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Supplementary Movie 2 from Committing Cytomegalovirus-Specific CD8 T Cells to Eliminate Tumor Cells by Bifunctional Major Histocompatibility Class I Antibody Fusion Molecules

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posted on 2023-04-03, 23:09 authored by Martina Schmittnaegel, Victor Levitsky, Eike Hoffmann, Guy Georges, Olaf Mundigl, Christian Klein, Hendrik Knoetgen

Time-lapse fluorescence microscopy of CMV-pp65-specific T cell mediated killing of MCSP+ tumor cells (Colo38) incubated with 15nM of EBV-pMHCI-IgG as control. Constructs are indicated in the respective panel. T cells are stained in red (PKH-26, Sigma), tumor cells in green (CMFDA, LifeTechnologies). Effector to target ratio was 3:1. Real-time is indicated in the lower left corner in h:m:s:ms (h=hours, m=minutes, s=seconds, ms=milliseconds), scale bar is shown in the lower right corner 0-50 µm (µm=micrometre).

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ARTICLE ABSTRACT

Tumor cells escape immune eradication through multiple mechanisms, including loss of antigenicity and local suppression of effector lymphocytes. To counteract these obstacles, we aimed to direct the unique cytomegalovirus (CMV)-specific immune surveillance against tumor cells. We developed a novel generation of fusion proteins composed of a tumor antigen–specific full immunoglobulin connected to a single major histocompatibility class I complex bearing a covalently linked virus-derived peptide (pMHCI–IgG). Here, we show that tumor antigen–expressing cancer cells, which are decorated with pMHCI–IgGs containing a HLA-A*0201 molecule associated with a CMV-derived peptide, are specifically eliminated through engagement of antigen-specific CD8+ T cells isolated from peripheral blood mononuclear cell preparations of CMV-infected humans. These CD8+ T cells act without additional expansion, preactivation, or provision of costimulatory signals. Elimination of tumor cells is induced at similar concentrations and with similar time kinetics as those seen with bispecific T-cell engagers (BiTE). However, while BiTE-like reagents indiscriminately activate T cells through binding to the T-cell receptor complex, pMHCI–IgGs selectively engage antigen-specific, constantly renewable, differentiated effector cytotoxic T lymphocytes to tumor cells, thereby representing a novel class of anticancer immunotherapeutics with potentially improved safety and efficacy profiles. Cancer Immunol Res; 3(7); 764–76. ©2015 AACR.

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