Blue Cone Monochromatism with Foveal Hypoplasia Caused by the Concomitant Effect of Variants in OPN1LW/OPN1MW and GPR143 Genes.

PurposeTo compare foveal hypoplasia and the appearance of the ellipsoid zone (EZ) at the fovea in patients with genetically confirmed achromatopsia (ACHM) and blue cone monochromacy (BCM).DesignRetrospective, multicenter observational study.ParticipantsMolecularly confirmed patients with ACHM (n = 89) and BCM (n = 33).MethodsWe analyzed high-resolution spectral-domain OCT (SD-OCT) images of the macula from patients with BCM. Three observers independently graded SD-OCT images for foveal hypoplasia (i.e., retention of ≥1 inner retinal layers at the fovea), and 4 observers judged the integrity of the EZ at the fovea, based on an established grading scheme. These measures were compared with previously published data from the patients with ACHM.Main Outcome MeasuresPresence of foveal hypoplasia and EZ grade.ResultsFoveal hypoplasia was significantly more prevalent in ACHM than in BCM (P < 0.001). In addition, we observed a significant difference in the distribution of EZ grades between ACHM and BCM, with grade II EZ being by far the most common phenotype in BCM (61% of patients). In contrast, patients with ACHM had a relatively equal prevalence of EZ grades I, II, and IV. Grade IV EZ was 2.6 times more prevalent in ACHM compared with BCM, whereas grade V EZ (macular atrophy) was present in 3% of both the ACHM and BCM cohorts.ConclusionsThe higher incidence of foveal hypoplasia in ACHM than BCM supports a role for cone activity in foveal development. Although there are differences in EZ grades between these conditions, the degree of overlap suggests EZ grade is not sufficient for definitive diagnosis, in contrast to previous reports. Analysis of additional OCT features in similar cohorts may reveal differences with greater diagnostic value. Finally, the extent to which foveal hypoplasia or EZ grade is prognostic for therapeutic potential in either group remains to be seen, but motivates further study.

An arginine to cysteine substitution at amino acid position 203 (C203R) is the most common missense mutation in human cone opsin. Linked to color blindness and blue cone monochromacy (BCM), C203 is involved in a crucial disulfide bond required for proper folding. It has previously been postulated that expression of mutant C203R cone opsin exerts a toxic effect on cone photoreceptors, similar to some well-characterized missense mutations in rhodopsin that lead to protein misfolding. In this study, we generated and characterized a BCM mouse model carrying the equivalent C203R mutation (Opn1mwC198ROpn1sw–/–) to investigate the disease mechanism and develop a gene therapy approach for this disorder. Untreated Opn1mwC198ROpn1sw–/– cones phenocopied affected cones in human patients with the equivalent mutation, exhibiting shortened or absent cone outer segments and loss of function. We determined that gene augmentation targeting cones specifically yielded robust rescue of cone function and structure when Opn1mwC198ROpn1sw–/– mice were treated at early ages. Importantly, treated cones displayed elaborated outer segments and replenished expression of crucial cone phototransduction proteins. Interestingly, we were unable to detect OPN1MWC198R mutant opsin at any age. We believe this is the first proof-of-concept study exploring the efficacy of gene therapy in BCM associated with a C203R mutation.

PurposeBlue cone monochromacy (BCM), a congenital X-linked retinal disease caused by mutations in the OPN1LW/OPN1MW gene cluster, is under consideration for intravitreal gene therapy. Difficulties with near vision tasks experienced by these patients prompted this study of reading performance as a potential outcome measure for a future clinical trial.MethodsClinically and molecularly diagnosed patients with BCM (n = 17; ages 15–63 years) and subjects with normal vision (n = 22; ages 18–72 years) were examined with the MNREAD acuity chart for both uniocular and binocular conditions. Parameters derived from the measurements in patients were compared with normal data and also within the group of patients. Intersession, interocular and between-subject variabilities were determined. The frequent complaint of light sensitivity in BCM was examined by comparing results from black text on a white background (regular polarity) versus white on black (reverse polarity) conditions.ResultsMNREAD curves of print size versus reading speed were right-shifted compared with normal in all patients with BCM. All parameters in patients with BCM indicated abnormal reading performance. Intersession variability was slightly higher in BCM than in normal, but comparable with results previously reported for other patients with maculopathies. There was a high degree of disease symmetry in reading performance in this BCM cohort. Reverse polarity showed better reading parameters than regular polarity in 82% of the patients.ConclusionsMNREAD measures of reading performance in patients with BCM would be a worthy and robust secondary outcome in a clinical trial protocol, given its dual purpose of quantifying macular vision and addressing an important quality of life issue.Translational RelevanceAssessment of an outcome for a clinical trial.

