Calcium-Enhanced Medium-Based Delivery of Splice Modulating Antisense Oligonucleotides in 2D and 3D hiPSC-Derived...

Area of Science:

• Neuroscience
• Stem Cell Biology
• Molecular Therapeutics

Background:

• Antisense technology is crucial for treating inherited brain diseases, with many ASO therapeutics in development.
• Human induced pluripotent stem cells (hiPSCs) are valuable for modeling neurological diseases and testing therapies.
• Delivering antisense oligonucleotides (ASOs) to hiPSC-derived neuronal models presents significant challenges.

Purpose of the Study:

• To evaluate various delivery methods for ASOs in human hiPSC-derived neuronal models.
• To identify an efficient and low-toxicity method for ASO delivery in 2D and 3D neural cultures.
• To optimize preclinical ASO strategies for neurological disorder treatment.

Main Methods:

• Utilized 2D and 3D hiPSC-derived neuronal models.
• Tested four ASO delivery methods: lipid-based transfection, gymnotic uptake, Ca(2+)-enhanced medium (CEM)-based delivery, and electroporation.
• Assessed delivery efficiency and cellular toxicity.

Main Results:

• CEM-based delivery demonstrated high efficiency and low toxicity in both 2D neuronal cultures and 3D brain organoids.
• CEM delivery was slightly more effective in neurons compared to astrocytes.
• Other tested methods showed varying degrees of success and toxicity.

Conclusions:

• CEM-based delivery is a promising method for introducing ASOs into hiPSC-derived neural models.
• This approach facilitates the study and treatment of neurological disorders using ASO therapeutics.
• Further optimization of CEM delivery holds potential for advancing preclinical ASO strategies.