Assay development for the assessment of pregnancy risks in early pregnancy

评估妊娠早期妊娠风险的测定方法开发

• 批准号: 10547072
• 负责人: Sascha Drewlo
• 金额: $ 36.82万
• 依托单位: AKNA HEALTH LLC
• 依托单位国家: 美国
• 财政年份: 2022
• 资助国家: 美国
• 起止时间: 2022-08-15 至 2024-08-14
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10547072
• 关键词: 2 year old; Abnormal placentation; Address; Biological Assay; Biological Markers; Birth; Birth Weight; Blood flow; Cell Lineage; Cell Separation; Cells; Cervical; Clinical; Clinical Research; Cytology; Cytometry; Data; Development; Diagnosis; Diagnostic Reagent Kits; Discipline of obstetrics; Disease; Early Diagnosis; Ensure; Evaluation; Feasibility Studies; Fetal Growth; Fetal Growth Retardation; Fetal Monitoring; Fetal health; Fetus; First Pregnancy Trimester; Foundations; Future; Gases; Genetic Fingerprintings; Genetic Transcription; Genome; Gestational Age; Goals; Government; Growth; HLA G antigen; Health; Health Benefit; Healthcare; High-Risk Pregnancy; Human; Human Chorionic Gonadotropin; Impairment; Intervention; Label; Link; Low Birth Weight Infant; Mass Spectrum Analysis; Maternal-Fetal Exchange; Medical History; Medical Records; Methodology; Methods; Molecular Profiling; Monitor; Mothers; Nutrient; Outcome; PGF gene; Pap smear; Pathology; Patient-Focused Outcomes; Patients; Pattern; Performance; Perinatal; Perinatal Disorder; Personal Satisfaction; Persons; Phase; Pilot Projects; Placenta; Placenta Diseases; Placental Biology; Placentation; Plant Roots; Point Mutation; Positioning Attribute; Pre-Eclampsia; Pregnancy; Pregnancy Complications; Pregnancy Outcome; Pregnancy Tests; Pregnant Women; Property; Proteins; Psychological Stress; Public Health; Publishing; Reaction; Research; Risk; Risk Assessment; Risk Management; Sampling; Second Pregnancy Trimester; Small for Gestational Age Infant; Specimen; Spontaneous abortion; Standardization; Technology; Testing; Third Pregnancy Trimester; Translating; Translational Research; Villous; adverse outcome; adverse pregnancy outcome; alpha-Fetoproteins; assay development; base; biomarker panel; cell type; cohort; cost; design; diagnostic tool; disability; early pregnancy; early pregnancy loss; experimental study; fetal; fetus cell; health management; healthy pregnancy; human fetal cells; implantation; in vivo; indexing; innovation; novel; perinatal period; pregnancy disorder; pregnant; premature; protein biomarkers; protein expression; screening; sex; tool; translational medicine; trophoblast; ultrasound

There is a lack of clinical information available on early human placentation when many pregnancy pathologies originate. To address this gap and to begin the development of robust diagnostic tools to manage pregnancy complications we propose a pilot study to analyze disease specific proteins in placental trophoblast cells. Using a safe pap smear between 5 to 20 wks of gestation we can non-invasively capture hundreds of homogeneous, HLA-G-and hCG expressing trophoblast cells. We propose that these placental cells are useful for assessing pregnancy status and assessing risk of perinatal disease in vivo. Our premise is based on our published data demonstrating that isolated placental cells obtained express extra villous trophoblast lineage markers (e.g., hCG, HLA-G), and have molecular profiles associated with pathology. Immunocytochemical (ICC) protein ex- pression analysis of the cells demonstrates altered levels of several key proteins in pregnancies that later develop miscarriage, fetal growth restriction (FGR) or preeclampsia. Since the applied method are difficult to translate into a clinical tool (standardization and multiplex capabilities), we will use now a robust and highly sen- sitive single cell mass cytometry assay that does not require prior cell isolation from the clinical sample. We hypothesize that, based on robust ICC data obtained in our published studies, that AFP and PGF levels are significantly altered in EVT cells from pregnancies linked to FGR, a key indicator of placenta-based perinatal disorders. Therefore, we will rigorously determine if expression of these proteins identified in our preliminary studies in fetal cells from cervical specimens correlate with fetal growth rates, considering fetal sex and other relevant factors. The specific aim of this application is to establish the fundamental relationship of cervical extra-villous trophoblast biomarker levels to pregnancy outcomes. We will quantify AFP and PGF and cell type specific proteins in placental cells in cervical specimens in the first trimester, comparing cohorts of subjects with normal term pregnancies to those with adverse outcomes associated with low birth weight for GA at birth. This goal will be accomplished in three milestones. Milestone 1, we will validate a superior high- sensitivity single cell mass spectrometry assay (MSA) to quantify AFP and PGF in trophoblast cells and opti- mize the assay with clinical specimens. Milestone 2, we will collect up to 100 specimens of trophoblast cells in the first trimester, along with de-identified medical records to determine pregnancy outcomes. Milestone 3, the remaining patient trophoblast samples will be assayed for AFP and PGF and other protein levels and compared to patient outcomes, particularly reduced birth weight for gestational age. These experiments will validate the testing platform and provide preliminary data for a larger clinical study to establish a perinatal risk scores and a test for placental disorders. Our long-term goal is to develop and commercialize testing kits and products with unique IP for early diagnosis of perinatal pathologies using EVT cells obtained safely during ongoing pregnancies. Building this assay will allow the creation of risk scores that will enable the identification of high as well as low risk patients that will aide to reduce psychological stress for patients and help shifting healthcare burden to patients in need.

