title: Automated manufacturing of chimeric antigen receptor T cells for adoptive immunotherapy using CliniMACS prodigy.
abstract: Novel cell therapies derived from human T lymphocytes are exhibiting enormous potential in early-phase clinical trials in patients with hematologic malignancies. Ex vivo modification of T cells is currently limited to a small number of centers with the required infrastructure and expertise. The process requires isolation, activation, transduction, expansion and cryopreservation steps. To simplify procedures and widen applicability for clinical therapies, automation of these procedures is being developed. The CliniMACS Prodigy (Miltenyi Biotec) has recently been adapted for lentiviral transduction of T cells and here we analyse the feasibility of a clinically compliant T-cell engineering process for the manufacture of T cells encoding chimeric antigen receptors (CAR) for CD19 (CAR19), a widely targeted antigen in B-cell malignancies. Using a closed, single-use tubing set we processed mononuclear cells from fresh or frozen leukapheresis harvests collected from healthy volunteer donors. Cells were phenotyped and subjected to automated processing and activation using TransAct, a polymeric nanomatrix activation reagent incorporating CD3/CD28-specific antibodies. Cells were then transduced and expanded in the CentriCult-Unit of the tubing set, under stabilized culture conditions with automated feeding and media exchange. The process was continuously monitored to determine kinetics of expansion, transduction efficiency and phenotype of the engineered cells in comparison with small-scale transductions run in parallel. We found that transduction efficiencies, phenotype and function of CAR19 T cells were comparable with existing procedures and overall T-cell yields sufficient for anticipated therapeutic dosing. The automation of closed-system T-cell engineering should improve dissemination of emerging immunotherapies and greatly widen applicability.
authors.affiliation: Institute of Child Health, Molecular and Cellular Immunology Unit, University College London, London, UK.
authors.forename: Ulrike
authors.initials: U
authors.lastname: Mock
authors.affiliation: Institute of Child Health, Molecular and Cellular Immunology Unit, University College London, London, UK. Electronic address: l.nickoay@ucl.ac.uk.
authors.forename: Lauren
authors.initials: L
authors.lastname: Nickolay
authors.affiliation: University College London Cancer Institute, London, UK.
authors.forename: Brian
authors.initials: B
authors.lastname: Philip
authors.affiliation: University College London Cancer Institute, London, UK.
authors.forename: Gordon Weng-Kit
authors.initials: GW
authors.lastname: Cheung
authors.affiliation: Institute of Child Health, Molecular and Cellular Immunology Unit, University College London, London, UK.
authors.forename: Hong
authors.initials: H
authors.lastname: Zhan
authors.affiliation: Research & Development, Miltenyi Biotec GmbH, Bergisch Gladbach, Germany.
authors.forename: Ian C D
authors.initials: ICD
authors.lastname: Johnston
authors.affiliation: Research & Development, Miltenyi Biotec GmbH, Bergisch Gladbach, Germany.
authors.forename: Andrew D
authors.initials: AD
authors.lastname: Kaiser
authors.affiliation: University College London Cancer Institute, London, UK.
authors.forename: Karl
authors.initials: K
authors.lastname: Peggs
authors.affiliation: University College London Cancer Institute, London, UK.
authors.forename: Martin
authors.initials: M
authors.lastname: Pule
authors.affiliation: Institute of Child Health, Molecular and Cellular Immunology Unit, University College London, London, UK.
authors.forename: Adrian J
authors.initials: AJ
authors.lastname: Thrasher
authors.affiliation: Institute of Child Health, Molecular and Cellular Immunology Unit, University College London, London, UK.
authors.forename: Waseem
authors.initials: W
authors.lastname: Qasim
mesh_terms: D000818:Animals; D018941:Antigens, CD19 / Q000235:genetics / Q000276:immunology / Q000378:metabolism; D057205:Automation, Laboratory / Q000295:instrumentation / Q000379:methods; D001402:B-Lymphocytes / Q000276:immunology; D018929:Cell Culture Techniques / Q000295:instrumentation / Q000379:methods; D060846:Cell Engineering / Q000295:instrumentation / Q000379:methods; D049109:Cell Proliferation; D002469:Cell Separation / Q000379:methods; D002478:Cells, Cultured; D017076:Computer-Aided Design; D006801:Humans; D016219:Immunotherapy, Adoptive / Q000379:methods; D051379:Mice; D016688:Mice, Inbred NOD; D016513:Mice, SCID; D011948:Receptors, Antigen, T-Cell / Q000737:chemistry / Q000235:genetics / Q000276:immunology* / Q000378:metabolism; D011993:Recombinant Fusion Proteins / Q000737:chemistry / Q000235:genetics / Q000276:immunology* / Q000378:metabolism; D013601:T-Lymphocytes / Q000276:immunology* / Q000378:metabolism; D014161:Transduction, Genetic; D023041:Xenograft Model Antitumor Assays
chemical_list: D018941:Antigens, CD19; D011948:Receptors, Antigen, T-Cell; D011993:Recombinant Fusion Proteins
keywords: CAR19; CliniMACS prodigy; GMP; T cells; T-cell transduction; automation; chimeric antigen receptors; immunotherapy