X-linked cone dysfunction disorders such as Blue Cone Monochromacy and X-linked Cone Dystrophy are characterized by complete loss (of) or reduced L- and M- cone function due to defects in the OPN1LW/OPN1MW gene cluster. Here we investigated 24 affected males from 16 families with either a structurally intact gene cluster or at least one intact single (hybrid) gene but harbouring rare combinations of common SNPs in exon 3 in single or multiple OPN1LW and OPN1MW gene copies. We assessed twelve different OPN1LW/MW exon 3 haplotypes by semi-quantitative minigene splicing assay. Nine haplotypes resulted in aberrant splicing of ≥20% of transcripts including the known pathogenic haplotypes (i.e. ‘LIAVA’, ‘LVAVA’) with absent or minute amounts of correctly spliced transcripts, respectively. De novo formation of the ‘LIAVA’ haplotype derived from an ancestral less deleterious ‘LIAVS’ haplotype was observed in one family with strikingly different phenotypes among affected family members. We could establish intrachromosomal gene conversion in the male germline as underlying mechanism. Gene conversion in the OPN1LW/OPN1MW genes has been postulated, however, we are first to demonstrate a de novo gene conversion within the lineage of a pedigree.

Blue cone monochromacy (BCM) is an X-linked retinal disorder characterized by low vision, photoaversion, and poor color discrimination. BCM is due to the lack of long-wavelength-sensitive and middle-wavelength-sensitive cone photoreceptor function and caused by mutations in the OPN1LW/OPN1MW gene cluster on Xq28. Here, we investigated the prevalence and the landscape of submicroscopic structural variants (SVs) at single-base resolution in BCM patients. We found that about one-third (n = 73) of the 213 molecularly confirmed BCM families carry an SV, most commonly deletions restricted to the OPN1LW/OPN1MW gene cluster. The structure and precise breakpoints of the SVs were resolved in all but one of the 73 families. Twenty-two families-all from the United States-showed the same SV, and we confirmed a common ancestry of this mutation. In total, 42 distinct SVs were identified, including 40 previously unreported SVs, thereby quadrupling the number of precisely mapped SVs underlying BCM. Notably, there was no "region of overlap" among these SVs. However, 90% of SVs encompass the upstream locus control region, an essential enhancer element. Its minimal functional extent based on deletion mapping in patients was refined to 358 bp. Breakpoint analyses suggest diverse mechanisms underlying SV formation as well as in one case the gene conversion-based exchange of a 142-bp deletion between opsin genes. Using parsimonious assumptions, we reconstructed the composition and copy number of the OPN1LW/OPN1MW gene cluster prior to the mutation event and found evidence that large gene arrays may be predisposed to the occurrence of SVs at this locus.

Novel therapeutic approaches for treating inherited retinal degenerations (IRDs) prompt a need to understand which patients with impaired vision have the anatomical potential to gain from participation in a clinical trial. We used supervised machine learning to predict foveal function from foveal structure in blue cone monochromacy (BCM), an X-linked congenital cone photoreceptor dysfunction secondary to mutations in the OPN1LW/OPN1MW gene cluster. BCM patients with either disease-associated large deletion or missense mutations were studied and results compared with those from subjects with other forms of IRD and various degrees of preserved central structure and function. A machine learning technique was used to associate foveal sensitivities and best-corrected visual acuities to foveal structure in IRD patients. Two random forest (RF) models trained on IRD data were applied to predict foveal function in BCM. A curve fitting method was also used and results compared with those of the RF models. The BCM and IRD patients had a comparable range of foveal structure. IRD patients had peak sensitivity at the fovea. Machine learning could successfully predict foveal sensitivity (FS) results from segmented or un-segmented optical coherence tomography (OCT) input. Application of machine learning predictions to BCM at the fovea showed differences between predicted and measured sensitivities, thereby defining treatment potential. The curve fitting method provided similar results. Given a measure of visual acuity (VA) and foveal outer nuclear layer thickness, the question of how many lines of acuity would represent the best efficacious result for each BCM patient could be answered. We propose that foveal vision improvement potential in BCM is predictable from retinal structure using machine learning and curve fitting approaches. This should allow estimates of maximal efficacy in patients being considered for clinical trials and also guide decisions about dosing.

