Journal
JOURNAL OF BIOLOGICAL CHEMISTRY
Volume 288, Issue 48, Pages 34671-34679Publisher
AMER SOC BIOCHEMISTRY MOLECULAR BIOLOGY INC
DOI: 10.1074/jbc.M113.496174
Keywords
Cell Differentiation; Cell Therapy; Hemoglobin; Homologous Recombination; Induced Pluripotent Stem (iPS) Cell; -Thalassemia; HBB; TALEN; Gene Correction; iPS
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Funding
- National Basic Research Program of China, 973 Program of China [2012CB966503]
- Strategic Priority Research Program of the Chinese Academy of Sciences [XDA01020202]
- 973 Program of China [2011CB965204]
- National S&T Major Special Project on Major New Drug Innovation [2011ZX09102-010]
- National Natural Science Foundation of China [31200970, 91213304]
- Bureau of Science and Technology of Guangzhou Municipality, China [2010U1-E00521]
- Hundred Talents Program of the Chinese Academy of Science
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Background: Gene editing in human patient-specific iPSCs is critical for regenerative medicine. Results: Nonintegrating -thalassemia iPSCs corrected by TALENs display undetectable off targets and can be differentiated into erythroblasts expressing normal -globin. Conclusion: TALENs can be used for HBB correction efficiently in -thalassemia iPSCs with different types. Significance: Our study extends TALENs for gene correction in patient-specific iPSCs and may have applications in cell therapy. -Thalassemia (-Thal) is a group of life-threatening blood disorders caused by either point mutations or deletions of nucleotides in -globin gene (HBB). It is estimated that 4.5% of the population in the world carry -Thal mutants (1), posing a persistent threat to public health. The generation of patient-specific induced pluripotent stem cells (iPSCs) and subsequent correction of the disease-causing mutations offer an ideal therapeutic solution to this problem. However, homologous recombination-based gene correction in human iPSCs remains largely inefficient. Here, we describe a robust process combining efficient generation of integration-free -Thal iPSCs from the cells of patients and transcription activator-like effector nuclease (TALEN)-based universal correction of HBB mutations in situ. We generated integration-free and gene-corrected iPSC lines from two patients carrying different types of homozygous mutations and showed that these iPSCs are pluripotent and have normal karyotype. We showed that the correction process did not generate TALEN-induced off targeting mutations by sequencing. More importantly, the gene-corrected -Thal iPS cell lines from each patient can be induced to differentiate into hematopoietic progenitor cells and then further to erythroblasts expressing normal -globin. Our studies provide an efficient and universal strategy to correct different types of -globin mutations in -Thal iPSCs for disease modeling and applications.
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