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Xia Wang, Olena Sivak, Kishor Wasan, Kevin Gregory-Evans, Cheryl Gregory-Evans; Pharmacological nonsense suppression therapy in the Rpe65-/- mouse model of Leber congenital amaurosis reverses the retinal deficits in vivo. Invest. Ophthalmol. Vis. Sci. 2018;59(9):5319.
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© ARVO (1962-2015); The Authors (2016-present)
Leber congenital amaurosis (LCA) is an inherited eye disease where more than 400 mutations in 19 different genes have been identified. This disease is characterized by severe and early vision loss, sensory nystagmus, and an un-recordable electroretinogram (ERG). In LCA approximately 35% of gene mutations are in-frame nonsense mutations. We hypothesized that inhibition of nonsense-mediated mRNA decay (NMD) combined with nonsense suppression therapy, using small molecule drugs Amlexanox and Ataluren, respectively, could be used in treating the Rd12 mouse that harbors a homozygous in-frame nonsense mutation in the Rpe65 gene to improve the ocular phenotype.
Rpe65-/- time-mated mice received daily subcutaneous injections of Ataluren (30 µg/g), Amlexanox (25 µg/g) or a combination of the two drugs from E12.5 until birth. The offspring then received daily subcutaneous injections from P4 to P60, P90, or P120. For postnatal treatment only, newborn mice received injections from P4 and continued to P60 or P90. Drug efficacy was tested by ELISA, TUNEL staining, ERG, optokinetic tracking (OKT), immunohistochemistry and quantitative real-time PCR.
Prenatal coupled with postnatal injections of Amlexanox alone or combinatorial Amlexanox and Ataluren showed significantly increased expression of Rpe65 protein and Rpe65 mRNA in the Rpe65-/- retina. TUNEL staining revealed the prenatal treatment rescued cell death in retina and RPE cells. The prenatal treatment also resulted in OKT spatial threshold frequencies of 0.4-0.5 cycles/degree at P90 and P120, equivalent to that in normal mice. Untreated Rpe6-/- mice exhibited reduced b-wave amplitudes in dark-adapted ERG response and a delayed light-adapted ERG response. However, the prenatally treated mice showed ~50% increase in dark-adapted rod responses, dark oscillatory potentials, and photopic cone responses at P120.
Treatment with a NMD inhibitor improved the efficacy of nonsense suppression therapy in the Rpe65-/- mutant eye. A NMD inhibitor would be predicted to keep the mutant mRNA available for longer allowing more nonsense suppression to occur. This could be important for recessive diseases, especially where low levels of readthrough might not be enough to rescue the phenotype. These results suggest that LCA patients may benefit from this safe and non-invasive novel therapeutic option.
This is an abstract that was submitted for the 2018 ARVO Annual Meeting, held in Honolulu, Hawaii, April 29 - May 3, 2018.
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