title: Anti-proliferative effects of T cells expressing a ligand-based chimeric antigen receptor against CD116 on CD34(+) cells of juvenile myelomonocytic leukemia.
abstract: BACKGROUND
  Juvenile myelomonocytic leukemia (JMML) is a fatal, myelodysplastic/myeloproliferative neoplasm of early childhood. Patients with JMML have mutually exclusive genetic abnormalities in granulocyte-macrophage colony-stimulating factor (GM-CSF) receptor (GMR, CD116) signaling pathway. Allogeneic hematopoietic stem cell transplantation is currently the only curative treatment option for JMML; however, disease recurrence is a major cause of treatment failure. We investigated adoptive immunotherapy using GMR-targeted chimeric antigen receptor (CAR) for JMML.
  
  
  METHODS
  We constructed a novel CAR capable of binding to GMR via its ligand, GM-CSF, and generated piggyBac transposon-based GMR CAR-modified T cells from three healthy donors and two patients with JMML. We further evaluated the anti-proliferative potential of GMR CAR T cells on leukemic CD34(+) cells from six patients with JMML (two NRAS mutations, three PTPN11 mutations, and one monosomy 7), and normal CD34(+) cells.
  
  
  RESULTS
  GMR CAR T cells from healthy donors suppressed the cytokine-dependent growth of MO7e cells, but not the growth of K562 and Daudi cells. Co-culture of healthy GMR CAR T cells with CD34(+) cells of five patients with JMML at effector to target ratios of 1:1 and 1:4 for 2 days significantly decreased total colony growth, regardless of genetic abnormality. Furthermore, GMR CAR T cells from a non-transplanted patient and a transplanted patient inhibited the proliferation of respective JMML CD34(+) cells at onset to a degree comparable to healthy GMR CAR T cells. Seven-day co-culture of GMR CAR T cells resulted in a marked suppression of JMML CD34(+) cell proliferation, particularly CD34(+)CD38(-) cell proliferation stimulated with stem cell factor and thrombopoietin on AGM-S3 cells. Meanwhile, GMR CAR T cells exerted no effects on normal CD34(+) cell colony growth.
  
  
  CONCLUSIONS
  Ligand-based GMR CAR T cells may have anti-proliferative effects on stem and progenitor cells in JMML.
authors.affiliation: Department of Pediatrics, Shinshu University School of Medicine, 3-1-1, Asahi, Matsumoto, 390-8621, Japan.
authors.forename: Yozo
authors.initials: Y
authors.lastname: Nakazawa
authors.affiliation: Department of Laboratory Medicine, Shinshu University Hospital, Matsumoto, Japan.
authors.forename: Kazuyuki
authors.initials: K
authors.lastname: Matsuda
authors.affiliation: Department of Pediatrics, Shinshu University School of Medicine, 3-1-1, Asahi, Matsumoto, 390-8621, Japan.
authors.forename: Takashi
authors.initials: T
authors.lastname: Kurata
authors.affiliation: Department of Laboratory Medicine, Shinshu University Hospital, Matsumoto, Japan.
authors.forename: Akane
authors.initials: A
authors.lastname: Sueki
authors.affiliation: Department of Pediatrics, Shinshu University School of Medicine, 3-1-1, Asahi, Matsumoto, 390-8621, Japan.
authors.forename: Miyuki
authors.initials: M
authors.lastname: Tanaka
authors.affiliation: Department of Pediatrics, Shinshu University School of Medicine, 3-1-1, Asahi, Matsumoto, 390-8621, Japan.|Division of Hematology/Oncology, Nagano Children's Hospital, Azumino, Japan.
authors.forename: Kazuo
authors.initials: K
authors.lastname: Sakashita
authors.affiliation: Department of Pediatrics, Niigata University School of Medicine, Niigata, Japan.
authors.forename: Chihaya
authors.initials: C
authors.lastname: Imai
authors.affiliation: Department of Medicine, Division of Nephrology and Hypertension, Vanderbilt University School of Medicine, Nashville, TN, USA.
authors.forename: Matthew H
authors.initials: MH
authors.lastname: Wilson
authors.affiliation: Department of Pediatrics, Shinshu University School of Medicine, 3-1-1, Asahi, Matsumoto, 390-8621, Japan. koikeken@shinshu-u.ac.jp.
authors.forename: Kenichi
authors.initials: K
authors.lastname: Koike
mesh_terms: D018952:Antigens, CD34 / Q000235:genetics / Q000276:immunology*; D045744:Cell Line, Tumor; D049109:Cell Proliferation / Q000187:drug effects; D002478:Cells, Cultured; D018920:Coculture Techniques; D005434:Flow Cytometry; D006801:Humans; D016219:Immunotherapy, Adoptive; D020014:K562 Cells; D054429:Leukemia, Myelomonocytic, Juvenile / Q000235:genetics / Q000276:immunology* / Q000628:therapy; D008024:Ligands; D009154:Mutation; D011948:Receptors, Antigen, T-Cell / Q000235:genetics / Q000276:immunology*; D016187:Receptors, Granulocyte-Macrophage Colony-Stimulating Factor / Q000235:genetics / Q000276:immunology*; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000276:immunology; D019089:Stem Cell Factor / Q000494:pharmacology; D013601:T-Lymphocytes / Q000276:immunology* / Q000378:metabolism / Q000637:transplantation; D013926:Thrombopoietin / Q000494:pharmacology
chemical_list: D018952:Antigens, CD34; D008024:Ligands; D011948:Receptors, Antigen, T-Cell; D016187:Receptors, Granulocyte-Macrophage Colony-Stimulating Factor; D011993:Recombinant Fusion Proteins; D019089:Stem Cell Factor; D013926:Thrombopoietin
keywords: Chimeric antigen receptor; GM-CSF receptor; Juvenile myelomonocytic leukemia; Leukemic stem cells; piggyBac transposon