Neurochemical Research

, Volume 36, Issue 4, pp 655–667 | Cite as

Validation of Molecular and Genomic Biomarkers of Retinal Drug Efficacy: Use of Ocular Fluid Sampling to Evaluate VEGF



The use of tissue- and cell-based methods in developing drugs for retinal diseases is inefficient. Consequently, many aspects of ocular drug therapy for retinal diseases are poorly understood. Biomarkers as prognostic indicators of change are needed to optimize the use of drugs. VEGF is considered an important target of drug therapy and VEGF levels in tissue are indicative of solid tumor growth. However, since many aspects of VEGF as a biomarker of ocular disease have not been validated, it has been difficult to ascertain without invasive procedures whether VEGF in the eye is a biomarker of response to drug therapy. Using published papers, registered clinical trials, and proteomic databases we assessed the earlier evidence for VEGF as an exploratory biomarker of proliferative and vasculopathic disease of the retina and asked whether the molecule has been rigorously validated in clinical trials. The emerging use of aqueous humor sampling has made it possible to explore biomarkers in oculo, and determine whether they are predictive of drug efficacy. We present data supporting the use of aqueous humor to validate drug-signaling pathways and biomarkers in the eye. In addition, we recommend convening a collaborative congress to help standardize the identification, validation, and use of biomarkers in retinal disease.


Vascular endothelial growth factor Ocular Retinal disease Biomarkers Aqueous humor Validation 


