Using Induced Pluripotent Stem Cells to Characterize Cystic Fibrosis

使用诱导多能干细胞来表征囊性纤维化

基本信息

批准号：
10408667
负责人：
Ruobing Wang
金额：
$ 16.79万
依托单位：
BOSTON CHILDREN'S HOSPITAL
依托单位国家：
美国
项目类别：
财政年份：
2021
资助国家：
美国
起止时间：
2021-06-01 至 2026-05-31
项目状态：
未结题

来源：
https://reporter.nih.gov/project-details/10408667
关键词：
3-Dimensional Agonist Air Archives Award Basal Cell Biological Assay Biological Models Biology Boston CRISPR/Cas technology Caring Caucasians Cells Childhood Chloride Channels Chlorides Clinical Cystic Fibrosis Cystic Fibrosis Transmembrane Conductance Regulator Defect Delta F508 mutation Derivation procedure Disease Disease model Drug Modelings Electrophysiology (science)Environment Epithelial Epithelial Cells Evaluation Exhibits Freezing Functional disorder Future Genes Genetic Genetic Diseases Genetic Polymorphism Genotype Goals Human In Vitro Individual Ion Channel Ions Liquid substance Lung diseases Manuscripts Measures Medicine Mendelian disorder Mentors Mentorship Modeling Mucociliary Clearance Mucous body substance Mutation Organoids Patients Pediatric Hospitals Pharmaceutical Preparations Pharmacology Phenotype Physicians Population Positioning Attribute Preparation Protocols documentation Regenerative Medicine Research Research Personnel Research Training Respiratory Disease Role Scientist Severity of illness Study models System Techniques Testing Time TimeLine Training Universities Variant Vertebral column Viscosity airway epithelium base candidate validation career career development cystic fibrosis airway epithelia cystic fibrosis patients disease phenotype drug discovery epithelial Na+ channel exome sequencing genetic variant improved in vitro Model individual patient individual response individualized medicine induced pluripotent stem cell inhibitor lung preservation meetings member molecular phenotype novel precision drugs pulmonary function pulmonary function decline response single-cell RNA sequencing stem cells therapeutic candidate therapeutic evaluation tool

项目摘要

Project Summary/Abstract This proposal details a 5-yr plan to prepare the candidate, Ruobing Wang, MD, for a career as an independent physician-scientist positioned to impact our understanding of pediatric lung diseases, particularly Cystic Fibrosis (CF). As a clinician taking care of CF patients, she has identified rare individuals with homozygous F508del CFTR mutations who met clinical criteria for inclusion as: 1) CF Long-term non-progressors (LTNP), with preserved lung function 2) CF Rapid progressors (RP), whose lung function declines rapidly. Whole exome sequencing uncovered rare missense polymorphisms in i) SCNN1 (which encodes epithelial sodium channel ENaC) in LTNPs, and ii) two genes in epithelial alternative chloride channels (ANO1 and SCL26A9) in RPs. The central hypothesis of the proposal is that the extreme phenotypes of CF are due to the alterations in epithelial ion and fluid transport driven by these gene defects, and that the extreme-phenotype disease severity and mechanism can be modeled and studied in vitro with a reprogrammed cell-based platform. Using a novel protocol, Dr. Wang generated airway basal-like cells from induced pluripotent stem cells (iPSCs) which can then differentiate into basal, multi-ciliated, and secretory lineages on air-liquid interface, forming a functioning airway epithelium with intact barrier function and aberrant trans-epithelial chloride transport. Normalization of chloride transport in these cells by CFTR gene editing confirms the reliability of her model to recapitulate CF phenotype. The central goals of the project are to establish this novel iPSC-platform for CF disease modeling and study the epithelial function of the extreme-phenotype patients. She is now uniquely poised to complete the aims to 1) To test whether iPSC-derived airway epithelia can serve as a platform to model airway epithelial ion and fluid transport and muco-ciliary transport in CF, 2) to model extreme-phenotype CF patient and interrogate the role of candidate modifier genes and their impact on ion and fluid transport, and 3) To establish the iPSC-platform for personalized drug response, and test the therapeutic role of pharmacologic targeting of alternative ion channel candidates. Dr. Wang has 80% protected time from Boston Children's Hospital (BCH) Division of Respiratory Diseases and the Department of Medicine. Her co-sponsors are 1) Dr. Darrell Kotton at the Center for Regenerative Medicine (CReM) at Boston University (BU) with whom she has trained for the past 1.5 years, and 2) Dr. Benjamin Raby, the chief of BCH Division of Respiratory Diseases. Furthermore, Dr. Wang has assembled a team of extraordinary scientific advisory members, each bringing their specific expertise, to assist her career development and scientific research. A detailed training plan is presented that includes mentored research, didactic coursework, presentations at meetings, and a timeline for completion of the research aims, preparation of manuscripts, and future R01 application. The proposed research, training plan, mentorship committee, and scientific-clinical environment at BCH and BU will position the candidate to transition to independence by the end of the award.

