B6-hTGFBI Mice

Catalog Number: C001546

Strain Name: C57BL/6JCya-Tgfbi^tm1(hTGFBI)/Cya

Genetic Background: C57BL/6JCya

Reproduction: Homozygote x Homozygote

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

Strain Description

Corneal dystrophy (CD) refers to a group of primary hereditary progressive corneal diseases. The typical clinical presentation involves gradual loss of corneal transparency in both eyes, often leading to recurrent corneal erosions and visual impairment. The TGFBI gene (also known as BIGH3) encodes an extracellular matrix protein called keratoepithelin (KE protein), which plays a role in cell growth, differentiation, wound healing, cell adhesion, migration, apoptosis, proliferation, and tumorigenesis ^[1]. Mutations in the TGFBI gene are associated with various types of corneal dystrophy. Abnormal accumulation of mutated TGFBI deposits in the corneal epithelium and stroma progressively affects corneal transparency, leading to visual impairment.

Currently, therapeutic pipelines targeting the TGFBI gene have entered preclinical research stages. For instance, SiSaf Ltd. is developing a siRNA drug pipeline called SIS-201-CD, which aims to treat the disease by specifically inhibiting abnormal TGFBI expression. Most gene therapies target human genes, but considering the genetic differences between animals and humans, humanizing mouse genes can accelerate the development of TGFBI-targeted gene therapies for clinical use. This strain is a mouse Tgfbi gene humanized model and can be used for research on CD. The homozygous B6-hTGFBI mice are viable and fertile. In addition, based on the independently developed TurboKnockout fusion BAC recombination technology, Cyagen can also generate hot mutation models based on this strain and provide customized services for specific mutations to meet the experimental needs in pharmacology and other fields related to CD.

Strain Strategy

Figure 1. Gene editing strategy of B6-hTGFBI mice. The sequences from aa.24 to TAG stop codon of mouse Tgfbi were replaced with the sequences from aa.24 to TAG stop codon of human TGFBI. The murine signal peptide (aa.1~23) was kept.

Application

• Research on corneal dystrophy (CD);
• Preclinical evaluation of TGFBI-targeted drugs.
  
  

Validation Data

1. Expression of human TGFBI gene and mouse Tgfbi gene

Figure 2. RT-qPCR detection of human TGFBI gene and mouse Tgfbi gene expression in the liver and eyes of 6-week-old male B6-hTGFBI mice and wild-type (WT) mice. The results indicate that B6-hTGFBI mice significantly express the human TGFBI gene in both the liver and eyes, while not expressing the mouse Tgfbi gene. In contrast, WT mice only exhibit expression of the mouse Tgfbi gene and do not show expression of the human TGFBI gene.

(ND: Not detected)

2. In vivo ocular phenotyping

a. Fundus, retina, cornea, and anterior chamber testing

Figure 3. Fundus morphology, retinal OCT, ocular surface, and corneal OCT results of wild-type and B6J-hTGFBI mice. The fundus morphology, retinal OCT, ocular surface, and corneal OCT results of heterozygous and homozygous B6J-hTGFBI  mice were consistent with those of wild-type.

b. Electroretinogram (ERG) testing

Figure 4. Electroretinogram (ERG) detection results of WT and B6J-hTGFBI mice. Compared with WT, the amplitudes of the a-wave and b-wave in both scotopic and photopic ERG recordings of heterozygous and homozygous B6J-hTGFBI mice were nearly identical to those of the WT. The retinal photoreceptor function of heterozygous and homozygous B6J-hTGFBI mice were normal.

Expanded Information: The Rare Disease Data Center (RDDC)

1. Basic information about the TGFBI gene

https://rddc.tsinghua-gd.org/gene/7045

2. TGFBI clinical variants

3. Disease introduction

Corneal dystrophy (CD) is a group of hereditary, non-inflammatory corneal diseases. Clinically, it is characterized by bilateral, slowly progressive loss of corneal transparency, often leading to recurrent corneal erosions and visual impairment. Mutations in the TGFBI gene are associated with various types of corneal dystrophy. The five most common CD types caused by TGFBI gene variants are Reis-Bucklers corneal dystrophy (RBCD), Thiel-Behnke corneal dystrophy (TBCD), granular corneal dystrophy type 1 (GCD1), granular corneal dystrophy type 2 (GCD2), and lattice corneal dystrophy type 1 (LCD1).

4. TGFBI gene and mutations

The TGFBI gene (also known as BIGH3) encodes an extracellular matrix protein called keratoepithelin (KE protein), which plays a role in cell growth, differentiation, wound healing, cell adhesion, migration, apoptosis, proliferation, and tumorigenesis ^[1]. Mutations in the TGFBI gene are associated with various types of corneal dystrophy. Abnormal accumulation of mutated TGFBI deposits in the corneal epithelium and stroma progressively affects corneal transparency, leading to visual impairment. 

The mutations in the TGFBI gene primarily include missense, nonsense, and frameshift mutations. Point mutations account for approximately 85%, while deletion mutations make up about 12%. The most common mutation types are R124 and R555, collectively representing 83%. R124C, R124H, and R124L located in exon 4, and R555Q and R555W located in exon 12 are the five most common mutations. Among these, R124H variation is responsible for granular corneal dystrophy type 2 (also known as Avellino corneal dystrophy, ACD), which is particularly prevalent in East Asian populations.

5. TGFBI-targeted gene therapy

Currently, gene therapy pipelines targeting the TGFBI gene have entered the preclinical stage. The pipeline SIS-201-CD, developed by SiSaf Ltd., utilizes siRNA drugs to inhibit abnormal TGFBI expression. Research has utilized corneal limbal biopsy tissue from patients with corneal dystrophy carrying the Arg124Cys mutation to establish an in vitro model for LCD1. It was found that siRNA can inhibit TGFBI mRNA and protein expression ^[2]. Additionally, research has corrected the R124H mutation in corneal cells of GCD patients using CRISPR gene editing technology ^[3].

6. Summary

The TGFBI gene is an important pathogenic gene for corneal dystrophy (CD). TGFBI gene humanized mice from Cyagen can be used for preclinical research on CD, and customized services can also be provided for different point mutations.

References

[1] Wang L, Zhao C, Zheng T, Zhang Y, Liu H, Wang X, Tang X, Zhao B, Liu P. Torin 1 alleviates impairment of TFEB-mediated lysosomal biogenesis and autophagy in TGFBI (p.G623_H626del)-linked Thiel-Behnke corneal dystrophy. Autophagy. 2022 Apr;18(4):765-782.

[2] Courtney DG, Atkinson SD, Moore JE, Maurizi E, Serafini C, Pellegrini G, Black GC, Manson FD, Yam GH, Macewen CJ, Allen EH, McLean WH, Moore CB. Development of allele-specific gene-silencing siRNAs for TGFBI Arg124Cys in lattice corneal dystrophy type I. Invest Ophthalmol Vis Sci. 2014 Feb 18;55(2):977-85.

[3] Taketani Y, Kitamoto K, Sakisaka T, Kimakura M, Toyono T, Yamagami S, Amano S, Kuroda M, Moore T, Usui T, Ouchi Y. Repair of the TGFBI gene in human corneal keratocytes derived from a granular corneal dystrophy patient via CRISPR/Cas9-induced homology-directed repair. Sci Rep. 2017 Dec 1;7(1):16713.