Truncation mutations in MYRF underlie primary angle closure glaucoma.

Mutations in myelin regulatory factor (MYRF), a gene mapped to 11q12-q13.3, are responsible for autosomal dominant high hyperopia and seem to be associated with angle closure glaucoma, which is one of the leading causes of irreversible blindness worldwide. Whether there is a causal link from the MYRF mutations to the pathogenesis of primary angle-closure glaucoma (PACG) remains unclear at this time. Six truncation mutations, including five novel and one previously reported, in MYRF are identified in seven new probands with hyperopia, of whom all six adults have glaucoma, further confirming the association of MYRF mutations with PACG. Immunofluorescence microscopy demonstrates enriched expression of MYRF in the ciliary body and ganglion cell layer in humans and mice. Myrfmut/+ mice have elevated IOP and fewer ganglion cells along with thinner retinal nerve fiber layer with ganglion cell layer than wild-type. Transcriptome sequencing of Myrfmut/+ retinas shows downregulation of Dnmt3a, a gene previously associated with PACG. Co-immunoprecipitation demonstrates a physical association of DNMT3A with MYRF. DNA methylation sequencing identifies several glaucoma-related cell events in Myrfmut/+ retinas. The interaction between MYRF and DNMT3A underlies MYRF-associated PACG and provides clues for pursuing further investigation into the pathogenesis of PACG and therapeutic target.

Defect of TIMP4 Is Associated with High Myopia and Participates in Rat Ocular Development in a Dose-Dependent Manner.

Thinning of the sclera happens in myopia eyes owing to extracellular matrix (ECM) remodeling, but the initiators of the ECM remodeling in myopia are mainly unknown. The matrix metalloproteinase (MMPs) and tissue inhibitors of matrix metalloproteinase (TIMPs) regulate the homeostasis of the ECM. However, genetic studies of the MMPs and TIMPs in the occurrence of myopia are poor and limited. This study systematically investigated the association between twenty-nine genes of the TIMPs and MMPs families and early-onset high myopia (eoHM) based on whole exome sequencing data. Two TIMP4 heterozygous loss-of-function (LoF) variants, c.528C>A in six patients and c.234_235insAA in one patient, were statistically enriched in 928 eoHM probands compared to that in 5469 non-high myopia control (p = 3.7 × 10-5) and that in the general population (p = 2.78 × 10-9). Consequently, the Timp4 gene editing rat was further evaluated to explore the possible role of Timp4 on ocular and myopia development. A series of ocular morphology abnormalities in a dose-dependent manner (Timp4-/- < Timp4+/- < Timp4+/+) were observed in a rat model, including the decline in the retinal thickness, the elongation in the axial length, more vulnerable to the form deprivation model, morphology changes in sclera collagen bundles, and the decrease in collagen contents of the sclera and retina. Electroretinogram revealed that the b-wave amplitudes of Timp4 defect rats were significantly reduced, consistent with the shorter length of the bipolar axons detected by HE and IF staining. Heterozygous LoF variants in the TIMP4 are associated with early onset high myopia, and the Timp4 defect disturbs ocular development by influencing the morphology and function of the ocular tissue.

New Insight into the Genotype-Phenotype Correlation of PRPH2-Related Diseases Based on a Large Chinese Cohort and Literature Review.

Variants in PRPH2 are a common cause of inherited retinal dystrophies with high genetic and phenotypic heterogeneity. In this study, variants in PRPH2 were selected from in-house exome sequencing data, and all reported PRPH2 variants were evaluated with the assistance of online prediction tools and the comparative validation of large datasets. All variants were classified based on the American College of Medical Genetics and Genomics/Association for Molecular Pathology (ACMG/AMP) guidelines. Individuals with pathogenic or likely pathogenic variants of PRPH2 were confirmed by Sanger sequencing. Clinical characteristics were summarized. Ten pathogenic or likely pathogenic variants of PRPH2 were identified in 14 families. In our cohort, the most frequent variant was p.G305Afs*19, accounting for 33.3% (5/15) of alleles, in contrast to the literature, where p.R172G (11.6%, 119/1028) was the most common variant. Nine in-house families (63.8%) were diagnosed with retinitis pigmentosa (RP), distinct from the phenotypic spectrum in the literature, which shows that RP accounts for 27.9% (283/1013) and macular degeneration is more common (45.2%, 458/1013). Patients carrying missense variants predicted as damaging by all seven prediction tools and absent in the gnomAD database were more likely to develop RP compared to those carrying missense variants predicted as damaging with fewer tools or with more than one allele number in the gnomAD database (p = 0.001). The population-specific genetic and phenotypic spectra of PRPH2 were explored, and novel insight into the genotype-phenotype correlation of PRPH2 was proposed. These findings demonstrated the importance of assessing PRPH2 variants in distinct populations and the value of providing practical suggestions for the genetic interpretation of PRPH2 variants.

