Annals of Surgical Oncology

, Volume 13, Issue 8, pp 1145–1155 | Cite as

Selective Blockade of Vascular Endothelial Growth Factor Receptor 2 With an Antibody Against Tumor-Derived Vascular Endothelial Growth Factor Controls the Growth of Human Pancreatic Adenocarcinoma Xenografts

  • Shane E. Holloway
  • Adam W. Beck
  • Latha Shivakumar
  • Jessica Shih
  • Jason B. Fleming
  • Rolf A. Brekken



Vascular endothelial growth factor (VEGF), a key regulator of angiogenesis, is critical for growth of human pancreatic adenocarcinoma. Preclinical studies demonstrate that blockade of VEGF activity can control the growth of pancreatic tumors in mice. In this study, we evaluated the efficacy of 2C3, an antibody that inhibits VEGF receptor 2 activation by human VEGF, to inhibit the growth of human pancreatic adenocarcinoma in mice.


Human pancreatic cancer cell lines (MiaPaca-2, Panc-1, and Capan-1) were used to establish xenografts in nu/nu mice. The expression of VEGF and its receptors was determined in each cell line. Proliferation of tumor cells in vitro and tumor growth in vivo in the presence of 2C3 or a control antibody was evaluated. The effect of 2C3 on tumor weight, total vessel density, number of pericyte-associated vessels, and tumor perfusion was determined, and the level of 2C3 in the serum of animals was measured by enzyme-linked immunosorbent assay.


2C3 did not affect the proliferation of cells in culture. 2C3 was present and active in the serum of tumor-bearing animals treated with 2C3, and these animals showed a decrease in tumor burden compared with control-treated mice. Therapy with 2C3 resulted in reduced vascular function, measured by a decrease in vessel density and in the percentage of vessels associated with pericytes. Furthermore, tumors derived from Capan-1 cells demonstrated decreased perfusion after treatment with 2C3.


Blockade of VEGF receptor 2 activation by tumor-derived VEGF decreases tumor vessel function and growth of some human pancreatic adenocarcinoma cell lines in mice.


