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P.K. Dioguardi, R.K. Schuler, J.M. Nickerson, C. Henjy, L.P. Cruysberg, H.F. Edelhauser; Transscleral Permeability of a Fluorescein Labeled Oligonucleotide . Invest. Ophthalmol. Vis. Sci. 2003;44(13):4427.
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Background: Treatment of diseases affecting the posterior segment such as age-related macular degeneration, diabetic retinopathy, and retinitis pigmentosa is limited by the difficulty in delivering therapeutic drugs and biologically active agents to tissues in the posterior segment. Developing more effective ocular drug delivery systems which can effectively carry potential therapeutic agents is essential to improving the treatment of posterior segment eye disease. During the past decade, intense efforts to develop and exploit antisense technology as a potential therapeutic option for posterior segment diseases have been mounted. Purpose: To determine the in vitro transscleral permeability of a fluorescein labeled therapeutic oligonucleotide for Oculocutaneous Albinism type 1. Methods: Scleral sections excised from moist-chamber stored human globes were mounted in a perfusion chamber. Transscleral permeability was measured by placing a small depot of a fluorescein labeled oligonucleotide (100 ul of 2.96 x 10-4 MW=7998.3) on the episcleral side of the sclera while perfusing the choroidal side with BSS. Fractions of choroidal perfusate were collected every hour for 24 hours and fluorescence was measured with a spectrofluorometer. From this data, scleral permeability Ktrans (cm/sec) was calculated (n=7). To assess possible degradation, permeability experiments were repeated using P-32 to label the oligonucleotide followed by PAGE analysis on fractions collected. Results: Ktrans for the transscleral diffusion of fluorescein labeled oligonucleotide was 7.13 +/- 0. x 107 cm/s (mean +/- SEM, n=7). After 24 hours 21.51 +/- 9.81% of the total amount of oligonucleotide had diffused across the sclera, leaving approximately 3.21 +/- 2.14% in the sclera itself and 75.28 +/- 8.05% in the donor chamber (mean +/- SEM). Conclusions: This study shows that transscleral diffusion of detectable amounts of a fluorescein labeled oligonucleotide can be achieved. PAGE analysis of P-32 oligonucleotide confirmed that the oligonucleotide diffused across the sclera. Delivering drugs to posterior ocular tissues across the sclera may avoid possible complications associated with intraocular delivery as well as limit systemic exposure, and, therefore the potential for systemic toxicity. The current results provide a promising initial step in the development of techniques to deliver antisense oligonucleotides to the posterior segment.
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