  1. 1.
    US Department of Health and Human Services (2008) Guidance for industry: E15 definitions for genomic biomarkers, pharmacogenomics, pharmacogenetics, genomic data and sample coding categories. Published April 2008. Accessed 28 July 2010
  2. 2.
    National Collaborating Centre for Cancer (2009) Advanced breast cancer: diagnosis and treatment. London (UK): National Institute for Health and Clinical Excellence (NICE). Published February 2009. Accessed 7 Nov 2010
  3. 3.
    Siljander HT, Simell S, Hekkala A (2009) Predictive characteristics of diabetes-associated autoantibodies among children with HLA-conferred disease susceptibility in the general population. Diabetes 58:2835–2842PubMedCrossRefGoogle Scholar
  4. 4.
    Murukesh N, Dive C, Jayson GC (2010) Biomarkers of angiogenesis and their role in the development of VEGF inhibitors. Br J Cancer 102:8–18PubMedCrossRefGoogle Scholar
  5. 5.
    Goodsaid F, Frueh F (2007) Biomarker qualification pilot process at the US food and drug administration. AAPS J 9:E105–E108PubMedCrossRefGoogle Scholar
  6. 6.
    Atkinson AJ, Colburn WA, DeGruttola VG et al (2001) Biomarkers and surrogate endpoints: preferred definitions and conceptual framework. Clin Pharmacol Ther 69:89–95CrossRefGoogle Scholar
  7. 7.
    Shen J, Samul R, Silva RL (2006) Suppression of ocular neovascularization with siRNA targeting VEGF receptor 1: knockdown of vegfr1 mRNA inhibits ocular neovascularization. Gene Ther 13: 225–234Google Scholar
  8. 8.
    Shams N, Ianchulev Y (2006) Role of vascular endothelial growth factor in ocular angiogenesis. Ophthalmol Clin North Am 19:335–344PubMedGoogle Scholar
  9. 9.
    Ciulla TA, Rosenfeld PJ (2009) Anti-vascular endothelial growth factor therapy for neovascular age-related macular degeneration. Curr Opin Ophthalmol 20:158–165PubMedCrossRefGoogle Scholar
  10. 10.
    Tolentino MJ (2009) Current molecular understanding and future treatment strategies for pathologic ocular neovascularization. Curr Mol Med 9:973–981PubMedCrossRefGoogle Scholar
  11. 11.
    Rudge JS, Holash J, Hylton D et al (2007) VEGF Trap complex formation measures production rates of VEGF, providing a biomarker for predicting efficacious angiogenic blockade. PNAS 104:18363–18370PubMedCrossRefGoogle Scholar
  12. 12.
    Ng Ew, Adamis AP (2005) Targeting angiogenesis, the underlying disorder in noevascular age-related macular degeneration. Can J Ophthalmol 40:352–368PubMedGoogle Scholar
  13. 13.
    Ferrara N, Damic L, Sham N et al (2006) Development of ranibizumab, an anti-vascular endothelial growth factor antigen binding fragment, as therapy for noevascular age-related macular degeneration. Retina 26:859–870PubMedCrossRefGoogle Scholar
  14. 14.
    Vemulakonda GA, Hariprasad SM, Mieler WF et al (2008) Aqueous and vitreous concentrations following topical administration of 1% voriconazole in humans. Arch Ophthalmol 126(1):18–22PubMedCrossRefGoogle Scholar
  15. 15.
    Van der Lelij A, Rothova A (1997) Diagnositic anterior chamber paracentesis in uveitis: a safe procedure? Br J Ophthalmol 81:976–979PubMedCrossRefGoogle Scholar
  16. 16.
    Campochiaro PA, Choy DF, Do DV et al (2009) Monitoring ocular drug therapy by analysis of aqueous samples. Ophthalmol 116(11):2158–2164CrossRefGoogle Scholar
  17. 17.
    Sharma RK, Rogojina AT, Chalam KV (2009) Multiplex immunoassay analysis of biomarkers in clinically accessible quantities of human aqueous humor. Mol Vis 15:60–69PubMedGoogle Scholar
  18. 18.
    Regenfuss B, Bock F, Parthasarathy A, Cursiefen C (2008) Corneal (lymph)angiogenesis–from bedside to bench and back: a tribute to Judah Folkman. Lymphat Res Biol 6:191–201PubMedCrossRefGoogle Scholar
  19. 19.
    Cross MJ, Dixelius J, Matsumoto T, Claesson-Welsh L (2003) VEGF-receptor signal transduction. Tren Biochem Sci 28:488–494CrossRefGoogle Scholar
  20. 20.
    Shibuya M (2009) Unique signal transduction of the VEGF family members VEGF-A and VEGF-E. Biochem Soc Trans 37:1161–1166PubMedCrossRefGoogle Scholar
  21. 21.
    Ranibizumab Full Prescribing Information, Genentech, South San Francisco 2007Google Scholar
  22. 22.
    Pegaptanib, Full Prescribing Information, OSI-Pfizer, NY, NY, 2004Google Scholar