Variants in HNRNPH1 are associated with high myopia in humans and ocular coloboma in zebrafish.

High myopia is one of the most common causes for blindness due to its associated complications. Genetic factor has been considered as the major cause for early-onset high myopia (eoHM), but exact genetic defects for most eoHM are yet to be identified. Through multi-step bioinformatics analysis of our in-house whole exome sequencing dataset from 6397 individuals, variants from 928 probands with eoHM were further compared with those from in-house controls as well as gnomAD database. The results showed that loss-of-function (LoF) variants in a novel gene HNRNPH1 were identified in two of 928 probands with eoHM but in none of 5469 probands with other eye conditions (p = 0.02). LoF variants in HNRNPH1 were extremely rare and intolerant, while two LoF variants in 928 eoHM were statistically higher than their frequency in gnomAD (p = 5.98 × 10-4 ). These two LoF variants, c.2dup/p.(M1?) and c.121dup/p.(Q41Pfs*20), were absent from existing database. Variants in HNRNPH1 have not been associated with any inherited eye disease before. Expression of HNRNPH1 was enriched in ganglion cell layer and inner nuclear layer in humans. Knockdown of hnrnph1 in zebrafish resulted in ocular coloboma. All these suggest that HNRNPH1 is potential contribution to eoHM when mutated.

CPSF1 mutations are associated with early-onset high myopia and involved in retinal ganglion cell axon projection.

High myopia is a severe form of nearsightedness, which can result in blindness due to its associated complications. While both genetic and environmental factors can cause high myopia, early-onset high myopia (eoHM), which is defined as high myopia that occurs before school age, is considered to be caused mainly by genetic variations, with minimal environmental involvement. Here we report six rare heterozygous loss-of-function (LoF) variants in CPSF1 that were identified in six of 623 probands with eoHM but none of 2657 probands with other forms of genetic eye diseases; this difference was statistically significant (P = 4.60 × 10-5, Fisher's exact test). The six variants, which were confirmed by Sanger sequencing, were c.3862_3871dup (p.F1291*), c.2823_2824del (p.V943Lfs*65), c.1858C>T (p.Q620*), c.15C>G (p.Y5*), c.3823G>T (p.D1275Y) and c.4146-2A>G. Five of these six variants were absent in existing databases, including gnomAD, 1000G and EVS. The remaining variant, c.4146-2A>G, was present in gnomAD with a frequency of 1/229918. Clinical data demonstrated eoHM in the six probands with these mutations. Knockdown of cpsf1 by morpholino oligonucleotide (MO) injection in zebrafish eggs resulted in small eye size in 84.38% of the injected larvae, and this phenotype was rescued in 61.39% of the zebrafish eggs when the cpsf1 MO and the cpsf1 mRNA were co-injected. The projection of retinal ganglion cell (RGC) towards the tectum was abnormal in cpsf1 morphants. Thus, we demonstrated that heterozygous LoF mutations in CPSF1 are associated with eoHM and that CPSF1 may play an important role in the development of RGC axon projection.

Confirming and expanding the phenotypes of FZD5 variants: Coloboma, inferior chorioretinal hypoplasia, and high myopia.

