Optimization of hCFTR lung expression in mice using DNA nanoparticles

Area of Science:

• Gene Therapy
• Molecular Biology
• Pulmonary Medicine

Background:

• Cystic fibrosis (CF) requires efficient and sustained expression of the human cystic fibrosis transmembrane conductance regulator (hCFTR) for effective lung therapy.
• Current gene therapy approaches face challenges in achieving prolonged hCFTR expression.

Purpose of the Study:

• To optimize a gene therapy vector for enhanced and prolonged hCFTR expression in the lungs of cystic fibrosis models.
• To investigate methods for extending the duration of hCFTR gene expression using novel vector modifications.

Main Methods:

• Development of a codon-optimized and CpG-reduced hCFTR synthetic gene (CO-CFTR).
• Formulation of CO-CFTR into compacted DNA nanoparticles using PEG-substituted lysine peptides.
• Incorporation of a novel prolonged expression (PE) element derived from the bovine growth hormone gene 3' flanking sequence into the expression vector.

Main Results:

• The CO-CFTR gene significantly increased hCFTR protein levels in cell culture compared to standard hCFTR cDNA.
• DNA nanoparticles containing CO-CFTR demonstrated improved lung mRNA expression in CF mouse models, though transient.
• The novel PE element successfully achieved prolonged CO-CFTR mRNA expression at biologically relevant levels in mouse lungs.
• Prolonged expression was dependent on PE element orientation and transcription, and not specific to the UbC promoter.

Conclusions:

• The optimized CO-CFTR gene and PE element represent a significant advancement in developing gene therapies for cystic fibrosis.
• This vector strategy holds potential for achieving sustained therapeutic levels of hCFTR in CF lung treatments.
• Further research is warranted to fully elucidate the mechanisms and optimize the application of this prolonged expression system.