  23. 23.
    Wu FT, Stefanini MO, Mac Gabhann F, Kontos CD, Annex BH et al (2010) VEGF and soluble VEGF receptor-1 (sFlt-1) distributions in peripheral arterial disease: an in silico model. Am J Physiol Heart Circ Physiol 298(6):H2174–H2191PubMedCrossRefGoogle Scholar
  24. 24.
    Miller J, Adamis AP, Shima D, D’Amore PA, Moulton RS et al (1994) Vascular endothelial growth factor/vascular permeability factor is temporally and spatially correlated with ocular angiogenesis in a primate model. Am J Pathol 145:574–584PubMedGoogle Scholar
  25. 25.
    Lopez PF, Sippy BD, Lambert HM, Thach AB, Hinton DR (1996) Transdifferentiated retinal pigment epithelial cells are immunoreactive for vascular endothelial growth factor in surgically excised age-related macular degeneration-related choroidal neovascular membranes. Invest Ophthalmol Vis Sci 37:855–868PubMedGoogle Scholar
  26. 26.
    Ishida S, Usui T, Yamashiro K, Kaji Y, Amano S et al (2003) VEGF164-mediated inflammation is required for pathological, but not physiological, ischemia-induced retinal neovascularization. J Exp Med 198:483–489PubMedCrossRefGoogle Scholar
  27. 27.
    Funk M, Karl D, Georgopoulos M, Benesch T, Sacu S et al (2010) Neovascular age-related macular degeneration: intraocular cytokines and growth factors and the influence of therapy with ranibizumab. Ophthalmol 116:2393–2399CrossRefGoogle Scholar
  28. 28.
    Wells JA, Murthy R, Chibber R, Nunn A, Molinatti PA et al (1996) Levels of vascular endothelial growth factor are elevated in the vitreous of patients with subretinal neovascularisation. Br J Ophthalmol 80(4):363–366PubMedCrossRefGoogle Scholar
  29. 29.
    Itakura H, Kishi S, Kotajima N, Murakami M (2004) Persistent secretion of vascular endothelial growth factor into the vitreous cavity in proliferative diabetic retinopathy after vitrectomy. Ophthalmology 111:1880–1884PubMedCrossRefGoogle Scholar
  30. 30.
    Brooks HL Jr, Caballero S Jr, Newell CK, Steinmetz RL, Watson D et al (2004) Vitreous levels of vascular endothelial growth factor and stromal-derived factor 1 in patients with diabetic retinopathy and cystoid macular edema before and after intraocular injection of triamcinolone. Arch Ophthalmol 122:1801–1807PubMedCrossRefGoogle Scholar
  31. 31.
    Funatsu H, Yamashita H, Sakata K, Noma H, Mimura T et al (2005) Vitreous levels of vascular endothelial growth factor and intercellular adhesion molecule 1 are related to diabetic macular edema. Ophthalmology 112(5):806–816PubMedCrossRefGoogle Scholar
  32. 32.
    Funatsu H, Yamashita H, Nakamura S, Mimura T, Eguchi S et al (2006) Vitreous levels of pigment epithelium-derived factor and vascular endothelial growth factor are related to diabetic macular edema. Ophthalmology 113:294–301PubMedCrossRefGoogle Scholar
  33. 33.
    Noma H, Funatsu H, Mimura T, Harino S, Hori S (2009) Vitreous levels of interleukin-6 and vascular endothelial growth factor in macular edema with central retinal vein occlusion. Ophthalmology 116:87–93PubMedCrossRefGoogle Scholar
  34. 34.
    Watanabe D, Suzuma K, Suzuma I, Ohashi H, Ojima T et al (2005) Vitreous levels of angiopoietin 2 and vascular endothelial growth factor in patients with proliferative diabetic retinopathy. Am J Ophthalmol 139:476–481PubMedCrossRefGoogle Scholar
  35. 35.
    Ishizaki E, Takai S, Ueki M, Maeno T, Maruichi M et al (2006) Correlation between angiotensin-converting enzyme, vascular endothelial growth factor, and matrix metalloproteinase-9 in the vitreous of eyes with diabetic. Am J Ophthalmol 141:129–134PubMedCrossRefGoogle Scholar
  36. 36.
    Sydorova M, Lee MS (2005) Vascular endothelial growth factor levels in vitreous and serum of patients with either proliferative diabetic retinopathy or proliferative vitreoretinopathy. Ophthalmic Res 37:188–190PubMedCrossRefGoogle Scholar
  37. 37.
    Funatsu H, Yamashita H, Mimura T, Nakamura S et al (2005) Aqueous humor levels of cytokines are related to vitreous levels and progression of diabetic retinopathy in diabetic patients. Graefes Arch Clin Exp Ophthalmol 243:3–8PubMedCrossRefGoogle Scholar
  38. 38.
