mRNA Aptamer Purification via Affinity Ligand Binding
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Solution Overview
Problem
Current methods for purifying messenger RNA (mRNA) for therapeutic use are inefficient and unreliable, leading to impurities such as proteins, non-RNA nucleic acids, and contaminants that hinder the production of highly pure and safe mRNA for large-scale manufacturing.
Innovation Solution
Incorporating RNA aptamers into the mRNA structure, specifically embedded in RNA scaffolds like tRNA or bioorthogonal scaffolds, which allows for affinity purification using immobilized affinity ligands on chromatography resins, effectively removing impurities while maintaining translation efficiency and minimizing immunogenicity.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Manufacturing precision
If conventional purification methods are used for mRNA, then the manufacturing process is simple, but the purity of the final mRNA product is insufficient
Solution Approach 1:
The patent incorporates RNA aptamers into the mRNA structure during the synthesis stage, before purification is needed. This preliminary incorporation of purification tags enables subsequent affinity-based purification to be highly effective, resolving the contradiction between achieving high purity and maintaining process simplicity.
Solution Approach 2:
The patent uses RNA aptamers as intermediary elements that are incorporated into the mRNA and enable specific binding to affinity ligands during purification. These aptamers act as mediators that facilitate the separation of mRNA from impurities, achieving high purity without complex purification protocols.
2Reliability
If RNA aptamers are incorporated into mRNA structure, then affinity purification effectiveness is improved, but the mRNA structure becomes more complex
Solution Approach 1:
The patent segments the mRNA structure by incorporating distinct RNA aptamer sequences at specific locations (such as the 3' end or within the 3' UTR region). This segmentation allows the aptamers to function as independent purification tags while preserving the functional integrity of the main mRNA structure, thus improving purification reliability without excessive structural complexity.
Solution Approach 2:
The patent applies local quality by placing RNA aptamers at specific locations within the mRNA molecule (such as the 3' end or 3' UTR region) rather than uniformly throughout. This localized incorporation ensures that the aptamers provide purification functionality only where needed, maintaining the overall simplicity and functionality of the mRNA while enabling effective affinity purification.
3Manufacturing precision
If multiple purification steps are added to remove contaminants, then mRNA purity increases, but production time and cost increase
Solution Approach 1:
The patent incorporates RNA aptamers into the mRNA during synthesis as a preliminary action, enabling a single highly effective affinity purification step to replace multiple conventional purification steps. This preliminary tagging approach achieves high purity while maintaining production efficiency, as the aptamer-mediated purification is faster and more selective than traditional methods.
Solution Approach 2:
The patent replaces complex mechanical purification systems (multiple chromatography steps, centrifugation, filtration) with a simpler aptamer-ligand binding system. This substitution uses specific molecular recognition rather than complex physical separation methods, achieving high purity with fewer steps and better productivity.
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
The method achieves high-purity mRNA with greater than 90% purity, effectively removing contaminants and maintaining translation efficiency, making it suitable for therapeutic applications.
Implementation Method 1
contacting a sample comprising the mRNA with an affinity ligand that is immobilized on a chromatography resin, wherein the RNA aptamer comprises binding affinity for the affinity ligand
Data Source
AI summary
The present disclosure provides methods and compositions for RNA affinity purification. In particular, the disclosure relates to compositions and methods of making and using mRNA comprising one or more aptamers which specifically bind an affinity ligand.