Newborn screening for PKU and BH4 responsiveness

新生儿 PKU 和 BH4 反应性筛查

• 批准号: 7107330
• 负责人: Steven F Dobrowolski
• 金额: $ 8.88万
• 依托单位: BIOFIRE DEFENSE, LLC.
• 依托单位国家: 美国
• 财政年份: 2006
• 资助国家: 美国
• 起止时间: 2006-05-15 至 2006-11-14
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7107330
• 关键词: aminoacid metabolism; clinical research; drug design /synthesis /production; gene expression; gene mutation; genetic mapping; genetic polymorphism; genetic screening; human genetic material tag; human subject; introns; messenger RNA; method development; newborn human (0-6 weeks); patient oriented research; phenylalanine; phenylketonurias; polymerase chain reaction; posttranscriptional RNA processing; reagent /indicator; tetrahydrobiopterin

DESCRIPTION (provided by applicant): In 1958, Dr Robert Guthrie was approached by Dr Robert Warner seeking aid to develop a better means to measure phenylalanine in the blood of newborns. Of this interaction was born the bacterial inhibition assay and the beginning of population-based newborn screening for phenylketonuria (PKU). Early identification of affected newborns avoided irreversible brain damage using a phenylalanine-restricted diet. While the BIA has been replaced by tandem mass spectrometry as the means to measure phenylalanine, PKU remains the paradigm for a disorder effectively treated by prospective identification of asymptomatic patients. PKU results from defects in phenylalanine hydroxylase (PAH) causing an inability to convert phenylalanine to tyrosine. Treating PKU by dietary means remained largely unchanged until several groups identified a sub- set of patients treatable using 6R-tetrahydrobiopterin (BH4), the obligatory co-factor of the PAH enzyme, without the phenylalanine restricted diet. While BH4-responsive patients are skewed to mild PKU and hyperphenyla'ianemia; classic PKU patients have been characterized as BH4 responsive. Analysis of the PAH gene is becoming an important aspect to determining BH4 response. Comprehensive analysis of the PAH gene may be performed using the universally collected newborn screening dried blood card as a source of DNA. Using the dried blood card and the emerging technology of high-resolution melt profiling, comprehensive analysis of PAH may be easily completed within 1.5 days of abnormal newborn screening results. Genotypic data will be in hand when results of the physiological Phe/ BH4 loading test are complete. Combining physiological analysis and genetic analysis will lead to effective identification of BH4 responsive PKU patients. Cataloging PAH mutations resulting in BH4 responsive disease is underway thus developing a sensitive, rapidly, and cost effective means to analyze the PAH gene will have utility to clinicians and researchers. Herein is proposed the use of high resolution melt profiling to develop a simplified and streamlined means of assessing the coding sequence and intronic regions critical to mRNA processing in the PAH gene. High resolution melt profiling is rapid with sensitivity at least equal to DNA sequence analysis and in excess of other pre-sequence scanning technologies. High resolution melt profiling will play a role to identify BH4 responsive PKU patients.

描述（由申请人提供）：1958 年，罗伯特·沃纳 (Robert Warner) 博士找到罗伯特·格思里 (Robert Guthrie) 博士寻求帮助，以开发一种更好的方法来测量新生儿血液中的苯丙氨酸。这种相互作用催生了细菌抑制试验，并开始了基于人群的新生儿苯丙酮尿症 (PKU) 筛查。通过限制苯丙氨酸饮食，早期识别受影响的新生儿可以避免不可逆转的脑损伤。虽然 BIA 已被串联质谱法取代作为测量苯丙氨酸的方法，但 PKU 仍然是通过前瞻性识别无症状患者来有效治疗疾病的范例。 PKU 是由苯丙氨酸羟化酶 (PAH) 缺陷导致无法将苯丙氨酸转化为酪氨酸造成的。通过饮食方式治疗 PKU 的方法基本上没有改变，直到几个研究小组确定了一组可以使用 6R-四氢生物蝶呤 (BH4)（PAH 酶的必需辅助因子）治疗且无需苯丙氨酸限制饮食的患者亚组。虽然 BH4 反应性患者倾向于轻度 PKU 和高苯血症；典型的 PKU 患者被描述为 BH4 反应性的。 PAH 基因分析正在成为确定 BH4 反应的一个重要方面。 PAH基因的综合分析可以使用普遍收集的新生儿筛查干血卡作为DNA来源来进行。利用干燥的血卡和新兴的高分辨率熔解分析技术，可以在新生儿筛查结果异常的1.5天内轻松完成PAH的综合分析。当生理 Phe/BH4 负载测试结果完成后，即可获得基因型数据。结合生理分析和遗传分析将有效识别 BH4 反应性 PKU 患者。对导致 BH4 反应性疾病的 PAH 突变进行编目正在进行中，因此开发一种灵敏、快速且具有成本效益的方法来分析 PAH 基因将对临床医生和研究人员有用。本文建议使用高分辨率熔解图谱来开发一种简化且精简的方法来评估对 PAH 基因中 mRNA 加工至关重要的编码序列和内含子区域。高分辨率熔解分析速度快，灵敏度至少等于 DNA 序列分析，并超过其他序列前扫描技术。高分辨率熔解图谱将有助于识别 BH4 反应性 PKU 患者。