Clinical & Experimental Metastasis

, Volume 22, Issue 8, pp 643–652 | Cite as

Downregulation of uPAR confirms link in growth and metastasis of osteosarcoma

  • Crispin R. Dass
  • Anne P. W. Nadesapillai
  • Daniel Robin
  • Monique L. Howard
  • Jane L. Fisher
  • Hong Zhou
  • Peter F. M. Choong


The uPA/uPAR system is involved in tumour progression and metastasis of a variety of cancers. Previously, we have shown that increased expression of urokinase plasminogen activator (uPA) correlated with malignancy grade in certain sarcomas. A study looking at in vivo inhibition of this system has not been done to date for osteosarcoma. More recently, this laboratory developed a clinically relevant mouse model where intratibial injection of UMR106-01 cells resulted in the development of osteosarcoma and lung metastases. Expression of uPA and its receptor (uPAR) were localised to the invading front of the tumours. Pulmonary metastasis is a predominant feature of the disease and is the major cause of death in patients. In the present study, the effects of down-regulating uPAR were observed in vitro and in vivo. UMR106-01 cells were transfected with either antisense-uPAR or vector control plasmids. Two antisense clones, exhibiting uPAR downregulation, demonstrated decreased adhesion, migration and invasion in cell-based assays in vitro (P<0.05). Cellular proliferation was not affected by uPAR downregulation. In␣vivo, a marked reduction of 80% in tibial tumour volumes (P<0.05), and total inhibition of pulmonary metastases were observed in mice injected with the more potent of the antisense clones. This study proves seminally the usefulness of uPAR antisense in curbing the growth and spread of osteosarcoma.


antisense cancer metastasis osteosarcoma urokinase plasminogen activator 


Unable to display preview. Download preview PDF.

Unable to display preview. Download preview PDF.



This work was supported by the Anti-Cancer Council of Victoria, and the Australian Orthopaedic Association. The authors thank Dr. Joseline Ojaimi for her critical reading of the manuscript.