Purpose: Two frameshift and two indel variants in FZD5 have been reported to cause coloboma in two families with incomplete penetrance and in two isolated cases in previous studies, respectively. This study aims to confirm this association and expand related specific phenotypes based on the genotype-phenotype analysis of FZD5 variants. Methods: Variants in FZD5 were collected from our in-house exome sequencing data of 5,845 probands with different eye conditions. Multistep bioinformatics analysis was used to classify the variants. Potential pathogenic variants and phenotypic variations were further evaluated based on family segregation and genotype-phenotype analysis. Results: In total, 63 rare variants were detected in FZD5. Multistep bioinformatics and genotype-phenotype analyses suggested that eight rare heterozygous variants in nine families should be considered potential pathogenic variants: three novel frameshift variants (c.350_356delCGCCGCT/p.Ser117*, c.1403_1406dupACCT/p.Tyr470Profs*130, and c.1428delG/p.Ser477Alafs*130) and five novel missense variants (c.388C>A/p.Arg130Ser, c.794G>T/p.Arg265Leu, c.1162G>A/p.Gly388Ser, c.1232A>G/p.Tyr411Cys, and c.1510A>T/p.Met504Leu). Among the nine families, carriers of these variants showed overlapping phenotypes, including typical uveal coloboma (12 eyes of seven patients from four families), inferior chorioretinal hypoplasia (ICH) or optic disc hypoplasia (ODH; 12 eyes of eight patients from six families), and high myopia (10 eyes of five patients from five families) within individual families or among different families. Conclusions: The data presented in this study confirmed that variants in FZD5, not only frameshift variants but also missense variants, are a common cause of uveal coloboma. In addition, ICH, ODH, and high myopia may be variant phenotypes that are frequently associated with FZD5 variants.

Heterozygous GJA1 variants with ocular phenotype: Missense in domain but truncation out of domain.

Purpose: Oculodentodigital dysplasia (ODDD) is a group disorder caused by GJA1 variants, of which glaucoma leading to blindness is a frequent complication of the ocular phenotype. In this study, the correlation of the GJA1 genotype with the ocular phenotype was analyzed systematically. Methods: GJA1 variants were collected from in-house whole-exome sequencing data of 5,307 individuals. Potentially pathogenic variants (PPVs) were defined based on prediction of multiple in silico tools, related phenotypes, and previously established evidence. The characteristics of GJA1 PPVs were evaluated based on our data, gnomAD, and HGMD. Results: In total, 21 rare variants in GJA1 were detected in 32 subjects from the study cohort. Four of the 21 variants were classified as PPVs, including two frameshift, one missense, and one in-frame deletion. The four PPVs were detected in four probands with microcornea or high hyperopia; two developed glaucoma. A systematic review of GJA1 variants in literature suggested that most heterozygous missense PPVs are located inside the connexin domain. All truncations downstream of the connexin domain are associated with autosomal dominant disease, while most truncations within the domain are associated with autosomal recessive ODDD. Ocular signs were present in 80.0% (116/145) of patients with GJA1 PPVs. Of the 116 patients, glaucoma was observed in 26.7% (31/116), among whom 77.4% (24/31) of cases occurred in patients ≥10 years old. Conclusions: Eye abnormalities are the most common signs associated with GJA1 PPVs, and they carry a high risk of developing glaucoma. The identification of GJA1 PPVs needs further attention and clarification.

Biallelic mutations in USP45, encoding a deubiquitinating enzyme, are associated with Leber congenital amaurosis.

BACKGROUND: Leber congenital amaurosis (LCA) is the earliest and most severe form of inherited retinal dystrophies. In approximately 56% of Chinese probands, genetic defects can be detected in known LCA-causing genes. In this study, the objective was to identify pathogenic variants in two unsolved Chinese families with LCA. METHODS: To identify the genetic defect, whole-exome sequencing (WES) and clinical analysis was performed in both probands with LCA as well as in 3011 in-house controls with other hereditary eye diseases. The expression profiles, as well as the phenotype analysis of knockdown zebrafish model and knockout mice model, were performed to investigate the function of USP45 in photoreceptors. RESULTS: By analysing WES data based on allele frequencies of in-house controls, population allele frequencies and in silico prediction tools, two rare homozygous mutations in USP45 were identified in two unrelated families. Immunohistochemistry of USP45 in the human and zebrafish retinal sections revealed enriched expression in the inner segments of photoreceptors. The knockdown of usp45 transcript in zebrafish led to abnormal retinal development with effects on photoreceptors, which could be successfully rescued by wild-type usp45 mRNA. Moreover, targeted knockout of Usp45 in mice caused abnormal electroretinography responses, similar to that seen in patients with LCA. CONCLUSIONS: Our study implicates that biallelic mutations in USP45 are associated with the occurrence of LCA. Moreover, our results indicate that USP45 is indispensable to the maintenance of photoreceptor function.

