Biologic properties of a bispecific single-chain antibody directed against 17-1A (EpCAM) and CD3: tumor cell-dependent T cell stimulation and cytotoxic activity.

Abstract

Anti-CD3 x anti-tumor-bispecific Abs have been used to redirect cytotoxic T cells to tumor cells in an MHC-unrestricted fashion and to induce their rejection in vivo. We have recently described a recombinant bispecific single-chain Ab that combines four different V regions of two Abs, anti-17-1A and anti-CD3, on one polypeptide chain. It folds correctly to a 60-kDa globular protein and is secreted in fully functional form by a high producer Chinese hamster ovary cell line. In this work, we report that its remarkable cytotoxicity against 17-1A+ tumor cells is exerted via T cells without an apparent engagement of a detectable costimulatory pathway. T cells are activated only by the bispecific Ab when coincubated with 17-1A+ target cells. In a chromium release assay, CD8+ T cells reach maximal tumor cell cytotoxicity within 4 h, while CD4+ T cells need about 20 h to reach similar levels of cytotoxicity. Addition of costimulatory CD28 Abs did not lead to a further increase in cytotoxicity. Its remarkable stability at 37 degrees in serum, the ease of production, and purification by affinity chromatography via polyhistidine tail make this smaller version of a bispecific Ab a promising candidate for a therapeutic trial in patients with solid tumor. Because adjuvant therapy with an intact, much less cytotoxic IgG2a Ab against the 17-1A target had already increased the 7-yr survival of colorectal cancer patients by 30%, the presented small bispecific construct lacking the immunogenic murine Fc region as well as autochthonous T lymphocyte stimulatory activity warrants a therapeutic trial in patients with minimal residual 17-1A+ cancer.