Developmental Neurotox Assay using Scalable Neurons and Astrocytes in High Content Imaging

There is unmet need for rapid, cost effective in vitro methods to identify chemicals that pose a hazard to the human brain. A number of in vitro assays utilizing primary cells have been tested in an effort to devise effective models to prioritize chemicals for further in depth toxicity studies. However, isolation of primary human neural tissue is cost and labor intensive, and cannot be produced in the quantities required for high throughput screening (HTS) and high content imaging (HCI). In addition, current systems do not include astrocytes which have a significant role in maintaining the health and function of the central nervous system. In Phase I, ArunA Biomedical (ArunA) addressed this deficit by developing hPSC derived neuron and glia co-culture system, containing progenitor cell derived neurons (hN2TM) and astrocytes (hAstroPro™), which more faithfully recapitulates early neural development. The use of progenitor cells to produce these cells enables scalable, HCI-amenable formats for toxicant screening. During Phase I, we successfully optimized the production of assay-ready neurons and astrocytes, developed protocols for cryopreservation and plating of co-culture systems, and demonstrated the feasibility of co-culturing astrocytes and neurons at assay-optimized ratios. ArunA showed that the co-culture system could be used to document the toxic effects of neurotoxins on neurite outgrowth, a neuro-developmentally relevant endpoint. Phase I studies reinforced the need for a more representative system by demonstrating that known neurotoxins do indeed evoke different neuronal responses in the presence of astrocytes. In Phase II, ArunA will develop protocols for human multi-cellular HCI & HTS neurotox assays in a ready-to-use kit format. ArunA outlines objectives aimed at optimizing scale-up production of astrocytes from its proprietary progenitor stem cells, and implementing further improvements to the assay system so that our novel co-culture system can be adopted for diverse use to address a broader market. Importantly, by providing a neuron and glia co-culture system, ArunA will solve the problem of more faithful representation of human neural tissue. For example, in the long term, ArunA’s multi-functional neural assays will enable electrophysiological multi-electrode array recordings which require a more physiologically representative environment that includes neurons and astrocytes. Moreover, developing products in which the end user can alter the ratio of neurons to astrocytes, will provide a unique system to examine the effects of compounds in a measured and more exacting system than the uncontrolled ratios of cells associated with all other commercially available systems.