Skip to main content

Predicting the Effects of Anti-angiogenic Agents Targeting Specific VEGF Isoforms

Abstract

Vascular endothelial growth factor (VEGF) is a key mediator of angiogenesis, whose effect on cancer growth and development is well characterized. Alternative splicing of VEGF leads to several different isoforms, which are differentially expressed in various tumor types and have distinct functions in tumor blood vessel formation. Many cancer therapies aim to inhibit angiogenesis by targeting VEGF and preventing intracellular signaling leading to tumor vascularization; however, the effects of targeting specific VEGF isoforms have received little attention in the clinical setting. In this work, we investigate the effects of selectively targeting a single VEGF isoform, as compared with inhibiting all isoforms. We utilize a molecular-detailed whole-body compartment model of VEGF transport and kinetics in the presence of breast tumor. The model includes two major VEGF isoforms, VEGF121 and VEGF165, receptors VEGFR1 and VEGFR2, and co-receptors Neuropilin-1 and Neuropilin-2. We utilize the model to predict the concentrations of free VEGF, the number of VEGF/VEGFR2 complexes (considered to be pro-angiogenic), and the receptor occupancy profiles following inhibition of VEGF using isoform-specific anti-VEGF agents. We predict that targeting VEGF121 leads to a 54% and 84% reduction in free VEGF in tumors that secrete both VEGF isoforms or tumors that overexpress VEGF121, respectively. Additionally, 21% of the VEGFR2 molecules in the blood are ligated following inhibition of VEGF121, compared with 88% when both isoforms are targeted. Targeting VEGF121 reduces tumor free VEGF and is an effective treatment strategy. Our results provide a basis for clinical investigation of isoform-specific anti-VEGF agents.

This is a preview of subscription content, access via your institution.

Fig. 1
Fig. 2
Fig. 3
Fig. 4
Fig. 5
Fig. 6

References

  1. Rennel ES, Harper SJ, Bates DO. Therapeutic potential of manipulating VEGF splice isoforms in oncology. Futur Oncol. 2009;5(5):703–12.

    Article  CAS  Google Scholar 

  2. Bates DO, Cui T-G, Doughty JM, Winkler M, Sugiono M, Shields JD, et al. VEGF165b, and inhibitory splice variant of vascular endothelial growth factor, is down-regulated in renal cell carcinoma. Cancer Res. 2002;62:4123–31.

    PubMed  CAS  Google Scholar 

  3. Nowak DG, Woolard J, Amin EM, Konopatskaya O, Saleem MA, Churchill AJ, et al. Expression of pro- and anti-angiogenic isoforms of VEGF is differentially regulated by splicing and growth factors. J Cell Sci. 2008;121:3487–95.

    PubMed  Article  CAS  Google Scholar 

  4. Dokun AO, Annex BH. The VEGF165b "ICE-o-form" puts a chill on the VEGF story. Circul Res. 2011;109:246–7.

    Article  CAS  Google Scholar 

  5. Manetti M, Guiducci S, Romano E, Ceccarelli C, Bellando-Randone S, Conforti ML, et al. Overexpression of VEGF165b, an inhibitory splice variant of vascular endothelial growth factor, leads to insufficient angiogenesis in patients with systemic sclerosis. Circul Res. 2011;109:e14–26.

    Article  CAS  Google Scholar 

  6. Catena R, Larzabal L, Larrayoz M, Molina E, Hermida J, Agorreta J, et al. VEGF121b and VEGF165b are weakly angiogenic isoforms of VEGF-A. Mol Cancer. 2010;9:320.

    PubMed  Article  CAS  Google Scholar 

  7. Soker S, Miao H-Q, Nomi M, Takashima S, Klagsbrun M. VEGF165 mediates formation of complexes containing VEGFR-2 and neuropilin-1 that enhance VEGF165-receptor binding. J Cell Biochem. 2002;85(2):357–68.

    PubMed  Article  CAS  Google Scholar 

  8. Fuh G, Garcia KC, De Vos AM. The interaction of neuropilin-1 with vascular endothelial growth factor and its receptor flt-1. J Biol Chem. 2000;275:26690–5.

    PubMed  CAS  Google Scholar 

  9. Park JE, Keller G-A, Ferrara N. The vascular endothelial growth factor (VEGF) isoforms: differential deposition into the subepithelial extracellular matrix and bioactivity of extracellular matrix-bound VEGF. Mol Biol Cell. 1993;4:1317–26.