项目概要/摘要该提案详细介绍了一个五年计划，旨在为候选人王若冰医学博士做好独立职业生涯的准备医生科学家致力于影响我们对儿科肺部疾病，特别是囊性肺部疾病的理解纤维化（CF）。作为一名照顾 CF 患者的临床医生，她发现了罕见的纯合子个体符合纳入临床标准的 F508del CFTR 突变：1) CF 长期非进展者 (LTNP)，肺功能保留 2) CF 快速进展者 (RP)，其肺功能迅速下降。全外显子组测序发现 i) SCNN1（编码上皮钠通道）中罕见的错义多态性 ENaC）位于 LTNP 中，ii）位于 RP 中上皮替代氯离子通道（ANO1 和 SCL26A9）中的两个基因。这该提案的中心假设是 CF 的极端表型是由于这些基因缺陷驱动的上皮离子和液体运输，以及极端表型疾病可以使用重新编程的基于细胞的平台在体外对严重性和机制进行建模和研究。使用 Wang 博士采用了一种新颖的方案，从诱导多能干细胞 (iPSC) 中生成了气道基底样细胞，然后可以在气液界面上分化为基底谱系、多纤毛谱系和分泌谱系，形成功能气道上皮具有完整的屏障功能和异常的跨上皮氯转运。标准化通过 CFTR 基因编辑在这些细胞中进行氯化物转运证实了她的模型重现 CF 的可靠性表型。该项目的中心目标是建立用于 CF 疾病建模的新型 iPSC 平台并研究极端表型患者的上皮功能。她现在完全准备好完成目的 1) 测试 iPSC 衍生的气道上皮细胞是否可以作为气道模型的平台 CF 中的上皮离子和液体转运以及粘膜纤毛转运，2) 模拟极端表型 CF 患者并询问候选修饰基因的作用及其对离子和液体运输的影响，以及 3) 建立个性化药物反应的iPSC平台，并测试药理学的治疗作用靶向替代离子通道候选物。王医生80%的时间受到波士顿儿童医院的保护医院 (BCH) 呼吸疾病科和医学部。她的共同发起人是 1) 博士。波士顿大学 (BU) 再生医学中心 (CReM) 的达雷尔·科顿 (Darrell Kotton) 与她一起过去 1.5 年的培训经历，以及 2) BCH 呼吸疾病科主任 Benjamin Raby 博士。此外，王博士还组建了一支由杰出的科学顾问组成的团队，每一位都带来了他们的具体专业知识，协助她的职业发展和科学研究。详细的训练计划是提出的内容包括指导研究、教学课程、会议演讲以及时间表研究目标的完成、手稿的准备以及未来R01的申请。拟议的 BCH 和 BU 的研究、培训计划、指导委员会和科学临床环境将定位候选人在颁奖结束时过渡到独立。