Abstract
With marketing approval of the first ocular gene therapy, and other gene therapies in clinical trial, treatments for inherited retinal degenerations (IRDs) have become a reality. Biallelic mutations in the tubby like protein 1 gene (TULP1) are causative of IRDs in humans; a mouse knock-out model (Tulp1−/−) is characterized by a similar disease phenotype. We developed a Tulp1 supplementation therapy for Tulp1−/− mice. Utilizing subretinal AAV2/5 delivery at postnatal day (p)2–3 and rhodopsin-kinase promoter (GRK1P) we targeted Tulp1 to photoreceptor cells exploring three doses, 2.2E9, 3.7E8, and 1.2E8 vgs. Tulp1 mRNA and TULP1 protein were assessed by RT-qPCR, western blot and immunocytochemistry, and visual function by electroretinography. Our results indicate that TULP1 was expressed in photoreceptors; achieved levels of Tulp1 mRNA and protein were similar to wild type levels at p20. However, the thickness of the outer nuclear layer (ONL) did not improve in treated Tulp1−/− mice. There was a small and transient electroretinography benefit in the treated retinas at 4 weeks of age (not observed by 6 weeks) when using 3.7E8 vg dose. Dark-adapted mixed rod and cone a- and b-wave amplitudes were 24.3 ± 13.5 μV and 52.2 ± 31.7 μV in treated Tulp1−/− mice, which were significantly different (p < 0.001, t-test), from those detected in untreated eyes (7.1 ± 7.0 μV and 9.4 ± 15.1 μV, respectively). Our results indicate that Tulp1 supplementation in photoreceptors may not be sufficient to provide robust benefit in Tulp1−/− mice. As such, further studies are required to fine tune the Tulp1 supplementation therapy, which, in principle, should rescue the Tulp1−/− phenotype.
Highlights
Biallelic mutations in tubby like protein 1 gene (TULP1) are responsible for rare forms of early onset, severe, autosomal recessive retinal degenerations, typically registered as retinitis pigmentosa (RP14) or Leber congenital amaurosis (LCA15) (Hagstrom et al, 1998; Mataftsi et al, 2007; Jacobson et al, 2014; RetNet - Retinal Information Network, 2019)
We found that the p2–3 timepoint resulted in the highest number of transduced photoreceptor cells when delivering AAV2/5 subretinally in Tulp1−/− mice
We studied the effects of photoreceptor-targeted associated virus (AAV)-delivered supplementation of Tulp1 in Tulp1−/− mouse retinas (Figure 2)
Summary
Biallelic mutations in tubby like protein 1 gene (TULP1) are responsible for rare forms of early onset, severe, autosomal recessive retinal degenerations, typically registered as retinitis pigmentosa (RP14) or Leber congenital amaurosis (LCA15) (Hagstrom et al, 1998; Mataftsi et al, 2007; Jacobson et al, 2014; RetNet - Retinal Information Network, 2019). Many preclinical studies have highlighted the value of adeno associated virus (AAV) as a means of gene delivery and have demonstrated benefit at molecular, histological and functional levels of AAV-delivered gene therapies (among others, Millington-Ward et al, 2011; Chadderton et al, 2013; Schön et al, 2017; Cideciyan et al, 2018). Such studies prompted us to explore of a similar therapeutic approach for TULP1-linked retinal degenerations
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