Novel truncation mutations in MYRF cause autosomal dominant high hyperopia mapped to 11p12-q13.3.

High hyperopia is a common and severe form of refractive error. Genetic factors play important roles in the development of high hyperopia but the exact gene responsible for this condition is mostly unknown. We identified a large Chinese family with autosomal dominant high hyperopia. A genome-wide linkage scan mapped the high hyperopia to chromosome 11p12-q13.3, with maximum log of the odds scores of 4.68 at theta = 0 for D11S987. Parallel whole-exome sequencing detected a novel c.3377delG (p.Gly1126Valfs*31) heterozygous mutation in the MYRF gene within the linkage interval. Whole-exome sequencing in other 121 probands with high hyperopia identified additional novel mutations in MYRF within two other families: a de novo c.3274_3275delAG (p.Leu1093Profs*22) heterozygous mutation and a c.3194+2T>C heterozygous mutation. All three mutations are located in the C-terminal region of MYRF and are predicted to result in truncation of that portion. Two patients from two of the three families developed angle-closure glaucoma. These three mutations were present in neither the ExAC database nor our in-house whole-exome sequencing data from 3280 individuals. No other truncation mutations in MYRF were detected in the 3280 individuals. Knockdown of myrf resulted in small eye size in zebrafish. These evidence all support that truncation mutations in the C-terminal region of MYRF are responsible for autosomal dominant high hyperopia in these families. Our results may provide useful clues for further understanding the functional role of the C-terminal region of this critical myelin regulatory factor, as well as the molecular pathogenesis of high hyperopia and its associated angle-closure glaucoma.

The Systemic Genotype-Phenotype Characterization of PAX6-Related Eye Disease in 164 Chinese Families.

Purpose: This study aims to evaluate the genetic and phenotypic characteristics and elucidate the genotype-phenotype correlations of a large Chinese cohort with PAX6-related disorders. Methods: Variants detected with exome sequencing were filtered through multistep bioinformatic and co-segregation analyses, and validated by Sanger sequencing. The related clinical data were collected, and cluster analysis and statistical analysis of the PAX6-related phenotypes across different variant groups were carried out. Parental mosaicism was investigated using cloning analysis and Droplet digital PCR. Results: A total of 119 pathogenic or likely pathogenic PAX6 variants, including 74 truncation, 31 missense, and 14 others, were identified in 228 patients from 164 unrelated families. The most common phenotypes were foveal hypoplasia (97.8%), nystagmus (92.6%), aniridia (76.7%), cataract (36.8%), and iris hypoplasia (22.4%). Mosaicism ranging from 13.9% to 18.8% was identified in 3 unrelated patients' parents with relatively mild phenotypes. Missense variants in the linker region of the paired domain were associated with high myopia, whereas truncation variants in the homeodomain and proline-serine-threonine-rich domain were associated with hyperopia. Similarly, the degree of iris defects, visual acuity, and associated ocular comorbidity varied among the different types and locations of PAX6 variants. Conclusions: Our data indicate that foveal hypoplasia but not aniridia is the most common sign of PAX6-related disorders, contributing to subtle iris changes that might easily be overlooked in clinical practice. Recognition of mosaicism in atypical cases or parents with very mild phenotypes is important in genetic counseling as their offspring are at increased risk of typical aniridia. Recognition of the genotype-phenotype relationship emphasizes involvement of PAX6 regulation in shaping complex ocular phenotypes.

Altered chromatin topologies caused by balanced chromosomal translocation lead to central iris hypoplasia.

Despite the advent of genomic sequencing, molecular diagnosis remains unsolved in approximately half of patients with Mendelian disorders, largely due to unclarified functions of noncoding regions and the difficulty in identifying complex structural variations. In this study, we map a unique form of central iris hypoplasia in a large family to 6q15-q23.3 and 18p11.31-q12.1 using a genome-wide linkage scan. Long-read sequencing reveals a balanced translocation t(6;18)(q22.31;p11.22) with intergenic breakpoints. By performing Hi-C on induced pluripotent stem cells from a patient, we identify two chromatin topologically associating domains spanning across the breakpoints. These alterations lead the ectopic chromatin interactions between APCDD1 on chromosome 18 and enhancers on chromosome 6, resulting in upregulation of APCDD1. Notably, APCDD1 is specifically localized in the iris of human eyes. Our findings demonstrate that noncoding structural variations can lead to Mendelian diseases by disrupting the 3D genome structure and resulting in altered gene expression.

