苯丙酮尿症(PKU)是由苯丙氨酸羟化酶(PAH)基因变异引起的遗传性疾病。在3369个报告的PAH变体中,33.7%是错义改变。不幸的是,这些错义变异中有30%被归类为未知意义的变异(VUS),对遗传风险评估提出了挑战。在我们的研究中,我们按照美国医学遗传学和基因组学学会/分子病理学协会(ACMG/AMP)指南,重点分析了由ClinGenPAH变异体固化专家小组(VCEP)标准规定的836种错义PAH变异体.我们使用并比较了诸如Franklin和Varsome之类的变体注释器工具,进行了PAH的3D结构分析,并检查了活跃和监管地点的热点。此外,我们评估了明显错义变异体的潜在剪接效应.通过评估22962例PKU患者的表型数据,我们的目的是重新评估错义变异的致病性.我们的综合方法成功地将836个错义变异中的309个VUS重新分类为可能的致病性或致病性(37%),将370种可能的致病变种升级为致病变种,并将以前认为可能的良性变异重新分类为可能的致病性。636个错义变体的表型信息可用,其中441例进行了180种变体的3D结构分析和活性位点热点鉴定。经过我们的分析,只有6%的错义变异被归类为VUS,和其中三个(c.23A>C/p。Asn8Thr,c.59_60delinsCC/p。Gln20Pro,和c.278A>T/p。Asn93Ile)可能受到异常剪接的影响。此外,致病性变异(c.168G>T/p。Glu56Asp)被确定为共有剪接位点修饰的风险超过98%,高分表明捐献者损失为0.94。ACMG/AMP指南与计算机结构分析和表型数据的整合显着减少了错义VUS的数量,为遗传咨询提供坚实的基础,并强调代谢表型信息在变异管理中的重要性。这项研究也揭示了PAH变体的当前景观。
Phenylketonuria (PKU) is a genetic disorder caused by variations in the phenylalanine hydroxylase (PAH) gene. Among the 3369 reported PAH variants, 33.7% are missense alterations. Unfortunately, 30% of these missense variants are classified as variants of unknown significance (VUS), posing challenges for genetic risk assessment. In our study, we focused on analyzing 836 missense PAH variants following the American College of Medical Genetics and Genomics/Association for Molecular Pathology (ACMG/AMP) guidelines specified by ClinGen PAH Variant Curation Expert Panel (VCEP) criteria. We utilized and compared variant annotator tools like Franklin and Varsome, conducted 3D structural analysis of PAH, and examined active and regulatory site hotspots. In addition, we assessed potential splicing effect of apparent missense variants. By evaluating phenotype data from 22962 PKU patients, our aim was to reassess the pathogenicity of missense variants. Our comprehensive approach successfully reclassified 309 VUSs out of 836 missense variants as likely pathogenic or pathogenic (37%), upgraded 370 likely pathogenic variants to pathogenic, and reclassified one previously considered likely benign variant as likely pathogenic. Phenotypic information was available for 636 missense variants, with 441 undergoing 3D structural analysis and active site hotspot identification for 180 variants. After our analysis, only 6% of missense variants were classified as VUSs, and three of them (c.23A>C/p.Asn8Thr, c.59_60delinsCC/p.Gln20Pro, and c.278A >T/p.Asn93Ile) may be influenced by abnormal splicing. Moreover, a pathogenic variant (c.168G>T/p.Glu56Asp) was identified to have a risk exceeding 98% for modifications of the consensus splice site, with high scores indicating a donor loss of 0.94. The integration of ACMG/AMP guidelines with in silico structural analysis and phenotypic data significantly reduced the number of missense VUSs, providing a strong basis for genetic counseling and emphasizing the importance of metabolic phenotype information in variant curation. This study also sheds light on the current landscape of PAH variants.