Pancreatic cancer Angiogenesis VEGF Therapy Monoclonal antibody 


  1. 1.
    Jemal A, Clegg LX, Ward E, et al. Annual report to the nation on the status of cancer, 1975-2001, with a special feature regarding survival. Cancer 2004;101:3–27PubMedCrossRefGoogle Scholar
  2. 2.
    Solorzano CC, Baker CH, Bruns CJ, et al. Inhibition of growth and metastasis of human pancreatic cancer growing in nude mice by PTK 787/ZK222584, an inhibitor of the vascular endothelial growth factor receptor tyrosine kinases. Cancer Biother Radiopharm 2001;16:359–70PubMedCrossRefGoogle Scholar
  3. 3.
    Bergers G, Benjamin LE. Tumorigenesis and the angiogenic switch. Nat Rev Cancer 2003;3:401–10PubMedCrossRefGoogle Scholar
  4. 4.
    Hicklin DJ, Ellis LM. Role of the vascular endothelial growth factor pathway in tumor growth and angiogenesis. J Clin Oncol 2005;23:1011–27PubMedCrossRefGoogle Scholar
  5. 5.
    Ferrara N. Vascular endothelial growth factor: basic science and clinical progress. Endocr Rev 2004;25:581–611PubMedCrossRefGoogle Scholar
  6. 6.
    Luo J, Guo P, Matsuda K, et al. Pancreatic cancer cell-derived vascular endothelial growth factor is biologically active in vitro and enhances tumorigenicity in vivo. Int J Cancer 2001;92:361–9PubMedCrossRefGoogle Scholar
  7. 7.
    Itakura J, Ishiwata T, Shen B, Kornmann M, Korc M. Concomitant over-expression of vascular endothelial growth factor and its receptors in pancreatic cancer. Int J Cancer 2000;85:27–34PubMedCrossRefGoogle Scholar
  8. 8.
    Niedergethmann M, Hildenbrand R, Wostbrock B, et al. High expression of vascular endothelial growth factor predicts early recurrence and poor prognosis after curative resection for ductal adenocarcinoma of the pancreas. Pancreas 2002;25:122–9PubMedCrossRefGoogle Scholar
  9. 9.
    Bruns CJ, Shrader M, Harbison MT, et al. Effect of the vascular endothelial growth factor receptor-2 antibody DC101 plus gemcitabine on growth, metastasis and angiogenesis of human pancreatic cancer growing orthotopically in nude mice. Int J Cancer 2002;102:101–8PubMedCrossRefGoogle Scholar
  10. 10.
    Bockhorn M, Tsuzuki Y, Xu L, Frilling A, Broelsch CE, Fukumura D. Differential vascular and transcriptional responses to anti-vascular endothelial growth factor antibody in orthotopic human pancreatic cancer xenografts. Clin Cancer Res 2003;9:4221–6PubMedGoogle Scholar
  11. 11.
    McBride G. Researchers optimistic about targeted drugs for pancreatic cancer. J Natl Cancer Inst 2004;96:1570–2PubMedCrossRefGoogle Scholar
  12. 12.
    Kindler HL, Friberg G, Stadler WM, et al. Bevacizumab (B) plus gemcitabine (G) in patient (pts) with advanced pancreatic cancer (PC): updated results of a multi-center phase II trial. J Clin Oncol (Meeting Abstracts) 2004;22(14 Suppl):4009Google Scholar
  13. 13.
    Brekken RA, Huang X, King SW, Thorpe PE. Vascular endothelial growth factor as a marker of tumor endothelium. Cancer Res 1998;58:1952–9PubMedGoogle Scholar
  14. 14.
    Brekken RA, Overholser JP, Stastny VA, Waltenberger J, Minna JD, Thorpe PE. Selective inhibition of vascular endothelial growth factor (VEGF) receptor 2 (KDR/Flk-1) activity by a monoclonal anti-VEGF antibody blocks tumor growth in mice. Cancer Res 2000;60:5117–24PubMedGoogle Scholar
  15. 15.
    Zhang W, Ran S, Sambade M, Huang X, Thorpe PE. A monoclonal antibody that blocks VEGF binding to VEGFR2 (KDR/Flk-1) inhibits vascular expression of Flk-1 and tumor growth in an orthotopic human breast cancer model. Angiogenesis 2002;5:35–44PubMedCrossRefGoogle Scholar
  16. 16.
    Stephan S, Datta K, Wang E, et al. Effect of rapamycin alone and in combination with antiangiogenesis therapy in an orthotopic model of human pancreatic cancer. Clin Cancer Res 2004;10:6993–7000PubMedCrossRefGoogle Scholar
  17. 17.
