Topical small molecule translational bypass therapy rescues Pax6 aniridia mutant phenotype

Abstract

Purpose: Congenital aniridia caused by haploinsufficiency of PAX6, is characterized at birth by absence of the iris, cataracts and foveal hypoplasia, with later development of glaucoma and corneal keratopathy. The commonest type of mutation in PAX6 is nonsense mutations leading to a premature stop codon, present in 40% of aniridia cases. In this study we tested the efficacy of the small molecule drug therapy aimed at bypassing stop mutations to ameliorate the phenotype of the heterozygous Pax6^Sey mouse model.

Methods: Pax6^Sey heterozygous pregnant mice were treated daily from E12.5 with subcutaneous injections of either gentamicin or Ataluren and then the offspring received the same treatment until P21. In an alternate postnatal only paradigm, Pax6^Sey heterozygous mice received daily drug therapy by either subcutaneous injection or topical eyedrops from P4-P60. Drug efficacy was tested by histology, electrophysiology, optokinetic tracking, and quantitative PCR.

Results: Treatment of Pax6^Sey pregnant mice with either gentamicin or Ataluren rescued the proliferative defect in the retina and normalized the lens, iris and cornea defects in the Pax6^Sey mutant offspring. Early postnatal treatment of Pax6^Sey mice either by systemic or eyedrop delivery similarly rescued the histological phenotype. Untreated Pax6^Sey mice do not exhibit ERG responses however, the treated mice showed characteristic dark-adapted rod responses, dark oscillatory potentials, photopic cone responses and a 30Hz flicker that were 70-90% of wildtype responses. In treated mice, optokinetic tracking demonstrated a spatial threshold frequency of 0.41 cycles/degree, equivalent to normal mice. In untreated mice, gene expression changes in Foxc1, Tgfβ2 and Bmp4 in the developing Pax6^Sey iris were normalized after drug treatment.

Conclusions: Translational bypass therapy used in a prenatal or postnatal paradigm rescued the histological, molecular and functional defects observed in the Pax6^Sey mutant eye. Using a targeted drug therapy to overcome the underlying Pax6 genetic defect by a postnatal strategy is the first evidence that it is possible to correct developmental abnormalities in congenital conditions such as aniridia. Since Ataluren is an orally bioavailable drug that does not have the ototoxic and nephrotoxic side effects associated with gentamicin, it may provide a treatment option for aniridia patients.

Keywords: 571 iris • 497 development • 739 transcription factors