scholarly journals TALEN-Mediated Gene Targeting for Cystic Fibrosis-Gene Therapy

Genes ◽  
2019 ◽  
Vol 10 (1) ◽  
pp. 39 ◽  
Author(s):  
Emily Xia ◽  
Yiqian Zhang ◽  
Huibi Cao ◽  
Jun Li ◽  
Rongqi Duan ◽  
...  
Keyword(s):  
Cystic Fibrosis ◽  
Gene Therapy ◽  
Gene Editing ◽  
Airway Epithelial Cells ◽  
In Vivo Studies ◽  
Cystic Fibrosis Gene ◽  
Lacz Reporter ◽  
Gene Correction ◽  
Transcription Activator ◽  
Stringent Requirement

Cystic fibrosis (CF) is an inherited monogenic disorder, amenable to gene-based therapies. Because CF lung disease is currently the major cause of mortality and morbidity, and the lung airway is readily accessible to gene delivery, the major CF gene therapy effort at present is directed to the lung. Although airway epithelial cells are renewed slowly, permanent gene correction through gene editing or targeting in airway stem cells is needed to perpetuate the therapeutic effect. Transcription activator-like effector nuclease (TALEN) has been utilized widely for a variety of gene editing applications. The stringent requirement for nuclease binding target sites allows for gene editing with precision. In this study, we engineered helper-dependent adenoviral (HD-Ad) vectors to deliver a pair of TALENs together with donor DNA targeting the human AAVS1 locus. With homology arms of 4 kb in length, we demonstrated precise insertion of either a LacZ reporter gene or a human cystic fibrosis transmembrane conductance regulator (CFTR) minigene (cDNA) into the target site. Using the LacZ reporter, we determined the efficiency of gene integration to be about 5%. In the CFTR vector transduced cells, we were able to detect CFTR mRNA expression using qPCR and function correction using fluorometric image plate reader (FLIPR) and iodide efflux assays. Taken together, these findings suggest a new direction for future in vitro and in vivo studies in CF gene editing.

scholarly journals TALEN Mediated Gene Targeting for CF Gene Therapy

2018 ◽  
Author(s):  
Emily Xia ◽  
Yiqian Zhang ◽  
Huibi Cao ◽  
Jun Li ◽  
Rongqi Duan ◽  
...  
Keyword(s):  
Gene Therapy ◽  
Gene Editing ◽  
Airway Epithelial Cells ◽  
In Vivo Studies ◽  
Lacz Reporter ◽  
Gene Correction ◽  
Transcription Activator ◽  
Stringent Requirement

Cystic Fibrosis (CF) is an inherited monogenic disorder, amenable to gene based therapies. Because CF lung disease is currently the major cause of mortality and morbidity, and lung airway is readily accessible to gene delivery, the major CF gene therapy effort at present is directed to the lung. Although airway epithelial cells are renewed slowly, permanent gene correction through gene editing or targeting in airway stem cells is needed to perpetuate the therapeutic effect. Transcription activator-like effector nuclease (TALEN) has been utilized widely for a variety of gene editing applications. The stringent requirement for nuclease binding target sites allows for gene editing with precision. In this study, we engineered helper-dependent adenoviral (HD-Ad) vectors to deliver a pair of TALENs together with donor DNA targeting the human AAVS1 locus. With homology arms of 4 kb in length, we demonstrated precise insertion of either a LacZ reporter gene or a human CFTR minigene into the target site. Using the LacZ reporter, we determined the efficiency of gene integration to be about 5%. In the CFTR vector transduced cells, we have detected both CFTR mRNA and protein expression by qPCR and Wetern analysis, respectively. We have also confirmed CFTR function correction by flurometric Image Plate Reader (FLIPR) and iodide efflux assays. Taking together, these findings suggest a new direction for future in vitro and in vivo studies in CF gene editing.

Aerosol delivery of DNA/liposomes to the lung for cystic fibrosis gene therapy

2014 ◽  
pp. 150127063140004
Author(s):  
Lee Adrian Davies ◽  
Graciela A Nunez-Alonso ◽  
Gerry McLachlan ◽  
Stephen C Hyde ◽  
Deborah Rebecca Gill
Keyword(s):  
Cystic Fibrosis ◽  
Gene Therapy ◽  
Cystic Fibrosis Gene ◽  
Aerosol Delivery

Biology students' understanding of cystic fibrosis, gene therapy, and gene screening

1998 ◽  
Vol 32 (2) ◽  
pp. 103-110 ◽  
Author(s):  
Ruaraidh Hill ◽  
Helen O'Sullivan ◽  
Martin Stanisstreet ◽  
Edward Boyes
Keyword(s):  
Cystic Fibrosis ◽  
Gene Therapy ◽  
Cystic Fibrosis Gene ◽  
Biology Students ◽  
Gene Screening

scholarly journals Cystic Fibrosis Gene Therapy: Looking Back, Looking Forward

Genes ◽  
2018 ◽  
Vol 9 (11) ◽  
pp. 538 ◽  
Author(s):  
Ashley Cooney ◽  
Paul McCray ◽  
Patrick Sinn
Keyword(s):  
Cystic Fibrosis ◽  
Gene Therapy ◽  
Anion Channel ◽  
Gene Replacement ◽  
Cystic Fibrosis Gene ◽  
Cftr Gene ◽  
System Disease ◽  
Cftr Mutations ◽  
Infection And Inflammation

Cystic fibrosis (CF) is an autosomal recessive disease caused by mutations in the cystic fibrosis transmembrane conductance regulator (CFTR) gene that encodes a cAMP-regulated anion channel. Although CF is a multi-organ system disease, most people with CF die of progressive lung disease that begins early in childhood and is characterized by chronic bacterial infection and inflammation. Nearly 90% of people with CF have at least one copy of the ΔF508 mutation, but there are hundreds of CFTR mutations that result in a range of disease severities. A CFTR gene replacement approach would be efficacious regardless of the disease-causing mutation. After the discovery of the CFTR gene in 1989, the in vitro proof-of-concept for gene therapy for CF was quickly established in 1990. In 1993, the first of many gene therapy clinical trials attempted to rescue the CF defect in airway epithelia. Despite the initial enthusiasm, there is still no FDA-approved gene therapy for CF. Here we discuss the history of CF gene therapy, from the discovery of the CFTR gene to current state-of-the-art gene delivery vector designs. While implementation of CF gene therapy has proven more challenging than initially envisioned; thanks to continued innovation, it may yet become a reality.

Sign in / Sign up

Export Citation Format

Share Document