Blue cone monochromacy and gene therapy

The human X chromosome contains ∼ 1600 genes, about 15% of which have been associated with a specific genetic condition, mainly affecting males. Blue cone monochromacy (BCM) is an X-linked condition caused by a loss-of-function of both the OPN1LW and OPN1MW opsin genes. The cone opsin gene cluster is composed of 2-9 paralogs with 99.8% sequence homology and is susceptible to deletions, duplications, and mutations. Current diagnostic tests employ polymerase chain reaction (PCR)-based technologies; however, alterations remain undetermined in 10% of patients. Furthermore, carrier testing in females is limited or unavailable. High-resolution X chromosome-targeted CGH microarray was applied to test for rearrangements in males with BCM and female carriers from three unrelated families. Pathogenic alterations were revealed in all probands, characterized by sequencing of the breakpoint junctions and quantitative real-time PCR. In two families, we identified a novel founder mutation that consisted of a complex 3-kb deletion that embraced the cis-regulatory locus control region and insertion of an additional aberrant OPN1MW gene. The application of high-resolution X-chromosome microarray in clinical diagnosis brings significant advantages in detection of small aberrations that are beyond the resolution of clinically available aCGH analysis and which can improve molecular diagnosis of the known conditions and unravel previously unrecognized X-linked diseases.

Blue cone monochromacy (BCM) is caused by the lack of expression of the normal proteins encoded by the OPN1LW and OPN1MW genes, resulting in the absence of red and green cone sensitivities. We analyzed two cases of BCM in two different families and identified deletion mutations in the locus control region upstream of the two genes. Deletion breakpoints were determined to an accuracy of one base for both cases.

Congenital nystagmus is characterized by involuntary, rhythmical, repeated oscillations of one or both eyes. We studied a large Chinese family with nystagmus as a prominent and consistent manifestation phenotype in nine patients to map and identify a disease-causing gene for nystagmus. X-linked recessive inheritance was observed in the family, and foveal hypoplasia was detected in some of the nine patients. The disease gene was mapped to an approximately 10.6 Mb region flanked by DXS996 and DXS7593 on Xp22 with a significant peak multipoint LOD score. Analysis of 21 candidate genes in the region revealed a novel p.S89F mutation in the second transmembrane domain of GPR143, a G protein-coupled receptor which causes ocular albinism when mutated. All male patients in the family were hemizygous for the mutation; the female carriers were heterozygous for the mutation. The p.S89F mutation was not identified in 100 normal females or 100 normal males. Our results indicate that a mutation in the GPR143 gene can cause a variant form of ocular albinism, with congenital nystagmus as the most prominent and only consistent finding in all patients in this Chinese family. These results expand the spectrum of clinical phenotypes associated with GPR143 mutations.

Blue cone monochromatism (BCM) is a rare cone dystrophy with recessive X-linked inheritance and therefore diagnosed in males whereas females are clinically unaffected. We present a female with clinically manifested BCM. The diagnosis was genetically verified with the identification of one single red-green OPN1LW/MW hybrid gene harboring a point mutation c.607C>G, p.Cys203Arg that associates with BCM and in addition a completely biased X-inactivation in DNA isolated from full blood and buccal mucosa. The present case illustrates that females may develop symptoms of recessive X-linked eye diseases in rare cases.

[Objective] To observe the clinical characteristics of ocular albinism type 1 (OA1) in China.[Methods] Sixteen patients with OA1 and eight female carriers (eight OA1 patients and four carriers were from the same family) were enrolled in this study.All subjects were examined for corrected visual acuity,slit-lamp microscopy and GPR143 gene mutation detection.Some subjects also received indirect ophthalmoscopy,photography of anterior segment and ocular fundus,retinoscopy and optical coherence tomography (OCT).Iris pigmentation was divided into type A,B and C according to the above findings.Type A:slight pigmentation; type B:uneven pigmentation; type C:large patch of hypo-pigmentation with hippus.Result The corrected visual acuity of 16 OA1 patients was from 0.1 to 0.3.All the OAI patients have bilateral horizontal nystagmus without strabismus.All eight subjects received retinoscopy have medium to low hypermetropia and astigmatism.Type A,B and C iris pigmentation were found in eight (50.0％),seven (43.7％) and one (6.3％) patients,respectively.Two (25.0％) carriers have Type A iris pigmentation,and other six carriers (75.0％) were normal.There is no typical albinism retinopathy,however macular foveal structures were never found in these patients.Four carriers have fundus striate or uneven hypo-pigmentation.The macular foveal structure of all the carriers was normal.The macular foveal structure was not found in three subjects who underwent examination of OCT.GPR143 gene mutation was found in all the OA1 patients (100.0％) and all the carriers.[Conclusion]s The major clinical characteristics of Chinese OA1 patients are foveal hypoplasia and GPR143 gene mutation. Key words: Albinism, ocular/complications; Disease attributes