    Noma H, Funatsu H, Mimura T, Shimada K (2010) Increase in aqueous inflammatory factors in macular edema with branch retinal vein occlusion: a case control study. J Inflam 7:44–50CrossRefGoogle Scholar
  39. 39.
    Tong JP, Chan WM, Liu DT, Lai TY, Choy KW et al (2006) Aqueous humor levels of vascular endothelial growth factor and pigment epithelium-derived factor in polypoidal choroidal vasculopathy and choroidal neovascularization. Am J Ophthalmol 141:456–462PubMedCrossRefGoogle Scholar
  40. 40.
    Missotten GS, Notting IC, Schlingemann RO, Zijlmans HJ, Lau C et al (2006) Vascular endothelial growth factor a in eyes with uveal melanoma. Arch Ophthalmol 124:1428–1434PubMedCrossRefGoogle Scholar
  41. 41.
    Jonas JB, Neumaier M (2007) Vascular endothelial growth factor and basic fibroblast growth factor in exudative age-related macular degeneration and diffuse diabetic macular edema. Ophthalmic Res 39:139–142PubMedCrossRefGoogle Scholar
  42. 42.
    Roh MI, Kim HS, Song JH, Lim JB, Kwon OW (2009) Effect of intravitreal bevacizumab injection on aqueous humor cytokine levels in clinically significant macular edema. Ophthalmol 116:80–86CrossRefGoogle Scholar
  43. 43.
    Matsuyama K, Ogata N, Jo N, Shima C, Matsuoka M et al (2009) Levels of vascular endothelial growth factor and pigment epithelium-derived factor in eyes before and after intravitreal injection of bevacizumab. Jpn J Ophthalmol 53:243–248PubMedCrossRefGoogle Scholar
  44. 44.
    Lai TY, Liu DT, Chan KP, Luk FO, Pang CP et al (2009) Visual outcomes and growth factor changes of two dosages of intravitreal bevacizumab for neovascular age-related macular degeneration: a randomized, controlled trial. Retina 29:1218–1226PubMedCrossRefGoogle Scholar
  45. 45.
    Park SP, Ahn JK, Mun GH (2010) Aqueous vascular endothelial growth factor levels are associated with serous macular detachment secondary to branch retinal vein occlusion. Retina 30:281–286PubMedCrossRefGoogle Scholar
  46. 46.
    Forooghian F, Kertes PJ, Eng KT, Agrón E, Chew EY (2010) Alterations in the intraocular cytokine milieu after intravitreal bevacizumab. Invest Ophthalmol Vis Sci 51:2388–2392PubMedCrossRefGoogle Scholar
  47. 47.
    Simo R, Carrasco E, Garcia-Ramirez M et al (2006) Angiogenic and antiangiogenic factors in proliferative diabetic retinopathy. Curr Diabetes Rev 2:71–98PubMedCrossRefGoogle Scholar
  48. 48.
    Bulcao C, Ferreira SR, Giuffrida FM et al (2006) The new adipose tissue and adipocytokines. Curr Diabetes Rev 2:19–28PubMedCrossRefGoogle Scholar
  49. 49.
    Kim SK, Novak RF (2007) The role of intracellular signaling in insulin-mediated regulation of drug metabolizing enzyme gene and protein expression. Pharmacol Ther 113:88–120PubMedCrossRefGoogle Scholar
  50. 50.
    Rabinovitch A (1998) An update on cytokines in the pathogenesis of insulin-dependent diabetes mellitus. Diabetes Metab Rev 14:129–151PubMedCrossRefGoogle Scholar
  51. 51.
    ILSI Health and Enviromental Sciences Institute (2009) 2009 Annual Report. Accessed 28 July 2010
  52. 52.
    Apple FS, Murakami MM, Ler R et al. for the HESI Technical Committee of Biomarkers Working Group on Cardiac Troponins (2008) Analytical characteristics of commercial cardiac troponin I and T immunoassays in serum from rats, dogs, and monkeys with induced acute myocardial injury. Clin Chem 54:1982–1998Google Scholar
  53. 53.
    Amur S, Frueh F, Lesko L et al (2008) Integration and use of biomarkers in drug development, regulation and clinical practice: a US regulatory perspective. Biomark Med 2:305–311PubMedCrossRefGoogle Scholar
  54. 54.
    Welge-Lussen U, May CA, Neubauer AS et al (2001) Role of tissue growth factors in aqueous humor homeostasis. Curr Opin Ophthalmol 12:94–99PubMedCrossRefGoogle Scholar
  55. 55.
    Tripathi RC, Li J, Chan WF et al (1994) Aqueous humor in glaucomatous eyes contains an increased level of TGF-beta 2. Exp Eye Res 59:723–727PubMedCrossRefGoogle Scholar

Copyright information

© Springer Science+Business Media, LLC 2010

Authors and Affiliations

  1. 1.Department of OphthalmologyUniversity of FloridaJacksonvilleUSA
  2. 2.Omar Consulting Group, LLC73 Sararoga DrivePrinceton JunctionUSA

Personalised recommendations