Abstract
Background & Purpose
Rett syndrome (RTT) and CDKL5 deficiency disorder (CDD) are rare X-linked neurodevelopmental disorders characterized by cognitive deficits, developmental delays, and seizures. Existing 2D cultures and animal models fail to capture the complex human neuronal activity needed for effective drug screening. 28bio developed microBrain™, a human iPSC-derived cortical organoid platform, designed for high-throughput functional screening of disease-modifying therapeutics. This study screens RTT and CDD organoids to identify novel drug candidates and biological targets.
Methods
- Human iPSC-Derived Cortical Organoid Culture:
- RTT and CDD patient-derived iPSCs were differentiated into neural progenitor cells and grown into 3D cortical organoids.
- High-throughput formats (up to 384-well plates) were used for large-scale compound screening.
- Drug Screening & Functional Analysis:
- RTT Screening: 296 compounds from the SMART library (curated by the International Rett Syndrome Foundation) were tested over four weeks to distinguish disease-modifying vs. symptomatic treatments.
- CDD Screening: Over 5000 compounds were screened to identify novel targets and molecules that restore neural activity.
- Calcium Imaging & Electrophysiology:
- 28bio’s AnalytiX software quantified peak shape, strength, and activity bursts to assess compound efficacy.
Results
- RTT Drug Candidates:
- Acetylcholinesterase (AChE) inhibitors (Donepezil) and histone deacetylase (HDAC) inhibitors (Vorinostat) outperformed existing clinical drugs (Trofinetide & Anavex 2-73).
- RTT organoids displayed irregular calcium oscillations, which were normalized by AChE and HDAC inhibitors.
- CDD Drug Candidates:
- 22 novel compounds spanning 15 biological targets were identified within one year of screening.
- CDD organoids showed hyperexcitability (high calcium peak frequency), which was reduced by selective drug candidates.
- Compounds were assessed for disease selectivity and safety, identifying molecules with high therapeutic potential.
Conclusion
The microBrain™ platform successfully models functional deficits in RTT and CDD and enables high-throughput drug discovery. Novel AChE and HDAC inhibitors showed superior efficacy in RTT, while CDD screening identified 22 new drug candidates in under a year. Future efforts will validate leading candidates and expand screening to additional neurodevelopmental disorders.
