RNAi Molecules with 5'-Triphosphates for Interferon Induction
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Solution Overview
Problem
Current RNAi-mediated gene silencing methods often induce interferon expression, leading to non-specific changes in gene expression and potential anti-viral and anti-cancer effects are not fully utilized, with the role of triphosphates in RNA molecules for inducing interferon and anti-viral responses remaining unexplored.
Innovation Solution
Introducing RNA molecules, specifically siRNAs and ssRNAs, with maintained or removed 5'-triphosphates to exploit interferon induction properties for anti-viral and anti-cancer effects, using in vitro transcription by phage polymerases like T7 RNA polymerase to produce RNAi molecules with 5'-triphosphates, which induce interferon α and β and elicit strong antiviral responses.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Manufacturing precision
If RNAi molecules are used for gene silencing, then post-transcriptional gene silencing is achieved, but interferon induction occurs causing non-specific changes in gene expression
Solution Approach 1:
The patent extracts and removes the 5'-triphosphate group from RNAi molecules to eliminate the harmful interferon induction effect while preserving the desired gene silencing function. This is achieved through enzymatic treatment with phosphatases or chemical modification methods that specifically remove the triphosphate moiety without affecting the RNAi activity of the molecule.
Solution Approach 2:
The patent modifies the chemical parameter of the RNA 5' end by changing the phosphate group configuration. By converting the 5'-triphosphate to 5'-monophosphate or 5'-hydroxyl forms, the molecule's ability to induce interferon is altered while maintaining its gene silencing capability through the RNA interference pathway.
2Reliability
If 5'-triphosphate RNA molecules are used to induce interferon, then strong antiviral responses are elicited, but the mechanism for controlled interferon induction is not established
Solution Approach 1:
The patent creates a dynamic system where the 5' end modification state of RNA molecules can be adjusted to control interferon induction. By providing methods to reversibly modify the 5' end (between triphosphate, monophosphate, and hydroxyl forms), the system allows flexible control over the degree of interferon response based on therapeutic needs.
Solution Approach 2:
The patent utilizes changes in the phosphate group parameter at the 5' end of RNA molecules to modulate interferon induction. The presence of triphosphate groups serves as a reliable indicator for strong interferon induction, while removal to monophosphate or hydroxyl forms provides controlled or reduced induction, enabling titration of the antiviral response.
Applied Scientific Principles
This section explains which scientific principles are used to turn an abstract innovation direction into a practical engineering solution.
Function Achieved in This Case
The approach effectively induces interferon and anti-viral responses, providing a potent means to prevent viral infections and potentially treat cancer, while allowing for controlled reduction of interferon induction by removing triphosphates from RNAi molecules.
Implementation Method 1
These siRNAs are made by in vitro transcription mediated by bacteriophage promoters from linearized DNA templates. In vitro transcription using bacteriophage T7 RNA polymerase has been shown to produce highly active siRNAs
Implementation Method 2
The presence of an initiating triphosphate on in vitro transcribed-RNAs can potently induce interferon α and β, as well as elicit a strong, non-sequence-specific antiviral response to viral challenge
Implementation Method 3
siRNAs are double-stranded molecules typically 21 to 25 nucleotides (nt) in length, which trigger RNA interference (RNAi), resulting in post-transcriptional message degradation
Implementation Method 4
Long double-stranded (ds) RNAs are processed into siRNAs by dicer, a ribonuclease of the Rnase III family
Data Source
AI summary
Double-stranded and single-stranded RNA molecules, and their use in methods for inducing interferon are provided. The interferon induction provides anti-viral and other medically useful effects, such as anti-cancer effects. Also provided are methods for reducing or inhibiting interferon induction exhibited by such molecules, particularly siRNA and shRNA molecules produced in vitro.