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分析：血清蛋白测量、超声胎儿成像和染色体分析。在这一应用中，我们建议通过发展对无细胞核酸作为不同胎龄的正常和异常胎儿功能发育的新的生物标志物的理解，为胎儿评估增加一个新的维度。在过去的四年里，这一领域呈指数级增长，包括对孕妇血液、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对胎儿恒河猴D状态进行无创性诊断，已经从工作台过渡到床边。我们现在提出新的研究，这些研究将建立在项目第1-4年的成果的基础上。在目标1中，我们将检验无细胞DNA(总DNA和胎儿DNA)在妊娠期间具有生物学意义和临床应用的假设。我们将研究X连锁条件下胎儿性别检测在先天性肾上腺增生症风险胎儿和生殖器不明确胎儿中的临床应用。我们还将检查总无细胞DNA水平作为组织缺氧和炎症的非侵入性标记物的意义。在目标2中，我们将检验这一假设，即利用母体体液(如全血和羊水)中的无细胞mRNA可以区分正常和异常的胎儿基因表达。我们的目标是开发一组区分正常和异常胎儿的关键基因，这将允许为特定的产前诊断应用开发定制的微阵列。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。利用现有的软件包，我们将分析在目标2中获得的差异调控基因列表，以检查特定的胎儿基因在哪里以及何时表达。我们将确定与正常和异常胎儿发育有关的关键生物学途径，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，以了解复杂的生物关系。拟议的实验计划的成功完成将显著扩大目前产前诊断的范围，超出解剖学、非整倍体和单基因紊乱检测的范围。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，最终将导致合理和全新的胎儿治疗方法。公共卫生相关性：这个翻译项目的总体目标是发展对无细胞核酸作为胎儿发育的新生物标记物的理解。这项拟议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。拟议目标中的基因组分析将导致对胎儿生物学过程的新见解，这将导致全新的胎儿治疗方法。

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.