Circular RNA Vector Architecture for Prolonged Eukaryotic Translation
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Solution Overview
Problem
The short half-life of messenger RNA (mRNA) in biological systems limits its therapeutic and engineering applications, necessitating a solution to extend protein expression duration.
Innovation Solution
A vector is designed to produce circular RNA (circRNA) by arranging specific elements such as 5' and 3' homology arms, group I intron fragments, and internal ribosome entry sites (IRES) to enable translation and biological activity within eukaryotic cells, allowing for the production of biologically active miRNA sponges or long non-coding RNAs.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Duration of action of moving object
If messenger RNA is used for protein expression, then translation and protein production are achieved, but the half-life is short and duration is limited
Solution Approach 1:
The patent applies circularization of the RNA molecule to resolve the contradiction between mRNA stability and protein expression duration. By converting linear mRNA into a circular RNA structure, the molecule gains resistance to exonuclease degradation while maintaining translation capability through IRES elements. This curvature transformation eliminates the vulnerable 5' and 3' ends of linear RNA, thereby extending half-life and duration of protein expression.
Solution Approach 2:
The patent changes the structural parameter of RNA from linear to circular configuration, and incorporates specific sequence elements (IRES, homology arms) that alter the biochemical properties of the molecule. These parameter changes enable the RNA to resist degradation while maintaining translational activity, thus improving both stability and duration of protein expression.
2Reliability
If circular RNA is produced with homology arms and intron fragments, then stability and translation efficiency are enhanced, but vector design complexity increases
Solution Approach 1:
The patent segments the circular RNA precursor into distinct functional modules: 5' homology arm, 3' homology arm, group I intron fragments, IRES element, and coding sequence. Each segment serves a specific function in circularization, stability, or translation. This modular segmentation simplifies the design process by allowing independent optimization of each component while maintaining overall system functionality.
Solution Approach 2:
The patent incorporates group I intron fragments and homology arms in advance within the linear precursor sequence before circularization occurs. These elements are pre-positioned to facilitate self-circularization through splice-site recognition and homology-directed recombination, eliminating the need for complex post-transcriptional processing or additional enzymatic steps.
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 vector enhances the stability and translation efficiency of circRNA, enabling prolonged protein expression and activity in eukaryotic cells, facilitating therapeutic and diagnostic applications.
Implementation Method 1
a 3′ group I intron fragment containing a 3′ splice site dinucleotide, c.) a 5′ spacer sequence, d.) an internal ribosome entry site (IRES), e.) a protein coding or noncoding region, f.) a 5′ group I intron fragment containing a 5′ splice site dinucleotide
Implementation Method 2
an internal ribosome entry site (IRES), e.) a protein coding or noncoding region
Data Source
AI summary
Circular RNA and methods and constructs for engineering circular RNA are disclosed. In some embodiments, the circular RNA includes the following elements arranged in the following sequence: a) a 3′ Group I self-splicing intron fragment, b) an internal ribosome entry site (IRES), c) a protein coding region or noncoding region, and d) a 5′ Group I self-splicing intron fragment.