B6-hCFTR Mice

Figure 1. Gene editing strategy of B6-hCFTR mice. The region from ~9.5 kb upstream to ~1.2 kb downstream of mouse Cftr gene was replaced with the region from ~40.1 kb upstream to ~25 kb downstream of human CFTR gene.

1. Detection of human CFTR gene expression

Figure 2. Human CFTR gene expression in the colon and testis of wild-type mice (WT) and B6-hCFTR mice (hCFTR). RT-qPCR results showed that there was expression of the human CFTR gene in the colon and testis of B6-hCFTR mice, while there was no expression in the WT mice.

2. Detection of mouse Cftr gene expression

Figure 3. Mouse Cftr gene expression in the colon and testis of wild-type mice (WT) and B6-hCFTR mice (hCFTR). RT-qPCR results showed that there was the expression of the mouse Cftr gene in the colon and testis of WT mice, while there was no expression in the B6-hCFTR mice.

1. Basic information about the CFTR gene

https://rddc.tsinghua-gd.org/gene/1080

2. CFTR clinical variants

https://rddc.tsinghua-gd.org/ai/pathogenicity/5c477f5adfbe4d5babef43b15819df8a

3. Disease introduction

Cystic Fibrosis (CF) is an autosomal recessive genetic disorder that causes severe damage to the lungs, digestive system, and other organs. It thickens normal mucus, sweat, and digestive fluids by affecting cells, preventing them from acting as lubricants and blocking ducts and channels. The impact on the lungs and pancreas is particularly severe. The disease has five clinical characteristics. Firstly, there is persistent productive cough and hyperinflation of the lung lobes in the respiratory system. Secondly, there is chronic nasal congestion, headaches, chronic postnasal drip cough, and sleep disorders in the sinuses. Thirdly, there are digestive system disorders such as pancreatic exocrine dysfunction and increased intestinal viscosity leading to intestinal obstruction. Fourthly, there are reproductive system disorders such as sperm transport defects leading to infertility in male CF patients, although spermatogenesis is unaffected. Fifthly, due to the reduced mineral content in the bones of CF patients, there may be nutritional and growth development disorders, such as an increased incidence of fractures and scoliosis.

4. CFTR gene and mutations

The CFTR gene encodes the Cystic Fibrosis Transmembrane Conductance Regulator, a cAMP-dependent chloride ion channel protein distributed in the airways, pancreatic ducts, digestive tract, reproductive system, sweat glands, etc., driving the secretion of chloride ions and bicarbonate. Abnormal CFTR function can cause transmembrane transport disorders of chloride ions and bicarbonate, leading to widespread mucus obstruction in exocrine glands throughout the body, affecting multiple systems such as respiration, digestion, endocrine, and reproduction. More than 2000 mutations of the CFTR gene have been discovered so far, with varying incidence rates reported in different countries and regions. The incidence rate in newborns is approximately 1/25000 to 1/1800. The number of cases in Asia and Africa is far less than in Europe and North America. In addition, 85% to 90% of Caucasian CF patients carry at least one F508del mutation ^[1].

5. Function of non-coding DNA sequences

Researchers have found numerous miRNA binding sites in the CFTR mRNA. Among them, miR-145-5p can inhibit the synthesis of CFTR through interactions with CFTR and other factors. To further investigate, researchers synthesized a peptide nucleic acid (PNAs) R8-PNA-a145 targeting miR-145-5p. Experimental results showed that this PNA can inhibit the expression of miR-145-5p and upregulate the expression of CFTR mRNA. This finding suggests that the 3’UTR of CFTR has a potential drug target value ^[2].

6. CFTR-targeted gene therapy gene therapy

The current CF treatment pipeline is primarily focused on small-molecule drugs. In recent years, numerous pipelines related to gene therapy have also emerged. One such pipeline is Eluforsen, a clinical phase 1 ASO-related pipeline from ProQR company. This pipeline targets the region near the F508dcl mutation of the CFTR gene to restore the function of the CFTR protein ^[3-4]. The CFTR-F508del mouse model was used in its preclinical research.

In summary, the CFTR gene is a significant pathogenic gene for Cystic Fibrosis (CF). Humanizing mouse genes can help accelerate CFTR-targeted therapies into clinical stages. CFTR gene humanized mice from Cyagen can be used for preclinical research on CF and customized services can also be provided for different point mutations.