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N. R. Thakkar, P. K. Karla, A. Ray, R. Vadlapatla, D. Pal, A. K. Mitra; Mechanism of Nucleotide Drug Efflux by Human Corneal Epithelial Cells. Invest. Ophthalmol. Vis. Sci. 2009;50(13):4616.
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© ARVO (1962-2015); The Authors (2016-present)
A nucleoside efflux transporter system (MRP4/MRP5) was identified on the human cornea recently in our laboratory. Various nucleoside drugs like acyclovir, used to treat herpes keratitis are employed in the treatment of corneal opacities. The primary objective of this project was to prove the mechanism of drug efflux in corneal epithelial cells.
Thin layer chromatography followed by autoradiography was used. A prodrug of nucleotide PMEA, bis(POM)PMEA was employed. Transfected human corneal epithelial cells and MDCKII-MRP5 cells were employed as cell culture models.
Following preloading the cells with bis(POM)PMEA under ATP depleting conditions for 1 hr, ~80% of drug converted to PMEA. Significant functional efflux of PMEA was observed in the supernatant solution. Functional efflux of PMEA was suppressed under ATP depleting conditions. Distinct bands corresponding to prodrug and active drug were noticed on X-ray films.
Significant conversion of ester prodrug to active nucleotide, PMEA proved the mechanism of efflux by corneal epithelial cells. This method confirms the functional expression of the transporter system on corneal epithelium.
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