Enter the 'Ten Deadly Sins of Rare Diseases' column to unlock the mechanisms of rare disease occurrence and development, industry research advancements (such as gene therapy), and innovative strategies for advancing clinical outcomes (including model construction and drug screening). For previous reviews, click here:

1. Exploring RHO-Related Pathogenic Mechanisms and RHO Gene Therapy Research Progress [Ten Deadly Sins of Rare DiseasesⅠ]

2. Why are humanized mice more suitable for hemophilia research, which has a higher prevalence in men? [Ten Deadly Sins of Rare Diseases Ⅱ]

3. The TARDBP gene is closely associated with pathogenesis of amyotrophic lateral sclerosis (ALS), but is it an angel or a demon? [Ten Deadly Sins of Rare Diseases Ⅲ]

4. Reviewing Muscular Dystrophy: Types, Pathology, and Gene Therapy Research [Ten Deadly Sins of Rare Diseases Ⅳ]

5. Is Smn2 Gene Therapy the Key to Treating Spinal Muscular Atrophy? [Ten Deadly Sins of Rare Diseases Ⅴ]

In this issue, we focus on a category of diseases caused by repetitive DNA sequences, al
so known as repeat expansion diseases. Repetitive DNA sequences are quite common in the human genome, especially within genes and their regulatory regions. Repeated sequences can also lead to differences at the mRNA and protein levels through processes like transcription and translation. Although these repetitive elements have been considered an evolutionary mechanism, they may also be associated with human genetic diseases and cancers [1].

There are several diseases caused by DNA repeat expansions, but our focus will be on exploring preclinical research applications for two specific conditions. Spinocerebellar ataxia type 3 (SCA3) is linked to CAG repeat expansions in the ATXN3 gene, while Fuchs endothelial corneal dystrophy is caused by CTG repeat expansions in the TCF4 gene. In the following sections, we will examine the pathogenic mechanisms and disease models to understand these types of diseases and their corresponding preclinical research.

CAG repeat expansions in ATXN3 Gene Lead to Spinocerebellar Ataxia Type 3 (SCA3)

Spinocerebellar ataxias (SCAs) are a group of hereditary diseases characterized by chronic progressive ataxia. To date, dozens of causative genes have been identified for SCAs, and many of them are inherited in an autosomal dominant manner. In China, one of the more common SCA subtypes is Spinocerebellar Ataxia Type 3, also known as Machado-Joseph Disease (MJD).

The causative gene for this disease is ATXN3, which encodes the ataxin-3 protein. The (CAG)n repeat sequence in exon 10 of the ATXN3 gene can undergo abnormal expansion mutations. This results in the abnormal aggregation of the polyglutamine product within neural tissues, leading to damage to nerve cells in the brain and the neural fibers that transmit information. In normal individuals, the CAG repeat number ranges from 10 to 44, while in SCA3 patients, it can reach as high as 61 to 87 repeat units. Typically, the severity of the disease is associated with the number of CAG repeats, with more repeats leading to more severe disease and an earlier age of  symptom onset [2].

In the field of preclinical research for SCA3, one of the more common disease models is the MJD84.2 transgenic mouse (SCA3-YAC-84Q) [3]. The SCA3-YAC-84Q transgenic mice harbor a YAC transgene that expresses a human ATXN3 modified with a human MJD/SCA3-associated 84 CAG repeat expansion. This transgenic  model has provided a basis for studying the progressive neurodegenerative processes underlying SCA3 pathogenesis and other polyglutamine and trinucleotide repeat disorders. To improve the success rate of drug development, Cyagen has independently developed a whole-genome humanized ATXN3 (hATXN3) mouse and a similar SCA3-YAC-84Q transgenic mouse model to evaluate the disease modeling improvements provided by a fully humanized genomic ortholog (HUGO) mouse model.

The CTG repeat expansions in the TCF4 gene lead to Fuchs endothelial corneal dystrophy

The transcription factor 4 (TCF4) encoded by the TCF4 gene is a basic helix-loop-helix transcription factor. TCF4, as a 'transcription factor' gene, regulates the activity of at least hundreds of other genes. If disrupted from the beginning of development, it can lead to various developmental abnormalities. The TCF4 gene is an important causative gene for conditions such as Pitt-Hopkins syndrome (with R580W as a mutation hotspot), Fuchs endothelial corneal dystrophy, autism, and schizophrenia [4].

In the general population, the CTG repeat sequence within intron 2 of the TCF4 gene typically contains 10-37 repeat units. However, when the CTG repeat sequence expands to 50 repeat units, it can lead to the development of Fuchs endothelial corneal dystrophy.

Currently, there is no commercially available humanized mouse model that simulates the CTG repeat mechanism in the TCF4 gene that leads to diseases. To advance the treatment of TCF4 gene-related disorders, such as through ASO and siRNA-based gene therapies, Cyagen has been independently developing fully humanized genomic ortholog (HUGO) mouse models including hTCF4 whole-genome humanized mice, hTCF4-n*(CTG), and hTCF4(R580W), among others.

