Circular RNA Constructs for Non-Integrating Gene Therapy
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Solution Overview
Problem
Conventional gene therapy using DNA can lead to integration into the host genome, causing mutations, disrupting essential gene function, and triggering immune responses, while viral vectors are costly and difficult to deliver effectively.
Innovation Solution
The use of circular RNA (oRNA) as a gene therapy agent, which does not integrate into the genome and requires no strong promoters, allowing for safer and more targeted protein expression.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Duration of action of stationary object
If DNA is used for gene therapy to achieve long-lasting action, then the duration of therapeutic effect is improved, but the risk of genomic integration and harmful mutations increases
Solution Approach 1:
The invention extracts the therapeutic gene expression function from DNA and implements it using RNA molecules. The circular RNA contains the coding sequence for the therapeutic protein but deliberately excludes integration capabilities into the host genome, thereby separating the beneficial long-lasting expression from the harmful integration risk.
Solution Approach 2:
The patent uses circular RNA as an intermediary molecule between DNA and protein expression. Instead of directly introducing DNA that would integrate into the genome, the circular RNA serves as a temporary mediator that translates the therapeutic gene into protein without permanent genomic alteration, thus mediating between long-term expression needs and safety concerns.
2Power
If strong promoter sequences are included in DNA-based gene therapy for effective gene expression, then the expression level of therapeutic protein is improved, but the risk of disrupting normal gene regulation increases
Solution Approach 1:
The invention extracts the gene expression function from the DNA-promoter system and implements it using circular RNA with internal ribosome entry sites (IRES). The IRES elements enable cap-independent translation initiation without requiring strong promoter sequences, thereby achieving effective protein expression while avoiding disruption of normal gene regulation.
Solution Approach 2:
The patent replaces the DNA-based promoter-driven transcriptional regulation system with an RNA-based IRES-mediated translational initiation system. This substitution changes the mechanism from promoter-dependent transcription to IRES-dependent translation, achieving effective expression without the regulatory disruptions associated with strong promoters.
3Productivity
If viral vectors are used for delivery of genetic material, then the efficiency of gene delivery is improved, but the cost and complexity of production increase
Solution Approach 1:
The invention uses circular RNA molecules that can be synthesized in vitro as copies of the therapeutic gene sequence. These circular RNA copies can be directly delivered to cells without requiring complex viral vector production processes, thereby maintaining delivery efficiency while dramatically simplifying production and reducing costs.
Solution Approach 2:
The patent employs circular RNA as a disposable, non-integrating therapeutic molecule that can be synthesized relatively simply and delivered directly. Unlike viral vectors that require complex production and purification, the circular RNA serves as a temporary but effective delivery vehicle that accomplishes its purpose without the associated complexity and cost.
4Ease of operation
If viral vectors are used for gene delivery, then the ability to deliver genetic material is improved, but the risk of inducing immune response increases
Solution Approach 1:
The invention extracts the gene delivery function from viral vectors and implements it using circular RNA molecules. By removing the viral components that trigger immune responses while retaining the ability to deliver and express therapeutic genes, the circular RNA achieves delivery capability without the harmful immune activation associated with viral vectors.
Solution Approach 2:
The patent converts the potential harm of viral vector-induced immune responses into a benefit by using circular RNA that deliberately avoids triggering immune pathways. The circular RNA structure and composition are designed to be immunologically inert while maintaining therapeutic efficacy, thus converting the problem of immune recognition into an advantage of reduced immunogenicity.
Data Source
AI summary
Circular RNA, along with related compositions and methods are described herein. In some embodiments, the inventive circular RNA comprises intron segments, spacers, an IRES, duplex forming regions, and an expression sequence. 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, the disclosed methods and constructs result in improved translation when compared to existing RNA approaches. In some embodiments, the disclosed methods and constructs result in improved circularization efficiency, splicing efficiency, and/or purity when compared to existing RNA circularization approaches.