A new diagnostic tool for rapid detection and characterization of REPEAT SEQUENCES in inherited diseases

一种新的诊断工具，用于快速检测和表征遗传性疾病中的重复序列

基本信息

批准号：
10354657
负责人：
Jason C Reed
金额：
$ 25.57万
依托单位：
VIRGINIA COMMONWEALTH UNIVERSITY
依托单位国家：
美国
项目类别：
财政年份：
2022
资助国家：
美国
起止时间：
2022-08-15 至 2024-07-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/10354657
关键词：
Affect Age of Onset Ataxia Atomic Force Microscopy Bar Codes Benchmarking Bioinformatics CGG repeat Capillary Electrophoresis Child Clinic Clinical Complex Coupled DNA DNA Sequence Alteration Decision Making Detection Development Diagnosis Diagnostic Disease Disease Management Early Diagnosis Elderly Equilibrium FMR1 FXTAS Family Family Planning Family member Fragile X Syndrome Generations Genes Genetic Genetic Anticipation Genetic Counseling Genetic Phenomena Genetic Risk Genome Genomic DNA Genomics Goals Healthcare Hereditary Disease Hospitals Individual Inherited Insurance Carriers Label Lead Length Lesion MJD1 protein Measures Medical Medical Genetics Meiosis Memory Disorders Methodology Methods Molecular Monitor Mood Disorders Movement Mutation Nature Patient Monitoring Patients Performance Persons Phenotype Physicians Polymerase Polymerase Chain Reaction Procedures Process Product Labeling Prognosis Relative Risks Risk Sampling Severities Skeletal Muscle Southern Blotting Speed Spinocerebellar Ataxias Stretching Symptoms System Techniques Technology Testing Time Tremor Trinucleotide Repeat Expansion Trinucleotide Repeats United States Variant Walking base biobank clinically relevant cost design diagnostic tool genetic analysis genetic disorder diagnosis genetic testing health care economics improved member multiplex assay multiplex detection mutant nanoparticle nanopore nervous system disorder new therapeutic target next generation sequencing non-genetic novel diagnostics novel strategies offspring patient stratification rapid detection risk stratification screening sequencing platform success targeted treatment tool virtual

项目摘要

=. PROJECT SUMMARY …... Ataxia is a discoordination of voluntary muscle movement and can be seen as the primary symptom of multiple disorders including genetic and non-genetic etiologies. Several genes are implicated in causing Spinocerebellar ataxia through trinucleotide repeat expansions (TREs) and are thought to affect between 1.5 to 4 per 10,000 people globally. Similar trinucleotide repeat expansions in the FMR1 gene, including potentially symptomatic premutations, can be found in around 1 in 300 people in the United States. Detection of repeat expansion disorders in a family are important for understanding genetic risks and early detection in at risk members, especially since unstable repeat expansions lead to the phenomenon of genetic anticipation, where symptoms can present earlier or more severely across generations. Additionally, several recent studies have suggested development of targeted therapies specifically targeting expanded repeats as a novel therapeutic target. As such, earlier and rapid detection of these disorders is crucial for advancing their medical management. Traditionally, short TRE targets are assessed via repeat-primed PCR (PR-PCR) and capillary electrophoresis, while longer mutants are confirmed via Southern blotting. This is clinically important, especially where repeat lengths outside of the range quantifiable by PR-PCR are diagnostically relevant. Testing for most TRE genes remains difficult for short read sequencing platforms, where repeat tract lengths exceed average read length (~100 bp for Illumina systems). Long-read sequencing technologies like PacBio and Oxford Nanopore have shown success in measuring TREs, however a combination of high costs, large quantities of DNA input, complex bioinformatics, problems determining repeat region boundaries, and high error rates makes them an unlikely solution for widespread screening in their current state We developed a solution, PRECYSE, based on high-speed atomic force microscopy (HSAFM) paired with unique ‘nanoparticle barcoding’ that can potentially characterize complex structural variants for numerous ataxia conditions simultaneously at much lower cost than possible with next generation sequencing (NGS) sequencing and other emerging approaches. This extremely sensitive technique can be conducted without using polymerase chain reaction (PCR) and can span a very wide range of possible target sizes, thereby allowing extension to virtually unlimited molecular lengths. Our hypothesis is that this technology can be easily adapted to multiplexed detection of TRE targets of practically any length. If successfully developed, our new approach will change the way clinicians identify, understand, and monitor changes in the genome caused by trinucleotide repeat expansion diseases through multiplexed panels. At the end of this R21 project, we will have a platform for multiplexed genomic analysis that successfully purifies and detects trinucleotide repeat targets. However, a follow-on larger scale (50-60 samples) planned R01 project will provide statistical significance and further refine the methodology.

=。共济失调是对自愿运动的障碍，可以看出是多重的主要症状包括遗传和非遗传病因的疾病。通过三核苷酸重复扩展（TRE）的共济失调，影响到每10,000次1.5至4 在全球范围内。在美国的300人中，可以发现大约1人的预言家庭中的疾病对于理解遗传风险和AT风险成员的早期发现很重要，尤其是因为不稳定的猫扩张导致遗传化的themenon是症状可以在几代人的早期或更严格的情况下提出。针对性疗法的发展是针对性的，如新颖的目标这些疾病的早期和快速检测对于推进其医疗管理至关重要。传统上，通过重复提交的PCR（PR-PCR）和毛细管评估短TRE目标电泳，而通过Southern印迹证实了更长的突变体。在PR-PCR可量化的范围之外的麻痹长度与大多数相关的测试 TRE基因仍然很困难堡垒读取测序平台，重复道超过平均值读取长度（Illumina系统的100 bp）。纳米孔在测量TE上表现出成功 DNA输入，复杂的生物信息学，确定重复重复重复重复界限的问题以及高错误率使制造它们是在当前状态的广泛筛选的精心解决方案我们基于配对的高速原子力显微镜（HSAFM）开发了一种解决方案，具有独特的“纳米颗粒条形码”，可以潜在地表征复杂的结构变体共济失调条件同时以下一代测序（NGS）的成本低得多可以在不使用的情况下进行测序和其他新兴方法。聚合酶链反应（PCR）可以跨越非常广泛的可能目标大小，从而允许扩展到虚拟的无限分子长度。为了多长时间的tre目标，我们的新方法实际上是任何长度的。将改变临床医生识别，理解和监测由三核苷酸引起的基因组变化在Theis R21项目的末端重复通过多路复用面板扩展对于成功净化和检测三核苷酸重复靶标的多重基因组分析随后的大规模（50-60个样本）计划的R01项目将统计显着性，并进一步完善方法。