Restored glyoxylate metabolism after AGXT gene correction and direct reprogramming of primary hyperoxaluria type 1 fibroblasts
Virginia Nieto-Romero,
Aida García-Torralba,
Andrea Molinos-Vicente,
Francisco José Moya,
Sandra Rodríguez-Perales,
Ramón García-Escudero,
Eduardo Salido,
José-Carlos Segovia,
María García-Bravo
Affiliations
Virginia Nieto-Romero
Cell Technology Division, Biomedical Innovation Unit, CIEMAT (Centro de Investigaciones Energéticas, Medioambientales y Tecnológicas), Centro de Investigación Biomédica en Red de Enfermedades Raras (CIBERER)-ISCIII, Instituto de Investigación Sanitaria Fundación Jiménez Díaz (IIS-FJD, UAM), 28040 Madrid, Spain
Aida García-Torralba
Cell Technology Division, Biomedical Innovation Unit, CIEMAT (Centro de Investigaciones Energéticas, Medioambientales y Tecnológicas), Centro de Investigación Biomédica en Red de Enfermedades Raras (CIBERER)-ISCIII, Instituto de Investigación Sanitaria Fundación Jiménez Díaz (IIS-FJD, UAM), 28040 Madrid, Spain
Andrea Molinos-Vicente
Cell Technology Division, Biomedical Innovation Unit, CIEMAT (Centro de Investigaciones Energéticas, Medioambientales y Tecnológicas), Centro de Investigación Biomédica en Red de Enfermedades Raras (CIBERER)-ISCIII, Instituto de Investigación Sanitaria Fundación Jiménez Díaz (IIS-FJD, UAM), 28040 Madrid, Spain
Francisco José Moya
Molecular Cytogenetics and Genome Editing Unit, Human Cancer Genetics Program, Centro Nacional de Investigaciones Oncológicas (CNIO), 28029 Madrid, Spain
Sandra Rodríguez-Perales
Molecular Cytogenetics and Genome Editing Unit, Human Cancer Genetics Program, Centro Nacional de Investigaciones Oncológicas (CNIO), 28029 Madrid, Spain
Ramón García-Escudero
Molecular Oncology Unit, CIEMAT (Centro de Investigaciones Energéticas, Medioambientales y Tecnológicas), Centro de Investigación Biomédica en Red de Cáncer (CIBERONC)-ISCIII, Research Institute Hospital 12 de Octubre (imas12)-University Hospital 12 de Octubre, 28040 Madrid, Spain
Eduardo Salido
Pathology Department, Hospital Universitario de Canarias, Universidad La Laguna, Centro de Investigación Biomédica en Red de Enfermedades Raras (CIBERER)-ISCIII, 38320 Tenerife, Spain
José-Carlos Segovia
Cell Technology Division, Biomedical Innovation Unit, CIEMAT (Centro de Investigaciones Energéticas, Medioambientales y Tecnológicas), Centro de Investigación Biomédica en Red de Enfermedades Raras (CIBERER)-ISCIII, Instituto de Investigación Sanitaria Fundación Jiménez Díaz (IIS-FJD, UAM), 28040 Madrid, Spain; Corresponding author
María García-Bravo
Cell Technology Division, Biomedical Innovation Unit, CIEMAT (Centro de Investigaciones Energéticas, Medioambientales y Tecnológicas), Centro de Investigación Biomédica en Red de Enfermedades Raras (CIBERER)-ISCIII, Instituto de Investigación Sanitaria Fundación Jiménez Díaz (IIS-FJD, UAM), 28040 Madrid, Spain; Corresponding author
Summary: Primary hyperoxaluria type 1 (PH1) is a rare inherited metabolic disorder characterized by oxalate overproduction in the liver, resulting in renal damage. It is caused by mutations in the AGXT gene. Combined liver and kidney transplantation is currently the only permanent curative treatment. We combined locus-specific gene correction and hepatic direct cell reprogramming to generate autologous healthy induced hepatocytes (iHeps) from PH1 patient-derived fibroblasts. First, site-specific AGXT corrected cells were obtained by homology directed repair (HDR) assisted by CRISPR-Cas9, following two different strategies: accurate point mutation (c.731T>C) correction or knockin of an enhanced version of AGXT cDNA. Then, iHeps were generated, by overexpression of hepatic transcription factors. Generated AGXT-corrected iHeps showed hepatic gene expression profile and exhibited in vitro reversion of oxalate accumulation compared to non-edited PH1-derived iHeps. This strategy set up a potential alternative cellular source for liver cell replacement therapy and a personalized PH1 in vitro disease model.