Assessment of Local Functional Improvement following Gene Therapy in the RPE65 Null Mutation Dog Model

Abstract

Abstract: : Purpose: Current studies in a large group of RPE65-/- dogs with early onset blindness show a significant improvement in visual function 4-6 weeks after local gene transfer (Narfstrom et al., IOVS, In press). In this work, we show that multifocal ERG (mfERG) can be used to study the topographic distribution of restored retinal function following the localized, subretinal injection of the rAAV.RPE65 gene construct. Methods: Seven RPE65-deficient Briard-Beagle dogs and 3 normal controls, treated at the age of 4 months to 2.5 years, were examined 3-6 months after surgery. Following general anesthesia, intubation and artificial ventilation, mfERGs were recorded using continuous stimulus position monitoring via an SLO by two methods as previously described (Seeliger et al., Doc Ophthalmol 2000; 100: 167-184). In short, while the infrared channel provided the retinal image in both cases, the RetiScan mfERG system used the 515 nm (green) wavelength of the Argon laser to stimulate the retina, whereas the VERIS system employed a stimulator prototype to introduce a CRT stimulus into the optical pathway of the SLO. Eyes were positioned with stay sutures and Jet contact lens electrodes were used to record the evoked signals at the corneal level. Results: Vision was restored to some degree in all affected animals included, documented by loss of nystagmus and Ganzfeld ERG improvement. With both mfERG methods of stimulation, the largest responses were obtained at the site of the subretinal injections, and also a marked difference in amplitudes was observed between the treated and the untreated eye. Furthermore, it was possible to obtain a focal 'response intensity series' in the treated eye, whereas the untreated eye did not show such an increase with stimulus intensity. Conclusions: The results of both Ganzfeld and mfERGs underline the objective improvement of visual function following gene replacement therapy in the Briard dog model of RPE65 deficiency. In particular, the mfERG allowed to detect the regional restoration of retinal function, and is thus a valuable tool for the objective local evaluation of the long-term effects of gene therapy.

Keywords: gene transfer/gene therapy • electroretinography: non-clinical • animal model