Clinical and Genetic Landscape of Ectopia Lentis Based on a Cohort of Patients From 156 Families.

Purpose: To extend the mutation spectrum and explore the characteristics of genotypes and ocular phenotypes in ectopia lentis (EL). Methods: Variants in all 14 reported EL-associated genes were selected from in-house data sets as well as literature review, and available clinical data were analyzed. Results: Likely pathogenic variants in three genes were identified in 156 unrelated families with EL from the in-house cohort, of which 97.4% resulted from variants in FBN1, whereas the remaining were caused by variants in ADAMTSL4 (1.3%) and LTBP2 (1.3%). A comparative analysis of the in-house data and literature review suggested several characteristics: (1) a higher proportion of cysteine involvement variants in FBN1, either variants introducing or eliminating cysteine, and an earlier diagnosis age were presented in our cohort than in published literature; (2) the axial length (AL) and refractive error increased more rapidly with age in preschool EL children than normal children, and the increased rate of AL was slower in patients with surgery than those without surgery; (3) aberrant astigmatism was common in EL; and (4) worse vision and earlier onset age were observed in patients with non-FBN1 variants (all P < 0.05). Conclusions: Variants in FBN1 are the predominant cause of EL, with the most common cysteine involvement variants. Early-stage EL manifests refractive error but gradually converts to axial myopia through defocus introduced by lens dislocation. Aberrant astigmatism is a suggestive sign of EL. Non-FBN1 variants cause early-onset and severe phenotypes. These results provide evidence for early diagnosis as well as timely treatment for EL.

Clinical and genetic features of retinoschisis in 120 families with RS1 mutations.

BACKGROUND/AIMS: X-linked retinoschisis (XLRS), associated with RS1, is the most common type of X-linked retinopathy in children. This study aimed to identify clinical and genetic features of retinoschisis in 120 families with RS1 variants in China. METHODS: RS1 variants were collected from our in-house exome data and were predicted by multiple-step bioinformatics analysis. Clinical data of 122 patients from 120 families with potential pathogenic RS1 variants were analysed and summarised, respectively. RESULT: Totally, 79 hemizygous variants (53 missense, 25 truncation and 1 indel), were detected. All except one (78/79, 98.7%), including 22 novels, were classified as potential pathogenic and detected exclusively in 120 families with retinoschisis. Clinical data demonstrated an average age of presentation at 5 years (1 month-41 years). Macular changes were classified as macular schisis (87.5%), macular atrophy (10.7%), normal (0.9%) and unclassified (0.9%). Patients with macular atrophy had older age but similar visual acuity compared with macular schisis. Peripheral retinal changes included flat retinoschisis (52.4%), bullous retinoschisis (BRS) (10.7%) and normal-like (36.9%) patients. Spontaneous regression was observed in two patients with BRS on follow-up examination. Visual acuity in the peripheral retinoschisis group was worse than that without peripheral retinoschisis. CONCLUSION: Almost all rare RS1 variants were potential pathogenic. All patients with RS1 pathogenic variants showed detectable characteristics in the macula and/or peripheral retina. Our data on RS1 variants and associated clinical phenotypes may be of value for clinical diagnosis and genetic test of retinoschisis.

FDXR-Associated Oculopathy: Congenital Amaurosis and Early-Onset Severe Retinal Dystrophy as Common Presenting Features in a Chinese Population.

Variants in FDXR reportedly cause autosomal recessive auditory neuropathy and optic atrophy, expanding to retinal dystrophy. This study aimed to further clarify associated phenotypes. FDXR variants were selected from our in-house whole-exome sequencing dataset of 6397 families with different eye conditions. The clinical data of the identified patients were summarized. Biallelic pathogenic or likely pathogenic FDXR variants were identified in 11 unrelated patients, including 14 missense variants of which 10 were novel. Fundus observation showed complete optic disc pallor, silver wiring or severe attenuation of retinal vessels, and varying degrees of generalized retinal degeneration. Before the detection of FDXR variants, four patients were clinically diagnosed as congenital amaurosis due to the presence of nystagmus a few months after birth, while seven were diagnosed as early-onset severe retinal dystrophy due to the presence of nyctalopia and/or poor vision in early childhood. Biallelic FDXR variants are a frequent cause of congenital or early-onset severe retinal dystrophy, especially for patients with severe optic atrophy and retinal dystrophy in early childhood.

