Feto-maternal DNA/RNA Trafficking: Biology and Application

描述（由申请人提供）：目前的产前诊断和筛查手段包括三种不同类型的检测：血清蛋白测量、超声胎儿成像和染色体分析。在这个应用程序中，我们建议通过发展对无细胞核酸作为不同胎龄正常和异常胎儿功能发育的新生物标志物的理解，为胎儿评估增加一个新的维度。在过去的四年中，这一领域呈指数级扩展，包括母体血液、羊水、脑脊液和尿液中胎儿和胎盘DNA和mRNA的评估。最近，利用母体血液中的无细胞胎儿DNA对胎儿D型恒河猴状态进行无创诊断，已经从实验室过渡到临床。我们现在建议在项目1-4年的成果基础上进行新的研究。在目标1中，我们将检验无细胞DNA（总和胎儿）在怀孕期间具有生物学意义和临床应用的假设。我们将研究胎儿性别检测在x连锁条件下的临床应用，在有先天性肾上腺增生风险的胎儿中，以及在生殖器模糊的胎儿中。我们还将研究总游离DNA水平作为组织缺氧和炎症的非侵入性标志物的意义。在目标2中，我们将检验一个假设，即正常和异常的胎儿基因表达可以通过母体体液（如全血和羊水）中的无细胞mRNA相互区分。我们的目标是开发一组区分正常和异常胎儿的关键基因，这将允许开发用于特定产前诊断应用的定制微阵列。在目标3中，我们将确定是否可以使用基因组方法来更好地了解胎儿发育。使用现有的软件包，我们将分析目标2中获得的差异调节基因列表，以检查特定胎儿基因在何时何地表达。我们将确定参与正常和异常胎儿发育的关键生物学途径，并受胎儿治疗程序的影响。我们还将使用新的基因组方法，如网络分析，来理解复杂的生物关系。该实验计划的成功完成将大大扩大目前产前诊断的范围，超出解剖，非整倍体和单基因疾病检测。基因组分析将导致对胎儿染色体和解剖异常的病理生理学的新见解，这将最终导致合理的和全新的胎儿治疗方法。公共卫生相关性：这个转化项目的总体目标是发展对无细胞核酸作为胎儿发育的新型生物标志物的理解。这项工作的成功完成将大大扩大目前产前诊断的范围，从而产生可以发现胎儿基因表达异常的测试。基因组分析在拟议的目标将导致新的见解关于胎儿生物学过程，这将导致胎儿治疗的全新方法。

DESCRIPTION (provided by applicant): Current means of prenatal diagnosis and screening involve three different types of assays: measurements of serum proteins, fetal imaging through sonography, and chromosome analysis. In this application we propose to add a new dimension to fetal evaluation by developing an understanding of cell-free nucleic acids as novel biomarkers of normal and abnormal fetal functional development at different gestational ages. Over the past four years this field has expanded exponentially, encompassing evaluation of fetal and placental DNA and mRNA in maternal blood, amniotic fluid, cerebrospinal fluid, and urine. Recently, noninvasive diagnosis of fetal Rhesus D status, using cell-free fetal DNA in maternal blood, has made the transition from bench to bedside. We now propose new studies that will build upon the achievements in project years 1-4. In aim 1 we will test the hypothesis that cell-free DNA (total and fetal) has biological significance and clinical applications during pregnancy. We will examine the clinical utility of fetal gender detection in X-linked conditions, in fetuses at risk for congenital adrenal hyperplasia, and in fetuses with ambiguous genitalia. We will also examine the significance of levels of total cell-free DNA as a noninvasive marker of tissue hypoxia and inflammation. In aim 2 we will test the hypothesis that normal and abnormal fetal gene expression can be distinguished from each other using cell-free mRNA in maternal body fluids such as whole blood and amniotic fluid. Our goal is to develop a panel of key genes that distinguish between normal and abnormal fetuses, which will permit development of custom microarrays for specific prenatal diagnostic applications. In aim 3 we will determine whether one can use genomic approaches to better understand fetal development. Using existing software packages we will analyze the lists of differentially-regulated genes acquired in aim 2 to examine where and when specific fetal genes are expressed. We will identify key biological pathways that are involved in normal and abnormal fetal development, and are affected by fetal treatment procedures. We will also use new genomic approaches, such as network analyses, to understand complex biological relationships. Successful completion of the proposed experimental plan will significantly expand the current scope of prenatal diagnosis beyond anatomy, aneuploidy, and single gene disorder detection. Genomic analysis will lead to new insights into the pathophysiology of fetal chromosomal and anatomic abnormalities, which will ultimately lead to rational and entirely novel approaches to fetal therapy. PUBLIC HEALTH RELEVANCE: The overall goal of this translational project is to develop an understanding of cell-free nucleic acids as novel biomarkers of fetal development. Successful completion of the proposed work will significantly expand the current scope of prenatal diagnosis, resulting in tests that can find abnormalities in fetal gene expression. Genomic analysis in the proposed aims will lead to novel insights regarding fetal biological processes, which will result in entirely new approaches to fetal therapy.