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Epithelial cell adhesion molecule EpCAM is a transmembrane glycoprotein that is frequently overexpressed in a variety of carcinomas. This pan-carcinoma antigen has served as the target for a plethora of immunotherapies. Innovative therapeutic approaches include the use of trifunctional antibodies (trAbs) that recruit and activate different types of immune effector cells at the tumour site. The trAb catumaxomab has dual specificity for EpCAM and CD3. In patients with malignant ascites, catumaxomab significantly increased the paracentesis-free interval, corroborating the high efficacy of this therapeutic antibody. Here, we characterised the monoclonal antibody (mAb) HO-3, that is, the EpCAM-binding arm of catumaxomab. Peptide mapping indicated that HO-3 recognises a discontinuous epitope, having three binding sites in the extracellular region of EpCAM. Studies with glycosylation-deficient mutants showed that mAb HO-3 recognised EpCAM independently of its glycosylation status. High-affinity binding was not only detected for mAb HO-3, but also for the monovalent EpCAM-binding arm of catumaxomab with an excellent K(D) of 5.6 x 10(-10) M. Furthermore, trAb catumaxomab was at least a 1000-fold more effective in eliciting the eradication of tumour cells by effector peripheral blood mononuclear cells compared with mAb HO-3. These findings suggest the great therapeutic potential of trAbs and clearly speak in favour of EpCAM-directed cancer immunotherapies.

Background Catumaxomab, the approved therapy for treatment of malignant ascites, exerts its anti-tumor effects via T-cell-mediated lysis, antibody-dependent cell-mediated cytotoxicity, and phagocytosis via activation of FcγR-positive accessory cells. It has been reported that FcγR-polymorphisms influence the IgG binding capacity of the receptor and that ethnic groups may differ with respect to responsiveness of immune effector cells to antibody-based therapies. The present nonclinical study therefore investigated potential inter-ethnic differences (Caucasian vs. Japanese, Korean, Black African and Hispanic Donors) and FcγR-gene polymorphisms with regard to the potency of catumaxomab in vitro. Methods Blood samples of healthy subjects of different ethnic background (25 donors of Caucasian, Korean, Black African and Hispanic ethnicity and 15 Japanese donors) were collected for comparative analyses. PBMCs were purified from whole blood samples and used as effector cells evaluating catumaxomab-mediated cytotoxicity towards EpCAM+ tumor target cells (HCT-8; colon) in vitro. Furthermore, the samples were genotyped for FcγR IIa and FcγR IIIa gene polymorphisms by PCR. Results The comparative analysis of catumaxomab-mediated cytotoxicity obtained for the individual ethnic subgroups did not reveal major inter-ethnic differences although genetic polymorphisms for FcγR IIa/IIIa had been identified among donors. Even though a wider variability was observed among Caucasians, the potency range observed with immune effector cells of Japanese, Korean, Black African and Hispanic was found to be within the potency range observed for the Caucasian ethnicity. Conclusions No impact of ethnic background or FcγRIIa/IIIa polymorphism on the immune cell-mediated pharmacological activity of the trifunctional antibody catumaxomab could be identified in vitro. Therefore, it can be assumed that patients with a different ethnic background may also benefit from catumaxomab therapy as already demonstrated for Caucasian patients. Disclosure F. Marme: consultant for Fresenius Biotech GmbH. K. Hasegawa: financial support for clinical studies from Fresenius Biotech GmbH. H. Martinius: works for Fresenius Biotech GmbH. J. Atz: works for Fresenius Biotech GmbH. K. Fujiwara: financial support for clinical studies from Fresenius Biotech GmbH.

e13031 Background: The anti-tumor activity of the trifunctional antibody catumaxomab (anti-EpCAM x anti-CD3) relies on the activation of immune effector cells against EpCAM+ tumor cells. It has bee...

e13043 Background: The trifunctional antibody catumaxomab (anti-EpCAM x anti-CD3) is approved in the EU for the intraperitoneal treatment of malignant ascites. Since chemotherapy may impair the fun...