Nanoparticles that deliver triplex-forming peptide nucleic acid molecules correct F508del CFTR in airway epithelium.
Cystic fibrosis (CF) is a lethal genetic disorder most commonly caused by the F508del mutation in the cystic fibrosis transmembrane conductance regulator (CFTR) gene. It is not readily amenable to gene therapy because of its systemic nature and challenges including in vivo gene delivery and transient gene expression. Here we use triplex-forming peptide nucleic acids and donor DNA in biodegradable polymer nanoparticles to correct F508del. We confirm modification with sequencing and a functional chloride efflux assay. In vitro correction of chloride efflux occurs in up to 25% of human cells. Deep-sequencing reveals negligible off-target effects in partially homologous sites. Intranasal delivery of nanoparticles in CF mice produces changes in the nasal epithelium potential difference assay, consistent with corrected CFTR function. Also, gene correction is detected in the nasal and lung tissue. This work represents facile genome engineering in vivo with oligonucleotides using a nanoparticle system to achieve clinically relevant levels of gene editing without off-target effects.
Department of Biomedical Engineering, Yale University, New Haven, Connecticut 06520, USA.
Nicole Ali
NA
McNeer
1] Department of Biomedical Engineering, Yale University, New Haven, Connecticut 06520, USA [2] Yale College, Department of Biomedical Engineering, New Haven, Connecticut 06520, USA.
Kavitha
K
Anandalingam
Department of Biomedical Engineering, Yale University, New Haven, Connecticut 06520, USA.
Rachel J
RJ
Fields
Department of Pediatrics, Yale School of Medicine, New Haven, Connecticut 06510, USA.
Christina
C
Caputo
Department of Cellular and Molecular Physiology, Yale School of Medicine, New Haven, Connecticut 06510, USA.
Sascha
S
Kopic
Department of Therapeutic Radiology and Genetics, Yale School of Medicine, New Haven, Connecticut 06510, USA.
Anisha
A
Gupta
Department of Biomedical Engineering, Yale University, New Haven, Connecticut 06520, USA.
Elias
E
Quijano
Department of Pediatrics, Yale School of Medicine, New Haven, Connecticut 06510, USA.
Lee
L
Polikoff
1] Yale Department of Molecular Biophysics and Biochemistry, New Haven, Connecticut 06520, USA [2] Yale University, Department of Bioinformatics, W.M Keck Foundation Biotechnology Resource Laboratory, New Haven, Connecticut 06511, USA.
Yong
0000000228815274
Y
Kong
Department of Therapeutic Radiology and Genetics, Yale School of Medicine, New Haven, Connecticut 06510, USA.
Raman
R
Bahal
1] Department of Cellular and Molecular Physiology, Yale School of Medicine, New Haven, Connecticut 06510, USA [2] Department of Surgery, Yale School of Medicine, New Haven, Connecticut 06510, USA.
John P
JP
Geibel
Department of Therapeutic Radiology and Genetics, Yale School of Medicine, New Haven, Connecticut 06510, USA.
Peter M
PM
Glazer
Department of Biomedical Engineering, Yale University, New Haven, Connecticut 06520, USA.
W Mark
WM
Saltzman
1] Department of Pediatrics, Yale School of Medicine, New Haven, Connecticut 06510, USA [2] Department of Cellular and Molecular Physiology, Yale School of Medicine, New Haven, Connecticut 06510, USA.
Marie E
ME
Egan
D000818:Animals; D002460:Cell Line; D002712:Chlorides / Q000378:metabolism; D003550:Cystic Fibrosis / Q000628:therapy*; D019005:Cystic Fibrosis Transmembrane Conductance Regulator / Q000235:genetics*; D004268:DNA-Binding Proteins; D015316:Genetic Therapy / Q000379:methods*; D059014:High-Throughput Nucleotide Sequencing; D006801:Humans; D019344:Lactic Acid; D008810:Mice, Inbred C57BL; D053758:Nanoparticles / Q000008:administration & dosage; D020135:Peptide Nucleic Acids / Q000627:therapeutic use*; D011100:Polyglycolic Acid; D000077182:Polylactic Acid-Polyglycolic Acid Copolymer; D011108:Polymers; D020545:Respiratory Mucosa / Q000378:metabolism
D002712:Chlorides; D004268:DNA-Binding Proteins; C475074:MPG peptide; D020135:Peptide Nucleic Acids; D011108:Polymers; C507253:poly(beta-amino ester); D019005:Cystic Fibrosis Transmembrane Conductance Regulator; D000077182:Polylactic Acid-Polyglycolic Acid Copolymer; D011100:Polyglycolic Acid; D019344:Lactic Acid