A novel exon-specific U1snRNA therapeutic strategy to prevent retinal degeneration in familial dysautonomia

Abstract

Purpose : Familial dysautonomia (FD) is an autosomal recessive neurodegenerative disorder caused by a splice mutation in the gene encoding Elongator complex protein 1 (ELP1). A T-to-C base change in the 5’ splice site of ELP1 gene results in exon 20 skipping with tissue specific reduction of ELP1 protein predominantly in the nervous system. In addition to a complex neurological phenotype, FD patients also exhibit progressive retinal degeneration severely affecting their quality of life. To test novel splicing-targeted therapeutic approaches, we have developed a phenotypic mouse model, TgFD9; Ikbkap^△20/flox which exhibits most of clinical features of the disease while displaying the same tissue specific mis-splicing observed in patients.

Methods : Spectral domain optical coherence tomography (SD-OCT) was used to evaluate thickness of the retinal layers. Retinal whole mounts were performed to count the number of retinal ganglion cells (RGCs). Immunohistochemical staining was performed to detect degeneration of the optic nerve in FD mice. Adeno Associated Viral (AAV) vectors expressing ExspeU1s were generated to correct ELP1 splicing.

Results : We have comprehensively characterized the retinas of our FD mouse using SD-OCT and immunohistochemical assays during disease progression. Our findings showed a significant decrease in the thickness of the retinal nerve fiber layer (RNFL) and the ganglion cell layer (GCL) starting from 3 months of age (p < 0.009 and 0.005). Retinal whole-mount analysis showed reduction of RGC cell counts from 6 months of age (p< 0.002). Neurofilament (NF) staining analysis of the optic nerve from FD mice indicated diffuse degeneration of axonic bundles, demonstrating that our mouse model correctly recapitulates the retinal degeneration observed in patients.To correct ELP1 splicing defect, we have designed a novel splice targeted therapy using modified version of the spliceosomal U1 snRNAs (ExSpeU1s) that permit targeted binding to intronic sequences downstream of the mutant 5' splice site enabling efficient recruitment of spliceosomal machinery. In vivo delivery of FD ExSpeU1 using AAV led to successful correction of ELP1 splicing.

Conclusions : Our findings demonstrate that our novel FD mouse model exhibits most of retinal degeneration pathology observed in FD patients and highlight the valuable therapeutic potential of ExSpeU1 delivery to treat retinal degeneration in FD.

This is a 2021 ARVO Annual Meeting abstract.