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Purpose: To identify the genetic cause of autosomal-dominant pattern dystrophy (PD) of the retinal pigment epithelium (RPE) in two families., Methods and Results: Two families with autosomal-dominant PD were identified. Eight members of family 1 (five affected) were subjected to whole-genome SNP genotyping; multipoint genome-wide linkage analysis identified 7 regions of potential linkage, and genotyping four additional individuals from family 1 resulted in a maximum logarithm of odds score of 2.09 observed across four chromosomal regions. Exome sequencing of two affected family 1 members identified 15 shared non-synonymous rare coding sequence variants within the linked regions; candidate genes were prioritised and further analysed. Sanger sequencing confirmed a novel heterozygous missense variant (E79K) in orthodenticle homeobox 2 (OTX2) that segregated with the disease phenotype. Family 2 with PD (two affected) harboured the same missense variant in OTX2. A shared haplotype of 19.68 cM encompassing OTX2 was identified between affected individuals in the two families. Within the two families, all except one affected demonstrated distinct 'patterns' at the macula. In vivo structural retinal imaging showed discrete areas of RPE-photoreceptor separation at the macula in all cases. Electroretinogram testing showed generalised photoreceptor degeneration in three cases. Mild developmental anomalies were observed, including optic nerve head dysplasia (four cases), microcornea (one case) and Rathke's cleft cyst (one case); pituitary hormone levels were normal., Conclusions: This is the first report implicating OTX2 to underlie PD. The retinal disease resembles conditional mice models that show slow photoreceptor degeneration secondary to loss of Otx2 function in the adult RPE., (Published by the BMJ Publishing Group Limited. For permission to use (where not already granted under a licence) please go to http://group.bmj.com/group/rights-licensing/permissions.)

Leber congenital amaurosis (LCA) is an autosomal recessive retinal dystrophy that manifests with genetic heterogeneity. We sequenced the exome of an individual with LCA and identified nonsense (c.507G>A, p.Trp169*) and missense (c.769G>A, p.Glu257Lys) mutations in NMNAT1, which encodes an enzyme in the nicotinamide adenine dinucleotide (NAD) biosynthesis pathway implicated in protection against axonal degeneration. We also found NMNAT1 mutations in ten other individuals with LCA, all of whom carry the p.Glu257Lys variant.

Background: Ocular albinism type 1 (OA1) is an X-linked ocular disorder characterized by a severe reduction in visual acuity, nystagmus, hypopigmentation of the retinal pigmented epithelium, foveal hypoplasia, macromelanosomes in pigmented skin and eye cells, and misrouting of the optical tracts. This disease is primarily caused by mutations in the OA1 gene., Methods: The ophthalmologic phenotype of the patients and their family members was characterized. We screened for mutations in the OA1 gene by direct sequencing of the nine PCR-amplified exons, and for genomic deletions by PCR-amplification of large DNA fragments., Results: We sequenced the nine exons of the OA1 gene in 72 individuals and found ten different mutations in seven unrelated families and three sporadic cases. The ten mutations include an amino acid substitution and a premature stop codon previously reported by our team, and eight previously unidentified mutations: three amino acid substitutions, a duplication, a deletion, an insertion and two splice-site mutations. The use of a novel Taq polymerase enabled us to amplify large genomic fragments covering the OA1 gene. and to detect very likely six distinct large deletions. Furthermore, we were able to confirm that there was no deletion in twenty one patients where no mutation had been found., Conclusion: The identified mutations affect highly conserved amino acids, cause frameshifts or alternative splicing, thus affecting folding of the OA1 G protein coupled receptor, interactions of OA1 with its G protein and/or binding with its ligand.

Ocular albinism type 1 (OA1) is an X-linked disorder, mainly characterized by a severe reduction in visual acuity, foveal hypoplasia, nystagmus, hypopigmentation of the retina, the presence of macromelanosomes in the skin and eyes, and the misrouting of optic pathways, resulting in the loss of stereoscopic vision. We screened the OA1 gene for mutations in three unrelated Canadian and French families and in two isolated patients with OA1. We found three different missense mutations and two different nonsense mutations, three of which were novel. To date, 41 mutations (including missense mutations, insertions, and deletions) have been reported in the OA1 gene. Mutation and polymorphism data for this gene are available from the international albinism center albinism database website: http://www.cbc.umn.edu/tad/oa1map.htm.

Timely molecular diagnosis of RB1 mutations enables earlier treatment, lower risk, and better health outcomes for patients with retinoblastoma; empowers families to make informed family-planning decisions; and costs less than conventional surveillance. However, complexity has hindered clinical implementation of molecular diagnosis. The majority of RB1 mutations are unique and distributed throughout the RB1 gene, with no real hot spots. We devised a sensitive and efficient strategy to identify RB1 mutations that combines quantitative multiplex polymerase chain reaction (QM-PCR), double-exon sequencing, and promoter-targeted methylation-sensitive PCR. Optimization of test order by stochastic dynamic programming and the development of allele-specific PCR for four recurrent point mutations decreased the estimated turnaround time to <3 wk and decreased direct costs by one-third. The multistep method reported here detected 89% (199/224) of mutations in bilaterally affected probands and both mutant alleles in 84% (112/134) of tumors from unilaterally affected probands. For 23 of 27 exons and the promoter region, QM-PCR was a highly accurate measure of deletions and insertions (accuracy 95%). By revealing those family members who did not carry the mutation found in the related proband, molecular analysis enabled 97 at-risk children from 20 representative families to avoid 313 surveillance examinations under anesthetic and 852 clinic visits. The average savings in direct costs from clinical examinations avoided by children in these families substantially exceeded the cost of molecular testing. Moreover, health care savings continue to accrue, as children in succeeding generations avoid unnecessary repeated anaesthetics and examinations.