15353-j-veluchamy

7 Summary | 165 induce effective killing of colon cancer cells irrespective of EGFR expression levels and RAS status. PBNK induced killing of EGFR + cells was significantly higher when colon cancer cells were coated with cetuximab, mediating ADCC. Importantly, PBNK cells were also highly cytotoxic to EGFR - colon cancer cells, which will obviously not respond to cetuximab therapy. The superior cytolysis observed for both cell lines and primary colon tumors, with variations in EGFR expression levels and RAS mutation status, indicates the potential of combined PBNK and cetuximab application in the treatment of cetuximab-resistant colon cancer. In Chapter 5, we monitored the frequency and function of NK cells in mCRC patients before and after the first cycle of chemotherapy, and found that mCRC patients not only had a 20% lower frequency of NK cells in peripheral blood before the initiation of chemotherapy, but that this percentage further declined during chemotherapy. In addition to this quantitative defect in NK cells of mCRC patients, their cytotoxic capacity was also impaired. Of interest, though the cytolytic activity of NK cells of mCRC patients could be increased by cetuximab through ADCC, the level of cytotoxicity was still markedly reduced compared with that mediated by NK cells from healthy adult volunteers. These data suggest that adoptive transfer of fully functional NK cells might be of benefit to restore the NK effector cell pool in mCRC patients. For this purpose, the cytotoxic effects of two allogeneic NK cell products, i.e. activated PBNK and UCB-NK cells, were tested and compared in vitro against a panel of colon cancer cell lines. UCB-NK cells were found to exert superior cytotoxicity compared to PBNK cells, their cytotoxicity being comparable to that achieved when PBNK were additionally stimulated using cetuximab. These superior cytotoxic effects of UCB-NK cells were verified in vivo, where treatment with UCB-NK cells alone significantly reduced the tumor load in mice inoculated with EGFR + RAS mut colon cancer cells. Of interest, this effect was not increased by the addition of cetuximab in vivo , which could be due to a sub- optimal up-regulation of CD16 cell surface expression levels on the adoptively transferred UCB-NK in the immunodeficient mice that was used for these studies. Importantly, as a clinical study in AML patients revealed that CD16a expression levels steadily increased on UCB-NK cells post-infusion, synergy between both approaches may be expected when applied clinically. To summarize, through the studies described in this thesis we have demonstrated that UCB- NK cells have superior anti-tumor efficacy against epidermoid, colon and cervical cancers as compared to activated PBNK cells, and are equally cytotoxic as the combination of PBNK and cetuximab, demonstrating significant anti-tumor benefit against EGFR + RAS mutant and EGFR + BRAF mutant tumors. This novel UCB-NK expansion and differentiation technique from Glycostem allows the generation of large numbers of cytolytic UCB-NK cells that may overcome the limitations of current NK cell based adoptive transfer strategies and supplement the immune system with sufficient numbers of NK cells to mount an effective