    PubMed  CAS  Google Scholar 

  10. Houck K, Leung DW, Rowland AM, Winer J, Ferrara N. Dual regulation of vascular endothelial growth factor bioavailability by genetic and proteolytic mechanisms. J Biol Chem. 1992;268(36):26031–7.

    Google Scholar 

  11. Grunstein J, Masbad JJ, Hickey R, Giordano F, Johnson RS. Isoforms of vascular endothelial growth factor act in a coordinate fashion to recruit and expand tumor vasculature. Mol Cell Biol. 2000;20(19):7292–1.

    Article  Google Scholar 

  12. Tozer GM, Akerman S, Cross NA, Barber PR, Bjorndahl MA, Greco O, et al. Blood vessel maturation and response to vascular-disrupting therapy in single vascular endothelial growth factor-A isoform-producing tumors. Cancer Res. 2008;68:2301–11.

    PubMed  Article  CAS  Google Scholar 

  13. Yu JL, Rak JW, Klement G, Kerbel RS. Vascular endothelial growth factor isoform expression as a determinant of blood vessel patterning in human melanoma xenografts. Cancer Res. 2002;62:1838–46.

    PubMed  CAS  Google Scholar 

  14. Yuan A, Yu C-J, Kuo S-H, Chen W-J, Lin F-Y, Luh K-T, et al. Vascular endothelial growth factor 189 mRNA isoform expression specifically correlates with tumor angiogenesis, patient survival, and postoperative relapse in non-small-cell lung cancer. J Clin Oncol. 2001;19(2):432–41.

    PubMed  CAS  Google Scholar 

  15. Cheng S-Y, Nagane M, Huang H-JS, Cavenee WK. Intracerebral tumor-associated hemorrhage caused by overexpression of the vascular endothelial growth factor isoforms VEGF121 and VEGF165 but not VEGF189. Proc Natl Acad Sci U S A. 1997;94(22):12081–7.

    PubMed  Article  CAS  Google Scholar 

  16. Yuan A, Lin C-Y, Chou C-H, Shih C-M, Chen C-Y, Cheng H-W, et al. Functional and structural characteristics of tumor angiogenesis in lung cancers overexpressing different VEGF isoforms assessed by DCE- and SSCE-MRI. PLoS One. 2011;6(1):e16062.

    PubMed  Article  CAS  Google Scholar 

  17. Tokunaga T, Oshika Y, Abe Y, Ozeki Y, Sadahiro S, Kijima H, et al. Vascular endothelial growth factor (VEGF) mRNA isoform expression pattern is correlated with liver metastasis and poor prognosis in colon cancer. Br J Cancer. 1998;77(6):998–1002.

    PubMed  Article  CAS  Google Scholar 

  18. Fenton BM, Paoni SF, Liu W, Cheng S-Y, Hu B, Ding I. Overexpression of VEGF121, but not VEGF165 or FGF-1, improves oxygenation in MCF-7 breast tumors. Br J Cancer. 2004;90(2):430–5.

    PubMed  Article  CAS  Google Scholar 

  19. Verhoeff JJC, Stalpers LJA, Claes A, Hovinga KE, Musters GD, Vandertop WP, et al. Tumour control by whole brain irradiation of anti-VEGF-treated mice bearing intracerebral glioma. Eur J Cancer. 2009;45(17):3074–80.

    PubMed  Article  CAS  Google Scholar 

  20. Ruckman J, Green LS, Beeson J, Waugh S, Gillette WL, Henninger DD, et al. 2′-Fluoropyrimidine RNA-based aptamers to the 165-amino acid form of vascular endothelial growth factor (VEGF165). Inhibition of receptor binding and VEGF-induced vascular permeability through interactions requiring the exon 7-encoded domain. J Biol Chem. 1998;273:20556–67.

    PubMed  Article  CAS  Google Scholar 

  21. Ng EWM, Shima DT, Calias P, Cunningham Jr ET, Guyer DR, Adamis AP. Pegaptanib, a targeted anti-VEGF aptamer for ocular vascular disease. Nat Rev Drug Discov. 2006;5:123–32.

    PubMed  Article  CAS  Google Scholar 

  22. Ishida S, Usui T, Yamashiro K, Kaji Y, Amano S, Ogura Y, et al. VEGF164-mediated inflammation is required for pathological, but not physiological, ischemia-induced retinal neovascularization. J Exp Med. 2003;198(3):483–9.

