Background Trifunctional antibodies, such as catumaxomab (anti-EpCAManti-CD3) and ertumaxomab (anti-HER-2/neuanti-CD3), transiently link immune effector cells to tumour cells, which results in cellular cytotoxicity towards the tumour cells. A functional immune system is therefore essential for effective anti-tumour activity. However, the commonly observed haematotoxicity of chemotherapeutics and radiation therapy may be associated with some degree of immunosuppression. Combining chemotherapy and trifunctional antibodies in cancer treatment requires understanding of the impact of chemotherapeutics on immune cell function and, thus, on the activity of trifunctional antibodies. Methods The effect of chemotherapeutic treatment on trifunctional antibody-mediated anti-tumour activity was assessed in vitro. Blood samples were collected from 12 head and neck squamous cell carcinoma patients after chemotherapy (5-fl uorouracil, cisplatin) and radiotherapy, and from one healthy control donor. The immune cell status was analysed and mononuclear cells (MNC) were isolated. The potency of catumaxomab and ertumaxomab was assessed in a cytotoxicity assay using MNC isolated from each patient sample in co-culture with a tumour target cell line. The release of infl ammatory cytokines was also monitored in the cell culture supernatant. Results Most patients included in this study had decreased immune cell counts during the course of chemotherapy. Nonetheless, an effective and concentration-dependent anti-tumour activity mediated by trifunctional antibodies was demonstrated using these patient immune effector cells. The immune response activity of the patient immune cells was not impaired one week after cisplatin administration or even three days after the last 5-fl uorouracil treatment. Conclusion This study shows for the fi rst time that immune effector cells from cancer patients undergoing standard chemotherapy and radiotherapy can be activated by trifunctional antibodies for effi cient killing of tumour cells.
Introduction In patients, a transient decrease in peripheral blood lymphocyte counts was observed following intraperitoneal administration of the trifunctional monoclonal antibody catumaxomab (anti-human EpCAM x anti-human CD3). The aim of this study was to clarify the observed effect in a preclinical mouse model and to analyse the related mechanism of action in vitro. Materials and methods A related antibody, BiLu (antihuman EpCAM x anti-mouse CD3), was administered to mice and blood leukocytes were analysed. In vitro studies measured activation and cytokine secretion from human peripheral blood mononuclear cells (PBMC). For the analysis of T cell adhesion, PBMC were preincubated with catumaxomab and then co-cultured with human endothelial cells (HUVEC); T cell adhesion was assessed in the presence or absence of endothelial cell preactivation by TNF. Adherent T cells were determined by fl ow cytometry. Results Treatment of mice with BiLu resulted in a dosedependent transient decrease in CD3+ T cells (both CD4+ and CD8+) that returned to the normal range within 48 h. Catumaxomab physiologically activated T cells in vitro (increased CD69 expression) and induced cytokine release (TNF, IFN). TNF increased expression of adhesion molecules CD54 and CD62E on endothelial cells. Furthermore, catumaxomab dose-dependently enhanced adhesion of T cells to endothelial cells. Adhesion was further increased when endothelial cells were preactivated with TNF.Conclusions Catumaxomab increases adhesion of T cells to endothelial cells due to antibody-mediated activation of T cells and production of T cell cytokines that up-regulate endothelial cell adhesion molecules. These results provide a mechanistic rationale for the transient, reversible decrease in lymphocyte counts observed following catumaxomab administration in patients, which is likely to be due to redistribution of lymphocytes.
Surgical intervention and radiotherapy still represent the gold standard in the therapy of head and neck squamous cell carcinoma (SCC), although often with unsatisfactory results. Radiation might induce the expression and secretion of vascular endothelial growth factor (VEGF) and basic fibroblast growth factor (bFGF) by unknown mechanisms. These two highly active proangiogenic and cytoprotective factors might contribute to a limited therapeutic success by promoting revascularization and cytoprotection of the radiated tumor. The aim of the present study was to analyze the potential of the cyclooxygenase inhibitor flurbiprofen to reduce radiation-induced increase of VEGF and bFGF secretion of tumor cells. We analyzed the expression of VEGF and bFGF at 72 h after radiation with 30 Gy in four SCC cell lines (De-pt, Hun, Lau, and A549) in cell culture with or without added flurbiprofen. Controls were not exposed to radiation and were analyzed at the same time after culture in the same media. We observed increased VEGF levels in all and increased bFGF levels in three of four lines after radiation. In irradiated cultures with flurbiprofen, VEGF was reduced between 13% and 26% and bFGF was reduced between 84% and 93% compared with radiated cultures without flurbiprofen. We found no reduction of VEGF and bFGF secretion in the unirradiated cultures despite added flurbiprofen. We conclude that flurbiprofen is able to alter the radiation-induced secretion of these two growth factors and might be useful in decreasing the resistance of SCC to radiation.
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