title: Genetically engineered fixed K562 cells: potent "off-the-shelf" antigen-presenting cells for generating virus-specific T cells.
abstract: OBJECTIVE
  The human leukemia cell line K562 represents an attractive platform for creating artificial antigen-presenting cells (aAPC). It is readily expandable, does not express human leukocyte antigen (HLA) class I and II and can be stably transduced with various genes.
  
  
  METHODS
  In order to generate cytomegalovirus (CMV) antigen-specific T cells for adoptive immunotherapy, we transduced K562 with HLA-A∗0201 in combination with co-stimulatory molecules.
  
  
  RESULTS
  In preliminary experiments, irradiated K562 expressing HLA-A∗0201 and 4-1BBL pulsed with CMV pp65 and IE-1 peptide libraries failed to elicit antigen-specific CD8⁺ T cells in HLA-A∗0201⁺ peripheral blood mononuclear cells (PBMC) or isolated T cells. Both wild-type K562 and aAPC strongly inhibited T cell proliferation to the bacterial superantigen staphylococcal enterotoxin B (SEB) and OKT3 and in mixed lymphocyte reaction (MLR). Transwell experiments suggested that suppression was mediated by a soluble factor; however, MLR inhibition was not reversed using transforming growth factor-β blocking antibody or prostaglandin E2 inhibitors. Full abrogation of the suppressive activity of K562 on MLR, SEB and OKT3 stimulation was only achieved by brief fixation with 0.1% formaldehyde. Fixed, pp65 and IE-1 peptide-loaded aAPC induced robust expansion of CMV-specific T cells.
  
  
  CONCLUSIONS
  Fixed gene-modified K562 can serve as effective aAPC to expand CMV-specific cytotoxic T lymphocytes for therapeutic use in patients after stem cell transplantation. Our findings have implications for broader understanding of the immune evasion mechanisms used by leukemia and other tumors.
authors.affiliation: Stem Cell Allogeneic Transplantation Section, Hematology Branch, National Heart, Lung and Blood Institute, National Institutes of Health, Bethesda, Maryland, USA. Electronic address: kazshi.tanimoto@gmail.com.
authors.forename: Kazushi
authors.initials: K
authors.lastname: Tanimoto
authors.affiliation: Stem Cell Allogeneic Transplantation Section, Hematology Branch, National Heart, Lung and Blood Institute, National Institutes of Health, Bethesda, Maryland, USA.
authors.forename: Pawel
authors.initials: P
authors.lastname: Muranski
authors.affiliation: Stem Cell Allogeneic Transplantation Section, Hematology Branch, National Heart, Lung and Blood Institute, National Institutes of Health, Bethesda, Maryland, USA.
authors.forename: Samantha
authors.initials: S
authors.lastname: Miner
authors.affiliation: Department of Bioregulatory Medicine, Ehime University Graduate School of Medicine, Ehime, Japan.
authors.forename: Hiroshi
authors.initials: H
authors.lastname: Fujiwara
authors.affiliation: Stem Cell Allogeneic Transplantation Section, Hematology Branch, National Heart, Lung and Blood Institute, National Institutes of Health, Bethesda, Maryland, USA.
authors.forename: Sachiko
authors.initials: S
authors.lastname: Kajigaya
authors.affiliation: Stem Cell Allogeneic Transplantation Section, Hematology Branch, National Heart, Lung and Blood Institute, National Institutes of Health, Bethesda, Maryland, USA.
authors.forename: Keyvan
authors.initials: K
authors.lastname: Keyvanfar
authors.affiliation: Stem Cell Allogeneic Transplantation Section, Hematology Branch, National Heart, Lung and Blood Institute, National Institutes of Health, Bethesda, Maryland, USA.
authors.forename: Nancy
authors.initials: N
authors.lastname: Hensel
authors.affiliation: Stem Cell Allogeneic Transplantation Section, Hematology Branch, National Heart, Lung and Blood Institute, National Institutes of Health, Bethesda, Maryland, USA.
authors.forename: A John
authors.initials: AJ
authors.lastname: Barrett
authors.affiliation: Stem Cell Allogeneic Transplantation Section, Hematology Branch, National Heart, Lung and Blood Institute, National Institutes of Health, Bethesda, Maryland, USA; Abramson Cancer Center, Department of Pathology and Laboratory Medicine, Perelman School of Medicine, University of Pennsylvania, Philadelphia, Pennsylvania, USA.
authors.forename: J Joseph
authors.initials: JJ
authors.lastname: Melenhorst
mesh_terms: D000938:Antigen-Presenting Cells / Q000276:immunology*; D018414:CD8-Positive T-Lymphocytes / Q000276:immunology; D003586:Cytomegalovirus Infections / Q000276:immunology* / Q000628:therapy; D005818:Genetic Engineering; D006680:HLA Antigens / Q000276:immunology; D006801:Humans; D020014:K562 Cells; D007963:Leukocytes, Mononuclear / Q000276:immunology*; D008213:Lymphocyte Activation / Q000276:immunology; D013602:T-Lymphocytes, Cytotoxic / Q000276:immunology
chemical_list: D006680:HLA Antigens
keywords: artificial APC; cytomegalovirus; cytotoxic T cell; fixation