title: Generation of autologous tumor-specific T cells for adoptive transfer based on vaccination, in vitro restimulation and CD3/CD28 dynabead-induced T cell expansion.
abstract: Adoptive cell transfer (ACT) of in vitro expanded autologous tumor-infiltrating lymphocytes (TIL) has been shown to exert therapeutic efficacy in melanoma patients. We aimed to develop an ACT protocol based on tumor-specific T cells isolated from peripheral blood and in vitro expanded by Dynabeads® ClinExVivo™CD3/CD28. We show here that the addition of an in vitro restimulation step with relevant peptides prior to bead expansion dramatically increased the proportion of tumor-specific T cells in PBMC-cultures. Importantly, peptide-pulsed dendritic cells (DCs) as well as allogeneic tumor lysate-pulsed DCs from the DC vaccine preparation could be used with comparable efficiency to peptides for in vitro restimulation, to increase the tumor-specific T-cell response. Furthermore, we tested the use of different ratios and different types of Dynabeads® CD3/CD28 and CD3/CD28/CD137 T-cell expander, for optimized expansion of tumor-specific T cells. A ratio of 1:3 of Dynabeads® CD3/CD28 T-cell expander to T cells resulted in the maximum number of tumor-specific T cells. The addition of CD137 did not improve functionality or fold expansion. Both T-cell expansion systems could generate tumor-specific T cells that were both cytotoxic and effective cytokine producers upon antigen recognition. Dynabeads®-expanded T-cell cultures shows phenotypical characteristics of memory T cells with potential to migrate and expand in vivo. In addition, they possess longer telomeres compared to TIL cultures. Taken together, we demonstrate that in vitro restimulation of tumor-specific T cells prior to bead expansion is necessary to achieve high numbers of tumor-specific T cells. This is effective and easily applicable in combination with DC vaccination, by use of vaccine-generated DCs, either pulsed with peptide or tumor-lysate.
authors.affiliation: Department of Hematology 54P4, Center for Cancer Immune Therapy, University Hospital Herlev, Herlev, Denmark.
authors.forename: Marie Klinge
authors.initials: MK
authors.lastname: Brimnes
authors.forename: Anne Ortved
authors.initials: AO
authors.lastname: Gang
authors.forename: Marco
authors.initials: M
authors.lastname: Donia
authors.forename: Per
authors.initials: P
authors.lastname: Thor Straten
authors.forename: Inge Marie
authors.initials: IM
authors.lastname: Svane
authors.forename: Sine Reker
authors.initials: SR
authors.lastname: Hadrup
mesh_terms: D018106:CD28 Antigens / Q000276:immunology / Q000378:metabolism; D017252:CD3 Complex / Q000276:immunology / Q000378:metabolism; D019496:Cancer Vaccines / Q000276:immunology / Q000378:metabolism; D018929:Cell Culture Techniques / Q000379:methods*; D002986:Clinical Trials as Topic; D005434:Flow Cytometry; D006801:Humans; D016219:Immunotherapy, Adoptive / Q000379:methods*; D013601:T-Lymphocytes / Q000166:cytology* / Q000276:immunology / Q000378:metabolism
chemical_list: D018106:CD28 Antigens; D017252:CD3 Complex; D019496:Cancer Vaccines