    PubMed  Article  CAS  Google Scholar 

  23. Arrell DK, Terzic A. Network systems biology for drug discovery. Clin Pharmacol Ther. 2010;8(1):120–5.

    Article  Google Scholar 

  24. Laubenbacher R, Hower V, Jarrah A, Torti SV, Shulaev V, Mendes P, et al. A systems biology view of cancer. Biochimica et Biophysica Acta (BBA) - Reviews on Cancer. 2009;1796(2):129–39.

    Article  CAS  Google Scholar 

  25. Finley SD, Engel-Stefanini MO, Imoukhuede PI, Popel AS. Pharmacokinetics and pharmacodynamics of VEGF-neutralizing antibodies. BMC Syst Biol. 2011;5:193.

    PubMed  Article  CAS  Google Scholar 

  26. Stefanini MO, Wu FTH, Mac Gabhann F, Popel AS. Increase of plasma VEGF after intravenous administration of bevacizumab is predicted by a pharmacokinetic model. Cancer Res. 2010;70(23):9886–94.

    PubMed  Article  CAS  Google Scholar 

  27. Padera TP, Kadambi A, Di Tomaso E, Carreira CM, Brown EB, Boucher Y, et al. Lymphatic metastasis in the absence of functional intratumor lymphatics. Science. 2002;296(5574):1883–6.

    PubMed  Article  CAS  Google Scholar 

  28. Leu AJ, Berk DA, Lymboussaki A, Alitalo K, Jain RK. Absence of functional lymphatics within a murine sarcoma: a molecular and functional evaluation. Cancer Res. 2000;60:4324–7.

    PubMed  CAS  Google Scholar 

  29. Imoukhuede PI, Popel AS. Quantification and cell-to-cell variation of vascular endothelial growth factor receptors. Exp Cell Res. 2011;317(7):955–65.

    PubMed  Article  CAS  Google Scholar 

  30. Genentech, Inc. Avastin prescribing information [cited September2011]; Available from: http://www.avastin.com/avastin/hcp/overview/about/dosing/index.html.

  31. Gordon MS, Margolin K, Talpaz M, Sledge Jr GW, Holmgren E, Benjamin R, et al. Phase I safety and pharmacokinetic study of recombinant human anti-vascular endothelial growth factor in patients with advanced cancer. J Clin Oncol. 2001;19(3):843–50.

    PubMed  CAS  Google Scholar 

  32. Hsei V, DeGuzman GG, Nixon A, Gaudreault J. Complexation of VEGF with bevacizumab decreases VEGF clearance in rats. Pharm Res. 2002;19(11):1753–6.

    PubMed  Article  CAS  Google Scholar 

  33. Liang W-C, Wu X, Peale FV, Lee CV, Meng YG, Gutierrez J, et al. Cross-species vascular endothelial growth factor (VEGF)-blocking antibodies completely inhibit the growth of human tumor xenografts and measure the contribution of stromal VEGF. J Biol Chem. 2006;281:951–61.

    PubMed  Article  CAS  Google Scholar 

  34. Guo P, Xu L, Pan S, Brekken RA, Yang S-T, Whitaker GB, et al. Vascular endothelial growth factor isoforms display distinct activities in promoting tumor angiogenesis at different anatomic sites. Cancer Res. 2001;61:8569–77.

    PubMed  CAS  Google Scholar 

  35. Catena R, Muniz-Medina V, Moralejo B, Javierre B, Best CJM, Emmert-Buck MR, et al. Increased expression of VEGF121/VEGF165-189 ratio results in a significant enhancement of human prostate tumor angiogenesis. Int J Cancer. 2007;120:2096–109.

    PubMed  Article  CAS  Google Scholar 

  36. Zhang H-T, Scot PAE, Morbidelli L, Peak S, Moore J, Turley H, et al. The 121 amino acid isoform of vascular endothelial growth factor is more strongly tumorigenic than other splice variants in vivo. Br J Cancer. 2000;83(1):63–8.

    PubMed  Article  CAS  Google Scholar 

  37. Stimpfl M, Tong D, Fasching B, Schuster E, Obermair A, Leodolter S, et al. Vascular endothelial growth factor splice variants and their prognostic value in breast and ovarian cancer. Clin Cancer Res. 2002;8(7):2253–9.

    PubMed  CAS  Google Scholar 

  38. Yuan A, Yu CJ, Luh KT, Lin FY, Kuo SH, Yang PC. Quantification of VEGF mRNA expression in non-small cell lung cancer using a real-time quantitative reverse transcription-PCR assay and a comparison with quantitative competitive reverse transcription-PCR. Lab Invest. 2000;2000(80):11.

