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Mohamed Ali, Dhani Tracey-White, Matthew Smart, Mariya Moosajee; Combined nonsense mediated decay inhibition and suppression as a treatment approach for choroideremia. Invest. Ophthalmol. Vis. Sci. 2018;59(9):5314.
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© ARVO (1962-2015); The Authors (2016-present)
Nonsense-mediated decay (NMD) is a cellular surveillance mechanism which degrades nonsense-containing mRNA. This occurs at differing rates depending on a variety of factors including the position of the mutation. Therapies based on nonsense suppression require mRNA as the substrate for drug efficacy, hence inhibiting NMD may have the potential to improve translational readthrough with subsequent production of functional protein. Aim: To test amlexanox, an anti-inflammatory drug with dual NMD inhibition and nonsense suppression, for its ability to restore functional rab escort protein 1 (REP1) in two choroideremia (CHM) patient fibroblast cell lines.
Skin biopsies were taken from two affected male CHM patients with p.Tyr42* (Exon 3 UAG stop) and p.Arg270* (Exon 6 UGA stop) nonsense mutations. Dermal fibroblast cell lines were established and baseline qRT-PCR was undertaken to measure mRNA levels as an indicator of NMD activity. Dose-response experiments were conducted to determine the safest and most efficacious dose of amlexanox. A single dose of 300 µM was administered to fibroblasts, 24 hours post-treatment, qRT-PCR was repeated with functional prenylation assays (n=3 independent experiments, all values are mean±SEM).
CHM p.Tyr42* and p.Arg270* fibroblasts showed mean 83±2% and 18±1% baseline mRNA levels, respectively. Amlexanox-treated p.Tyr42* and p.Arg270* cell lines showed increases to 96±1% and 73±5% mRNA levels post-treatment suggesting NMD inhibition, particularly in the latter. Functional prenylation assay showed 100% accumulation of unprenylated rabs in both untreated CHM fibroblasts, but a significant mean (±SEM) reduction of 39±4% for p.Tyr42*, and 26±5% for p.Arg270*, following treatment with amlexanox.
The CHM p.Tyr42* fibroblasts had a higher baseline level of mRNA with less unprenylated rab accumulation post-treatment, indicating baseline mRNA may be a prognostic indicator for treatment response in future clinical trials. The p.Arg270* mRNA levels were boosted through amlexanox treatment enabling effective readthrough. Drugs with either combined NMD inhibition and nonsense suppression, or complementary actions may be the best approach for providing personalised medicine for treating nonsense-mediated choroideremia and other related inherited retinal dystrophies.
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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