    Kern SE. Advances from genetic clues in pancreatic cancer. Curr Opin Oncol 1998;10:74–80PubMedGoogle Scholar
  18. 18.
    Rozenblum E, Schutte M, Goggins M, et al. Tumor-suppressive pathways in pancreatic carcinoma. Cancer Res 1997;57:1731–4PubMedGoogle Scholar
  19. 19.
    Brekken RA, Puolakkainen P, Graves DC, Workman G, Lubkin SR, Sage EH. Enhanced growth of tumors in SPARC null mice is associated with changes in the ECM. J Clin Invest 2003;111:487–95PubMedCrossRefGoogle Scholar
  20. 20.
    Hallmann R, Mayer DN, Berg EL, Broermann R, Butcher EC. Novel mouse endothelial cell surface marker is suppressed during differentiation of the blood brain barrier. Dev Dyn 1995;202:325–32PubMedGoogle Scholar
  21. 21.
    Feng D, Nagy JA, Brekken RA, et al. Ultrastructural localization of the vascular permeability factor/vascular endothelial growth factor (VPF/VEGF) receptor-2 (FLK-1, KDR) in normal mouse kidney and in the hyperpermeable vessels induced by VPF/VEGF-expressing tumors and adenoviral vectors. J Histochem Cytochem 2000;48:545–56PubMedGoogle Scholar
  22. 22.
    Goertz DE, Yu JL, Kerbel RS, Burns PN, Foster FS. High-frequency Doppler ultrasound monitors the effects of antivascular therapy on tumor blood flow. Cancer Res 2002;62:6371–5PubMedGoogle Scholar
  23. 23.
    Parikh AA, Liu WB, Fan F, et al. Expression and regulation of the novel vascular endothelial growth factor receptor neuropilin-1 by epidermal growth factor in human pancreatic carcinoma. Cancer 2003;98:720–9PubMedCrossRefGoogle Scholar
  24. 24.
    Folkman J. What is the evidence that tumors are angiogenesis dependent? J Natl Cancer Inst 1990;82:4–6PubMedGoogle Scholar
  25. 25.
    Reinmuth N, Parikh AA, Ahmad SA, et al. Biology of angiogenesis in tumors of the gastrointestinal tract. Microsc Res Tech 2003;60:199–207PubMedCrossRefGoogle Scholar
  26. 26.
    Robinson CJ, Stringer SE. The splice variants of vascular endothelial growth factor (VEGF) and their receptors. J Cell Sci 2001;114(Pt 5):853–65PubMedGoogle Scholar
  27. 27.
    Ahmed SI, Thomas AL, Steward WP. Vascular endothelial growth factor (VEGF) inhibition by small molecules. J Chemother 2004;16(Suppl 4):59–63PubMedGoogle Scholar
  28. 28.
    Ruggeri B, Singh J, Gingrich D, et al. CEP-7055: a novel, orally active pan inhibitor of vascular endothelial growth factor receptor tyrosine kinases with potent antiangiogenic activity and antitumor efficacy in preclinical models. Cancer Res 2003;63:5978–91PubMedGoogle Scholar
  29. 29.
    Solorzano CC, Baker CH, Bruns CJ, et al. Inhibition of growth and metastasis of human pancreatic cancer growing in nude mice by PTK 787/ZK222584, an inhibitor of the vascular endothelial growth factor receptor tyrosine kinases. Cancer Biother Radiopharm 2001;16:359–70PubMedCrossRefGoogle Scholar
  30. 30.
    Kim KJ, Li B, Winer J, et al. Inhibition of vascular endothelial growth factor-induced angiogenesis suppresses tumour growth in vivo. Nature 1993;362:841–4PubMedCrossRefGoogle Scholar
  31. 31.
    Ferrara N, Hillan KJ, Gerber HP, Novotny W. Discovery and development of bevacizumab, an anti-VEGF antibody for treating cancer. Nat Rev Drug Discov 2004;3:391–400PubMedCrossRefGoogle Scholar
  32. 32.
    Presta LG, Chen H, O’Connor SJ, et al. Humanization of an anti-vascular endothelial growth factor monoclonal antibody for the therapy of solid tumors and other disorders. Cancer Res 1997;57:4593–9PubMedGoogle Scholar
  33. 33.
    Tammela T, Enholm B, Alitalo K, Paavonen K. The biology of vascular endothelial growth factors. Cardiovasc Res 2005;65:550–63PubMedCrossRefGoogle Scholar
  34. 34.
    Tong RT, Boucher Y, Kozin SV, Winkler F, Hicklin DJ, Jain RK. Vascular normalization by vascular endothelial growth factor receptor 2 blockade induces a pressure gradient across the vasculature and improves drug penetration in tumors. Cancer Res 2004;64:3731–6PubMedCrossRefGoogle Scholar
