Splicing-Dependent Gene Silencing With ASO Isoform Control
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Solution Overview
Problem
Existing methods struggle to effectively regulate gene expression by modulating splicing, particularly in cases where introns and exons require precise control for therapeutic purposes, such as in conditions like myotonic dystrophy type I, Huntington's disease, and certain cancers, where specific isoforms need to be silenced or activated.
Innovation Solution
The use of antisense oligonucleotides (ASOs) and small molecules targeted to internal exons or silencing elements in genes to inhibit or activate transcription start sites, including the introduction of exogenous splice sites and exons to enhance gene expression, using morpholino oligonucleotides, phosphorothioate ASOs, and other specific ASO types.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Manufacturing precision
If antisense oligonucleotides target internal exons to inhibit splicing, then specific isoform expression is reduced, but transcription from proximal start sites is also suppressed
Solution Approach 1:
The patent segments the gene expression control by targeting specific internal exons (e.g., exon 12 in TSKU gene) with antisense oligonucleotides, allowing selective inhibition of particular isoforms while preserving expression from other start sites. This enables precise spatial control at the molecular level.
Solution Approach 2:
The invention applies local quality by designing ASOs with specific sequences complementary to particular exon regions, creating localized effects at targeted splice sites without affecting the entire gene. The ASO concentration and binding affinity are optimized for specific genomic locations.
2Adaptability or versatility
If multiple transcription start sites exist in a gene, then regulatory flexibility is increased, but controlling specific isoform expression becomes more difficult
Solution Approach 1:
The patent uses antisense oligonucleotides as intermediary molecules that bridge the gap between transcriptional start sites and splicing outcomes. By binding to specific exon sequences, ASOs mediate the connection between proximal start site activity and downstream isoform production, enabling indirect control of alternative start site usage.
Solution Approach 2:
The invention implements feedback control where ASO-mediated splicing inhibition of internal exons provides information about proximal start site activity, which then feeds back to regulate overall transcription. The splicing outcome serves as a readout that influences further transcriptional regulation.
Applied Scientific Principles
This section explains which scientific principles are used to turn an abstract innovation direction into a practical engineering solution.
Function Achieved in This Case
This approach allows for precise regulation of gene expression by inhibiting or activating specific isoforms, enhancing therapeutic protein production in conditions like Duchenne muscular dystrophy and cancers, by targeting internal exons or silencing elements to modulate splicing patterns.
Implementation Method 1
antisense oligonucleotides (ASOs) and small molecules targeted to internal exons or silencing elements in genes
Implementation Method 2
using morpholino oligonucleotides, phosphorothioate ASOs, and other specific ASO types
Data Source
AI summary
Disclosed herein are methods for inhibiting or activating the transcription of a gene of interest, or inhibiting or activating the transcription of specific mRNA isoforms of a gene by using antisense oligonucleotides and/or small molecules. Also described herein are methods for activating transcription from a promoter and increasing overall gene expression by creating of a new splice site in a gene of a cell.