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Development and Characterization of Human iPSC-derived Neurons for Drug Discovery Applications
Lucas Chase1, Monica Strathman1, Jeff Grinager1, David Majewski1, Regina Whitemarsh2, Sabine Pellett2, Oksana Sirenko3, Jayne Hesley3, Penny Tavormina3, Casey Stankewicz1, Matt George1, Ning Liu1, Nathan Meyer1, Matthew Riley1, Xuezhu Feng1, Eric Johnson2, Wen Bo Wang1 and Brad Swanson1

1Cellular Dynamics International, Inc., Madison, WI 53711;
2Department of Bacteriology, University of Wisconsin, Madison, WI 53706; 3Molecular Devices, Sunnyvale, CA 94089
Sponsored by Cellular Dynamics CDI
The human brain represents a complex organ that has consistently been proven difficult to model in vitro. Current models including primary rodent tissue and immortalized cell lines have served as mainstays in both academic research and the pharmaceutical industry. These models, while providing a means for numerous landmark discoveries, have suffered from various issues including biological relevance, reproducibility and scalability. Considerable efforts have been made specifically within the pharmaceutical industry to reduce late-stage drug attrition through the development of more relevant in vitro human model systems. One area given significant attention has been the development of platforms than can enable the modeling of human degenerative (e.g. Alzheimer’s and Parkinson’s disease) and genetic (i.e. Huntington's disease and muscular dystrophy) diseases as well as neurotoxicity. The recent discovery of induced pluripotent stem cells (iPSCs) not only overcomes the ethical and logistical issues associated with human embryonic stem cells, but also provides a flexible platform for generating various differentiated cell types from diseased individuals. Using this platform, we have developed a highly consistent and scalable protocol to differentiate and cryopreserve purified human iPSC-derived neurons, called iCell® Neurons. Phenotypically, these cells are >90% pure as measured using flow cytometry for presence of the neuronal marker class III beta tubulin (TuJ1) and absence of the progenitor marker nestin. Within 24hrs of thawing, these neurons display a typical neuronal morphology including a dense network of neurites. Detailed phenotypic analyses reveal that these neurons are comprised of a mix of predominantly GABAergic and Glutamatergic subtypes as measured at both the gene expression and protein levels and form characteristic synaptic connections. Functionally, these cells reveal typical electrophysiological characteristics as measured using single-cell patch clamp to detect both spontaneous and evoked action potentials as well as functional ion channels. Finally, when applied to high throughput applications, including cytotoxicity assays, iCell Neurons reveal characteristic pharmacological responses to known toxic compounds. The results demonstrate not only a novel cell model for use in various academic and pharmaceutical applications, but they also support the use of the iPSC technology as a platform capable of generating neurons against diverse genetic backgrounds.
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