Circular RNA Therapeutics for Stable Gene Expression
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Solution Overview
Problem
Conventional gene therapy using DNA can cause mutations, disrupt essential gene functions, and trigger immune responses due to integration into the host genome, and is costly and difficult to deliver effectively, while RNA-based therapies are safer but limited by existing methods for producing stable forms of RNA.
Innovation Solution
Development of circular RNA polynucleotides with specific structures, including post-splicing group I intron fragments, Internal Ribosome Entry Sites (IRES), and expression sequences, which improve expression, stability, and manufacturing ease, and reduce immunogenicity, using methods like the 2A self-cleaving peptide for separate protein expression.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Duration of action of stationary object
If DNA based gene therapy is used for delivery of secreted proteins, then long-lasting action is achieved, but integration into host genome may cause mutations and disrupt essential gene functions
Solution Approach 1:
The invention extracts the essential function of long-term gene expression while removing the harmful integration step by using circular RNA that remains cytoplasmic and does not integrate into the host genome, thus achieving sustained protein expression without genotoxicity
Solution Approach 2:
The patent employs circular RNA as a temporary, non-integrating therapeutic agent that provides sustained expression during its functional lifetime without permanent genomic alteration, allowing the body to clear it safely after therapeutic effect is achieved
2Ease of operation
If viral vectors are used for gene therapy delivery, then targeted delivery can be achieved, but adverse immune response and integration risks increase
Solution Approach 1:
The invention extracts the beneficial targeted delivery capability while removing the harmful viral components by using non-viral circular RNA constructs that can be delivered via non-viral methods, eliminating immunogenicity associated with viral vectors
Solution Approach 2:
The patent creates a functional copy of viral vector capabilities using synthetic circular RNA that mimics the sustained expression properties without using actual viral materials, thereby achieving similar therapeutic effects with reduced immunogenicity
3Object-affected harmful factors
If linear RNA is used for gene therapy, then safety from genome integration is achieved, but stability and half-life are limited
Solution Approach 1:
The invention applies the principle of circularization (curvature) to linear RNA to create circular RNA structures that resist exonuclease degradation, thereby dramatically extending half-life while maintaining the safety advantage of non-integration
4Ease of manufacture
If conventional RNA circularization methods are used, then circular RNA can be produced, but size limitations restrict therapeutic application
Solution Approach 1:
The patent employs ribozyme-based self-cleavage and ligation mechanisms that enable circularization of RNA molecules across a broad size range, overcoming the size limitations of previous methods and expanding therapeutic applicability
Data Source
AI summary
Circular RNA, along with related compositions and methods are described herein. In some embodiments, the inventive circular RNA comprises post splicing group I in iron fragments, spacers, an IRES, optional duplex forming regions, and more than one expression sequence. In some embodiments, the expression sequences are separated by one or more polynucleotide sequences encoding a cleavage site. In some embodiments, circular RNA of the invention has improved expression, functional stability, immunogenicity, ease of manufacturing, and/or half-life when compared to linear RNA. In some embodiments, inventive methods and constructs result in improved circularization efficiency, splicing efficiency, and/or purity when compared to existing RNA circularization approaches.