5′-Cap Compounds Stabilizing RNA Against Decapping Enzymes
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Solution Overview
Problem
The instability of RNA in vivo, particularly due to decapping enzymes, and recognition by proteins such as IFIT1, limits the effectiveness of RNA-based therapies by reducing protein expression.
Innovation Solution
The use of 5′-cap compounds with phosphorothioate, phosphoroselenoate, and boranophosphate moieties, along with modifications like 2′-O methylation, to stabilize RNA and enhance translation efficiency.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Reliability
If RNA is used for protein expression in therapy, then transient expression and non-transforming character are achieved, but RNA instability due to decapping enzymes reduces effectiveness
Solution Approach 1:
The patent modifies the chemical structure of the 5′-cap by introducing phosphorothioate, phosphoroselenoate, and boranophosphate moieties in the phosphate bridge, along with 2′-O methylation of the second nucleotide. These parameter changes in chemical composition and structure significantly increase resistance to decapping enzymes while maintaining translation efficiency, thereby resolving the contradiction between RNA stability and duration of action.
Solution Approach 2:
The invention creates a composite 5′-cap structure combining multiple modified components: phosphorothioate/phosphoroselenoate/boranophosphate moieties and 2′-O methylated nucleotides. This composite structure provides enhanced enzymatic stability while preserving biological function, effectively addressing the RNA instability problem without sacrificing expression duration.
2Reliability
If 5′-cap compounds with phosphorothioate, phosphoroselenoate, and boranophosphate moieties are used, then RNA stability is increased, but device complexity increases
Solution Approach 1:
The patent divides the 5′-cap structure into distinct functional segments: the phosphorothioate/phosphoroselenoate/boranophosphate moiety in the phosphate bridge provides stability, while the 2′-O methylated second nucleotide maintains translation efficiency. This segmentation allows each component to perform its specific function independently, managing complexity through functional modularity.
3Productivity
If standard 5′-cap structure is used, then RNA is recognized by translation machinery, but decapping enzymes and proteins like IFIT1 reduce protein expression
Solution Approach 1:
The patent applies preliminary anti-action by pre-modifying the 5′-cap structure with phosphorothioate, phosphoroselenoate, and boranophosphate moieties, along with 2′-O methylation, before RNA enters the cell. These pre-installed modifications create steric and electronic barriers that prevent decapping enzymes and IFIT1 proteins from recognizing and degrading the RNA, thereby protecting protein expression in advance.
Data Source
AI summary
The present invention relates to 5′-cap compounds, in particular the stabilization of RNA by such 5′-cap compounds, and provides compositions, such as pharmaceutical compositions, and cells comprising an RNA which is modified with such a 5′-cap compound, as well as methods for producing a peptide or protein of interest using the compositions or cells according to the present invention. Furthermore, the present invention provides the RNA, compositions, or cells for use in therapy, in particular for use in a method of treating a disease or disorder by protein replacement therapy, genome engineering, genetic reprogramming, and immunotherapy; a method for increasing the stability of RNA in cells; a method for increasing the expression of RNA in cells; and a method for providing an RNA with a 5′-cap structure.