{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,5,1]],"date-time":"2026-05-01T20:50:37Z","timestamp":1777668637621,"version":"3.51.4"},"reference-count":78,"publisher":"SAGE Publications","issue":"15-16","license":[{"start":{"date-parts":[[2025,8,1]],"date-time":"2025-08-01T00:00:00Z","timestamp":1754006400000},"content-version":"vor","delay-in-days":0,"URL":"https:\/\/creativecommons.org\/licenses\/by\/4.0\/"},{"start":{"date-parts":[[2025,8,1]],"date-time":"2025-08-01T00:00:00Z","timestamp":1754006400000},"content-version":"tdm","delay-in-days":0,"URL":"https:\/\/journals.sagepub.com\/page\/policies\/text-and-data-mining-license"}],"content-domain":{"domain":["journals.sagepub.com"],"crossmark-restriction":true},"short-container-title":["Human Gene Therapy"],"published-print":{"date-parts":[[2025,8]]},"abstract":"<jats:p>\n                    Cystic fibrosis (CF) is caused by mutations in the\n                    <jats:italic toggle=\"yes\">cystic fibrosis transmembrane conductance regulator<\/jats:italic>\n                    (\n                    <jats:italic toggle=\"yes\">CFTR<\/jats:italic>\n                    ) gene. While CRISPR-based\n                    <jats:italic toggle=\"yes\">CFTR<\/jats:italic>\n                    editing approaches have shown proof-of-concept for functional rescue in primary airway basal cells, induced pluripotent stem cells, and organoid cultures derived from patients with CF, their efficacy remains suboptimal. Here, we developed the CuFi\n                    <jats:sup>Cas9(Y66S)eGFP<\/jats:sup>\n                    reporter system by integrating spCas9 and a non-fluorescent Y66S eGFP mutant into CuFi-8 cells, an immortalized human airway epithelial cell line derived from a patient with CF with homozygous F508del mutations. These cells retain the basal cell phenotype in proliferating cultures and can differentiate into polarized airway epithelium at an air-liquid interface (ALI), enabling both visualized detection of gene editing and electrophysiological assessment of\n                    <jats:italic toggle=\"yes\">CFTR<\/jats:italic>\n                    functional restoration. Using this system, recombinant adeno-associated virus (rAAV)-mediated homology-directed repair (HDR) was evaluated in proliferating cultures. A correction rate of 13.5 \u00b1 0.8% was achieved in a population where 82.3 \u00b1 5.6% of cells were productively transduced by AAV.eGFP630g2-CMVmCh, an rAAV editing vector with an mCherry reporter. Dual-editing of F508del\n                    <jats:italic toggle=\"yes\">CFTR<\/jats:italic>\n                    and Y66S\n                    <jats:italic toggle=\"yes\">eGFP<\/jats:italic>\n                    was explored using AAV.HR-eGFP630-F508(g03) to deliver two templates and single guide RNAs. eGFP\n                    <jats:sup>+<\/jats:sup>\n                    (Y66S-corrected) cells and eGFP\n                    <jats:sup>\u2212<\/jats:sup>\n                    (non-corrected) cells were sorted via fluorescence-activated cell sorting and differentiated at an ALI to assess the recovery of CFTR function. Despite a low F508 correction rate of 2.8%, ALI cultures derived from the eGFP\n                    <jats:sup>\u2212<\/jats:sup>\n                    population exhibited 25.2% of the CFTR-specific transepithelial Cl\n                    <jats:sup>\u2212<\/jats:sup>\n                    transport observed in CuFi-ALI cultures treated with CFTR modulators. Next-generation sequencing revealed frequent co-editing at both genomic loci, with sixfold higher F508 correction rate in the eGFP\n                    <jats:sup>+<\/jats:sup>\n                    cells than eGFP\n                    <jats:sup>\u2212<\/jats:sup>\n                    cells. In both populations, non-homology end joining predominated over HDR. This reporter system provides a valuable platform for optimizing editing efficiencies in proliferating airway basal cells, particularly for development of strategies to enhance HDR through modulation of DNA repair pathways.\n                  <\/jats:p>","DOI":"10.1089\/hum.2024.260","type":"journal-article","created":{"date-parts":[[2025,5,13]],"date-time":"2025-05-13T12:42:38Z","timestamp":1747140158000},"page":"1067-1082","update-policy":"https:\/\/doi.org\/10.1177\/sage-journals-update-policy","source":"Crossref","is-referenced-by-count":2,"title":["Recombinant Adeno-Associated Virus Vector Mediated Gene Editing in Proliferating and Polarized Cultures of Human Airway Epithelial Cells"],"prefix":"10.1177","volume":"36","author":[{"given":"Soo Yeun","family":"Park","sequence":"first","affiliation":[{"name":"Department of Anatomy and Cell Biology, Carver College of Medicine, University of Iowa, Iowa City, Iowa, USA"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Zehua","family":"Feng","sequence":"additional","affiliation":[{"name":"Department of Anatomy and Cell Biology, Carver College of Medicine, University of Iowa, Iowa City, Iowa, USA"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Soon H.","family":"Choi","sequence":"additional","affiliation":[{"name":"Department of Anatomy and Cell Biology, Carver College of Medicine, University of Iowa, Iowa City, Iowa, USA"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Xiujuan","family":"Zhang","sequence":"additional","affiliation":[{"name":"Department of Microbiology, Molecular Genetics and Immunology, University of Kansas Medical Center, Kansas City, Kansas, USA."}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Yinghua","family":"Tang","sequence":"additional","affiliation":[{"name":"Department of Anatomy and Cell Biology, Carver College of Medicine, University of Iowa, Iowa City, Iowa, USA"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-2393-2674","authenticated-orcid":false,"given":"Grace N.","family":"Gasser","sequence":"additional","affiliation":[{"name":"Department of Anatomy and Cell Biology, Carver College of Medicine, University of Iowa, Iowa City, Iowa, USA"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Donovan","family":"Richart","sequence":"additional","affiliation":[{"name":"Department of Microbiology, Molecular Genetics and Immunology, University of Kansas Medical Center, Kansas City, Kansas, USA."}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Feng","family":"Yuan","sequence":"additional","affiliation":[{"name":"Department of Anatomy and Cell Biology, Carver College of Medicine, University of Iowa, Iowa City, Iowa, USA"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"ORCID":"https:\/\/orcid.org\/0000-0001-9850-1695","authenticated-orcid":false,"given":"Jianming","family":"Qiu","sequence":"additional","affiliation":[{"name":"Department of Microbiology, Molecular Genetics and Immunology, University of Kansas Medical Center, Kansas City, Kansas, USA."}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"John F.","family":"Engelhardt","sequence":"additional","affiliation":[{"name":"Department of Anatomy and Cell Biology, Carver College of Medicine, University of Iowa, Iowa City, Iowa, USA"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"ORCID":"https:\/\/orcid.org\/0000-0001-8210-5567","authenticated-orcid":false,"given":"Ziying","family":"Yan","sequence":"additional","affiliation":[{"name":"Department of Anatomy and Cell Biology, Carver College of Medicine, University of Iowa, Iowa City, Iowa, 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