| Purposes To study the USH2A gene variant profile in a large cohort of Chinese patients with nonsyndromic retinitis pigmentosa(nsRP)or Usher syndrome type Ⅱ(USH2)and to explore the genotype-phenotype correlation.Methods 284 patients from 160 pedigrees with USH2A variants were recruited at the Ophthalmic Genetics Clinic at Peking Union Medical College Hospital and the Ophthalmic Genetics Division of The Eye Hospital of Wenzhou Medical University from 2010 to 2019.Both personal medical history and family histories were reviewed.Comprehensive ocular examinations were performed,including best-corrected Snellen visual acuity(BCVA),colour vision,slit-lamp biomicroscopy,dilated indirect ophthalmoscopy,fundus photography,fundus autofluorescence(FAF),optical coherence tomography(OCT),visual field and electroretinogram(ERG).Audiograms were administered if hearing loss was suspected.5 ml peripheral blood from each patient and their relatives was collected,and genomic DNA was extracted.Targeted exome capture plus next-generation sequencing(NGS)was used to detect the candidate variants.Suspected causative variants were validated by Sanger sequencing and segregation analysis.The genotype-phenotype correlation was evaluated through statistical analyses(SPSS version 25.0.).Results A total of 284 patients from 260 unrelated Chinese families were collected,with age range 1-69 and mean age 38.3.Among them,131 patients in 116 pedigrees were diagnosed with USH2 and 163 patients in 144 pedigrees were nsRP.4 consanguineous families and 3 pseudo-dominant families were identified.1、Clinical findings:① The age range of the 131 USH2 patients was 5-63 years and the mean age was 35.1 years.The age range of nyctalopia onset was 1—40 years and the mean age was 15.4 years.The age range of hearing loss onset was 1-45 years and the mean age was 13.1 years.The onset age of nyctalopia was positively correlated with that of hearing loss(p<0.05,r=0.219).The age range of the low vision was 24-58 years and the median age was 36 years.The age range of the legal blindness was 25-62 years and the median age was 47.5 years.The BCVAs of 242 eyes were recorded,with the distribution as follows:≥ 0.5(55.4%,134/242),0.3～0.5(11.2%,27/242),0.05～0.3(18.6%,45/242),<0.05(14.9%,36/242).The prevalence of cataract,cystoid retinopathy and extinguished ERG were 60%(20/50),52.5%(21/40)and 81%(21/26),respectively.② The age range of the 153 nsRP patients was 1-69 years and the mean age was 40.9 years.The age range of nyctalopia onset was 1-46 years and the mean age was 21.1 years.The age range of the low vision was 27-66 years and the median age was 44.5 years.The age range of the legal blindness was 34-69 years and the median age was 57 years.The BCVAs of 278 eyes were recorded,with the distribution as follows:≥ 0.5(56.5%,157/278),0.3～0.5(12.6%,35/278),0.05～0.3(21.6%,60/278),<0.05(9.4%,26/278).The prevalence of cataract,cystoid retinopathy and extinguished ERG were 54%(34/63),52.5%(31/59)and 65%(24/37),respectively.③The mean age for presentation to our clinic and onset of night blindness for USH2 patients was 5.8 years and 5.7 years earlier than that of nsRP patients,respectively(p<0.001).The median age of legal blindness for USH2 patients was 10 years younger than that of nsRP patients(p<0.01).The BCVA distribution did not show significant differences between RP and USH2 group.The prevalence of cataract between two groups was close and the prevalence of cystoid retinopathy was same.The extinguished ERG was more common in USH2 patients(p<0.01).2、Molecular genetics results:① A total of 284 patients from 260 unrelated Chinese families were identified harboring USH2A disease-associated variants,including 240 probands with biallelic variants and 20 with heterozygous variant.A total of 230 variants in the USH2A gene were identified,including 125(54.4%)missense variants,44(19.1%)stop-gain variants,24(10.4%)splice-site changes and 37(16.1%)frameshift alterations.Most variants(72.6%,167 of 230)were observed only once.Ninety(39.1%,90/230)variants have not been reported previously,of which 50(55.6%)were missense variants,10(11.1%)were stop-gain variants,10(11.1%)were splice-site variants,and 20(22.2%)were frameshift changes.The most frequent allele was p.Tyr2854_Arg2894del(12.7%,66/520),followed by variant p.Cys934Trp(9.6%,50/520).The most frequent location was exon 43(13.8%,72/520),followed by exon 13(11.3%,59/520).The most prevalent domain was the FN3-15 domain(14.0%,73/520),followed by the LE-8 domain(10.2%,53/520).② A total of 116 probands with USH2 were collected,including 103(89%)with biallelic variants and 13(11%)with heterozygous variant.Among the 103 probands,16(15%)with biallelic missense variants,46(45%)with biallelic truncating variants,and 41(40%)with one missense allele and one truncating allele.A total of 232 alleles of 116 probands with USH2 were identified,including 75(32.3%)missense alleles,49(21.1%)stop-gain alleles,57(24.6%)splice-site alleles,38(16.4%)frameshift alleles and 13(5.6%)unsolved alleles.The most frequent allele was p.Tyr2854_Arg2894del(15.9%,37/232),followed by p.Cys934Trp(4.7%,11/232).The most frequent location was exon 43(17.2%,40/232),followed by exon 13(6.5%,15/232).The most prevalent domain was the FN3-15 domain(17.7%,41/232),followed by the LE-8 domain(4.7%,11/232).③A total of 144 probands with nsRP were collected,including 137(95%)with biallelic variants and 7(5%)with heterozygous variant.Among the 137 probands,67(49%)with biallelic missense variants,7(5%)with biallelic truncating variants,and 63(46%)with one missense allele and one truncating allele.A total of 288 alleles of 144 probands with nsRP were identified,including 197(68.4%)missense alleles,19(6.6%)stop-gain alleles,46(16.0%)splice-site alleles,17(5.9%)frameshift alleles and 9(3.1%)unsolved alleles.The most frequent allele was p.Cys934Trp(13.5%,39/288),followed by p.Tyr2854_Arg2894del(10.0%,29/288).The most frequent location was exon 13(15.3%,44/288),followed by exon 43(11.1%,32/288).The most prevalent domain was the LE-8 domain(14.9%,42/288),followed by the FN3-15 domain(11.5%,33/288).④The differences between USH2 group and nsRP group include:the most frequent variants,the distribution of variant type and the percentage of genotype subgroups.NsRP patients have more missense variants but USH2 patients have more truncating variants(p<0.001);nsRP group have more patients with biallelic missense variants but USH2 group have more patients with biallelic truncating variants(p<0.001).3、Genotype-phenotype correlation:The mean age for presentation to our clinic,onset of night blindness and hearing loss for patients harbored biallelic truncating variants was 5.3 years,3.3 years earlier and 8.8 years younger than that of patients carried biallelic missense variants,respectively(p<0.001).The median age of low vision and legal blindness for patients harbored biallelic truncating variants was 8 years and 8.5 years earlier than that of patients carried biallelic missense variants,respectively(p<0.001).And patients harbored biallelic truncating variants had more eyes with BCVA<0.03 than patients carried biallelic missense variants(p<0.05).Conclusions This study enrolled the largest cohort of Chinese USH2A patients and expanded the variant spectrum of USH2A gene.The most frequent variants and variant atlas between USH2 and nsRP groups were different.USH2 patients have more truncating variants and experience an earlier decline in visual function. |
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