JAUNDICE NEXT: A diagnostic tool for cholestatic liver disease.

黄疸下一个：胆汁淤积性肝病的诊断工具。

• 批准号: 8312819
• 负责人: JORGE A. BEZERRA
• 金额: $ 34.48万
• 依托单位: P2D, INC.
• 依托单位国家: 美国
• 财政年份: 2012
• 资助国家: 美国
• 起止时间: 2012-07-01 至 2013-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8312819
• 关键词: Accounting; Adult; Algorithms; Base Sequence; Biliary; Biological Assay; Blood capillaries; ChIP-seq; Child; Child Care; Cholelithiasis; Cholestasis; Chronic; Clinical; DNA; DNA Sequence; DNA amplification; Detection; Development; Diagnosis; Diagnostic; Disease; Fibrosis; Future; Gene Chips; Gene Targeting; Genes; Genetic; Genotype; Healthcare; Hepatocyte; Housing; Icterus; Inherited; Intrahepatic Cholestasis; Knowledge; Life; Link; Liver diseases; Malignant neoplasm of liver; Medical; Methodology; Missense Mutation; Mutation; Mutation Detection; Mutation Spectra; Names; Nucleotides; Output; Pathologic; Patients; Phase; Phenotype; Phospholipids; Physicians; Play; Population; Positioning Attribute; Preparation; Procedures; Protocols documentation; Public Health; Role; Sampling; Screening procedure; Series; Small Business Technology Transfer Research; Surveys; Syndrome; System; Target Populations; Technology; Testing; Variant; Work; analytical tool; base; bile formation; capillary; cholangiocyte; chronic liver disease; clinical care; clinical phenotype; cohort; cost; disease-causing mutation; improved; innovative technologies; insertion/deletion mutation; intrahepatic cholestasis of pregnancy; next generation; patient population; prototype; research clinical testing; research study; tool

DESCRIPTION (provided by applicant): Cholestasis (or pathologic jaundice) is the most common sign of liver disease in children and often results from mutations in genes that play a critical role in bile formation and transport. In adults, mutations in the same genes also cause chronic cholestasis that may manifest as non-specific portal fibrosis, intrahepatic cholestasis of pregnancy, and low-phospholipid-associated cholelithiasis. Using recent knowledge of the genetic basis of these disorders, we developed a high-throughput gene chip that identifies mutations in the genes that cause the most common forms of inherited syndromes of chronic cholestasis. The chip is now a clinical test that is increasingly used by physicians as part of diagnostic algorithms in symptomatic patients. Although the chip remarkably facilitates specific diagnoses, its accuracy is limited by the inability to reliably detect insertions or deletions, whih account for 10-20% of the disease-causing mutations, thus decreasing the sensitivity of the chip. In this application, we propose to solve this technological gap by developing the next version of a mutation-screening tool that we are naming "JAUNDICENEXT". This tool customizes NextGen sequencing to accurately survey the nucleotide composition of target genes to diagnose disease-causing mutations. Our Specific Aim is "to determine the accuracy of JAUNDICENEXT to sequence the target genes." To test Hypothesis-1 that "JAUNDICENEXT sequences the target genes at >99% accuracy," we will compare the nucleotide sequence produced by the JaundiceNext with the sequence produced by standard sequencing methodology. To test Hypothesis-2 that "JAUNDICENEXT identifies insertion and deletion mutations in patients with cholestasis," we will examine whether the sequence output produced by JAUNDICENEXT detects insertions, deletions, or indels in DNA known to carry these mutations. Collectively, these experiments will validate the technical merits of JAUNDICENEXT and position us for future experiments to further bench-test the assay and develop an automated detection algorithm to in a future Phase-II application. PUBLIC HEALTH RELEVANCE: Jaundice is a clinical sign that is common to several types of chronic liver diseases in children and adults. In a previous STTR, we developed a gene chip to identify mutations in the genes that cause the most common forms of inherited syndromes of persistent jaundice (also known as cholestasis). The chip is now a clinical test that is increasingly used by physicians as part of diagnostic algorithms in symptomatic patients. Although the chip remarkably facilitates specific diagnoses, its accuracy is limited by the inabiliy to reliably detect insertions or deletions, which account for 10-20% of the disease-causing mutations, thus decreasing the sensitivity of the chip. We propose to solve this technological gap by developing the next version of a mutation-screening tool that we are naming "JAUNDICENEXT". This new tool customizes NextGen, a powerful sequencing technology, to accurately detect the wider spectrum of mutations in four genes simultaneously. These studies will improve the sensitivity and accuracy of a non-invasive diagnostic tool that significantly improves the diagnostic algorithm in patients with liver disease.

描述（由申请人提供）：胆汁淤积（或病理性黄疸）是儿童肝病最常见的体征，通常是由在胆汁形成和运输中发挥关键作用的基因突变引起的。在成人中，相同基因的突变也会导致慢性胆汁淤积，可能表现为非特异性门静脉纤维化、妊娠期肝内胆汁淤积和低磷脂相关胆石症。利用这些疾病的遗传基础的最新知识，我们开发了一种高通量基因芯片，可以识别导致最常见形式的慢性胆汁淤积遗传综合征的基因突变。该芯片现在是一种临床测试，越来越多地被医生用作有症状患者诊断算法的一部分。尽管该芯片显着促进了特定诊断，但其准确性受到无法可靠检测插入或缺失的限制，插入或缺失占致病突变的 10-20%，从而降低了芯片的灵敏度。在此应用中，我们建议通过开发下一版本的突变筛选工具（我们将其命名为“JAUNDICENEXT”）来解决这一技术差距。该工具可定制 NextGen 测序，以准确调查目标基因的核苷酸组成，从而诊断致病突变。我们的具体目标是“确定 JAUNDICENEXT 对目标基因进行测序的准确性”。为了测试假设 1“JAUNDICENEXT 以 >99% 的准确度对目标基因进行测序”，我们将比较 JaundiceNext 产生的核苷酸序列与标准测序方法产生的序列。为了检验假设 2“JAUNDICENEXT 识别胆汁淤积患者的插入和缺失突变”，我们将检查 JAUNDICENEXT 产生的序列输出是否检测到已知携带这些突变的 DNA 中的插入、缺失或插入缺失。总的来说，这些实验将验证 JAUNDICENEXT 的技术优点，并为我们未来的实验做好准备，以进一步对测定进行基准测试并开发自动检测算法以用于未来的 II 期应用。 公共卫生相关性：黄疸是儿童和成人几种慢性肝病的常见临床症状。在之前的 STTR 中，我们开发了一种基因芯片来识别导致最常见形式的持续性黄疸（也称为胆汁淤积）遗传综合征的基因突变。该芯片现在是一种临床测试，越来越多地被医生用作有症状患者诊断算法的一部分。尽管该芯片显着促进了特定诊断，但其准确性受到无法可靠检测插入或缺失的限制，插入或缺失占致病突变的 10-20%，从而降低了芯片的灵敏度。我们建议通过开发下一版本的突变筛选工具（我们将其命名为“JAUNDICENEXT”）来解决这一技术差距。这款新工具定制了 NextGen（一种强大的测序技术），可同时准确检测四个基因中更广泛的突变。这些研究将提高非侵入性诊断工具的灵敏度和准确性，从而显着改善肝病患者的诊断算法。