CRISPR-Edited Exon 44 Deletion Mouse Model for DMD
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Solution Overview
Problem
Current models for Duchenne muscular dystrophy (DMD) lack representation of various known and new mutations, limiting the availability of effective animal models for therapeutic testing and validation.
Innovation Solution
Creation of a mouse model with a deletion of exon 44 in the dystrophin gene, mimicking the second most common DMD mutation, using CRISPR/Cas9-mediated genome editing, which includes a reporter gene and protease sequence for enhanced expression and therapeutic assessment.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Adaptability or versatility
If existing mouse models are used for DMD research, then research can proceed with available models, but the models lack representation of various known and new DMD mutations limiting therapeutic testing
Solution Approach 1:
The invention creates separate mouse models for different DMD mutation types (exon 44 deletion model for out-of-frame mutations, and exon 43 deletion model for in-frame mutations). This segmentation allows each model to specifically represent particular mutation categories, enabling more accurate and targeted therapeutic testing for different patient populations with distinct mutation types.
2Adaptability or versatility
If a mouse model with exon 44 deletion is created to represent the second most common DMD mutation, then the model will represent ~12% of DMD patients, but this requires developing new animal models rather than using existing ones
Solution Approach 1:
The invention uses CRISPR/Cas9 genome editing technology as an intermediary tool to create the exon 44 deletion mouse model. This molecular editing system enables precise deletion of specific exons in the dystrophin gene, facilitating the creation of accurate DMD mutation models without requiring complex traditional breeding programs or extensive model development procedures.
3Measurement precision
If diverse DMD mutation models are developed, then therapeutic assessment can be optimized, but the complexity of maintaining multiple models increases
Solution Approach 1:
The invention creates mouse models with modular design features including loxP sites and reporter genes that can serve multiple functions. These models can be used for assessing different therapeutic approaches (exon skipping, read-through, gene replacement), for monitoring dystrophin expression levels, and for evaluating therapeutic efficacy across various treatment modalities, reducing the need for entirely separate model systems.
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
Provides a valuable system for assessing therapeutic efficacy and optimizing exon-skipping approaches, allowing for rapid testing of candidate substances and modalities in a 'humanized' model, thereby addressing the lack of diverse DMD mutation representation in existing models.
Implementation Method 1
using CRISPR/Cas9-mediated genome editing
Data Source
AI summary
Duchenne muscular dystrophy (DMD), which affects 1 in 5,000 male births, is one of the most common genetic disorders of children. This disease is caused by an absence or deficiency of dystrophin protein in striated muscle. The major DMD deletion “hot spots” are found between exon 6 to 8, and exons 45 to 53. Here, a “humanized” mouse model is provided that can be used to test a variety of DMD exon skipping strategies. Among these are, CRISPR/Cas9 oligonucleotides, small molecules or other therapeutic modalities that promote exon skipping or micro dystrophin mini genes or cell based therapies. Methods for restoring the reading frame of exon 44 deletion via CRISPR-mediated exon skipping in the humanized mouse model, in patient-derived iPS cells and ultimately, in patients using various delivery systems are also contemplated. The impact of CRISPR technology on DMD is that gene editing can permanently correct mutations.