RNA Expression Stability via Interferon Signaling Inhibition
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Solution Overview
Problem
Repetitive RNA-based gene transfer for expressing reprogramming transcription factors is hindered by the induction of the interferon response, leading to instability and cytotoxicity, which limits the continuous expression necessary for somatic cell reprogramming into induced pluripotent stem cells.
Innovation Solution
Preventing engagement of the interferon receptor by extracellular interferon and inhibiting intracellular interferon signaling through the use of viral binding agents and inhibitors such as vaccinia virus B18R, E3, and K3, which are introduced into cells as nucleic acids, thereby stabilizing RNA expression and enhancing cell viability.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Duration of action of stationary object
If repetitive RNA-based gene transfer is performed to achieve continuous expression of reprogramming transcription factors, then the duration and stability of RNA expression is improved, but the interferon response is induced leading to cytotoxicity and reduced cell viability
Solution Approach 1:
The patent applies preliminary anti-action by introducing interferon response inhibitors (such as viral proteins or small molecules) before or concurrent with RNA transfection to preemptively block the harmful interferon signaling pathway. This prevents the cytotoxic effects while allowing the RNA to maintain stable expression over multiple transfection cycles, directly resolving the contradiction between prolonged expression duration and cellular toxicity
Solution Approach 2:
The patent uses interferon response inhibitors as intermediary substances that mediate between the exogenous RNA and the cell's immune defense system. These inhibitors act as protective intermediaries that block the activation of interferon signaling pathways, thereby preventing cytotoxicity while permitting sustained RNA expression. The intermediary compounds enable the system to tolerate repetitive transfections without triggering harmful immune responses
2Productivity
If exogenous single-stranded RNA is introduced into cells for gene transfer, then transient protein expression is achieved, but cellular defense mechanisms are activated leading to RNA degradation and reduced expression stability
Solution Approach 1:
The patent converts the harmful cellular defense response into a beneficial situation by using interferon response inhibitors to block the degradation pathways. Instead of allowing the natural immune response to degrade the RNA, the inhibitors transform the scenario by selectively blocking only the harmful degradation mechanisms while permitting productive protein expression to proceed, thereby simultaneously improving both expression levels and RNA stability
Solution Approach 2:
The patent applies parameter changes by modifying the cellular environment through the introduction of interferon response inhibitors. These inhibitors alter the biochemical parameters of the cellular context, specifically changing the activity state of interferon signaling pathways and associated degradation enzymes. This parameter modification creates a more favorable environment for RNA stability and sustained protein production
Data Source
AI summary
The present invention relates to expressing RNA in cells and, in particular, enhancing viability of cells in which RNA is to be expressed. Specifically, the present invention provides methods for expressing RNA in cells comprising the steps of preventing engagement of IFN receptor by extracellular IFN and inhibiting intracellular IFN signalling in the cells. Thus, preventing engagement of IFN receptor by extracellular IFN and inhibiting intracellular IFN signalling in the cells allows repetitive transfer of RNA into the cells.