丁酰基肉碱代谢异常新生儿的基因型和生化表型分析

Analysis of genotypes and biochemical phenotypes of neonates with abnormal metabolism of butyrylcarnitine

目的:探讨丁酰基肉碱(C4)代谢异常新生儿的基因型和生化表型特征。方法:收集2018年1月至2023年6月在浙江大学医学院附属儿童医院经串联质谱法筛查单纯C4增高的120例新生儿初筛和召回复查的C4、C4/C3检测数据,并换算为C4增高倍数。采用液相捕获技术靶向捕获酰基辅酶A脱氢酶8(ACAD8)和短链酰基辅酶A脱氢酶(ACADS)基因的外显子及邻近50 bp区域,通过高通量测序和生物信息学分析获取基因变异信息,参考美国医学遗传学与基因组学学会分类标准进行致病性评估。采用威尔科克森秩和检验分析不同基因型新生儿C4增高倍数的差异。结果:共检出32种ACAD8基因变异型,其中7种变异型未见报道;检出41种ACADS基因变异型,其中17种变异型未见报道。ACAD8双等位基因变异39例,ACAD8单等位基因变异3例,ACADS双等位基因变异34例,ACADS单等位基因变异36例,ACAD8和ACADS双基因变异5例。ACAD8双等位基因变异组、ACADS双等位基因变异组C4增高倍数初筛值和召回值均高于ACADS单等位基因变异组(均P<0.01),且ACAD8双等位基因变异组C4增高倍数初筛值和召回值较ACADS双等位基因变异组更高(均P<0.01)。所有携带ACAD8或ACADS双等位基因变异新生儿的初筛C4增高倍数均大于1.5;而仅有25%(9/36)携带ACADS单等位基因变异的新生儿初筛C4增高倍数大于1.5。结论:ACAD8和(或)ACADS基因变异是浙江地区新生儿C4增高的主要遗传学原因,其变异型具有高度异质性。双等位基因变异者C4水平高于单等位基因变异者。常规串联质谱法新生儿筛查结果为单纯的C4增高时,其“筛查切值”可以适当提升。

Objective:To investigate the genotypes and biochemical phenotypes of neonates with abnormal metabolism of butyrylcarnitine(C4).Methods:One hundred and twenty neonates with increased C4 levels detected by tandem mass spectrometry in the neonatal screening at Children’s Hospital,Zhejiang University School of Medicine from January 2018 to June 2023 were included.The initial screening data and recalled data of C4 and C4/C3 were collected and converted into multiples of C4 reference range.Next generation sequencing was performed and the exons with adjacent 50 bp regions of ACAD8 and ACADS genes were captured by liquid phase capture technique.Variant information was obtained by bioinformatic analysis and the pathogenicity were classified according to the American College of Medical Genetics and Genomics criteria.The Wilcoxon rank sum test was used to analyze the differences in C4 levels among neonates with different variation types.Results:In total,32 variants in ACAD8 gene were detected,of which 7 variants were reported for the first time;while 41 variants of ACADS gene were detected,of which 17 variants have not been previously reported.There were 39 cases with ACAD8 biallelic variations and 3 cases with ACAD8 monoallelic variations;34 cases with ACADS biallelic variations and 36 cases with ACADS monoallelic variations.Furthermore,5 cases were detected with both ACAD8 and ACADS gene variations.Inter group comparison showed that the multiples of C4 reference range in initial screening and re-examination of the ACAD8 biallelic variations and ACADS biallelic variations groups were significantly higher than those of the ACADS monoallelic variations group(all P<0.01),while the multiples in the ACAD8 biallelic variations group were significantly higher than those in the ACADS biallelic variations group(all P<0.01).The multiples of C4 reference range in the initial screening greater than 1.5 times were observed in all neonates carrying ACAD8 or ACADS biallelic variations,while only 25%(9/36)in neonates carrying ACADS monoallelic variations.Conclusions:ACAD8 and/or ACADS gene variants are the main genetic causes for elevated C4 in newborns in Zhejiang region with high genotypic heterogeneity.The C4 levels of neonates with biallelic variations are significantly higher than those of neonates with monoallelic variations.The cut-off value for C4 level could be modestly elevated,which could reduce the false positive rate in tandem mass spectrometry neonatal screening.