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

> **NIH NIH R44** · STEM PHARM, INC. · 2021 · $930,433

## Abstract

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.
.

## Key facts

- **NIH application ID:** 10259033
- **Project number:** 2R44ES029898-02A1
- **Recipient organization:** STEM PHARM, INC.
- **Principal Investigator:** Connie S Lebakken
- **Activity code:** R44 (R01, R21, SBIR, etc.)
- **Funding institute:** NIH
- **Fiscal year:** 2021
- **Award amount:** $930,433
- **Award type:** 2
- **Project period:** 2019-05-15 → 2023-02-28

## Primary source

NIH RePORTER: https://reporter.nih.gov/project-details/10259033

## Citation

> US National Institutes of Health, RePORTER application 10259033, Hydrogel-enabled self-assembled human brain organoids for neurotoxicity applications (2R44ES029898-02A1). Retrieved via AI Analytics 2026-06-11 from https://api.ai-analytics.org/grant/nih/10259033. Licensed CC0.

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