Abstract
Background & Purpose
Rett syndrome (RTT) is a severe neurodevelopmental disorder caused by MECP2 mutations, with no disease-modifying treatments currently available. While 2D cultures and animal models have been used for drug discovery, they fail to translate effectively to humans. 28bio developed microBrain™, a human iPSC-derived cortical organoid platform, designed for high-throughput, reproducible compound screening. This study utilizes the microBrain™ RTT model to identify and analyze potential therapeutic candidates, with a focus on acetylcholinesterase (AChE) and histone deacetylase (HDAC) inhibitors.
Methods
- Cortical Organoid Culture:
- RTT patient-derived iPSCs were differentiated into 3D cortical organoids.
- Organoids were grown in high-throughput plate formats (up to 384 wells) for scalable drug screening.
- Functional activity was analyzed using multi-electrode arrays (MEA) and fluorometric imaging (FLIPR) assays.
- Drug Screening:
- A library of compounds was screened, including:
- AChE inhibitors: Donepezil, Rivastigmine, Galantamine
- HDAC inhibitors: Panobinostat, Belinostat, Vorinostat
- Clinical trial candidates: Trofinetide, Anavex 2-73
- Electrophysiological and phenotypic activity was quantified, focusing on bursting patterns, waveform characteristics, and toxicity.
Results
- Functional Phenotypic Recovery:
- RTT organoids displayed clustered bursts of hyperactivity, distinguishing them from control organoids.
- AChE and HDAC inhibitors significantly improved functional activity, while clinical candidates Trofinetide and Anavex 2-73 showed minimal recovery.
- Vorinostat (HDAC inhibitor) improved function but exhibited toxicity.
- AChE inhibitors provided strong, dose-dependent recovery without overt toxicity.
- High-Throughput Scalability & Data Reproducibility:
- Both RTT and control organoids showed similar neuron-to-astrocyte ratios, making the model ideal for consistent comparisons.
- Smart library screening data revealed distinct recovery fingerprints, correlating with biological target specificity.
Conclusion
The microBrain™ platform successfully models Rett syndrome functional deficits and enables human-relevant drug discovery. AChE and HDAC inhibitors emerged as the most promising therapeutic candidates, outperforming clinical trial drugs. Future directions include:
- Investigating combination therapies for enhanced recovery.
- Expanding microBrain™ models to include microglia for neuroinflammation studies.
- Further optimizing high-throughput screening to accelerate therapeutic development for RTT and other neurodevelopmental disorders.
