Circular RNA with Group I Intron Fragments for Sustained 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 approaches are safer but limited by existing methodologies for circularizing RNA.
Innovation Solution
Development of circular RNA polynucleotides comprising group I intron fragments, Internal Ribosome Entry Sites (IRES), and expression sequences, optimized for improved expression, stability, and reduced immunogenicity, using specific spacers and termini, and formulated in lipid nanoparticles for enhanced delivery.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Duration of action of stationary object
If DNA based gene therapy is used to achieve long-lasting action, then sustained expression of desired gene product is improved, but risk of genome integration and mutation increases
Solution Approach 1:
The patent segments the genetic material into two separate components: DNA that remains extrachromosomal (plasmid) and RNA that is transcribed and translated. This segmentation allows the DNA to serve as a stable template without integrating into the genome, while the RNA performs the therapeutic function temporarily, thus resolving the contradiction between sustained expression and genome integration risk.
Solution Approach 2:
The patent introduces RNA as an intermediary between the extrachromosomal DNA template and the host cell's protein synthesis machinery. The DNA serves as a stable, non-integrating template that continuously produces RNA, which then acts as the actual therapeutic agent. This intermediary approach enables long-lasting action through continuous RNA production without the risks of DNA genome integration.
2Reliability
If viral vectors are used for delivery, then targeting delivery is improved, but immune response and production cost increase
Solution Approach 1:
The patent replaces expensive, complex viral vectors with a simpler, cheaper non-viral delivery system using extrachromosomal DNA and RNA. The system uses transient, non-integrating DNA that is degraded after serving its purpose as a template, eliminating the need for expensive viral vector production while reducing immune response risks associated with viral components.
3Object-affected harmful factors
If linear RNA is used for gene therapy, then safety from genome integration is improved, but stability and half-life are reduced
Solution Approach 1:
The patent establishes a continuous production system where extrachromosomal DNA continuously transcribes RNA, maintaining therapeutic levels of RNA expression. The circular structure of the DNA plasmid enables continuous, high-level transcription without degradation, ensuring sustained RNA production and prolonged therapeutic effect while maintaining the safety advantages of non-integrating RNA-based therapy.
Data Source
AI summary
Circular RNA, along with related compositions and methods are described herein. In some embodiments, the inventive circular RNA comprises group I intron fragments, spacers, an IRES, duplex forming regions, and an expression sequence. In some embodiments, the expression sequence encodes an antigen. 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.