Format

Send to

Choose Destination
Stem Cell Reports. 2015 Dec 8;5(6):1053-1066. doi: 10.1016/j.stemcr.2015.10.002. Epub 2015 Nov 5.

Generation of a High Number of Healthy Erythroid Cells from Gene-Edited Pyruvate Kinase Deficiency Patient-Specific Induced Pluripotent Stem Cells.

Author information

1
Hematopoietic Innovative Therapies Division, Centro de Investigaciones Energéticas, Medioambientales y Tecnológicas (CIEMAT), Centro de Investigaciones Biomédicas en Red de Enfermedades Raras (CIBERER), Madrid 28040, Spain; Advanced Therapies Mixed Unit, Instituto de Investigación Sanitaria-Fundación Jiménez Díaz (IIS-FJD, UAM), Madrid 28040, Spain; Center for Stem Cell and Regenerative Medicine, Brown Foundation Institute of Molecular Medicine, University of Texas Health Science Center, Houston, TX 77030, USA.
2
Hematopoietic Innovative Therapies Division, Centro de Investigaciones Energéticas, Medioambientales y Tecnológicas (CIEMAT), Centro de Investigaciones Biomédicas en Red de Enfermedades Raras (CIBERER), Madrid 28040, Spain; Advanced Therapies Mixed Unit, Instituto de Investigación Sanitaria-Fundación Jiménez Díaz (IIS-FJD, UAM), Madrid 28040, Spain. Electronic address: oscar.quintana@ciemat.es.
3
Center for Stem Cell and Regenerative Medicine, Brown Foundation Institute of Molecular Medicine, University of Texas Health Science Center, Houston, TX 77030, USA.
4
Institute of Cardiovascular and Medical Sciences, University of Glasgow, Glasgow G12 8QQ, UK.
5
Cellectis, Paris 75013, France.
6
Agios Pharmaceuticals, Cambridge, MA 02139-4169, USA.
7
Hematology and Cell Therapy, Clinica Universidad de Navarra and CIMA, Pamplona 31008, Spain.
8
Hematopoietic Innovative Therapies Division, Centro de Investigaciones Energéticas, Medioambientales y Tecnológicas (CIEMAT), Centro de Investigaciones Biomédicas en Red de Enfermedades Raras (CIBERER), Madrid 28040, Spain; Advanced Therapies Mixed Unit, Instituto de Investigación Sanitaria-Fundación Jiménez Díaz (IIS-FJD, UAM), Madrid 28040, Spain.
9
JST PRESTO and Ophthalmology, Keio University, Tokyo 108-8345, Japan.
10
Histocompatibility and Molecular Biology Laboratory, Centro de Transfusion de Madrid, Madrid 28032, Spain.
11
Serviço de Hematologia, Centro Hospitalar e Universitario de Coimbra, Coimbra 3000-075, Portugal.
12
Hospital Universitario Niño Jesús, Madrid 28009, Spain.
13
Hematopoietic Innovative Therapies Division, Centro de Investigaciones Energéticas, Medioambientales y Tecnológicas (CIEMAT), Centro de Investigaciones Biomédicas en Red de Enfermedades Raras (CIBERER), Madrid 28040, Spain; Advanced Therapies Mixed Unit, Instituto de Investigación Sanitaria-Fundación Jiménez Díaz (IIS-FJD, UAM), Madrid 28040, Spain. Electronic address: jc.segovia@ciemat.es.

Abstract

Pyruvate kinase deficiency (PKD) is a rare erythroid metabolic disease caused by mutations in the PKLR gene. Erythrocytes from PKD patients show an energetic imbalance causing chronic non-spherocytic hemolytic anemia, as pyruvate kinase defects impair ATP production in erythrocytes. We generated PKD induced pluripotent stem cells (PKDiPSCs) from peripheral blood mononuclear cells (PB-MNCs) of PKD patients by non-integrative Sendai viral vectors. PKDiPSCs were gene edited to integrate a partial codon-optimized R-type pyruvate kinase cDNA in the second intron of the PKLR gene by TALEN-mediated homologous recombination (HR). Notably, we found allele specificity of HR led by the presence of a single-nucleotide polymorphism. High numbers of erythroid cells derived from gene-edited PKDiPSCs showed correction of the energetic imbalance, providing an approach to correct metabolic erythroid diseases and demonstrating the practicality of this approach to generate the large cell numbers required for comprehensive biochemical and metabolic erythroid analyses.

PMID:
26549847
PMCID:
PMC4682065
DOI:
10.1016/j.stemcr.2015.10.002
[Indexed for MEDLINE]
Free PMC Article

Supplemental Content

Full text links

Icon for Elsevier Science Icon for PubMed Central
Loading ...
Support Center