In vivo selection of human embryonic stem cell-derived cells expressing methotrexate-resistant dihydrofolate reductase

• Published in: Gene therapy Vol. 17; no. 2; pp. 238 - 249
• Main Authors: Gori, J L, Tian, X, Swanson, D, Gunther, R, Shultz, L D, McIvor, R S, Kaufman, D S
• Format: Journal Article
• Language: English
• Published: London Nature Publishing Group UK 01.02.2010; Nature Publishing Group
• Subjects: Anesthesia. Intensive care medicine. Transfusions. Cell therapy and gene therapy; Animals; Applied cell therapy and gene therapy; Biological and medical sciences; Biomedical and Life Sciences; Biomedicine; Biotechnology; Bone Marrow; Bone marrow transplantation; CD34 antigen; CD45 antigen; Cell Biology; Cell Differentiation; Cell Line; Chemotherapy; Diabetes mellitus; Dihydrofolate reductase; Drug Resistance; Embryo cells; Embryonic stem cells; Embryonic Stem Cells - metabolism; Embryos; Endothelial cells; Fundamental and applied biological sciences. Psychology; Gene Expression; Gene Therapy; Genetic aspects; Genetic Therapy - methods; Graft Survival; Health aspects; Health. Pharmaceutical industry; Human Genetics; Humans; Immunodeficiency; Industrial applications and implications. Economical aspects; Interleukin 2 receptors; Medical sciences; Methotrexate; Methotrexate - pharmacology; Mice; Mice, Inbred NOD; Mice, SCID; Nanotechnology; original-article; Peripheral blood; Physiological aspects; Pluripotency; Rodents; Stem cell transplantation; Stem Cell Transplantation - methods; Stem cells; Teratoma - genetics; Tetrahydrofolate Dehydrogenase - genetics; Tetrahydrofolate Dehydrogenase - metabolism; Transfusions. Complications. Transfusion reactions. Cell and gene therapy; Xenografts; United States; methotrexate; embryonic stem cells; dihydrofolate reductase; Human; In vivo; Enzyme; Embryonic cell; Stem cell; Selection; Oxidoreductases; Dihydrofolate reductase; Methotrexate; Gene therapy
• ISSN: 0969-7128; 1476-5462
• DOI: 10.1038/gt.2009.131

Human embryonic stem cells (hESCs) provide a novel source of hematopoietic and other cell populations suitable for gene therapy applications. Preclinical studies to evaluate engraftment of hESC-derived hematopoietic cells transplanted into immunodeficient mice demonstrate only limited repopulation. Expression of a drug-resistance gene, such as Tyr22-dihydrofolate reductase (Tyr22-DHFR), coupled to methotrexate (MTX) chemotherapy has the potential to selectively increase the engraftment of gene-modified, hESC-derived cells in mouse xenografts. Here, we describe the generation of Tyr22-DHFR–GFP-expressing hESCs that maintain pluripotency, produce teratomas and can differentiate into MTXr-hemato-endothelial cells. We demonstrate that MTX administered to nonobese diabetic/severe combined immunodeficient/IL-2Rγc null (NSG) mice after injection of Tyr22-DHFR-hESC-derived cells significantly increases human CD34 + and CD45 + cell engraftment in the bone marrow (BM) and peripheral blood of transplanted MTX-treated mice. These results demonstrate that MTX treatment supports selective, long-term engraftment of Tyr22-DHFR cells in vivo , and provides a novel approach for combined human cell and gene therapy.