July 2019
Volume 60, Issue 9
Open Access
ARVO Annual Meeting Abstract  |   July 2019
In Vitro Degradation of Human Vascular Endothelial Growth Factor
Author Affiliations & Notes
  • Ryan A Shields
    Stanford Hospital, Palo Alto, California, United States
  • Jose Davila
    Stanford Hospital, Palo Alto, California, United States
  • Ira H Schachar
    Stanford Hospital, Palo Alto, California, United States
  • Footnotes
    Commercial Relationships   Ryan Shields, None; Jose Davila, None; Ira Schachar, None
  • Footnotes
    Support  NIH P30-026877
Investigative Ophthalmology & Visual Science July 2019, Vol.60, 5375. doi:
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      Ryan A Shields, Jose Davila, Ira H Schachar; In Vitro Degradation of Human Vascular Endothelial Growth Factor. Invest. Ophthalmol. Vis. Sci. 2019;60(9):5375.

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      © ARVO (1962-2015); The Authors (2016-present)

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Purpose : Intravitreal injection of vascular endothelial growth factor (VEGF) in rabbits shows rapid initial vitreous clearance. It has been theorized that this is due to consumption or degradation of VEGF intravitreally. We hypothesized that rapid clearance is primarily due to enzymatically driven proteolysis of VEGF in vivo. We sought to test this hypothesis by assessing rates of enzymatic VEGF degradation in vitro.

Methods : Recombinant human VEGF A-165 was incubated at physiologic temperature (37C) with either phosphate buffered saline (PBS), bovine serum albumin (BSA), porcine vitreous, or porcine vitreous with a broad protease inhibitor (cOmpleteTM Mini, Sigma-Aldrich). Aliquots were removed at 24 hours, 48 hours, 4 days, 7 days, and 14 days and stored at -80C. A human VEGF ELISA Kit (KHG0111, Invitrogen) was used to quantify the concentration of VEGF in each sample.

Results : At 24 hours there was a 6.1% reduction in the VEGF concentration in PBS, 3.8% increase in BSA, 5.5% reduction in porcine vitreous, and an 11.4% reduction in protease-inhibited porcine vitreous. At day 14 VEGF concentrations were reduced by 15.9% in PBS, 1.4% in BSA, 22.6% in porcine vitreous, and 18.8% in protease-inhibited vitreous Though both the vitreous and protease-inhibited vitreous demonstrated a statistically significant reduction in VEGF concentration at 14 days (p = 0.01 and p = 0.02, respectively), approximately 80% of VEGF still remained in both samples.

Conclusions : Enzymatic degradation is unlikely to play a major role in the clearance of VEGF from the vitreous. Rapid clearance of VEGF observed in pharmacokinetic studies in rabbits may be due to inadequate pharmacokinetic sampling resulting in overestimation of clearance rates or as of yet unknown factors which bind or clear VEGF faster than macromolecules of similar size. An accurate understanding of the rate of intraocular VEGF clearance is necessary to predict the rate of VEGF production in various disease states and critical to the development of effective sustained delivery anti-VEGF therapeutics.

This abstract was presented at the 2019 ARVO Annual Meeting, held in Vancouver, Canada, April 28 - May 2, 2019.


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