Pathogenic Variants in CEP290 or IQCB1 Cause Earlier-Onset Retinopathy in Senior-Loken Syndrome Compared to Those in INVS, NPHP3, or NPHP4.

PURPOSE: Senior-Loken syndrome (SLSN) is an autosomal recessive disorder characterized by retinopathy and nephronophthisis. This study aimed to evaluate whether different phenotypes are associated with different variants or subsets of 10 SLSN-associated genes based on an in-house data set and a literature review. DESIGN: Retrospective case series. METHODS: Patients with biallelic variants in SLSN-associated genes, including NPHP1, INVS, NPHP3, NPHP4, IQCB1, CEP290, SDCCAG8, WDR19, CEP164, and TRAF3IP1, were recruited. Ocular phenotypes and nephrology medical records were collected for comprehensive analysis. RESULTS: Variants in 5 genes were identified in 74 patients from 70 unrelated families, including CEP290 (61.4%), IQCB1 (28.6%), NPHP1 (4.2%), NPHP4 (2.9%), and WDR19 (2.9%). The median age at the onset of retinopathy was approximately 1 month (since birth). Nystagmus was the most common initial sign in patients with CEP290 (28 of 44, 63.6%) or IQCB1 (19 of 22, 86.4%) variants. Cone and rod responses were extinguished in 53 of 55 patients (96.4%). Characteristic fundus changes were observed in CEP290- and IQCB1-associated patients. During follow-up, 70 of the 74 patients were referred to nephrology, among whom nephronophthisis was not detected in 62 patients (88.6%) at a median age of 6 years but presented in 8 patients (11.4%) aged approximately 9 years. CONCLUSIONS: Patients with pathogenic variants in CEP290 or IQCB1 presented early with retinopathy, whereas other patients with INVS, NPHP3, or NPHP4 variants first developed nephropathy. Therefore, awareness of the genetic and clinical features may facilitate the clinical management of SLSN, especially early intervention of kidney problems for patients with eyes affected first.

The Genetic Confirmation and Clinical Characterization of LOXL3-Associated MYP28: A Common Type of Recessive Extreme High Myopia.

Purpose: In previous studies, biallelic LOXL3 variants have been shown to cause autosomal recessive Stickler syndrome in one Saudi Arabian family or autosomal recessive early-onset high myopia (eoHM, MYP28) in two Chinese families. The current study aims to elucidate the clinical and genetic features of LOXL3-associated MYP28 in seven new families and two previously published families. Methods: LOXL3 variants were detected based on the exome sequencing data of 8389 unrelated probands with various ocular conditions. Biallelic variants were identified through multiple online bioinformatic tools, comparative analysis, and co-segregation analysis. The available clinical data were summarized. Results: Biallelic LOXL3 variants were exclusively identified in nine of 1226 families with eoHM but in none of the 7163 families without eoHM (P = 2.97 × 10-8, Fisher's exact test), including seven new and two previously reported families. Seven pathogenic variants were detected, including one nonsense (c.1765C>T/p.Arg589*), three frameshift (c.39dupG/p.Leu14Alafs*21; c.544delC/p.Leu182Cysfs*3, c.594delG/p.Gln199Lysfs*35), and three missense (c.371G>A/p.Cys124Tyr; c.1051G>A/p.Gly351Arg; c.1669G>A/p.Glu557Lys) variants. Clinical data of nine patients from nine unrelated families revealed myopia at the first visit at about 5 years of age, showing slow progression with age. Visual acuity at the last visit ranged from 0.04 to 0.9 (median age at last visit = 5 years, range 3.5-15 years). High myopic fundus changes, observed in all nine patients, were classified as tessellated fundus (C1) in five patients and diffuse choroidal atrophy (C2) in four patients. Electroretinograms showed mildly reduced cone responses and normal rod responses. Except for high myopia, no other specific features were shared by these patients. Conclusions: Biallelic LOXL3 variants exclusively presenting in nine unrelated patients with eoHM provide firm evidence implicating MYP28, with an estimated prevalence of 7.3 × 10-3 in eoHM and of about 7.3 × 10-5 in the general population for LOXL3-associated eoHM. So far, MYP28 represents a common type of autosomal recessive extreme eoHM, with a frequency comparable to LRPAP1-associated MYP23.

