SUMMARY Nephronophthisis-related ciliopathies (NPHP-RC) are degenerative recessive diseases that affect kidney, retina and brain. Genetic defects in NPHP gene products that localize to cilia and centrosomes defined them as ‘ciliopathies’. However, disease mechanisms remain poorly understood. Here we identify by whole exome resequencing, mutations of MRE11, ZNF423, and CEP164 as causing NPHP-RC. All three genes function within the DNA damage response (DDR) pathway, hitherto not implicated in ciliopathies. We demonstrate that, upon induced DNA damage, the NPHP-RC proteins ZNF423, CEP164 and NPHP10 colocalize to nuclear foci positive for TIP60, known to activate ATM at sites of DNA damage. We show that knockdown of CEP164 or ZNF423 causes sensitivity to DNA damaging agents, and that cep164 knockdown in zebrafish results in dysregulated DDR and an NPHP-RC phenotype. We identify TTBK2, CCDC92, NPHP3 and DVL3 as novel CEP164 interaction partners. Our findings link degenerative diseases of kidney and retina, disorders of increasing prevalence, to mechanisms of DDR.

Retinal dystrophy (RD) is a heterogeneous group of hereditary diseases caused by loss of photoreceptor function and contributes significantly to the etiology of blindness globally but especially in the industrialized world. The extreme locus and allelic heterogeneity of these disorders poses a major diagnostic challenge and often impedes the ability to provide a molecular diagnosis that can inform counseling and gene-specific treatment strategies. In a large cohort of nearly 150 RD families, we used genomic approaches in the form of autozygome-guided mutation analysis and exome sequencing to identify the likely causative genetic lesion in the majority of cases. Additionally, our study revealed six novel candidate disease genes (C21orf2, EMC1, KIAA1549, GPR125, ACBD5, and DTHD1), two of which (ACBD5 and DTHD1) were observed in the context of syndromic forms of RD that are described for the first time.

MERTK is an essential component of the signaling network that controls phagocytosis in retinal pigment epithelium (RPE), the loss of which results in photoreceptor degeneration. Previous proof-of-concept studies have demonstrated the efficacy of gene therapy using human MERTK (hMERTK) packaged into adeno-associated virus (AAV2) in treating RCS rats and mice with MERTK deficiency. The purpose of this study was to assess the safety of gene transfer via subretinal administration of rAAV2-VMD2-hMERTK in subjects with MERTK-associated retinitis pigmentosa (RP). After a preclinical phase confirming the safety of the study vector in monkeys, six patients (aged 14 to 54, mean 33.3 years) with MERTK-related RP and baseline visual acuity (VA) ranging from 20/50 to <20/6400 were entered in a phase I open-label, dose-escalation trial. One eye of each patient (the worse-seeing eye in five subjects) received a submacular injection of the viral vector, first at a dose of 150 µl (5.96 × 10(10)vg; 2 patients) and then 450 µl (17.88 × 10(10)vg; 4 patients). Patients were followed daily for 10 days at 30, 60, 90, 180, 270, 365, 540, and 730 days post-injection. Collected data included (1) full ophthalmologic examination including best-corrected VA, intraocular pressure, color fundus photographs, macular spectral domain optical coherence tomography and full-field stimulus threshold test (FST) in both the study and fellow eyes; (2) systemic safety data including CBC, liver and kidney function tests, coagulation profiles, urine analysis, AAV antibody titers, peripheral blood PCR and ASR measurement; and (3) listing of ophthalmological or systemic adverse effects. All patients completed the 2-year follow-up. Subretinal injection of rAAV2-VMD2-hMERTK was associated with acceptable ocular and systemic safety profiles based on 2-year follow-up. None of the patients developed complications that could be attributed to the gene vector with certainty. Postoperatively, one patient developed filamentary keratitis, and two patients developed progressive cataract. Of these two patients, one also developed transient subfoveal fluid after the injection as well as monocular oscillopsia. Two patients developed a rise in AAV antibodies, but neither patient was positive for rAAV vector genomes via PCR. Three patients also displayed measurable improved visual acuity in the treated eye following surgery, although the improvement was lost by 2 years in two of these patients. Gene therapy for MERTK-related RP using careful subretinal injection of rAAV2-VMD2-hMERTK is not associated with major side effects and may result in clinical improvement in a subset of patients.

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Purpose:Retinal dystrophies (RD) are heterogeneous hereditary disorders of the retina that are usually progressive in nature. The aim of this study was to clinically and molecularly characterize a large cohort of RD patients. Methods:We have developed a next-generation sequencing assay that allows known RD genes to be sequenced simultaneously. We also performed mapping studies and exome sequencing on familial and on syndromic RD patients who tested negative on the panel. Results:Our panel identified the likely causal mutation in >60% of the 292 RD families tested. Mapping studies on all 162 familial RD patients who tested negative on the panel identified two novel disease loci on Chr2:25,550,180-28,794,007 and Chr16:59,225,000-72,511,000. Whole-exome sequencing revealed the likely candidate as AGBL5 and CDH16, respectively. We also performed exome sequencing on negative syndromic RD cases and identified a novel homozygous truncating mutation in GNS in a family with the novel combination of mucopolysaccharidosis and RD. Moreover, we identified a homozygous truncating mutation in DNAJC17 in a family with an apparently novel syndrome of retinitis pigmentosa and hypogammaglobulinemia. Conclusion:Our study expands the clinical and allelic spectrum of known RD genes, and reveals AGBL5, CDH16, and DNAJC17 as novel disease candidates.