Etiology And Genetic Analysis Of 140 Children With Developmental Delay/intellectual Disability By Trio-WES

Objective:Etiology and genetic analysis of 140 children with developmental delay(DD)or intellectual disability(ID)were performed using Trio Whole Exome Sequencing(Trio-WES).Pathogenic and likely pathogenic gene variations or chromosomal copy number variations(CNVs)and corresponding clinical phenotype were used as references to find pathogenic gen es at relatively high frequency,unreported mutation loci and possible pathogenic candidate genes,which can clarify the pathogenic cause and enrich gene spectrum and phenotype spectrum of DD/ID.The correlation between pathogenic gene mutation loci and DD/ID with different organ abnormalities can be further clarified by analyzing the genetic mode of positive mutation loci,the biogenic analysis of unreported loci and the association between gene mutation sites and clinical phenotype.This study can improve the etiological cognition,promote clinicians and researchers to conduct joint biogenic analysis of unreported variants based on data mining and basic scientific research,and propose more effective molecular diagnosis and individualized treatment programs,so as to provide genetic guidance for next pregnancy of these families.In addition,this study can also provide research basis for improving the distribution of pathogenic gene mutation loci and corresponding phenotype of DD/ID,which has important guiding significance for the prevention and control of birth defects and the improvement of population quality in Shanxi Province.Methods:Clinical data,including basic information,clinical phenotype,genetic data,etc,were collected from children who were diagnosed with developmental delay or intellectual disability and who had undergone karyotyping at the Shanxi children’s Hospital(Shanxi maternal and child health hospital)from 2019 to 2022.Patient peripheral blood was collected for Trio-WES and Sanger sequencing and the results were analyzed by National Human Genetic Resources Sharing Service Platform and Zhiyin Oriental Genetic Disease Analysis Platform.The results of Trio-WES were analyzed by using databases,such as Clin Var,HGMD,OMIM and DGV.The pathogenicity of gene variations and CNVs was assessed based on ACMG guideline and relevant reported papers were retrieved through Pub Med database.The conservation of mutation loci among different species were analyzed by Uniprot and Clustal Omega.The protein harmfulness of some unreported pathogenic gene mutation loci were predicted by SIFT and Poly Phen-2,and the changes of protein advanced structure were predicted by Swiss and Phyre2.Correlation analysis was conducted for the pathogenic gene variation loci and related clinical phenotype and the follow-up about rebirth were conducted in these parents whose children had positive mutation loci.Results:Trio-WES detected 140 children with DD/ID,and the results showed 75 cases of pathonic or likely pathogenic variation,including 65 cases of gene variations(one case was diagnosed with pathogenic variation due to retrospective analysis)and 10 cases of CNVs,and the positive detection rate was 53.57%(75/140).Among the 65 cases of gene variants,77 mutation loci of 56 pathogenic genes were detected,including a pathogenic mutation site of CUL3 gene by retrospective analysis.PAH,FOXG1,CTNNB1,POGZ and SHANK3 were detected with relatively high frequency,and the highest frequency were PAH and FOXG1.Among the 10 cases of CNVs,11 pathogenic or likely pathogenic CNVs were detected,including 2 replicates,1 homozygous deletion and 8 heterozygous deletion.These CNVs ranged in size from 1.25 Kb for single exon to 4.79 Mb for large fragments,with 8 CNVs less than 1Mb in length and 3 CNVs greater than 1Mb.According to the classification of Mendelian inheritance mode,40 cases were autosomal dominant inheritance(AD),15 cases were autosomal recessive inheritance(AR),3 cases were X chromosome dominant inheritance(XD),and 7 cases were X chromosome recessive inheritance(XR).In the 77 mutation loci,40 loci were de nove mutations and 37 loci were paternal/maternal mutations according to the classification of the mutation loci origin.11 families had performed prenatal diagnosis or preimplantation diagnosis of In vitro fertilization embryo transfer(IVF-ET)during follow-up.37 unreported loci were discovered by searching Clin Var and other databases,and the unreported loci pathogenicity of IARS1 and SPATA5 was confirmed by biogenic analysis.The detection rate of DD/ID combined system abnormalities was higher than that of DD/ID by analyzing gene variation loci-clinical phenotype association in 75 patients.DD is often associated with neurological,musculoskeletal,head and neck,and genitourinary abnormalities.In addition to the common clinical phenotype such as short stature and motor/language retardation,epilepsy,brain MRI abnormalities,gait abnormalities,hypotonia and special facial features also happen in DD/ID patients.In addition,the detected P/LP mutation loci were relatively common in the AD,XR and AR genetic modes and P/LP CNVs,among which AR was the most common in metabolic and CNVs was the most common in the immune system.It is worth noting that ATP8A2 and CTBP1 pathogenic genes are not associated with epilepsy phenotype,FRMPD4,SHANK3 and UBE3 B genes are not associated with gait abnormalities,and TBL1XR1 genes are not associated with undescended testis phenotype in OMIM at present.Conclusion:In this study,the positive detection rate of Trio-WES was 53.57% in 140 DD/ID patients,including 65 cases of gene variations and 10 cases of CNVs.CNVs less than10 Kb can be identified in two cases.PAH,FOXG1,CTNNB1,POGZ and SHANK3 were detected with relatively high frequency,and the highest frequency were PAH,which was consistent with the epidemiological characteristics of Shanxi Province.AD accounted for the highest proportion in DD/ID,and the mutation loci of AD and XD were mainly new mutations,while the mutation loci of AR and XR were mostly inherited from parents in the 65 patients with gene variants.In addition,c.217A>G of TUBB was first reported to be associated with CDCBM and c.887G>A of MED12 was first reported to be associated with clened hand,overlapping fingers,and clubfoot phenotype.37 unreported loci were identified,and bioinformatics analysis was performed in IARS1 and SPATA5,which enriched the mutant spectrum of DD/ID pathogenic genes.The correlation between positive mutation loci and DD/ID with different systems abnormalities were further clarified by gene variation loci-clinical phenotype association analysis,which can be used as the basis for phenotypic prediction.It is worth noting that ATP8A2 and CTBP1 pathogenic genes are not associated with epilepsy phenotype,FRMPD4,SHANK3 and UBE3 B genes are not associated with gait abnormalities and TBL1XR1 genes are not associated with undescended testis phenotype in OMIM at present,which enriched the phenotype spectrum of DD/ID pathogenic genes.