RECODE: Synergistic Genetic and Microenvironmental Engineering Platforms For Directed Liver Organoid Differentiation

RECODE：用于定向肝脏类器官分化的协同遗传和微环境工程平台

• 批准号: 2134986
• 负责人: Salman Khetani
• 金额: $ 150万
• 依托单位: University of Illinois at Chicago
• 依托单位国家: 美国
• 项目类别: Standard Grant
• 财政年份: 2021
• 资助国家: 美国
• 起止时间: 2021-09-01 至 2025-08-31
• 项目状态: 未结题
• 来源: https://www.nsf.gov/awardsearch/showAward?AWD_ID=2134986&HistoricalAwards=false
• 关键词: RECODE; Synergistic; Genetic; Microenvironmental; Engineering

Injury to the liver due to toxic drugs is a leading cause of acute liver failures. Unfortunately, testing drugs on animals before human clinical trials is inadequate due to significant differences between animals and humans in liver function. Therefore, human liver organoids generated from stem cells are being increasingly utilized to mitigate limitations with animal testing; however, current organoids are not reproducibly manufactured, and their functions do not approximate those in the human body. This effort will engineer new cell culture devices and genetically edit the cells in specific ways to make liver organoids more reproducible for routine drug testing. The approaches and technologies developed can be broadly applicable beyond liver to other organ types. Additionally, underrepresented minority high school and undergraduate students, as well as high school teachers from underserved districts in Chicago, will be provided hands-on opportunities to engage in the research topics of this effort, including novel curriculum development for high schools using the concepts developed here.This RECODE project will synergize advances in microfabricated cell culture devices, induced pluripotent stem cell (iPSC) biology, synthetic biology, single cell transcriptomics, and computational biology to address a critical question: What are the design rules and underlying mechanisms that lead to functionally mature and reproducible 3-dimensional organoids within scalable culture platforms? This effort will utilize iPSC-derived human liver cells with validation against primary human liver cells and whole human livers. Specifically, this project will develop an unprecedented pipeline of microenvironmental engineering, clustered regularly interspaced short palindromic repeats (CRISPR)-Cas transcriptional activation techniques that can activate introduced and endogenous genes with precise temporal control, and computational biology approaches that can infer transcriptional factor activity from single cell RNA sequencing data on liver organoids. The human liver organoids developed here can be used to develop safer drugs, industrial chemicals, and vaccines for humans, and for elucidating the underlying principles of human liver development, physiology, and disease. The microfluidic, synthetic biology, and computational approaches/platforms developed here will serve as a broader resource to investigators developing reproducible organoids for various applications. The research efforts will be integrated with sustainable hands-on educational efforts aimed at training high school and undergraduate underrepresented minority students as well as high school teachers through summer internship programs in the approaches developed here. Such efforts will introduce cutting-edge research concepts earlier in high school, thereby preparing students better for a rigorous engineering/bioengineering curriculum at the college level. This award is co-funded by the Systems and Synthetic Biology Cluster in the Division of Molecular and Cellular Biosciences and the Engineering Biology and Health Cluster in the Division of Chemical, Bioengineering, Environmental, and Transport Systems.This award reflects NSF's statutory mission and has been deemed worthy of support through evaluation using the Foundation's intellectual merit and broader impacts review criteria.

毒性药物对肝脏的损伤是急性肝功能衰竭的主要原因。不幸的是，在人类临床试验之前对动物进行药物测试是不够的，因为动物和人类之间在肝功能方面存在显着差异。因此，由干细胞产生的人类肝脏类器官越来越多地被用于减轻动物试验的限制;然而，目前的类器官不能重复制造，它们的功能与人体中的功能不接近。这项工作将设计新的细胞培养设备，并以特定的方式对细胞进行遗传编辑，使肝脏类器官在常规药物测试中更具重现性。开发的方法和技术可以广泛应用于肝脏以外的其他器官类型。此外，代表性不足的少数民族高中和本科生，以及来自芝加哥服务不足地区的高中教师，将有机会参与这项工作的研究课题，包括使用这里开发的概念为高中开发新的课程。合成生物学、单细胞转录组学和计算生物学来解决一个关键问题：在可扩展的培养平台中，导致功能成熟和可重复的三维类器官的设计规则和潜在机制是什么？这项工作将利用iPSC衍生的人类肝细胞，并对原代人类肝细胞和整个人类肝脏进行验证。具体来说，该项目将开发一个前所未有的微环境工程管道，聚类规则间隔短回文重复序列（CRISPR）-Cas转录激活技术，可以通过精确的时间控制激活引入和内源性基因，以及计算生物学方法，可以从肝脏类器官的单细胞RNA测序数据中推断转录因子活性。这里开发的人类肝脏类器官可用于为人类开发更安全的药物，工业化学品和疫苗，并阐明人类肝脏发育，生理学和疾病的基本原理。这里开发的微流体，合成生物学和计算方法/平台将为研究人员开发可再生的类器官提供更广泛的资源。研究工作将与可持续的实践教育工作相结合，旨在通过这里开发的方法，通过暑期实习计划培训高中和本科代表性不足的少数民族学生以及高中教师。这些努力将在高中早期引入尖端的研究概念，从而使学生更好地为大学阶段严格的工程/生物工程课程做好准备。该奖项由分子和细胞生物科学部的系统和合成生物学集群以及化学、生物工程、环境和运输系统部的工程生物学和健康集群共同资助。该奖项反映了NSF的法定使命，并通过使用基金会的知识价值和更广泛的影响审查标准进行评估，被认为值得支持。