In vitro expansion of CD34(+)CD38(-) cells under stimulation with hematopoietic growth factors on AGM-S3 cells in juvenile myelomonocytic leukemia

Leukemia. 2015 Mar;29(3):606-14. doi: 10.1038/leu.2014.239. Epub 2014 Aug 8.

Abstract

Using serum-containing culture, we examined whether AGM-S3 stromal cells, alone or in combination with hematopoietic growth factor(s), stimulated the proliferation of CD34(+) cells from patients with juvenile myelomonocytic leukemia (JMML). AGM-S3 cells in concert with stem cell factor plus thrombopoietin increased the numbers of peripheral blood CD34(+) cells to approximately 20-fold of the input value after 2 weeks in nine JMML patients with either PTPN11 mutations or RAS mutations, who received allogeneic hematopoietic transplantation. Granulocyte-macrophage colony-stimulating factor (GM-CSF) also augmented the proliferation of JMML CD34(+) cells on AGM-S3 cells. The expansion potential of CD34(+) cells was markedly low in four patients who achieved spontaneous hematological improvement. A large proportion of day-14-cultured CD34(+) cells were negative for CD38 and cryopreservable. Cultured JMML CD34(+)CD38(-) cells expressed CD117, CD116, c-mpl, CD123, CD90, but not CXCR4, and formed GM and erythroid colonies. Day-7-cultured CD34(+) cells from two of three JMML patients injected intrafemorally into immunodeficient mice stimulated with human GM-CSF after transplantation displayed significant hematopoietic reconstitution. The abilities of OP9 cells and MS-5 cells were one-third and one-tenth, respectively, of the value obtained with AGM-S3 cells. Our culture system may provide a useful tool for elucidating leukemogenesis and for therapeutic approaches in JMML.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • ADP-ribosyl Cyclase 1 / genetics
  • ADP-ribosyl Cyclase 1 / metabolism
  • Adolescent
  • Animals
  • Antigens, CD34 / genetics
  • Antigens, CD34 / metabolism
  • Cell Proliferation / drug effects
  • Clone Cells
  • Coculture Techniques
  • Embryonic Stem Cells / drug effects*
  • Embryonic Stem Cells / metabolism
  • Embryonic Stem Cells / pathology
  • GTP Phosphohydrolases / genetics
  • GTP Phosphohydrolases / metabolism
  • Gene Expression Regulation, Leukemic*
  • Granulocyte-Macrophage Colony-Stimulating Factor / pharmacology*
  • Hematopoietic Stem Cells / drug effects*
  • Hematopoietic Stem Cells / metabolism
  • Hematopoietic Stem Cells / pathology
  • Humans
  • Leukemia, Myelomonocytic, Juvenile / genetics*
  • Leukemia, Myelomonocytic, Juvenile / metabolism
  • Leukemia, Myelomonocytic, Juvenile / pathology
  • Membrane Proteins / genetics
  • Membrane Proteins / metabolism
  • Mice
  • Mice, Inbred NOD
  • Mice, SCID
  • Mutation
  • Neoplastic Stem Cells / metabolism
  • Neoplastic Stem Cells / pathology
  • Neoplastic Stem Cells / transplantation
  • Protein Tyrosine Phosphatase, Non-Receptor Type 11 / genetics
  • Protein Tyrosine Phosphatase, Non-Receptor Type 11 / metabolism
  • Proto-Oncogene Proteins / genetics
  • Proto-Oncogene Proteins / metabolism
  • Proto-Oncogene Proteins p21(ras)
  • Signal Transduction
  • Stromal Cells / drug effects*
  • Stromal Cells / metabolism
  • Stromal Cells / pathology
  • ras Proteins / genetics
  • ras Proteins / metabolism

Substances

  • Antigens, CD34
  • KRAS protein, human
  • Membrane Proteins
  • Proto-Oncogene Proteins
  • Granulocyte-Macrophage Colony-Stimulating Factor
  • PTPN11 protein, human
  • Protein Tyrosine Phosphatase, Non-Receptor Type 11
  • ADP-ribosyl Cyclase 1
  • GTP Phosphohydrolases
  • NRAS protein, human
  • Proto-Oncogene Proteins p21(ras)
  • ras Proteins