title: Optimization of lentiviral vector transduction into peripheral blood mononuclear cells in combination with the fibronectin fragment CH-296 stimulation.
abstract: Large scale T-cell expansion and efficient gene transduction are required for adoptive T-cell gene therapy. Based on our previous observations, human peripheral blood mononuclear cells (PBMCs) can be expanded efficiently while conserving a naïve phenotype by stimulating with both recombinant human fibronectin fragment (CH-296) and anti-CD3 monoclonal antibodies. In this article, we explored the possibility of using this co-stimulation method to generate engineered T cells using lentiviral vector. Human PBMCs were stimulated with anti-CD3 together with immobilized CH-296 or anti-CD28 antibody as well as anti-CD3/anti-CD28 conjugated beads and transduced with lentiviral vector simultaneously. Co-stimulation with CH-296 gave superior transduction efficiency than with anti-CD28. Next, PBMCs were stimulated and transduced with anti-CD3/CH-296 or with anti-CD3/CD28 beads. T-cell expansion, gene transfer efficiencies and immunophenotypes were analysed. Stimulation with anti-CD3/CH-296 resulted in more than 10-times higher cell expansion and higher gene transfer efficiency with conservation of the naïve phenotype compared with anti-CD3/CD28 stimulation method. Thus, lentiviral transduction with anti-CD3/CH-296 co-stimulation is an efficient way to generate large numbers of genetically modified T cells and may be suitable for many gene therapy protocols that use adoptive T-cell transfer therapy.
authors.affiliation: Center for Cell and Gene Therapy, Takara Bio Inc. Seta 3-4-1, Otsu, Shiga, Japan.
authors.forename: Hideto
authors.initials: H
authors.lastname: Chono
authors.forename: Yumi
authors.initials: Y
authors.lastname: Goto
authors.forename: Satoko
authors.initials: S
authors.lastname: Yamakawa
authors.forename: Shinya
authors.initials: S
authors.lastname: Tanaka
authors.forename: Yasuhiro
authors.initials: Y
authors.lastname: Tosaka
authors.forename: Ikuei
authors.initials: I
authors.lastname: Nukaya
authors.forename: Junichi
authors.initials: J
authors.lastname: Mineno
mesh_terms: D000911:Antibodies, Monoclonal / Q000276:immunology / Q000494:pharmacology; D018106:CD28 Antigens / Q000276:immunology; D017252:CD3 Complex / Q000276:immunology; D005353:Fibronectins / Q000737:chemistry / Q000494:pharmacology*; D005434:Flow Cytometry; D005822:Genetic Vectors / Q000235:genetics*; D006801:Humans; D016219:Immunotherapy, Adoptive; D016086:Lentivirus / Q000235:genetics*; D007963:Leukocytes, Mononuclear / Q000166:cytology / Q000187:drug effects* / Q000378:metabolism*; D008213:Lymphocyte Activation / Q000187:drug effects; D013601:T-Lymphocytes / Q000166:cytology / Q000187:drug effects / Q000378:metabolism; D014161:Transduction, Genetic / Q000379:methods*
chemical_list: D000911:Antibodies, Monoclonal; D018106:CD28 Antigens; D017252:CD3 Complex; D005353:Fibronectins