Non-invasive Aneuploidy Screening of Circulating Fetal Cells for Prenatal Diagnos

描述(由申请人提供)：在怀孕期间，医生和患者希望获得尽可能多的关于胎儿健康的信息。出于情感和医疗方面的原因，在尽可能早的时候寻求这一信息，并将母亲和孩子的风险降至最低。虽然广泛使用的早孕绒毛取样(CVS)和中孕羊膜穿刺术是相对安全的，但这两种方法都不是没有风险的。为了完全避免这些风险，研究人员转向从母血中分离循环中的胎儿有核红细胞(FNRBCs)，作为一种替代的、非侵入性的胎儿组织来源。尽管发展了FNRBC扩增方法，但在与随后的非整倍体筛查相结合方面取得的成功有限，仍需克服几个挑战，例如检测单个胎儿细胞的24染色体非整倍体，确认分离细胞的来源(胎儿与母体)，同时筛查单核苷酸变异或微内/缺失引起的疾病。我们创新的Parental SupportTM技术为所有这些挑战提供了解决方案，而开发一种怀孕早期的非侵入性产前诊断测试是这项赠款申请的最终目标。在第一阶段，我们首先计划优化单细胞裂解和全基因组扩增方案，专门针对抗体染色的FNRBCs。单细胞分析的方案优化属于GSN的核心能力，因为我们之前已经成功地将一种创新的单细胞分子核型分析方案商业化，从而能够在24小时内对单个卵裂球进行基因分析。然后，我们将系统地评估哪种现有FNRBC浓缩方法的组合能够提供最大的产量和纯度，适合后续的基于父母支持的遗传分析，使用预定义的胎儿和成人血液混合物。第二阶段的主要目标将是从预定义的胎儿和成人血液混合物过渡到实际的产妇血液样本。我们将首先进行一项初步研究，以确定第一阶段确定的最好的FNRBC分离方法(S)中的哪一种应该成为主导方法。然后，我们将利用这一领先方法进行一项更大规模的研究，以评估Parental SupportTM的非整倍体诊断与羊膜穿刺术或绒毛采样的核型鉴定之间的一致性。如果成功，我们预计这些目标的完成将对产前诊断领域产生重大影响，改善全球数百万夫妇和儿童的生活，并将非侵入性诊断带入产前医学的主流。 与公共卫生相关：在没有产前诊断的情况下，每50名婴儿中就有1名患有严重的身体或智力残疾，每30名婴儿中就有1名患有某种形式的先天性畸形，每200名婴儿中就有1名患有明显的染色体异常。虽然这些异常可以通过羊膜穿刺术或绒毛取样等技术来诊断，但这两种手术都会增加对母亲和胎儿的伤害风险。我们的创新技术有可能通过简单地分析母亲的血液来评估未出生婴儿的健康，从而将该程序的风险降至最低，并将产前筛查扩大到普通人群。

DESCRIPTION (provided by applicant): During the course of a pregnancy, physicians and patients desire as much information as possible regarding the health of the fetus. For both emotional and medical reasons, this information is sought as early in term as possible, and with the fewest possible risks to both mother and child. Although the widely used first trimester chorionic villus sampling (CVS) and second trimester amniocentesis are relatively safe, both procedures are not without negligible risks. In efforts to avoid these risks altogether, researchers have turned toward isolating circulating fetal nucleated red blood cells (FNRBCs) from maternal blood as an alternative, non- invasive source of fetal tissue. Despite the development of FNRBC enrichment methods, there has been limited success with their coupling to subsequent aneuploidy screening and several challenges still must be overcome such as ability to test single fetal cells for 24-chromosome aneuploidy, confirm the isolated cell's origin (fetal versus maternal) and simultaneously screen for diseases caused by single nucleotide variants or micro in/dels. Our innovative Parental SupportTM technology provides a solution to all of these challenges and the development of a first trimester non-invasive prenatal diagnostic test is the ultimate goal of this grant application. In Phase I, we first plan to optimize single cell lysis and whole genome amplification protocols specifically for antibody-stained FNRBCs.. Protocol optimization for single cell analysis falls within the core competencies of GSN as we have previously successfully commercialized an innovative single cell molecular karyotyping protocol to enable genetic analysis of single blastomeres within 24 hours. We will then systematically evaluate which combination of existing FNRBC enrichment methods provides maximum yield and purity suitable for subsequent Parental Support"-based genetic analysis using predefined mixtures of fetal and adult blood. The main objective of Phase II will be to transition from the predefined blood mixtures of fetal and adult blood to actual maternal blood samples. We will first conduct a pilot study to determine which of the best FNRBC isolation method(s) identified in Phase I should become the lead method. Using this lead method, we will then conduct a larger study to evaluate concordance between aneuploidy diagnosis by Parental SupportTM and karyotyping by amniocentesis or chorionic villus sampling. If successful, we expect that the completion of these Aims would have a major impact on the field of prenatal diagnosis, improve the lives of millions of couples and children worldwide, and bring non-invasive diagnosis to the mainstream of prenatal medicine. PUBLIC HEALTH RELEVANCE: In the absence of prenatal diagnosis, up to 1 in 50 babies have serious physical or mental handicaps, up to 1 in 30 babies have some form of congenital malformation, and up to 1 in 200 have a phenotypically significant chromosome abnormality Although these abnormalities can be diagnosed with techniques such as amniocentesis or chorionic villus sampling, both procedures carry an increased risk of harm to both the mother and fetus. Our innovative technology has the potential to evaluate the health of an unborn child by simply analyzing the mother's blood, thereby minimizing the risks of the procedure and expanding prenatal screening to the general population.