15353-j-veluchamy

24 | Chapter 1 Scope of this thesis In this thesis, we explore the potential benefits of allogeneic NK cells either alone or in combination with the therapeutic monoclonal antibody cetuximab in the fight against solid cancer. We set out to study NK cell killing mechanisms and the effect of ADCC in solid epithelial malignancies, functionally comparing two clinically applicable NK cell products, i.e. peripheral blood NK cells (PBNK) and umbilical cord blood stem cell derived NK cells (UCB-NK), against epidermoid, colorectal and cervical tumors. In chapter 2 we report on the development of two standardized eight color NK cell specific flowcytometry panels that enable monitoring of NK cell subset frequency, functionality, and phenotype in peripheral blood mononuclear cells under cytokine activated/non-activated and target cell stimulated/non-stimulated conditions in both fresh and cryopreserved samples. Using these standardized NK cell flowcytometry panels we conclude that cryopreserved PBMC samples are optimal for the assessment of NK cell functionality and NK cell receptor expression studies across multiple centers in both healthy donors and cancer patients. In advanced cervical cancer, targeted intervention therapies have limited success in controlling tumor growth. About 80% of the cervical cancer cases express epidermal growth factor receptor (EGFR), thus making EGFR an attractive surface antigen to target cervical tumors. However, treatment with the anti-EGFR monoclonal antibody cetuximab failed to demonstrate therapeutic efficacy. Further, apparent down-regulation of HLA class-I surface expression levels also makes tumor cells escape T cell mediated killing. In these cases, NK cell-based therapies may offer a viable alternative to T cell-based approaches. Therefore, in this study, the anti-tumor effects of two different allogeneic NK cell products, i.e. UCB-NK cells and activated PBNK cells, were studied either alone or in combination with the anti- EGFR mAb cetuximab in cervical cancer (Chapter 3). Similarly to cervical cancer, cetuximab monotherapy was also proven ineffective in EGFR + RAS mut advanced colorectal cancer (CRC). About 40-50% of CRC patients have mutations in the RAS gene, thus leaving nearly half of the CRC patients with no standard treatment option. In the study described in Chapter 4, we tested whether activated allogeneic peripheral blood NK cells (PBNK) together with cetuximab could overcome these limitations. In most of the patients with advanced CRC the NK cell response is highly impaired, as previously reported 81,82 . These findings were further substantiated by our data, the results of which revealed that CRC patients NK cell percentages and functionality were significantly lower than their healthy counterparts with a further reduction post-chemotherapy. In Chapter 5 we therefore investigated whether adoptively transferred allogeneic UCB-NK cells could effectively target colon cancer cells in vivo. Altogether, our in vitro and in vivo