Allele-Selective Thiomorpholino Antisense Oligonucleotides as a Therapeutic Approach for Fused-in-Sarcoma Amyotrophic...

Area of Science:

• Genetics and Molecular Biology
• Neuroscience
• Oligonucleotide Therapeutics

Background:

• Pathogenic variations in the fused in sarcoma (FUS) gene are linked to aggressive forms of amyotrophic lateral sclerosis (ALS).
• FUS-ALS is a dominant disease, necessitating allele-selective therapeutic strategies.
• Antisense oligonucleotides (AOs) represent a promising platform for gene-targeted therapies.

Purpose of the Study:

• To explore the potential of allele-selective knockdown of the FUS gene using antisense oligonucleotides (AOs).
• To compare the efficacy and selectivity of methoxyethyl (MOE) and thiomorpholino (TMO) modified AOs.
• To evaluate the impact of AO length and gap length on allele selectivity and efficiency.

Main Methods:

• Design and evaluation of gapmer-type AOs targeting neutral polymorphisms in the FUS gene.
• Testing of AOs with methoxyethyl (MOE) or thiomorpholino (TMO) modifications in human fibroblasts.
• Assessment of FUS transcript levels, nuclear inclusions, SFPQ aggregation, and off-target gene knockdown.

Main Results:

• TMO-modified AOs demonstrated superior allele selectivity and efficacy compared to MOE-modified AOs.
• TMO-modified gapmer knockdown resulted in up to 93% of detected FUS transcripts originating from the non-target allele.
• TMO-modified AOs showed reduced formation of nuclear inclusions and SFPQ aggregation compared to MOE-modified AOs.

Conclusions:

• Allele-selective knockdown of FUS is a viable therapeutic strategy for FUS-ALS.
• The TMO modification offers significant advantages for allele-selective AO applications.
• Optimized AO design, including length and gap length, is crucial for maximizing therapeutic potential.