  1. 1.
    Wittig JC, Bickels J, Priebat D, et al. Osteosarcoma: a multidisciplinary approach to diagnosis and treatment Am Fam Physician 2002; 65(6): 1123–32PubMedGoogle Scholar
  2. 2.
    Rabbani SA, Mazar AP The role of the plasminogen activation system in angiogenesis and metastasis Surg Oncol Clin N Am 2001; 10(2): 393–415PubMedGoogle Scholar
  3. 3.
    Mazzieri R, Masiero L, Zanetta L et al. Control of type IV collagenase activity by components of the urokinase-plasmin system: a regulatory mechanism with cell-bound reactants EMBO J 1997; 16(9): 2319–32PubMedCrossRefGoogle Scholar
  4. 4.
    Ferrara N Role of vascular endothelial growth factor in the regulation of angiogenesis Kidney Int 1999; 56: 794–814PubMedCrossRefGoogle Scholar
  5. 5.
    Ganesh S, Sier CF, Heerding MM et al. Urokinase receptor and colorectal cancer survival Lancet 1994; 344(8919): 401–2PubMedCrossRefGoogle Scholar
  6. 6.
    Kuhn W, Pache L, Schmalfeldt B et al. Urokinase (uPA) and PAI-1 predict survival in advanced ovarian cancer patients (FIGO III) after radical surgery and platinum-based chemotherapy Gynecol Oncol 1994; 55(3 Pt 1): 401–9PubMedCrossRefGoogle Scholar
  7. 7.
    Itaya T, Motai H, Suzuki K, Baba S Immunohistological study of fibrinolytic factors of head and neck squamous cell carcinomas Nippon Jibiinkoka Gakkai Kaiho 1995; 98(2): 197–202PubMedGoogle Scholar
  8. 8.
    Hofmann R, Lehmer A, Buresch M et al. Clinical relevance of urokinase plasminogen activator, its receptor, and its inhibitor in patients with renal cell carcinoma Cancer 1996; 78(3): 487–92PubMedCrossRefGoogle Scholar
  9. 9.
    Fisher JL, Field CL, Zhou H et al. Urokinase plasminogen activator system gene expression is increased in human breast carcinoma and its bone metastases – a comparison of normal breast tissue, non-invasive and invasive carcinoma and osseous metastases Breast Cancer Res Treat 2000; 61(1): 1–12PubMedCrossRefGoogle Scholar
  10. 10.
    McCabe NP, Angwafo FF 3rd, Zaher A et al. Expression of soluble urokinase plasminogen activator receptor may be related to outcome in prostate cancer patients Oncol Rep 2000; 7(4): 879–82PubMedGoogle Scholar
  11. 11.
    He C, He P, Liu LP, Zhu YS Analysis of expressions of components in the plasminogen activator system in high- and low-metastatic human lung cancer cells J Cancer Res Clin Oncol 2001; 127(3): 180–6PubMedCrossRefGoogle Scholar
  12. 12.
    Choong PF, Ferno M, Akerman M et al. Urokinase-plasminogen-activator levels and prognosis in 69 soft-tissue sarcomas Int J Cancer 1996; 69(4): 268–72PubMedCrossRefGoogle Scholar
  13. 13.
    Hackel C, Ueda Y, Tsuchya H et al. Plasminogen activators and their inhibitors in osteosarcomas and other bone tumours Zentralbl Pathol 1994; 140: 363–9Google Scholar
  14. 14.
    Hackel C, Ayala AG, Radig K, et al. Protease expression in dedifferentiated parosteal osteosarcoma Arch Pathol Lab Med 1999; 123(3): 213–21Google Scholar
  15. 15.
    Hackel CG, Krueger S, Grote HJ, et al. Over expression of cathepsin B and urokinase plasminogen activator is associated with increased risk of recurrence and metastasis in patients with chondrosarcoma Cancer 2000; 89(5): 995–1003PubMedCrossRefGoogle Scholar
  16. 16.
    Fisher JL, Mackie PS, Howard ML, et al. The expression of the urokinase plasminogen activator system in metastatic murine osteosarcoma: an in vivo mouse model Clin Cancer Res 2001; 7(6): 1654–60PubMedGoogle Scholar
  17. 17.
    Mackie PS, Fisher JL, Zhou H, Choong PF Bisphosphonates regulate cell growth and gene expression in the UMR106-01 clonal rat osteosarcoma cell line Br J Cancer 2001; 84(7): 951–8PubMedCrossRefGoogle Scholar
  18. 18.
    Berlin O, Samid D, Donthineni-Rao R et al. Development of a novel spontaneous metastasis model of human osteosarcoma transplanted orthotopically into bone of athymic mice Cancer Res 1993; 3(20): 4890–5Google Scholar
  19. 19.
    Aznavoorian S, Murphy AN, Stetler-Stevenson WG, Liotta LA Molecular aspects of tumour cell invasion and metastasis Cancer 1993; 71(4): 1368–83PubMedCrossRefGoogle Scholar
  20. 20.
    Reich R, Thompson EW, Iwamoto Y et al. Effects of inhibitors of plasminogen activator, serine proteinases, and collagenase IV on the invasion of basement membranes by metastatic cells Cancer Res 1981; 48(12): 3307–12Google Scholar
  21. 21.
    Wang Y The role and regulation of urokinase-type plasminogen activator receptor gene expression in cancer invasion and metastasis Med Res Rev 2001; 21(2): 146–70PubMedCrossRefGoogle Scholar
  22. 22.
    Kook YH, Adamski J, Zelent A, Ossowski L The effect of antisense inhibition of urokinase receptor in human squamous cell carcinoma on malignancy EMBO J 1994; 13(17): 3983–91PubMedGoogle Scholar
