Date of Award

Spring 5-18-2016

Degree Type

Thesis

Degree Name

Master of Science in Vision Science

Committee Chair

Deepa Rao, PhD

Second Advisor

Len Koh, PhD, OD

Third Advisor

Ryan Bulson, OD

Abstract

Pterygium is a fleshy, inflamed, overgrowth of the conjunctiva that can invade the visual axis and cause vision loss. Histologically, pterygia tissue shows overexpression of angiogenic factors. Potassium dobesilate (KD) and resveratrol (RES) are known to inhibit angiogenesis and thereby may be useful agents in reducing pterygia size and inflammation.

This study aimed at developing and characterizing three ophthalmic formulations each of KD, RES, or KD:RES 10:1. The formulations for the individual and dual drug combination are prepared in normal saline (NS) with a viscosity enhancer (10% polyethylene glycol 400; PEG 400) or as a sol-gel using 10% PEG 400 and 21% Pluronic® F127. All formulations contained either 25 mg of KD, 2.5 mg of RES, or a combination of KD:RES 10:1 ratio and were stored at room temperature in the dark. The two types of formulations are tested to provide flexible dosing schedules of once a day (Pluronic ® F127) or twice a day (NS+PEG 10%).

The physical and chemical stability of these formulations was assessed over a 28-day period, in triplicate, on days 1, 7, 14, 21, and 28. Physical stability was determined by visual inspection and optical density measurements at 295 nm (KD) and 306 nm (RES). Chemical stability was assessed by Reverse Phase High Performance Liquid Chromatography (RP-HPLC) using a C18 column and a DAD detector at 295 nm (KD) and 306 nm (RES). The effect of the compounds on the proliferation of human umbilical vein endothelial cells (HUVEC) was used to assess the anti-angiogenic properties of KD and RES. Cell viability of HUVEC was determined using Cell-Titer Blue® 24-hours post-treatment with KD, RES, or the combination.

Formulations 5 and 6 demonstrated physical stability for 28 days when stored at room temperature protected from light. Formulation 1 in NS underwent a color change from slight yellow to darker yellow by day 7. Optical density measurements were significantly lower at day 21 and 28 when compared to day 1. The PEG 400 and sol-gel formulations did not undergo a color or optical changes during storage. All formulations were chemically stable for 28 days as assessed by RP-HPLC using United States Pharmacopeia (USP) guidelines for ophthalmic formulations. The anti-proliferative effect of KD, RES, and KD:RES combination in HUVEC indicated that RES is a more potent anti-angiogenic agent as compared to KD. Combination index analysis of the KD:RES formulation indicates the combination is more potent inhibitor of angiogenesis as compared to the individual compounds.

Our study indicates that KD in combination with RES may be a viable alternative treatment to reduce pterygium size and inflammation. The sol-gel formulation may enhance patient compliance as compared to the solution formulations with once a day, overnight, dosing. Future work for this project lies in assessing the microbiological stability of these formulations in vitro and determining effectiveness in vivo.

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