Lentiviral gene therapy and vitamin B3 treatment enable granulocytic differentiation of G6PC3-deficient induced pluripotent stem cells

• Published in: Gene therapy 2020-06, Vol.27 (6), p.297-306
• Main Authors: Hoffmann, Dirk, Kuehle, Johannes, Lenz, Daniela, Philipp, Friederike, Zychlinski, Daniela, Lachmann, Nico, Moritz, Thomas, Steinemann, Doris, Morgan, Michael, Skokowa, Julia, Klein, Christoph, Schambach, Axel
• Format: Article
• Language: English
• Subjects: 13/100; 13/31; 13/44; 38/77; 631/532/2435; 692/699/249; Analysis; Apoptosis; Biomedical and Life Sciences; Biomedicine; Brief Communication; Care and treatment; Cell Biology; Cell Differentiation; Codon; Colonies; Colony-stimulating factor; Gene Expression; Gene Therapy; Genes; Genetic disorders; Genetic Therapy; Genetic vectors; Glucose-6-Phosphatase; Granulocyte Colony-Stimulating Factor; Granulocytes; Granulopoiesis; Health aspects; Human Genetics; Humans; Induced Pluripotent Stem Cells; Interleukin 3; Leukocytes (granulocytic); Leukocytes (neutrophilic); Macrophage colony stimulating factor; Macrophages; Monocytes; Myeloid cells; Nanotechnology; Neutropenia; Neutrophils; Niacin; Niacinamide; Nicotinic acid; Nonsense mutation; Phenotypes; Phosphatases; Pluripotency; Stem cells; Transplantation
• ISSN: 0969-7128; 1476-5462
• DOI: 10.1038/s41434-020-0127-y

Induced pluripotent stem cells (iPSCs) from patients with genetic disorders are a valuable source for in vitro disease models, which enable drug testing and validation of gene and cell therapies. We generated iPSCs from a severe congenital neutropenia (SCN) patient, who presented with a nonsense mutation in the glucose-6-phosphatase catalytic subunit 3 ( G6PC3 ) gene causing profound defects in granulopoiesis, associated with increased susceptibility of neutrophils to apoptosis. Generated SCN iPSC clones exhibited the capacity to differentiate into hematopoietic cells of the myeloid lineage and we identified two cytokine conditions, i.e., using granulocyte-colony stimulating factor or granulocyte-macrophage colony stimulating factor in combination with interleukin-3, to model the SCN phenotype in vitro. Reduced numbers of granulocytes were produced by SCN iPSCs compared with control iPSCs in both settings, which reflected the phenotype in patients. Interestingly, our model showed increased monocyte/macrophage production from the SCN iPSCs. Most importantly, lentiviral genetic correction of SCN iPSCs with a codon-optimized G6PC3 transgene restored granulopoiesis and reduced apoptosis of in vitro differentiated myeloid cells. Moreover, addition of vitamin B3 clearly induced granulocytic differentiation of SCN iPSCs and increased the number of neutrophils to levels comparable with those obtained from healthy control iPSCs. In summary, we established an iPSC-derived in vitro disease model, which will serve as a tool to test the potency of alternative treatment options for SCN patients, such as small molecules and gene therapeutic vectors.