苯丙氨酸羟化酶缺乏症患儿基因型和表型关系及其临床应用的研究

Relationship between genotype and phenotype of phenylalanine hydroxylase deficiency in children and its clinical application

目的构建苯丙氨酸羟化酶(PAH)缺乏症患儿的PAH基因变异谱,研究基因型和表型的关系。方法回顾分析76例经新生儿筛查确诊的PAH缺乏症患儿的PAH基因变异位点,建立基因型-表型预测模型。对等位基因进行赋值,设定变异位点特定值(AV),并计算AV评分之和,进而预测临床表型。结果应用第二代高通量测序技术和多重连接酶探针依赖扩增检测(MLPA)技术,在76例新生儿152个PAH等位基因中共检测出146个变异位点,频率为96.1%。70例新生儿检测出2个变异等位基因,其中3例纯合变异,67例复合杂合变异;另外6例检测出1个变异等位基因。共检测出变异类型45种,变异频率较高的位点为c.728G>A(26/146,17.8%)、c.158G>A(13/146,8.9%)、c.1068C>A(11/146,7.5%)、c.721C>T(10/146,6.8%),c.1238G>C(8/146,5.5%)、c.331C>T(8/146,5.5%)、c.1301C>A(7/146,4.8%)。其中c.95A>G位点未曾被BIOPKU数据库报道。患儿治疗前血苯丙氨酸(Phe)浓度与AV评分之和呈显著负相关(r=-0.83,P<0.001)。随访发现2例轻度高苯丙氨酸血症患儿血Phe浓度<120μmol/L者无需治疗。结论构建常见PAH基因变异谱,验证了PAH基因突变和表型的相关性,有助于PAH确诊及分型、早期判断预后以及遗传咨询。

Objective To explore the PAH gene variation spectrum of phenylalanine hydroxylase(PAH)deficiency in children,and to explore the relationship between genotype and phenotype.Method The PAH gene mutation sites in 76 children with PAH deficiency confirmed by neonatal screening were analyzed retrospectively,and the genotype-phenotype prediction model was established.Alleles were assigned,specific values of mutation sites(AV)were set,and the sum of AV scores was calculated to predict the clinical phenotype.Results Using the second-generation high-throughput sequencing technology and multiplex ligation-dependent probe amplification technology,146 mutation sites were detected in 152 PAH alleles in 76 neonates and the frequency was 96.1%.Two variant alleles were detected in 70 newborns,including 3 homozygous and 67 compound heterozygous variants.One variant allele was detected in the other 6 newborns.A total of 45 types of variation were detected,and the locus with high frequency of variation was c.728G>A(26/146,17.8%),c.158G>A(13/146,8.9%),and c.1068C>A(11/146,7.5%),c.721C>T(10/146,6.8%),c.1238G>C(8/146,5.5%),c.331C>T(8/146,5.5%)and c.1301C>A(7/146,4.8%).The c.95A>G locus has not been reported by BIOPKU database.There was a significantly negative correlation between serum phenylalanine(Phe)concentration and the sum of AV scores before treatment(r=-0.83,P<0.001).During follow-up,it was found that 2 mild hyperphenylalaninemia children whose blood Phe concentration was less than 120μmol/L did not require treatment.Conclusion Constructing common PAH gene variation spectrum and verifying the correlation between PAH gene mutation and phenotype are conducive to diagnosis and typing of PAH,early prognosis judgment,and genetic counseling.