    Google Scholar 

  39. Cheung N, Wong MP, Yuen ST, Leung SY, Chung LP. Tissue-specific expression pattern of vascular endothelial growth factor isoforms in the malignant transformation of lung and colon. Hum Pathol. 1998;29(9):910–4.

    PubMed  Article  CAS  Google Scholar 

  40. Ljungberg B, Jacobsen J, Haggstrom-Rudolfssson S, Rasmuson T, Lindh G, Grankvist K. Tumor vascular endothelial growth factor (VEGF) mRNA in relation to serum VEGF protein levels and tumour progression in human renal cell carcinoma. Urol Res. 2003;31(5):335–40.

    PubMed  Article  CAS  Google Scholar 

  41. Zygalaki E, Tsaroucha EG, Kaklamanis L, Lianidou ES. Quantitative real-time reverse transcription-PCR study of the expression of vascular endothelial growth factor (VEGF) splice variants and VEGF receptors (VEGFR-1 and VEGFR-2) in non-small cell lung cancer. Clin Chem. 2007;53(8):1433–9.

    PubMed  Article  CAS  Google Scholar 

  42. Hata K, Watanabe Y, Nakai H, Hata T, Hoshiai H. Expression of the vascular endothelial growth factor (VEGF) gene in epithelial ovarian cancer: an approach to anti-VEGF therapy. Anticancer Res. 2011;31(2):731–7.

    PubMed  CAS  Google Scholar 

  43. Wiig H, Tenstad O, Iversen PO, Kalluri R, Bjerkvig R. Interstitial fluid: the overlooked component of the tumor microenvironment? Fibrogenesis Tissue Repair. 2010;3:12.

    PubMed  Article  Google Scholar 

  44. Huang J, Moore J, Soffer S, Kim E, Rowe D, Manley CA, et al. Highly specific antiangiogenic therapy is effective in suppressing growth of experimental Wilms tumors. J Pediatr Surg. 2001;36:357–61.

    PubMed  Article  CAS  Google Scholar 

  45. Chidlow Jr JH, Glawe JD, Pattillo CB, Pardue S, Zhang S, Kevil CG. VEGF164 isoform specific regulation of T-cell-dependent experimental colitis in mice. Inflamm Bowel Dis. 2011;17:1501–12.

    PubMed  Article  Google Scholar 

  46. Sawada O, Kawamura H, Kakinoki M, Sawada T, Ohji M. Vascular endothelial growth factor in aqueous humor before and after intravitreal injection of bevacizumab in eyes with diabetic retinopathy. Arch Ophthalmol. 2007;125(10):1363–6.

    PubMed  Article  CAS  Google Scholar 

  47. Tian XJ, Wu J, Meng L, Dong ZW, Shou CC. Expression of VEGF121 in gastric carcinoma MGC803 cell line. World J Gastroenterol. 2000;6(2):281–3.

    CAS  Google Scholar 

  48. Guo P, Fang Q, Tao H-Q, Schafer CA, Fenton BM, Ding I, et al. Overexpression of vascular endothelial growth factor by MCF-7 breast cancer cells promotes estrogen-independent tumor growth in vivo. Cancer Res. 2003;63:4684–91.

    PubMed  CAS  Google Scholar 

Download references

Acknowledgments

The authors thank Gang Liu and Spyridon Stamatelos for helpful discussions. This work was supported by National Institutes of Health grant R01 CA138264 (ASP) and fellowship F32 CA154213 (SDF), and the UNCF/Merck Postdoctoral Fellowship (SDF).

Author information

Authors and Affiliations

Authors

Corresponding author

Correspondence to Stacey D. Finley.

Electronic supplementary material

Below is the link to the electronic supplementary material.

ESM 1

(PDF 101 kb)

ESM 2

(XML 318 kb)

ESM 3

(PDF 405 kb)

Rights and permissions

Reprints and Permissions

About this article

Cite this article

Finley, S.D., Popel, A.S. Predicting the Effects of Anti-angiogenic Agents Targeting Specific VEGF Isoforms. AAPS J 14, 500–509 (2012). https://doi.org/10.1208/s12248-012-9363-4

Download citation

  • Received:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1208/s12248-012-9363-4

Key words

  • angiogenesis
  • cancer drug target
  • computational model
  • pharmacokinetic model
  • systems biology