RNA Booster Sequence for High Transient Vaccine Expression
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Solution Overview
Problem
Current RNA vectors, particularly RT-defective retroviral vectors, face challenges in achieving high transient expression levels in host cells without the need for high vector doses, due to low transfection rates and rapid degradation of RNA molecules, making them inefficient for in vivo applications.
Innovation Solution
Incorporation of an artificial 9-nucleotide RNA Booster sequence, such as mmsknkkkm or its reverse, upstream or downstream of a transgene, enhances transgene expression by improving stability and transfection efficiency in host cells.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Reliability
If RT-defective retroviral vectors are used for transient RNA expression, then transgene expression is achieved without DNA integration, but expression levels are low and high vector doses are required
Solution Approach 1:
The patent modifies the RNA vector by incorporating a specific 9-nucleotide sequence (5'-UURUUUURU-3' or reverse complement) at defined positions within the RNA genome. This sequence parameter change enhances cap-dependent translation initiation efficiency, thereby increasing transgene expression levels from the RNA vector without requiring integration into host DNA, thus maintaining safety while improving productivity
Solution Approach 2:
The patent introduces a specific 9-nucleotide sequence as an intermediary element that mediates between the RNA vector and the host cell's translation machinery. This sequence acts as a bridge to enhance cap-dependent translation initiation, allowing efficient protein expression from transient RNA vectors without requiring high vector doses or DNA integration
2Productivity
If high vector doses are administered to overcome low transfection efficiency, then transgene expression levels improve, but the complexity and cost of treatment increase
Solution Approach 1:
The patent changes the RNA sequence parameter by incorporating the 9-nucleotide motif (5'-UURUUUURU-3' or reverse complement) to enhance translation efficiency. This allows achieving high transgene expression levels with lower vector doses, thereby reducing treatment complexity and cost while maintaining effective expression
3Object-affected harmful factors
If RNA vectors are used instead of DNA vectors, then genotoxicity risk is eliminated, but RNA stability and transfection efficiency decrease
Solution Approach 1:
The patent introduces a specific 9-nucleotide sequence as a protective intermediary element within the RNA vector structure. This sequence enhances RNA stability and protects against degradation while maintaining the non-integrating nature of the vector, thus preserving the genotoxicity advantage of RNA vectors while improving their stability and transfection efficiency
Data Source
AI summary
A ribonucleic acid (RNA) or deoxyribonucleic acid (DNA) molecule, which includes an RNA Booster sequence and a sequence of interest encoding at least one antigen. Also, methods for robust transient RNA expression, in particular in the field of vaccination.