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Hydrogel-enabled self-assembled human brain organoids for neurotoxicity applications

用于神经毒性应用的水凝胶自组装人脑类器官

基本信息

批准号：
10374175
负责人：
Connie S Lebakken
金额：
$ 78.35万
依托单位：
STEM PHARM, INC.
依托单位国家：
美国
项目类别：
财政年份：
2019
资助国家：
美国
起止时间：
2019-05-15 至 2024-02-29
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/10374175
关键词：
Address Animal Model Animals Automation Biological Biological Assay Biological Models Biological Products Bioreactors Blood Vessels Brain Cell Communication Cell Density Cell Differentiation process Cells Cerebrovascular system Characteristics Chemicals Clinical Coculture Techniques Communicable Diseases Complex Contracts Cost Analysis Data Data Set Development Disease Drug Modelings Embryo Endothelial Cells Engineering Extracellular Matrix Failure Gene Expression Gene Expression Profiling Genes Genetic Transcription Goals Harvest Human Hydrogels Image Immune Immunofluorescence Immunologic Inflammation International Laboratories Liquid substance Manuals Methods Microelectrodes Microglia Modeling Monitor Neuraxis Neurons Organoids Output Pathology Pharmaceutical Preparations Phase Poison Protein Analysis Publishing Quality Control Reproducibility Risk Safety Signal Transduction Small Business Innovation Research Grant Source Suspensions System Techniques Testing Thinness Time Toxic effect Toxicology Toxin Transcript Tumor-Derived Universities Validation Variant Wisconsin Work archive data archived data cell type cost cost effective cytokine data integrity density developmental neurotoxicity drug candidate drug discovery experimental study feeding human embryonic stem cell in vitro Model induced pluripotent stem cell multi-electrode arrays multimodality multiplex assay nerve stem cell neural model neuroinflammation neurotoxicity novel pre-clinical relating to nervous system response screening single-cell RNA sequencing small molecule stem stem cells transcriptome sequencing

项目摘要

Project Summary/Abstract There is a critical need to move advanced Central Nervous System (CNS) models into screening applications for drug discovery and toxicology applications. Current in vitro models do not accurately reflect the complexity of cell types and important cell-cell interactions and animal models fail to recapitulate the human condition. There is also a great need for more accurate and scalable models for developmental neurotoxicity screenings as there are 86,405 compounds listed on the Toxic Substance Control Act inventory17 with little biological data to understand their risks. Recent advances in stem-cell derived neural organoids have led to use of these models to study developmental mechanisms, infectious diseases, and toxicology applications (18-26 and reviewed in27-29), but their cost, complexity, and workflow requirements make them challenging to transition to screening applications. Work performed in our successful Phase I activities at Stem Pharm with iPSC-derived precursor and differentiated cells has demonstrated that complex neural organoids containing a variety of neural subtypes can be developed reproducibly in a 96-well plate on engineered hydrogel substrates. Unlike organoids cultured in suspension systems, these organoids can be formed, cultured, and assayed in multi-well plates. RNA-seq analysis demonstrated high intraclass correlation and low coefficients of variation. Importantly, we demonstrated incorporation of microglia into the organoids and demonstrated their activation as a model of neural inflammation as well as their activation or depletion in response to compound treatment. In order to bring this novel model to the market we propose the following specific aims for the Phase II proposal: 1) To optimize timing and seeding densities with cells derived from a single iPSC-donor source, optimize incorporation of microglia to maintain robust activation signatures but decrease cost and maintain data integrity. To compare a less-costly transcriptional read-out, the TempO-Seq S1500 human panel, to our RNA-seq data obtained in Phase I activities and to validate a qPCR panel for product release quality control. 2) To validate organoids generated on our thin hydrogel coatings to enable better imaging options, microelectrode array analysis and liquid handling automation and 3) Validate multiplexed assays to assess multiple responses in single wells including MEA analysis, cytokine and LDH release and harvest for transcript or protein analysis. This work will lead to the first commercially available neural organoid containing vascular cells and microglia with broad applicability in both toxicology and drug discovery markets. .

项目总结/文摘