galvagni F, Pennacchini S, Salameh A, Rocchigiani M, Neri F, Orlandini M, Petraglia F, Gotta S, Sardone GL, Matteucci G et al (2010) Endothelial cell adhesion to the extracellular matrix induces c-Src-dependent VEGFR-3 phosphorylation without the activation of the receptor intrinsic kinase activity. Circ Res 106:1839–1848

    Article  PubMed  CAS  Google Scholar 

  28. Rosanò L, Cianfrocca R, Masi S, Spinella F, Di Castro V, Biroccio A, Salvati E, Nicotra MR, Natali PG, Bagnato A (2009) Beta-arrestin links endothelin A receptor to beta-catenin signaling to induce ovarian cancer cell invasion and metastasis. Proc Natl Acad Sci U S A 106:2806–2811

    Article  PubMed  Google Scholar 

  29. Cohen B, Addadi Y, Sapoznik S, Meir G, Kalchenko V, Harmelin A, Ben-Dor S, Neeman M (2009) Transcriptional regulation of vascular endothelial growth factor C by oxidative and thermal stress is mediated by lens epithelium-derived growth factor/p75. Neoplasia 11:921–933

    PubMed  CAS  Google Scholar 

  30. Min Y, Ghose S, Boelte K, Li J, Yang L, Lin PC (2011) C/EBP-δ regulates VEGF-C autocrine signaling in lymphangiogenesis and metastasis of lung cancer through HIF-1α. Oncogene 30:4901–4909

    Article  PubMed  CAS  Google Scholar 

  31. Garrafa E, Caprara V, Di Castro V, Rosanò L, Bagnato A, Spinella F (2012) Endothelin-1 cooperates with hypoxia to induce vascular-like structures through vascular endothelial growth factor-C, -D and -A in lymphatic endothelial cells. Life Sci. doi:10.1016/j.lfs.2012.03.033

  32. Paulis YW, Soetekouw PM, Verheul HM, Tjan-Heijnen VC, Griffioen AW (2010) Signalling pathways in vasculogenic mimicry. Biochim Biophys Acta 1806:18–28

    PubMed  CAS  Google Scholar 

  33. Liu Y, Ye F, Yamada K, Tso JL, Zhang Y, Nguyen DH, Dong Q, Soto H, Choe J, Dembo A et al (2011) Autocrine endothelin-3/endothelin receptor B signaling maintains cellular and molecular properties of glioblastoma stem cells. Mol Cancer Res 9:1668–1685

    Article  PubMed  CAS  Google Scholar 

  34. Cueni LN, Hegyi I, Shin JW, Albinger-Hegyi A, Gruber S, Kunstfeld R, Moch H, Detmar M (2010) Tumor lymphangiogenesis and metastasis to lymph nodes induced by cancer cell expression of podoplanin. Am J Pathol 177:1004–1016

    Article  PubMed  CAS  Google Scholar 

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Acknowledgments

We gratefully acknowledge Aldo Lupo for excellent technical assistance, Maria Vincenza Sarcone for secretarial support, and Dr. Tania Merlino for the formal revision of the manuscript. Grant support: Associazione Italiana Ricerca sul Cancro (AIRC, MFAG #8910).

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The authors declare no conflict of interests related to the study.

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Correspondence to Francesca Spinella or Anna Bagnato.

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Spinella, F., Caprara, V., Di Castro, V. et al. Endothelin-1 induces the transactivation of vascular endothelial growth factor receptor-3 and modulates cell migration and vasculogenic mimicry in melanoma cells. J Mol Med 91, 395–405 (2013). https://doi.org/10.1007/s00109-012-0956-2

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  • DOI: https://doi.org/10.1007/s00109-012-0956-2

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