title: A simplified method for the clinical-scale generation of central memory-like CD8+ T cells after transduction with lentiviral vectors encoding antitumor antigen T-cell receptors.
abstract: Adoptive transfer of antigen-specific CD8+ T cells can effectively treat patients with metastatic melanoma. Recent efforts have emphasized the in vitro generation of antitumor T cells by transduction of genes encoding antitumor T-cell receptors. At present, lentiviral vector-mediated transduction of CD8+ T cells relies on anti-CD3/CD28 bead stimulation; however, this method fails to efficiently expand CD8+ T cells. Herein we sought to establish a methodology for lentiviral vector transduction using optimal activating agents for efficient gene delivery and robust expansion of CD8+ T cells. To overcome the inability of anti-CD3/CD28 beads to efficiently expand CD8+ T cells, we evaluated alternative activating agents including feeder cells from allogeneic peripheral blood mononuclear cells and plate-bound anti-CD3 antibody. Analyses of gene transfer, cell phenotype, fold expansion, and biologic activities were used to determine the optimal methodology. Plate-bound anti-CD3 provided an ideal activation platform that afforded optimal lentiviral vector-mediated gene transfer efficiency (up to 90%), and coupled with peripheral blood mononuclear cells feeder cells yielded up to 600-fold expansion of CD8+ T cells within 12 days. The T-cell antigen receptor (TCR) engineered CD8+ T cells conferred specific antitumor activity and many displayed a central memory-like phenotype. The methodology described here could be readily applied for engineering CD8+ T cells with antitumor specificity for human adoptive immunotherapy.
authors.affiliation: Surgery Branch, Center for Cancer Research, National Cancer Institute, National Institutes of Health, Bethesda, MD 20892, USA.
authors.forename: Shicheng
authors.initials: S
authors.lastname: Yang
authors.forename: Mark E
authors.initials: ME
authors.lastname: Dudley
authors.forename: Steven A
authors.initials: SA
authors.lastname: Rosenberg
authors.forename: Richard A
authors.initials: RA
authors.lastname: Morgan
mesh_terms: D000911:Antibodies, Monoclonal / Q000276:immunology / Q000378:metabolism; D000951:Antigens, Neoplasm / Q000276:immunology; D017252:CD3 Complex / Q000276:immunology; D018414:CD8-Positive T-Lymphocytes / Q000276:immunology / Q000378:metabolism* / Q000473:pathology; D049109:Cell Proliferation; D015316:Genetic Therapy / Q000379:methods; D005822:Genetic Vectors / Q000235:genetics; D006801:Humans; D007156:Immunologic Memory; D016130:Immunophenotyping; D016219:Immunotherapy, Adoptive; D016086:Lentivirus / Q000235:genetics*; D008213:Lymphocyte Activation / Q000235:genetics; D008545:Melanoma / Q000235:genetics / Q000276:immunology / Q000628:therapy*; D009362:Neoplasm Metastasis; D011948:Receptors, Antigen, T-Cell / Q000235:genetics / Q000276:immunology / Q000378:metabolism*; D014161:Transduction, Genetic
chemical_list: D000911:Antibodies, Monoclonal; D000951:Antigens, Neoplasm; D017252:CD3 Complex; D011948:Receptors, Antigen, T-Cell