Photodegradable Crosslinking Agent for dsRNA Gene Expression Control

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Solution Overview

Problem

Current methods for regulating gene expression using RNAi struggle to effectively suppress the expression of target genes by crosslinking double-stranded RNA, as existing caged compounds fail to efficiently control gene expression when bound to siRNA.

Innovation Solution

A crosslinking agent with photodegradable protective groups at two ends is used to crosslink double-stranded RNA, allowing for precise control of gene expression by binding and irradiating the RNA with ultraviolet light.

Engineering Contradictions & Design Principles

VSEngineering Contradiction Analysis

1Reliability

If caged compounds are bound to single-stranded mRNA to regulate gene expression, then the expression can be conditionally controlled by light irradiation, but the method cannot effectively suppress RNAi effect by crosslinking double-stranded RNA

Engineering Contradiction:

Improvegene expression regulation effectivenessVSAvoidapplicability to double-stranded RNA

Core Design Contradiction:

ReliabilityVSAdaptability or versatility

Solution Approach 1:

The crosslinking agent is divided into two separate photodegradable protective groups (Q1 and Q2) that can independently bind to each strand of the double-stranded RNA. This segmentation allows each protective group to function as a caged compound while working cooperatively to crosslink both strands, thereby achieving effective RNAi suppression that single-stranded binding cannot accomplish.

Inventive Principle:

Principle #1Segmentation

Solution Approach 2:

The invention merges the functionality of two caged compounds into a single crosslinking agent structure where Q1 and Q2 are connected through a linker (A1-T1-A2). This combining enables simultaneous binding to both strands of dsRNA and formation of interstrand crosslinks, achieving the dual function of light-controlled release and effective RNAi suppression.

Inventive Principle:

Principle #5Merging (Combining)

2Ease of operation

If crosslinking agent with photodegradable protective groups is used to crosslink double-stranded RNA, then gene expression can be regulated at arbitrary timing and location, but the complexity of the crosslinking agent structure increases

Engineering Contradiction:

Improvetiming and location controlVSAvoidcrosslinking agent structure

Core Design Contradiction:

Ease of operationVSDevice complexity

Solution Approach 1:

The linker structure (A1-T1-A2) serves as an intermediary component that connects the two photodegradable protective groups Q1 and Q2. This intermediary maintains the appropriate spatial distance and orientation between the two binding sites, enabling effective crosslinking of dsRNA while keeping the overall molecular structure manageable and synthetically accessible.

Inventive Principle:

Principle #24Intermediary (Mediator)

3Device complexity

If conventional caged compounds are used to bind siRNA, then the structure is simple, but the RNAi effect cannot be effectively suppressed

Engineering Contradiction:

Improvecompound structureVSAvoidRNAi suppression efficiency

Core Design Contradiction:

Device complexityVSReliability

Solution Approach 1:

The crosslinking agent represents a composite molecular structure combining two photodegradable protective groups (Q1 and Q2) with a linker scaffold (A1-T1-A2). This composite design integrates the light-responsive properties of caged compounds with the crosslinking functionality needed for effective dsRNA binding, achieving both structural feasibility and high RNAi suppression efficiency.

Inventive Principle:

Principle #40Composite materials

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

This approach enables efficient and timely regulation of gene expression, allowing for the examination and identification of genes expressed at specific times and locations.

Implementation Method 1

Q1 and Q2 each independently represent a photodegradable protective group

Methodology Applied

Scientific EffectPhotodegradation: Photodissociation

Data Source

PatentUS8084625B2Crosslinking agent, crosslinking method, method of controlling gene expression, and method of examining gene function

Publication Date: 2011.12.27 FUJIFILM WAKO PURE CHEMICAL CORP

AI summary

The present invention provides a crosslinking agent which have photodegradable protective groups at two ends to crosslink double-stranded nucleic acid, a nucleic acid and a protein or a polypeptide, or proteins or polypeptides, in particular, double-stranded RNA; a method for crosslinking a double-stranded RNA or the like using the same; a method for regulating gene expression, which can control the expression of a target gene at an arbitrary timing and location; and a method for examining a gene function.According to the present invention, crosslinking between double-stranded nucleic acids between a nucleic acid and a protein or a polypeptide, or between proteins or polypeptides, in particular, between double-stranded RNA can be easily formed, and in addition, the crosslinking can also be easily removed, so that the expression of a target gene can be easily controlled at an arbitrary timing and location with high efficiency. Hence, as a result, function examination and/or identification of a gene that is expressed at a specific timing and location can be performed. In addition, the RNAi effect of a double-stranded RNA (siRNA) that cannot be easily inhibited by a conventional caged compound can be inhibited, and the expression of a target gene can be easily controlled at an arbitrary timing and location.