目前缺乏关于人类早期胎盘形成的临床资料， 起源。为了解决这一差距，并开始开发强大的诊断工具来管理怀孕 并发症我们提出了一个初步研究，以分析胎盘滋养层细胞中的疾病特异性蛋白质。使用 在妊娠5至20周之间的安全巴氏涂片，我们可以非侵入性地捕获数百个同质的， 表达HLA-G和hCG的滋养层细胞。我们认为这些胎盘细胞可用于评估 妊娠状况和评估体内围产期疾病的风险。我们的前提是基于我们公布的数据 证明获得的分离的胎盘细胞表达绒毛外滋养层谱系标记（例如， hCG、HLA-G），并具有与病理学相关的分子谱。免疫细胞化学（ICC）蛋白 细胞的表达分析表明，怀孕期间几种关键蛋白质的水平发生了变化， 发生流产、胎儿生长受限（FGR）或先兆子痫。由于应用的方法很难 转化为临床工具（标准化和多重功能），我们现在将使用一个强大的，高度敏感的， 不需要从临床样品中预先分离细胞的选择性单细胞团细胞计数测定。我们 假设，根据我们发表的研究中获得的可靠的ICC数据，AFP和PGF水平是 在与FGR相关的妊娠中，EVT细胞发生了显着变化，FGR是基于胎盘的围产期疾病的关键指标。 紊乱因此，我们将严格确定这些蛋白质的表达是否在我们的初步研究中鉴定。 考虑到胎儿性别和其他因素，对宫颈标本中胎儿细胞的研究与胎儿生长率相关 相关因素。本申请的具体目的是建立宫颈癌的基本关系， 绒毛外滋养层生物标志物水平与妊娠结局的关系。我们将定量AFP和PGF， 妊娠早期子宫颈标本中胎盘细胞中的细胞类型特异性蛋白，比较 正常足月妊娠受试者与GA低出生体重相关不良结局受试者 在出生时。这一目标将分三个阶段实现。里程碑1，我们将验证一个上级- 敏感性单细胞质谱分析（MSA）定量滋养层细胞中的AFP和PGF， 米泽测定与临床标本。里程碑2，我们将收集多达100个滋养层细胞标本 沿着去身份化的医疗记录，以确定怀孕结果。里程碑3， 剩余的患者滋养层样品将被分析AFP和PGF以及其它蛋白质水平， 与患者结局相比，特别是胎龄出生体重降低。这些实验将 验证测试平台，并为更大规模的临床研究提供初步数据，以确定围产期风险 分数和胎盘疾病的测试。我们的长期目标是开发和商业化检测试剂盒， 具有独特IP的产品，用于使用在妊娠期间安全获得的EVT细胞进行围产期病理早期诊断 正在怀孕。构建此测定将允许创建风险评分，从而能够识别 高风险和低风险患者，这将有助于减轻患者的心理压力， 对有需要的患者造成医疗负担。