Use of Anti-CD3 × 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 × 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-γ (IFN-γ), tumor necrosis factor-α (TNF-α), and granulocyte-macrophage colony-stimulating factor(GM-CSF) secreted was 1699, 922, and 3092 pg/ml/10⁶ 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.