Augmenting and Assaying Nav1.1 Protein Quantity for Dravet Syndrome Therapy

Area of Science:

• Neuroscience
• Genetics
• Stem Cell Biology

Background:

• Dravet Syndrome (DS) is a severe developmental and epileptic encephalopathy.
• DS is primarily caused by loss-of-function variants in the SCN1A gene, which encodes the Nav1.1 voltage-gated sodium channel.
• Restoring Nav1.1 function is a key therapeutic goal for SCN1A-related disorders.

Purpose of the Study:

• To validate conserved upstream open reading frames (uORFs) in SCN1A as regulators of Nav1.1 translation.
• To establish and validate a human induced pluripotent stem cell (iPSC) model for quantifying endogenous Nav1.1 translation.
• To demonstrate the therapeutic potential of modulating Nav1.1 levels using antisense oligonucleotides (ASOs).

Main Methods:

• Reporter assays were used to validate the regulatory role of SCN1A uORFs.
• A human iPSC line with endogenously HiBiT-tagged Nav1.1 was generated.
• Gapmer and splice-switching ASOs were employed to modulate Nav1.1 expression in iPSCs.
• Quantitative analysis of endogenous Nav1.1 protein levels was performed.

Main Results:

• Conserved uORFs in SCN1A were confirmed to regulate Nav1.1 translation.
• The iPSC model demonstrated dose-dependent downregulation of Nav1.1 with gapmer ASOs.
• Upregulation of Nav1.1 was achieved using splice-switching ASOs in the iPSC model.
• The developed platform proved sensitive for quantifying endogenous Nav1.1 translation.

Conclusions:

• Increasing Nav1.1 protein from a wild-type allele is a therapeutically viable strategy for SCN1A-related disorders.
• The HiBiT-tagged iPSC line serves as a valuable platform for screening translational modulators.
• This study validates a novel approach for developing and testing therapies for Dravet Syndrome and related conditions.