Unique Haplotypes in OPN1LW as a Common Cause of High Myopia With or Without Protanopia: A Potential Window Into Myopic Mechanism.

Purpose: Specific haplotypes (LVAVA, LIVVA, and LIAVA) formed by five polymorphisms (p.L153M, p.V171I, p.A174V, p.I178V, and p.S180A in exon 3 of OPN1LW) that cause partial or complete exon skipping have been reported as unique genetic causes of high myopia with or without colorblindness. This study aimed to identify the contribution of OPN1LW to early-onset high myopia (eoHM) and the molecular basis underlying eoHM with or without colorblindness. Methods: Comparative analysis of exome sequencing data was conducted for 1226 families with eoHM and 9304 families with other eye conditions. OPN1LW variants detected by targeted or whole exome sequencing were confirmed by long-range amplification and Sanger sequencing, together with segregation analysis. The clinical data were thoroughly analyzed. Results: Unique haplotypes and truncation variants in OPN1LW were detected exclusively in 68 of 1226 families with eoHM but in none of the 9304 families with other visual diseases (P = 1.63 × 10-63). Four classes of variants were identified: haplotypes causing partial splicing defects in OPN1LW (LVAVA or LIVVA in 31 families), LVAVA in OPN1LW-OPN1MW hybrid gene (in 3 families), LIAVA in OPN1LW (in 29 families), and truncations in OPN1LW (in 5 families). The first class causes partial loss of red photopigments, whereas the latter three result in complete loss of red photopigments. This is different from the replacement of red with green owing to unequal re-arrangement causing red-green colorblindness alone. Of the 68 families, 42 affected male patients (31 families) with the first class of variants (LVAVA or LIVVA in OPN1LW) had eoHM alone, whereas 37 male patients with the latter 3 classes had eoHM with protanopia. Adaptive optics retinal imaging demonstrated reduced cone regularity and density in men with eoHM caused by OPN1LW variants compared to those patients with eoHM and without OPN1LW variants. Conclusion: Based on the 68 families with unique variants in OPN1LW, our study provides firm evidence that the two different phenotypes (eoHM with or without colorblindness) are caused by two different classes of variants (partial splicing-effect haplotypes or complete splicing-effect haplotypes/truncation variants, respectively). The contribution of OPN1LW to eoHM (isolated and syndromic) was characterized by OPN1LW variants found in 5.5% (68/1226) of the eoHM families, making it the second most common cause of monogenic eoHM alone (2.4%) and a frequent cause of syndromic monogenic eoHM with colorblindness. Such haplotypes, in which each individual variant alone is considered a benign polymorphism, are potential candidates for other hereditary diseases with causes of missing genetic defects.

Different Phenotypes Represent Advancing Stages of ABCA4-Associated Retinopathy: A Longitudinal Study of 212 Chinese Families From a Tertiary Center.

Purpose: To evaluate the nature and association of different phenotypes associated with ABCA4 mutations in Chinese. Methods: All patients were recruited from our pediatric and genetic eye clinic. Detailed ocular phenotypes were characterized. The disease course was evaluated by long-term follow-up observation, with a focus on fundus changes. Cox regression was used to identify the factors associated with disease progression. Results: A systematic review of genetic and clinical data for 228 patients and follow-up data for 42 patients indicated specific features in patients with two ABCA4 variants. Of 185 patients with available fundus images, 107 (57.8%) showed focal lesions restricted to the central macula without flecks. Among these 107 patients, 30 patients (28.0%) initially presented with relatively preserved visual acuity and inconspicuous performance on routine fundus screening. A pigmentary change in the posterior pole was observed in 22 of 185 patients (11.9%), and this change mimicked retinitis pigmentosa in 10 cases (45.5%). Follow-up visits and sibling comparisons demonstrated disease progression from cone-rod dystrophy, Stargardt disease, to retinitis pigmentosa. An earlier age of onset was associated with a more rapid decrease in visual acuity (P = 0.03). Patients with two truncation variants had an earlier age of onset. Conclusion: Phenotypic variation in ABCA4-associated retinopathy may represent sequential changes in a single disease: early-stage Stargardt disease may resemble cone-rod dystrophy, whereas the presence of diffuse pigmentation in the late stage may mimic retinitis pigmentosa. Recognizing the natural progression of fundus changes, especially those visualized by wide-field fundus autofluorescence, is valuable for diagnostics and therapeutic decision-making.

