Modified Oligonucleotides for DMD Exon Skipping
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Solution Overview
Problem
Current oligonucleotides for treating Duchenne Muscular Dystrophy (DMD) have limitations in delivery, biostability, biodistribution, and cellular uptake, necessitating further optimization for enhanced therapeutic efficacy.
Innovation Solution
Development of oligonucleotides comprising 2'-O-methyl RNA monomers with a phosphorothioate backbone and incorporating 5-methylpyrimidine and/or 2,6-diaminopurine bases, which are designed to be shorter in length and optimized for improved binding affinity, stability, and reduced immunogenicity.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Stability of the object's composition
If oligonucleotides are chemically modified to resist endonucleases and exonucleases, then biostability is improved, but delivery and cellular uptake are reduced
Solution Approach 1:
The patent applies different chemical modifications to specific positions within the oligonucleotide sequence. The 2'-O-methyl modification is applied to all nucleotides providing backbone stability, while phosphorothioate linkages are strategically placed at specific positions (e.g., positions 2-18) to enhance stability without compromising cellular uptake. This localized differentiation of modification types resolves the contradiction between biostability and cellular uptake.
Solution Approach 2:
The oligonucleotide employs a composite chemical structure combining multiple modification types: 2'-O-methyl RNA monomers, phosphorothioate backbones, 5-methylpyrimidine bases, and 2,6-diaminopurine bases. This composite chemistry integrates the biostability benefits of phosphorothioate linkages with the enhanced cellular uptake properties of 2'-O-methyl modifications, simultaneously addressing both requirements.
2Length of moving object
If oligonucleotide length is reduced to improve delivery, then biodistribution is improved, but binding affinity is reduced
Solution Approach 1:
The patent changes the chemical parameters of the oligonucleotide by incorporating 5-methylpyrimidine and 2,6-diaminopurine bases, which increase binding affinity through enhanced base stacking and hydrophobic interactions. This allows the use of shorter oligonucleotide sequences (15-30 nucleotides) that improve delivery and biodistribution while maintaining high binding affinity to the target pre-mRNA through these modified base interactions.
Solution Approach 2:
The combination of 2'-O-methyl modifications with 5-methylpyrimidine and 2,6-diaminopurine bases creates a composite structure where the modified bases provide enhanced binding affinity per nucleotide. This compensates for the reduced total length, allowing shorter sequences to achieve equivalent or superior binding strength while improving delivery characteristics.
3Ease of manufacture
If standard oligonucleotide chemistry is used, then synthesis is simpler, but immunogenicity is increased
Solution Approach 1:
The patent modifies the chemical parameters by incorporating 5-methylpyrimidine and 2,6-diaminopurine bases, which reduce immunogenicity through altered base recognition by immune sensors. These modified bases change the chemical and physical properties of the oligonucleotide, reducing its recognition as foreign by the immune system while maintaining manufacturability through established solid-phase synthesis methods.
4Reliability
If oligonucleotides are optimized for binding affinity, then exon skipping activity is improved, but delivery efficiency is reduced
Solution Approach 1:
The patent applies 2'-O-methyl modifications to all nucleotides to enhance binding affinity and exon skipping activity at the target site, while simultaneously incorporating phosphorothioate linkages and 5-methylpyrimidine bases to improve delivery efficiency. The local differentiation of these modifications ensures high binding affinity where needed while maintaining favorable delivery properties throughout the sequence.
Solution Approach 2:
The composite chemistry combining 2'-O-methyl RNA with 5-methylpyrimidine and 2,6-diaminopurine bases creates an oligonucleotide that achieves both high binding affinity for effective exon skipping and improved delivery efficiency. The modified bases enhance cellular uptake and tissue distribution while the 2'-O-methyl backbone provides stable target binding, resolving the contradiction between these two requirements.
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
These modified oligonucleotides demonstrate enhanced exon skipping activity, improved biodistribution, and increased cellular uptake, leading to increased production of functional dystrophin protein, thereby alleviating DMD symptoms.
Implementation Method 1
antisense oligonucleotides (AONs) which bind to the exon flanking or containing the mutation and which interfere with its splicing signals
Implementation Method 2
chemically modified to resist endonucleases, exonucleases and RNaseH
Implementation Method 3
the dystrophin deficiency in DMD, resulting in damaged and thus more permeable fiber membranes, actually promotes uptake
Data Source
Figure 1a~1b
Figure 1c
Figure 2a~2b
AI summary
The current invention provides an improved oligonucleotide and its use for treating, ameliorating, preventing and/or delaying DMD or BMD.