title: Selection of adenovirus-specific and Epstein-Barr virus-specific T cells with major histocompatibility class I streptamers under Good Manufacturing Practice (GMP)-compliant conditions.
abstract: OBJECTIVE
  Despite antiviral drug therapies, human adenovirus (HAdV), cytomegalovirus (CMV) and Epstein-Barr virus (EBV) infections still contribute substantially to transplant-related death of patients after hematopoietic stem cell transplantation. Earlier clinical studies demonstrated successful adoptive transfer of magnetically selected CMV-specific T cells via removable, and thus Good Manufacturing Practice-compliant, major histocompatibility class I streptamers. Thus, the primary focus of the present study was the selection of HAdV-streptamer+ T cells, although in three experiments, EBV-streptamer+ T cells were also selected.
  
  
  METHODS
  Cells from leukaphereses of healthy donors were prepared in large (1-6 × 10(9)) and small (25 × 10(6)) cell batches. Whereas the larger batch was directly labeled with streptamers to select HAdV- and/or EBV-specific T cells (large-scale), the smaller batch was used to generate in vitro virus-specific T-cell lines before streptamer labeling for streptamer selection (small-scale). Isolation of HAdV- and/or EBV-specific T cells was performed with the use of the CliniMACS device.
  
  
  RESULTS
  The purity of HAdV- and EBV-streptamer+ T cells among CD3+ cells, obtained from large-scale selection, was up to 6.7% and 44%, respectively. If HAdV- and EBV-streptamers were applied simultaneously, the purity of antigen-specific T cells reached up to 50.7%. A further increase in purity reaching up to 98% was achieved by small-scale selection of HAdV-specific T cells. All final products fulfilled the microbiological and chemical release criteria. Interferon-γ-response indicating functional activity was seen in 6 of 9 HAdV and 2 of 3 EBV large-scale selections and in 2 of 3 HAdV small-scale selections.
  
  
  CONCLUSIONS
  HAdV-streptamers were shown to be clinically feasible for few patients after the large-scale approach but for larger patient numbers if combined with EBV-streptamers or after the small-scale approach.
authors: Department of Clinical Cell Biology and FACS Core Unit, Children's Cancer Research Institute (CCRI), Vienna, Austria.
authors: Christine
authors: C
authors: Freimüller
authors: Department of Clinical Cell Biology and FACS Core Unit, Children's Cancer Research Institute (CCRI), Vienna, Austria.
authors: Julia
authors: J
authors: Stemberger
authors: Department of Quality Management, CCRI, Vienna, Austria.
authors: Michaela
authors: M
authors: Artwohl
authors: Stage Cell Therapeutics, Göttingen, Germany.
authors: Lothar
authors: L
authors: Germeroth
authors: St. Anna Children's Hospital, Vienna, Austria.
authors: Volker
authors: V
authors: Witt
authors: Department of Blood Group Serology and Transfusion Medicine, Medical University of Vienna, Vienna, Austria.
authors: Gottfried
authors: G
authors: Fischer
authors: Institute of Transfusion Medicine, Hannover Medical School, Hannover, Germany.
authors: Sabine
authors: S
authors: Tischer
authors: Institute of Transfusion Medicine, Hannover Medical School, Hannover, Germany.
authors: Britta
authors: B
authors: Eiz-Vesper
authors: Department of Immune Modulation at the Department of Dermatology, Universitätsklinikum Erlangen, Germany.
authors: Ilka
authors: I
authors: Knippertz
authors: Department of Dermatology, Universitätsklinikum Erlangen, Erlangen, Germany.
authors: Jan
authors: J
authors: Dörrie
authors: Department of Dermatology, Universitätsklinikum Erlangen, Erlangen, Germany.
authors: Niels
authors: N
authors: Schaft
authors: Department Pediatrics, Medical University of Vienna, Vienna, Austria; Department of Molecular Microbiology, CCRI, Vienna, Austria.
authors: Thomas
authors: T
authors: Lion
authors: Department of Clinical Cell Biology and FACS Core Unit, Children's Cancer Research Institute (CCRI), Vienna, Austria; Department Pediatrics, Medical University of Vienna, Vienna, Austria.
authors: Gerhard
authors: G
authors: Fritsch
authors: Department of Clinical Cell Biology and FACS Core Unit, Children's Cancer Research Institute (CCRI), Vienna, Austria. Electronic address: rene.geyeregger@ccri.at.
authors: René
authors: R
authors: Geyeregger
mesh_terms: D000257:Adenoviridae Infections / Q000276:immunology; D000260:Adenoviruses, Human / Q000276:immunology*; D019264:Adoptive Transfer; D003587:Cytomegalovirus / Q000276:immunology*; D003586:Cytomegalovirus Infections / Q000276:immunology; D020031:Epstein-Barr Virus Infections / Q000276:immunology; D018380:Hematopoietic Stem Cell Transplantation / Q000009:adverse effects; D004854:Herpesvirus 4, Human / Q000276:immunology*; D015395:Histocompatibility Antigens Class I / Q000276:immunology; D006801:Humans; D007371:Interferon-gamma / Q000276:immunology; D008213:Lymphocyte Activation / Q000276:immunology; D008297:Male; D013194:Staining and Labeling / Q000379:methods*; D013601:T-Lymphocytes / Q000276:immunology*
chemical_list: D015395:Histocompatibility Antigens Class I; D007371:Interferon-gamma
keywords: Good Manufacturing Practice; HAdV-specific T cells; MHC streptamer technology; hematopoietic stem cell transplantation