B6-hDMD(E8-30) Mice

Catalog Number: I001224

Strain Name: C57BL/6NCya-Dmdtm4(hDMD Exon 8-30)/Cya

Genetic Background: C57BL/6NCya

One of Cyagen's HUGO-GT™ (Humanized Genomic Ortholog for Gene Therapy) Strains

 

Strain Description

Duchenne Muscular Dystrophy (DMD) is a severe, progressive, and disabling X-linked recessive genetic disorder characterized primarily by muscle atrophy. This disease leads to motor impairments, eventually requiring assisted ventilation, and often results in premature death. The primary cause of DMD is mutations in the DMD gene, which encodes the dystrophin protein. These mutations lead to a reduction or absence of dystrophin in muscle tissue, resulting in muscle atrophy and related complications [1]. The lack of dystrophin leads to the breakdown of the dystrophin-associated protein complex (DAPC) within the muscle membrane, disrupting the interaction between actin and the extracellular matrix, making the muscles more susceptible to damage. This susceptibility results in the gradual loss of muscle tissue and function, potentially leading to cardiomyopathy [2]. Researchers have identified thousands of different DMD gene mutations in patients with DMD. Deletion mutations account for approximately 60%–70%, while duplication mutations account for 5%–15%. These mutations are primarily concentrated in hotspot regions of the DMD gene, specifically between exons 45-55 (47%) and exons 3-9 (7%) [1].

Currently, gene therapy approaches for Duchenne Muscular Dystrophy (DMD) primarily include exon skipping and AAV supplementation, as well as emerging gene editing techniques like CRISPR. The exon skipping strategy involves using antisense oligonucleotide (ASO) drugs to bind to specific sequences of pre-mRNA, skipping the mutated exon and restoring the open reading frame (ORF) integrity, thus producing a truncated but partially functional dystrophin protein. Several ASO drugs targeting the DMD gene have been approved, such as Eteplirsen (targeting exon 51), Golodirsen (targeting exon 53), and Casimersen (targeting exon 45) developed by Sarepta, and Viltolarsen (targeting exon 53) developed by Nippon Shinyaku. Since most ASO and CRISPR-based gene editing therapies target the human DMD gene, humanizing mouse genes helps accelerate clinical applications for DMD therapies, considering the genetic differences between animals and humans.

The B6-hDMD(E8-30) mouse is a humanized model of exons 8-30 of the Dmd gene, used for researching Duchenne Muscular Dystrophy. Homozygotes are viable and fertile. In addition, based on the independently developed TurboKnockout fusion BAC recombination technology, Cyagen provides other humanized models such as [hE49-53], [hE49-53, del E50], [hE44-45], [hE44-45, del E44], and [hE44-45, c.6438+2 T to A], covering most popular research areas and offering customized services based on different mutation needs.

Strain Strategy


Figure 1. Gene editing strategy of B6-hDMD(E8-30) mice. The genomic region spanning intron 7 (~5 kb) to intron 30 (~5 kb) of the mouse Dmd gene is replaced with the corresponding sequence from the human DMD gene.

Application

  • Research on the pathogenesis of Duchenne Muscular Dystrophy (DMD);
  • Preclinical efficacy evaluation of DMD therapeutic drugs.

 

Validation Data

1. Gene Expression Analysis

Figure 2. RT-qPCR analysis of DMD expression in B6-hDMD(E8-30) and wild-type (WT) mice. Results demonstrate significant expression of human DMD in the skeletal muscle, cerebral cortex, and heart of B6-hDMD(E8-30) mice, while WT mice lack DMD expression. Conversely, the mouse Dmd gene is expressed in WT mice but not in B6-hDMD(E8-30) mice. Data are presented as mean ± SEM, n≥3.

Note: Primers targeting exon 13–14 regions of the human DMD and mouse Dmd genes were used for detection.

2. Protein Expression Analysis

Figure 3. Western blot analysis of dystrophin protein in B6-hDMD(E8-30) and WT mice. Western blot results show normal dystrophin (DMD) expression in the skeletal muscle and heart of B6-hDMD(E8-30) mice, comparable to WT mice.

Note: The antibody used recognizes mouse dystrophin residues 3550–3678, corresponding to the exon 78–79 region, which is located downstream of the humanized sequence.

 

References
[1]Duan D, Goemans N, Takeda S, Mercuri E, Aartsma-Rus A. Duchenne muscular dystrophy. Nat Rev Dis Primers. 2021 Feb 18;7(1):13.
[2]Babbs A, Chatzopoulou M, Edwards B, Squire SE, Wilkinson IVL, Wynne GM, Russell AJ, Davies KE. From diagnosis to therapy in Duchenne muscular dystrophy. Biochem Soc Trans. 2020 Jun 30;48(3):813-821.