Abstract
Purpose: :
While the field of ophthalmology continues to be revolutionized by new pharmacologic and biologic treatments, techniques for delivery of such therapies have evolved minimally. Phase I clinical trials investigating adeno-associated virus mediated gene replacement have employed a 3-port pars plana vitrectomy approach with subretinal injection of the viral complex through a 37 gauge cannula. We investigated the efficacy and safety of therapeutic delivery by an ab-externo approach using the iTtrackTM 250A microcatheter (iScience Surgical Corp., Menlo Park, CA). Further investigation is needed to assess whether this system could be used to safely infiltrate the subretinal space for drug delivery, and whether delivery of gene therapy vector into the suprachoroidal space can elicit the desired transfection of the retinal pigment epithelial cells.
Methods: :
Six adult Zealand white rabbits were treated in their left eye. The animals were used according to the ARVO Resolution concerning the use of animals in research. Following pupil dilation and appropriate anesthesia, a conjunctival peritomy was performed and a 2 mm radial scleral incision was made to enter the subretinal space. The microcatheter was introduced into the suprachoroidal or subretinal space accordingly. 150 microliters of phosphate-buffered saline with the green fluorescent protein (GFP) gene within an AAV2 viral vector were injected into the peripapillary space. Animals were dilated and photographed for green fluorescence after 6 weeks and later sacrificed.for histologic examination using laser scanning confocal microscopy to assess the level of GFP expression.
Results: :
Of the 6 eyes treated, vector delivery was obtained in all eyes without any complications including retinal detachment, suprachoroidal hemorrhage, or endophthalmitis. None of the rabbits exhibited signs of discomfort post-operatively. All surgeries were completed within 15 minutes and in all cases the exact site of viral gene delivery could be directly visualized at surgery by indirect ophthalmoscopy.
Conclusions: :
The 250 micron microcatheter, is a safe and easily performed alternative method of gene delivery to the retina using an ab-externo approach. and can offer a quicker and less technically challenging surgical option.
Keywords: gene transfer/gene therapy • microscopy: light/fluorescence/immunohistochemistry • retina