BackgroundBlue Cone Monochromacy (BCM) is an X-linked retinopathy caused by mutations in the OPN1LW / OPN1MW gene cluster, encoding long (L)- and middle (M)-wavelength sensitive cone opsins. Recent evidence shows sufficient structural integrity of cone photoreceptors in BCM to warrant consideration of a gene therapy approach to the disease. In the present study, the vision in BCM is examined, specifically seeking clinically-feasible outcomes for a future clinical trial.MethodsBCM patients (n = 25, ages 5–72) were studied with kinetic and static chromatic perimetry, full-field sensitivity testing, and eye movement recordings. Vision at the fovea and parafovea was probed with chromatic microperimetry.ResultsKinetic fields with a Goldmann size V target were generally full. Short-wavelength (S-) sensitive cone function was normal or near normal in most patients. Light-adapted perimetry results on conventional background lights were abnormally reduced; 600-nm stimuli were seen by rods whereas white stimuli were seen by both rods and S-cones. Under dark-adapted conditions, 500-nm stimuli were seen by rods in both BCM and normals. Spectral sensitivity functions in the superior retina showed retained rod and S-cone functions in BCM under dark-adapted and light-adapted conditions. In the fovea, normal subjects showed L/M-cone mediation using a 650-nm stimulus under dark-adapted conditions, whereas BCM patients had reduced sensitivity driven by rod vision. Full-field red stimuli on bright blue backgrounds were seen by L/M-cones in normal subjects whereas BCM patients had abnormally reduced and rod-mediated sensitivities. Fixation location could vary from fovea to parafovea. Chromatic microperimetry demonstrated a large loss of sensitivity to red stimuli presented on a cyan adapting background at the anatomical fovea and surrounding parafovea.ConclusionsBCM rods continue to signal vision under conditions normally associated with daylight vision. Localized and retina-wide outcome measures were examined to evaluate possible improvement of L/M-cone-based vision in a clinical trial.

Cones are responsible for daylight, central, high acuity and color vision. Three proteins found in human cones, i.e. long-wavelength (L)-, middle-wavelength (M)-, and short-wavelength sensitive (S)-opsins, are responsible for red, green and blue color recognition, respectively. Human blue cone monochromacy (BCM) is characterized by functional loss of both L- and M-cone opsins due to mutations in the OPN1LW/OPN1MW gene cluster on the X chromosome. BCM patients, who rely on their vision from only S-cones and rods, suffer severely reduced visual acuity and impaired color vision. Recent studies show that there is sufficient cone structure remaining in the central fovea of BCM patients to consider AAV-mediated gene augmentation therapy. In contrast, mouse retina has only two opsins, S-opsin and M-opsin, but no L-opsin. We generated an M-opsin knockout mouse (Opn1mw−/−) expressing only S-opsin as a model for human BCM. We show that recombinant M-opsin delivered by AAV5 vectors rescues M-cone function in Opn1mw−/− mice. We also show that AAV delivered M-opsin localizes in the dorsal cone outer segments, and co-localizes with S-opsin in the ventral retina. Our study demonstrates that cones without M-opsin remain viable and respond to gene augmentation therapy, thereby providing proof-of-concept for cone function restoration in BCM patients.

PurposeBlue cone monochromacy (BCM) is an X-linked retinopathy caused by mutations in the red and green cone opsin genes. The aim of this study was to establish the clinical, genetic, and electrophysiological characteristics of a specific form of BCM.MethodsPatients harboring mutations in the OPN1LW/OPN1MW genes underwent a full clinical examination, including ocular examination, color vision, full-field electroretinography, color fundus and autofluorescence photography, and optical coherence tomography. Genetic analysis was performed using whole-exome sequencing, duplex PCR, PCR/restriction fragment length polymorphism, and Sanger sequencing. IBM SPSS Statistics v. 21.0 was used for the data analysis.ResultsTwenty-five patients harboring various haplotypes in exon 3 of the OPN1LW/OPN1MW genes were recruited. They showed a milder incomplete phenotype of BCM than the typical BCM control group. They presented significantly better visual acuity (logarithm of the minimum angle of resolution [logMAR] 0.48 ± 0.26 vs. 1.10 ± 0.54; p < 0.0001) and a highly myopic refraction (−7.81 ± 5.81 D vs. −4.78 ± 5.27 D; p = 0.0222) compared with the BCM control group. The study group had higher 30-Hz cone flicker responses (28.60 ± 15.02 µv; n = 24), whereas the BCM group had none (0.66 ± 2.12 µv; n = 21; p < 0.0001). The Lanthony 15-HUE desaturated test was variable for the exon 3 haplotype group, with a tendency toward the deutan-protan axis.ConclusionsThe present study included genetic and clinical data from the largest cohort of patients with exon 3 haplotypes that were previously shown to cause missplicing of the OPN1LW and OPN1MW genes. Analysis of the clinical data revealed better best-corrected visual acuity, more severe myopia, and higher 30-Hz cone flicker responses in the patients with exon 3 haplotypes than in those with typical BCM.