Other DNA Repeat Expansion Disease-Associated Models

In addition to the models related to the ATXN3 and TCF4 genes, Cyagen has also independently developed multiple animal models associated with other repeat sequence-induced diseases. These models encompass several DNA repeat diseases, including myotonic dystrophy (caused by DMPK gene CTG repeats), Friedreich's ataxia (caused by FXN gene GAA repeats), SCA1 (caused by ATXN1 gene CAG repeats), Huntington's disease (caused by HTT gene CAG repeats), and more.

Gene Disease Repeat Sequences
ATXN3 Spinocerebellar Ataxia Type 3 CAG
TCF4 Fuchs Endothelial Corneal Dystrophy CTG
DMPK Myotonic Dystrophy CTG
FXN Friedreich's Ataxia GAA
ATXN1 Spinocerebellar Ataxia Type 1 CAG
HTT Huntington's Disease CAG

Next-Generation Humanized Mouse Models for Preclinical Gene Therapy Research: HUGO-GTTM

Conducting thorough research on the pathogenic mechanisms of diseases like Spinocerebellar Ataxia Type 3, Retinitis Pigmentosa (RP), Age-Related Macular Degeneration (AMD), Parkinson's Disease (PD), and others necessitates the use of humanized mice with extended genomic fragments. For even more accurate modeling of human pathology, mice with whole-genome humanization are preferred. However, the technology required for whole-genome replacement is challenging, and the large-scale introduction of exogenous sequences may potentially affect the normal expression and regulation of native genes.

To overcome these obstacles, Cyagen has launched the groundbreaking Humanized Genomic Ortholog for Gene Therapy (HUGO-GTTM) Program initiative - a remarkable leap in Next-Generation Humanized Mouse Model Development. Powered by our innovative TurboKnockout-Pro technology, this program enables the seamless in situ replacement of mouse genes. This breakthrough paves the way for the creation of fully humanized mice carrying entirely human genomic DNA segments, opening a realm of possibilities for diverse intervention targets and preclinical gene therapy research

➢Validated Next-Generation Humanized (HUGO-GT) Mouse Models for Gene Therapy Research

Product Number Product Name Strain Background Application
C001396 B6J-hRHO C57BL/6J Retinitis Pigmentosa (RP), Congenital Stationary Night Blindness (CSNB), and other retinal diseases.
C001410 B6-htau C57BL/6J Frontotemporal Dementia (FTD), Alzheimer's Disease (AD), and other neurodegenerative diseases.
C001418 B6-hTARDBP C57BL/6J Amyotrophic Lateral Sclerosis (ALS), Frontotemporal Dementia (FTD), and other neurodegenerative diseases.
C001427 B6-hSNCA C57BL/6N Parkinson's Disease.
C001428 B6-hCOL7A1 C57BL/6N Epidermolysis Bullosa.
C001437 B6-hIGHMBP2 C57BL/6N Spinal Muscular Atrophy with Respiratory Distress Type 1 (SMARD1) and Charcot-Marie-Tooth Disease Type 2S (CMT2S).

➢Upcoming Next-Generation Humanized (HUGO-GT) Mouse Models

Disease Type Disease Name Target Gene Target Type
Ophthalmology Leber Congenital Amaurosis Type 10 (LCA) CEP290 Humanization(WT, Mut)
Age-Related Macular Degeneration (AMD) VEGFA Humanization
ABCA4 Humanization(WT, Mut)
Neurology Amyotrophic Lateral Sclerosis (ALS) SOD1 Humanization
FUS Humanization(WT, Mut)
Rett Syndrome (RTT) MECP2 Humanization(WT, Mut)
Spinocerebellar Ataxias (SCAs) ATXN3 Humanization(WT, Mut), TG
Familial Dysautonomia (FD) ELP1 Humanization(WT, Mut)
Muscle Duchenne Muscular Dystrophy (DMD) DMD Humanization(WT, Mut, KO)
Spinal Muscular Atrophy (SMA) SMN1 Humanization
Metabolism Atherosclerosis (AS) APOE2 Humanization
APOE3 Humanization
APOE4 Humanization
Blood Hemophilia A (HA) F8 Humanization(WT, Mut)
Others Transthyretin Amyloidosis (ATTR) TTR Humanization(WT, Mut), TG



[1]Erwin GS, Gürsoy G, Al-Abri R ,et al.Recurrent repeat expansions in human cancer genomes[J]. Nature. 2023 Jan;613(7942):96-102. doi: 10.1038/s41586-022-05515-1. 

[2]Corral-Juan, M., et al., Clinical, genetic and neuropathological characterization of spinocerebellar ataxia type 37. Brain, 2018. 141(7): p. 1981-1997.

[3]Cemal C K , Carroll C J , Lorraine L ,et al.YAC transgenic mice carrying pathological alleles of the MJD1 locus exhibit a mild and slowly progressive cerebellar deficit[J].Human Molecular Genetics, 2002(9):1075-1094.DOI:10.1093/hmg/11.9.1075.

[4]Thaxton,Courtney,Kloth,et al.Common Pathophysiology in Multiple Mouse Models of Pitt-Hopkins Syndrome[J].Journal of Neuroscience the Official Journal of the Society for Neuroscience, 2018.