Liver-Specific Cis-Regulatory Elements for High-Expression Gene Therapy
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Solution Overview
Problem
Existing gene therapy methods face challenges in achieving liver-specific gene expression with minimal off-target effects, particularly when using vectors with limited capacity like AAV, as they often result in non-specific expression in tissues other than the liver.
Innovation Solution
Development of synthetic liver-specific cis-regulatory enhancer elements, such as V1 and V2, which can be combined with other regulatory sequences to enhance liver-specific gene expression, utilizing transcription factor binding sites in a specific order and orientation to drive high expression levels in the liver while minimizing off-target effects.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Productivity
If adenoviral vectors are used to target liver, then gene delivery efficiency is improved, but off-target infection of lung and skeletal muscle occurs
Solution Approach 1:
The patent applies local quality by designing liver-specific regulatory nucleic acid sequences that are active only in liver cells. These sequences include liver-specific promoters and enhancers that bind liver-enriched transcription factors, ensuring that gene expression occurs exclusively in the target organ (liver) while preventing off-target expression in other tissues like lung and skeletal muscle.
Solution Approach 2:
The patent employs parameter changes by modifying the regulatory elements of gene therapy vectors to achieve tissue-specific expression. By incorporating liver-specific promoters (e.g., albumin promoter, APOE promoter) and enhancer sequences, the system changes the expression parameters from constitutive to liver-specific, thereby maintaining high gene delivery efficiency while eliminating harmful off-target effects.
2Reliability
If liver-specific regulatory sequences are used to improve specificity, then off-target effects are reduced, but expression power may be compromised
Solution Approach 1:
The patent applies merging by combining multiple liver-specific regulatory elements into composite promoters and enhancers. By merging liver-specific promoters with powerful viral enhancers (e.g., CMV enhancer, RSV enhancer) or other liver-specific enhancer sequences, the system achieves both high liver-specificity and high expression levels, resolving the contradiction between specificity and expression power.
Solution Approach 2:
The patent uses composite materials by creating composite regulatory nucleic acid sequences that integrate different functional elements. These composite sequences include combinations of liver-specific promoter regions, enhancer elements, and transcription factor binding sites, which together provide both high liver-specificity and strong transcriptional activation, ensuring reliable and powerful liver-specific gene expression.
Data Source
AI summary
The present invention relates to regulatory nucleic acid sequences, in particular liver-specific cis-regulatory elements, cis-regulatory modules, promoters and other such nucleic acid sequences, that are capable of enhancing liver-specific expression of genes. The invention also relates to expression constructs, vectors and cells comprising such liver-specific regulatory nucleic acid sequences, and to methods of their use. The liver-specific regulatory nucleic acid sequences are of particular utility for gene therapy applications, but also find utility in other areas such as bioprocessing and biotechnology.