Study Of Molecular Genetic Characteristics In Chineses Population With LCA Based On Next-generation Sequencing

Purpose:Investigation on phenotypes and pathological changes in Chinese population with Leber congenital amaurosis(LCA),and further analysis on the correlations between genotype and phenotype by exploring new phenotype features with novel mutation sites.Comprehensive analysis and evaluation of potential novel causative genetic characteristics in Chinese population with LCA Method: Patients(2011-5 to 2015-5)were collected in our hospital with strictly diagnosed as LCA by their clinical features.All patients underwent fundus screening and full field electroretinogram(ERG)examination,and optical coherence tomography(OCT)and other imaging examination were performed on available patients.All enrolled patients underwent targeted next-generation sequencing for known disease genes screening and followed by pathogenic analysis(in silico prediction)on suspicious pathogenic site.Potential causative variants were verified by Sanger sequencing and co-segregation analysis.Families with no known genes detected underwent whole exome sequencing,homozygous mutations were selected as candidate loci for annotation.Pathogenic analysis and GO,KEGG pathway analysis and co-segregation analysis were performed subsequently.Expanded verification for potential pathogenic locus were performed on more samples.Result: Totally 65 patients with LCA were enrolled with age of 4 months ~ 8 years old.Retinal pigment changes are diverse,detected in 72% of total patients.42% patients presented macular degeneration,but the severity of maculopathy didn't correspond to the surrounding pigment changes.OCT shows macular various structural changes,with foveal thickening detected in 48% patients.All patients underwent targeted second-generation sequencing and new causative mutations were found in 35 patients with good or high confidence,who are all from the east coast.That is totally 46 new variants covering 15 known disease genes.Four patients carried the same AIPL1 mutations(c.C241 T p.Q81X),which were homozygous in patients 1 and 2.One patient with novel mutations in NMNAT1 showed peripherally exudative vasculopathy with fluorescein angiography.Another patient with RPE65 mutations showed peripheral vascular occlusion and white sheath.One patient with novel mutation in CRB1 showed proliferative membrane covered the macula and numerous fine,glistening crystals appearing in both eyes.Ten families underwent whole-exome sequencing by which homozygous mutations of KRT12 and CYP1A1 were found in three pedigrees.Enrichment analysis showed the potential causative genes mostly gathered in the adhesion and cell biological,vitamin A metabolism and eye development related functions.Conclusion: LCA is a highly heterogeneous disease,both clinically and genetically.There is a high frequency of novel mutations in Chinese population of LCA with a special phenotype and gene mutation spectrum.AIPL1 also has a high frequency of novel mutations in Chinese cohort and the novel mutation(c.C241 T p.Q81X)may be a founder mutation in the east coast Chinese population.Unreported mutations in KRT12,CYP1A1,etc.may be involved in the development of LCA.