title: Use of anti-CD3 x anti-HER2/neu bispecific antibody for redirecting cytotoxicity of activated T cells toward HER2/neu+ tumors.
abstract: Relapse after adjuvant chemotherapy or high-dose chemotherapy with stem cell transplant for high-risk breast cancer remains high and new strategies that provide additional antitumor effects are needed. This report describes methods to generate highly effective HER2/neu-specific cytotoxic T cells by arming activated T cells with anti-CD3 x anti-HER2/neu bispecific antibody (BsAb). OKT3 and 9184 (anti-HER2) monoclonal antibodies (mAb) were conjugated and used to arm T cells that were subsequently tested in binding, cytotoxicity, and cytokine secretion assays. Armed T cells aggregated and specifically killed HER2/neu(+) breast cancer cells. Cytotoxicity emerged after 6 days of culture, was higher in armed T cells than unarmed T cells at all effector to target ratios (E/T) tested, and increased as the arming dose was increased. At an E/T of 20:1, the mean cytotoxicity of armed activated T cells (ATC) from 10 normal subjects increased by 59 +/- 11% (+/-SD) over that seen in unarmed ATC (p < 0.001) and the mean cytotoxicity of armed ATC from 6 cancer patients increased by 32 +/- 9% above that seen for unarmed ATC (p < 0.0004). After arming, the BsAb persisted on ATC up to 72 h and armed ATC continued to be cytotoxic up to 54 h. The amount of interferon-gamma (IFN-gamma), tumor necrosis factor-alpha (TNF-alpha), and granulocyte-macrophage colony-stimulating factor (GM-CSF) secreted was 1699, 922, and 3092 pg/ml/10(6) cells per 24 h, respectively, when armed T cells were exposed to a HER2/neu(+) breast carcinoma cell line. These studies show the feasibility and clinical adaptability of this approach for generating large numbers of anti-HER2-specific, cytotoxic T cells for clinical trials.
authors.affiliation: Blood Center of Southeastern Wisconsin, Milwaukee, WI 53201, USA.
authors.forename: M
authors.initials: M
authors.lastname: Sen
authors.forename: D M
authors.initials: DM
authors.lastname: Wankowski
authors.forename: N K
authors.initials: NK
authors.lastname: Garlie
authors.forename: R E
authors.initials: RE
authors.lastname: Siebenlist
authors.forename: D
authors.initials: D
authors.lastname: Van Epps
authors.forename: A V
authors.initials: AV
authors.lastname: LeFever
authors.forename: L G
authors.initials: LG
authors.lastname: Lum
mesh_terms: D018033:Antibodies, Bispecific / Q000633:toxicity*; D001943:Breast Neoplasms / Q000276:immunology / Q000473:pathology; D002470:Cell Survival / Q000187:drug effects; D003602:Cytotoxicity, Immunologic; D005260:Female; D016178:Granulocyte-Macrophage Colony-Stimulating Factor / Q000096:biosynthesis; D006801:Humans; D007371:Interferon-gamma / Q000096:biosynthesis; D008213:Lymphocyte Activation; D016853:Muromonab-CD3 / Q000276:immunology*; D018719:Erb-b2 Receptor Tyrosine Kinases / Q000276:immunology*; D013601:T-Lymphocytes / Q000276:immunology*; D014407:Tumor Cells, Cultured; D014409:Tumor Necrosis Factor-alpha / Q000096:biosynthesis
chemical_list: D018033:Antibodies, Bispecific; D016178:Granulocyte-Macrophage Colony-Stimulating Factor; D007371:Interferon-gamma; D016853:Muromonab-CD3; D018719:Erb-b2 Receptor Tyrosine Kinases; D014409:Tumor Necrosis Factor-alpha