The role of apoptosis in bispecific antibody-mediated T-cell cytotoxicity.
In this report we describe the role of apoptosis in the process of tumour cell killing by bispecific monoclonal antibody (BsMAb)-redirected cytolytic T cells. The BsMAb used, BIS-1, has dual specificity for the CD3 complex on T cells and the pancarcinoma-associated 38 kDa transmembrane antigen EGP-2. BIS-1 allows activated T cells to specifically recognise and kill EGP-2-positive but not EGP-2-negative target cells. An assay was developed to quantify apoptosis in cells by separation of 3H-thymidine-labelled low-molecular, i.e. fragmented, from high-molecular, i.e. non-fragmented DNA. The presence of low molecular weight DNA was measured both within the target cells and in the cell-free supernatant. After exposure to BIS-1-redirected, -activated T cells, apoptosis was observed in EGP-2-positive target cells but not in EGP-2-negative target cells. Also no DNA fragmentation proved to be induced in the activated effector cells during assay. The degree of EGP-2-positive target DNA fragmentation depended on the concentration of BsMAb, the E/T ratio and the incubation time. Using a low E/T ratio (1/1), DNA fragmentation in and 51Cr release from target cells showed similar characteristics and kinetics. At higher E/T ratio (20/1), the 51Cr release from the target cells increased to a greater extent than the percentage fragmented target cell DNA. Inhibitors of DNA fragmentation added to the cytotoxicity assay inhibited not only DNA fragmentation, but also the release of chromium-51 from the target cells, suggesting that apoptosis and cell lysis are closely related in BsMAb-mediated cell killing.
Kroesen
B J
BJ
Department of Clinical Immunology, University Hospital Groningen, The Netherlands.
Wellenberg
G J
GJ
Bakker
A
A
Helfrich
W
W
The
T H
TH
de Leij
L
L
D018033:Antibodies, Bispecific / Q000276:immunology / Q000494:pharmacology*; D000911:Antibodies, Monoclonal / Q000276:immunology / Q000494:pharmacology*; D000920:Antibody-Dependent Cell Cytotoxicity / Q000187:drug effects / Q000276:immunology*; D017209:Apoptosis / Q000187:drug effects / Q000502:physiology*; D004247:DNA / Q000378:metabolism; D004249:DNA Damage; D006801:Humans; D008213:Lymphocyte Activation; D013602:T-Lymphocytes, Cytotoxic / Q000187:drug effects / Q000276:immunology* / Q000378:metabolism
D018033:Antibodies, Bispecific; D000911:Antibodies, Monoclonal; D004247:DNA