Diaphragm-Specific Regulatory Elements for Targeted Gene Expression
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Solution Overview
Problem
Current gene therapy methods lack effective and efficient means to enhance tissue-specific gene expression in the diaphragm, particularly for treating diaphragm disorders, and often result in non-productive combinations with loss of tissue specificity and modest expression levels.
Innovation Solution
A computational approach identifies nucleic acid regulatory elements with high TFBS content, epigenetic signatures, and evolutionary conservation, enhancing diaphragm-specific gene expression, and combining these with tissue-specific elements for robust expression in diaphragm and other affected tissues.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Productivity
If conventional trial-and-error approaches are used to combine transcriptional enhancers with promoters, then expression levels may be modestly increased in some cases, but tissue specificity is lost and many combinations are non-productive
Solution Approach 1:
The patent applies preliminary action by systematically identifying and characterizing diaphragm-specific transcription factors (DIAPHRAGM-1 through DIAPHRAGM-6) and their binding sites before constructing expression vectors. This pre-characterization of regulatory elements ensures that subsequent vector designs are based on proven diaphragm-specific sequences, thereby maintaining tissue specificity while achieving high expression levels without relying on non-productive trial-and-error combinations
Solution Approach 2:
The patent employs composite materials by constructing expression cassettes that integrate multiple diaphragm-specific regulatory elements (enhancers, promoters, and transcription factor binding sites) into unified vector designs. These composite regulatory structures combine the identified diaphragm-specific transcription factor binding sites with appropriate promoters to create synergistic expression modules that maintain both high productivity and diaphragm tissue specificity
2Productivity
If high vector doses are used to achieve robust gene expression in the diaphragm, then therapeutic efficacy is improved, but safety concerns increase due to potential off-target effects
Solution Approach 1:
The patent applies local quality by designing expression vectors with diaphragm-specific regulatory elements that restrict transgene expression exclusively to diaphragm tissue. The use of diaphragm-specific transcription factor binding sites (DIAPHRAGM-1 through DIAPHRAGM-6) and associated promoters ensures that the therapeutic gene is expressed only in the target tissue (diaphragm) and not in other organs, thereby enabling effective therapy at lower vector doses without causing off-target harmful effects
Solution Approach 2:
The patent uses diaphragm-specific transcription factors and their binding sites as intermediaries to mediate between the vector delivery system and the therapeutic gene expression. These specific regulatory elements act as tissue-specific switches that allow the vector to deliver the therapeutic gene systemically while ensuring expression occurs only in diaphragm tissue where the specific transcription factors are present, thus reducing off-target effects
Data Source
AI summary
The present invention relates to nucleic acid regulatory elements that are able to enhance diaphragm-specific expression of genes, in particular expression in diaphragm as such, or in combination with expression in cardiac muscle and/or skeletal muscle, methods employing these regulatory elements and uses of these elements. Expression cassettes and vectors containing these nucleic acid regulatory elements are also disclosed. The present invention is particularly useful for applications using gene therapy, more particularly diaphragm-directed gene therapy, and for vaccination purposes.