  35. 35.
    Shaheen RM, Tseng WW, Vellagas R, et al. Effects of an antibody to vascular endothelial growth factor receptor-2 on survival, tumor vascularity, and apoptosis in a murine model of colon carcinomatosis. Int J Oncol 2001;18:221–6PubMedGoogle Scholar
  36. 36.
    Fong GH, Rossant J, Gertsenstein M, Breitman ML. Role of the Flt-1 receptor tyrosine kinase in regulating the assembly of vascular endothelium. Nature 1995;376:66–70PubMedCrossRefGoogle Scholar
  37. 37.
    Fong GH, Zhang L, Bryce DM, Peng J. Increased hemangioblast commitment, not vascular disorganization, is the primary defect in flt-1 knock-out mice. Development 1999;126:3015–25PubMedGoogle Scholar
  38. 38.
    Hiratsuka S, Minowa O, Kuno J, Noda T, Shibuya M. Flt-1 lacking the tyrosine kinase domain is sufficient for normal development and angiogenesis in mice. Proc Natl Acad Sci U S A 1998;95:9349–54PubMedCrossRefGoogle Scholar
  39. 39.
    Luttun A, Autiero M, Tjwa M, Carmeliet P. Genetic dissection of tumor angiogenesis: are PlGF and VEGFR-1 novel anti-cancer targets? Biochim Biophys Acta 2004;1654:79–94PubMedGoogle Scholar
  40. 40.
    Hattori K, Heissig B, Wu Y, et al. Placental growth factor reconstitutes hematopoiesis by recruiting VEGFR1(+) stem cells from bone-marrow microenvironment. Nat Med 2002;8:841–9PubMedGoogle Scholar
  41. 41.
    Clauss M, Weich H, Breier G, et al. The vascular endothelial growth factor receptor Flt-1 mediates biological activities. Implications for a functional role of placenta growth factor in monocyte activation and chemotaxis. J Biol Chem 1996;271:17629–34PubMedCrossRefGoogle Scholar
  42. 42.
    Kim KJ, Li B, Houck K, Winer J, Ferrara N. The vascular endothelial growth factor proteins: identification of biologically relevant regions by neutralizing monoclonal antibodies. Growth Factors 1992;7:53–64PubMedGoogle Scholar
  43. 43.
    Rahimi N, Dayanir V, Lashkari K. Receptor chimeras indicate that the VEGFR-1 modulates mitogenic activity of VEGFR-2 in endothelial cells. J Biol Chem 2000; 275:16986–92PubMedCrossRefGoogle Scholar
  44. 44.
    Dunk C, Ahmed A. Vascular endothelial growth factor receptor-2-mediated mitogenesis is negatively regulated by vascular endothelial growth factor receptor-1 in tumor epithelial cells. Am J Pathol 2001;158:265–73PubMedGoogle Scholar
  45. 45.
    Yoshiji H, Harris SR, Thorgeirsson UP. Vascular endothelial growth factor is essential for initial but not continued in vivo growth of human breast carcinoma cells. Cancer Res 1997;57:3924–8PubMedGoogle Scholar
  46. 46.
    Kiessling F, Farhan N, Lichy MP, et al. Dynamic contrast-enhanced magnetic resonance imaging rapidly indicates vessel regression in human squamous cell carcinomas grown in nude mice caused by VEGF receptor 2 blockade with DC101. Neoplasia 2004;6:213–23PubMedCrossRefGoogle Scholar
  47. 47.
    Ljungkvist AS, Bussink J, Rijken PF, Kaanders JH, van der Kogel AJ, Denekamp J. Vascular architecture, hypoxia, and proliferation in first-generation xenografts of human head-and-neck squamous cell carcinomas. Int J Radiat Oncol Biol Phys 2002;54:215–28PubMedCrossRefGoogle Scholar

Copyright information

© The Society of Surgical Oncology, Inc. 2006

Authors and Affiliations

  • Shane E. Holloway
    • 1
  • Adam W. Beck
    • 1
  • Latha Shivakumar
    • 1
  • Jessica Shih
    • 1
  • Jason B. Fleming
    • 1
  • Rolf A. Brekken
    • 1
    • 2
  1. 1.Department of Surgery, Division of Surgical Oncology, and Hamon Center for Therapeutic Oncology ResearchUniversity of Texas Southwestern Medical CenterDallasUSA
  2. 2.Department of PharmacologyUniversity of Texas Southwestern Medical CenterDallasUSA

Personalised recommendations