Genotype-Phenotype Analysis of RPGR Variations: Reporting of 62 Chinese Families and a Literature Review.

PURPOSE: RPGR is the most common cause of X-linked retinitis pigmentosa (RP), of which female carriers are also frequently affected. The aim of the current study was to explore the RPGR variation spectrum and associated phenotype based on the data from our lab and previous studies. METHODS: Variants in RPGR were selected from exome sequencing data of 7,092 probands with different eye conditions. The probands and their available family members underwent comprehensive ocular examinations. Similar data were collected from previous reports through searches in PubMed, Web of Science, and Google Scholar. Systematic analyses of genotypes, phenotypes and their correlations were performed. RESULTS: A total of 46 likely pathogenic variants, including nine missense and one in-frame variants in RCC1-like domain and 36 truncation variants, in RPGR were detected in 62 unrelated families in our in-house cohort. In addition, a total of 585 variants, including 491 (83.9%) truncation variants, were identified from the literature. Systematic analysis of variants from our in-house dataset, literature, and gnomAD suggested that most of the pathogenic variants of RPGR were truncation variants while pathogenic missense and in-frame variants were enriched in the RCC1-like domain. Phenotypic variations were present between males and female carriers, including more severe refractive error but better best corrected visual acuity (BCVA) in female carriers than those in males. The male patients showed a significant reduction of BCVA with increase of age and males with exon1-14 variants presented a better BCVA than those with ORF15 variants. For female carriers, the BCVA also showed significant reduction with increase of age, but BCVA in females with exon1-14 variants was not significant difference compared with those with ORF15 variants. CONCLUSION: Most pathogenic variants of RPGR are truncations. Missense and in-frame variants located outside of the RCC1-like domain might be benign and the pathogenicity criteria for these variants should be considered with greater caution. The BCVA and refractive error are different between males and female carriers. Increase of age and location of variants in ORF15 contribute to the reduction of BCVA in males. These results are valuable for understanding genotypes and phenotypes of RPGR.

Novel BMP4 Truncations Resulted in Opposite Ocular Anomalies: Pathologic Myopia Rather Than Microphthalmia.

BMP4 variants have been reported to be associated with syndromic microphthalmia (MCOPS6, OMIM 607932). This study aims to describe BMP4 truncation mutations contributing to a novel phenotype in eight patients from four Chinese families. In this study, BMP4 variants were collected from a large dataset from in-house exome sequencing. Candidate variants were filtered by multiple in silico tools as well as comparison with data from multiple databases. Potential pathogenic variants were further confirmed by Sanger sequencing and cosegregation analysis. Four novel truncation variants in BMP4 were detected in four out of 7,314 unrelated probands with different eye conditions. These four mutations in the four families solely cosegregated in all eight patients with a specific form of pathologic myopia, characterized by significantly extended axial length, posterior staphyloma, macula patchy, chorioretinal atrophy, myopic optic neuropathy or glaucoma, vitreous opacity, and unique peripheral snow-grain retinopathy. The extreme rarity of the truncations in BMP4 (classified as intolerant in the gnomAD database, pLI = 0.96), the exclusive presence of these variants in the four families with pathologic myopia, variants fully co-segregated with the same specific phenotypes in eight patients from the four families, and the association of the pathogenicity of truncations with syndromic microphthalmia in previous studies, all support a novel association of BMP4 truncations with a specific form of pathologic myopia. The data presented in this study demonstrated that heterozygous BMP4 truncations contributed to a novel phenotype: pathologic myopia rather than microphthalmia. Mutations in the same gene resulting in both high myopia and microphthalmia have been observed for a few other genes like FZD5 and PAX6, suggesting bidirectional roles of these genes in early ocular development. Further studies are expected to elucidate the molecular mechanism of the bidirectional regulation.