Mutations in the retinal pigment epithelium gene encoding RPE65 are a cause of the incurable early-onset recessive human retinal degenerations known as Leber congenital amaurosis. Rpe65-deficient mice, a model of Leber congenital amaurosis, have no rod photopigment and severely impaired rod physiology. We analyzed retinoid flow in this model and then intervened by using oral 9-cis-retinal, attempting to bypass the biochemical block caused by the genetic abnormality. Within 48 h, there was formation of rod photopigment and dramatic improvement in rod physiology, thus demonstrating that mechanismbased pharmacological intervention has the potential to restore vision in otherwise incurable genetic retinal degenerations.

The visual process is initiated by the photoisomerization of 11-cis-retinal to all-trans-retinal. For sustained vision the 11-cis-chromophore must be regenerated from all-trans-retinal. This requires RPE65, a dominant retinal pigment epithelium protein. Disruption of the RPE65 gene results in massive accumulation of alltrans-retinyl esters in the retinal pigment epithelium, lack of 11-cis-retinal and therefore rhodopsin, and ultimately blindness. We reported previously (Van Hooser, J. P., Aleman, T. S., He, Y. G., Cideciyan, A. V., Kuksa, V., Pittler, S. J., Stone, E. M., Jacobson, S. G., and Palczewski, K. (2000) Proc. Natl. Acad. Sci. U. S. A. 97, 8623-8628) that in Rpe65؊/؊ mice, oral administration of 9-cis-retinal generated isorhodopsin, a rod photopigment, and restored light sensitivity to the electroretinogram. Here, we provide evidence that early intervention by 9-cis-retinal administration significantly attenuated retinal ester accumulation and supported rod retinal function for more than 6 months post-treatment. In single cell recordings rod light sensitivity was shown to be a function of the amount of regenerated isorhodopsin; high doses restored rod responses with normal sensitivity and kinetics. Highly attenuated residual rod function was observed in untreated Rpe65؊/؊ mice. This rod function is likely a consequence of low efficiency production of 11-cis-retinal by photo-conversion of alltrans-retinal in the retina as demonstrated by retinoid analysis. These studies show that pharmacological intervention produces long lasting preservation of visual function in dark-reared Rpe65؊/؊ mice and may be a useful therapeutic strategy in recovering vision in humans diagnosed with Leber congenital amaurosis caused by mutations in the RPE65 gene, an inherited group of early onset blinding and retinal degenerations. Leber congenital amaurosis (LCA)1 is a group of conditions that cause blindness or severe visual impairment from birth. All show both rod and cone dysfunction, a negligible (not recordable) electroretinogram (ERG), and nystagmus. They result in early onset retinal dystrophy (1), which over time may be accompanied by pigmentary changes in the retina, hence "amaurosis" (Greek for darken). LCA is caused by defects in at least five different genes that disrupt a variety of different cellular functions (2-6).In ϳ12% of all LCA cases the gene for a 65-kDa protein (RPE65) of retinal pigment epithelium cells (RPE) is disabled (7,8). RPE65 is heavily expressed in RPE cells, where it plays an essential role in the retinoid cycle. This is a set of tightly interconnected events that involve both photoreceptors and RPE cells. The photoisomerization of the visual pigment chromophore (11-cis-retinal) produces all-trans-retinal, which is reduced in the photoreceptor, transferred to the RPE, converted back to 11-cis-retinal, and then transferred back to the photoreceptor to regenerate the original visual pigment (reviewed in Ref. 9). The precise function of RPE65 in retinoid processing is unknown.Genetically enginee...