  23. 23.
    Mohanam S, Chintala SK, Go Y et al. In vitro inhibition of human glioblastoma cell line invasiveness by antisense uPA receptor Oncogene 1997; 14(11): 1351–9PubMedCrossRefGoogle Scholar
  24. 24.
    Lakka SS, Rajagopal R, Rajan MK et al. Adenovirus-mediated antisense urokinase-type plasminogen activator receptor gene transfer reduces tumour cell invasion and metastasis in non-small cell lung cancer cell lines Clin Cancer Res 2001; 7(4): 1087–93PubMedGoogle Scholar
  25. 25.
    Wang Y, Liang X, Wu S et al. Inhibition of colon cancer metastasis by a 3′-end antisense urokinase receptor mRNA in a nude mouse model Int J Cancer 2001; 92(2): 257–62PubMedCrossRefGoogle Scholar
  26. 26.
    Reuning U, Magdolen V, Wilhelm O et al. Multifunctional potential of the plasminogen activation system in tumour invasion and metastasis Int J Oncol 1998; 13(5): 893–906PubMedGoogle Scholar
  27. 27.
    Kusch A, Tkachuk S, Haller H et al. Urokinase stimulates human vascular smooth muscle cell migration via a phosphatidylinositol 3-kinase-Tyk2 interaction J Biol Chem 2000; 275(50): 39466–73PubMedCrossRefGoogle Scholar
  28. 28.
    Nguyen DH, Webb DJ, Catling AD et al. Urokinase-type plasminogen activator stimulates the Ras/extracellular signal-regulated kinase (ERK) signalling pathway and MCF-7 cell migration by a mechanism that requires focal adhesion kinase, Src, and Shc J Biol Chem 2000; 275(25): 19382–8PubMedCrossRefGoogle Scholar
  29. 29.
    Kjoller L, Hall A Rac mediates cytoskeletal rearrangements and increased cell motility induced by urokinase-type plasminogen activator receptor binding to vitronectin J Cell Biol 2001; 152(6): 1145–57PubMedCrossRefGoogle Scholar
  30. 30.
    Goncharova EA, Vorotnikov AV, Gracheva EO et al. Activation of p38 MAP-kinase and caldesmon phosphorylation are essential for urokinase-induced human smooth muscle cell migration Biol Chem 2002; 383(1): 115–26PubMedCrossRefGoogle Scholar
  31. 31.
    Adachi Y, Lakka SS, Chandrasekar N et al. Down-regulation of integrin alpha(v)beta(3) expression and integrin-mediated signalling in glioma cells by adenovirus-mediated transfer of antisense urokinase-type plasminogen activator receptor (uPAR) and sense p16 genes J Biol Chem 2001; 276(50): 47171–7PubMedCrossRefGoogle Scholar
  32. 32.
    Resnati M, Guttinger M, Valcamonica S et al. Proteolytic cleavage of the urokinase receptor substitutes for the agonist-induced chemotactic effect EMBO J 1996; 15(7): 1572–82PubMedGoogle Scholar
  33. 33.
    Go Y, Chintala SK, Mohanam S et al. Inhibition of in vivo tumorigenicity and invasiveness of a human glioblastoma cell line transfected with antisense uPAR vectors Clin Exp Metastasis 1997; 15(4): 440–6PubMedCrossRefGoogle Scholar
  34. 34.
    Yang JL, Seetoo D, Wang Y et al. Urokinase-type plasminogen activator and its receptor in colorectal cancer: independent prognostic factors of metastasis and cancer-specific survival and potential therapeutic targets Int J Cancer 2000; 89(5): 431–9PubMedCrossRefGoogle Scholar
  35. 35.
    Quan GM, Ojaimi J, Nadesapillai AP et al. Resistance of epiphyseal cartilage to invasion by osteosarcoma is likely to be due to expression of antiangiogenic factors Pathobiology 2003; 70(6): 361–7CrossRefGoogle Scholar
  36. 36.
    Carmeliet P, Bouche A, De Clercq C et al. Biological effects of disruption of the tissue-type plasminogen activator, urokinase-type plasminogen activator, and plasminogen activator inhibitor-1 genes in mice Ann NY Acad Sci 1995; 748: 367–82PubMedCrossRefGoogle Scholar
  37. 37.
    Dewerchin M, Nuffelen AV, Wallays G et al. Generation and characterization of urokinase receptor-deficient mice J Clin Invest 1996; 97(3): 870–8PubMedCrossRefGoogle Scholar
  38. 38.
    Dass CR Oligonucleotide delivery to tumours using macromolecular carriers Biotech Appl Biochem, 2004 Oct; 40(Pt 2): 113–22Google Scholar

Copyright information

© Springer Science+Business Media, Inc. 2006

Authors and Affiliations

  • Crispin R. Dass
    • 1
  • Anne P. W. Nadesapillai
    • 1
  • Daniel Robin
    • 1
  • Monique L. Howard
    • 1
  • Jane L. Fisher
    • 2
  • Hong Zhou
    • 3
  • Peter F. M. Choong
    • 1
    • 4
  1. 1.Department of OrthopaedicsThe University of Melbourne, St. Vincent’s Hospital MelbourneFitzroyAustralia
  2. 2.St. Vincent’s Institute of Medical ResearchFitzroyAustralia
  3. 3.Department of MedicineThe University of Melbourne, St. Vincent’s Hospital MelbourneFitzroyAustralia
  4. 4.Division of Surgical OncologyPeter MacCallum Cancer InstituteEast MelbourneAustralia

Personalised recommendations