15353-j-veluchamy

3 HLA independent killing of cervical tumors by UCB-NK cells | 83 (Tumor necrosis factor-related apoptosis-inducing ligand), and FAS (Fas-Ligand interactions) alsomight contribute to the observed target cell lysis 48,49 . Indeed, NKp44 has been previously reported as highly expressed on expanded UCB-NK, in sharp contrast to PB-NK cells, which in the steady state don’t express NKp44 33 . The known ligands for NKp44 have mostly been associated with microbial responses, whereas the identity of cancer associated ligands until recently has remained mostly obscure. A ligand for NKp44 has now been identified on tumor cells, designated NKp44L, which opens the way for further exploration of the relative importance of this activating receptor axis in NK-mediated tumor cytolysis 50 . Interestingly, in the present study we have shown the predominant effect of HLA class I expression on the functionality of PBNK. In contrast to PBNK, UCB-NK have the ability to overcome resistance to cytolysis due to HLA-ABC expression as demonstrated by the correlative studies with all ten cell lines which revealed efficient UCB-NK-mediated cytolysis of both HLA-ABC high and low expressing cell lines (figure 4c). A lack of expression of inhibitory KIRs on UCB-NK may provide a mechanistic explanation for their ability for HLA class I independent cytotoxicity. Indeed, whereas PBNK and UCB-NK expressed similar levels of NKG2A, inhibitory KIRs, as measured by a panKIR2D antibody, were completely lacking from the UCB-NK cell surface. In keeping with this observation, we previously published the profiling of UCB-NK using an expanded panel of antibodies to inhibitory KIR, which revealed low expression levels of KIR2DL1/DS1, KIR2DL2/DL3/DS2, and KIR3DL1/DS1 as compared to PBNK 33 . Cervical tumors have been shown to also have aberrant non-classical HLA class I expression which might help them to escape from NK cell killing 14 . Remarkably, in our hands NK cytotoxicity was not impaired by higher levels of HLA-E or HLA-G expression. The apparent ability of UCB-NK to overcome the possible resistance related to expression of both inhibitory classical and non-classical HLA molecules may offer an excellent treatment modality for cervical cancer. NK cells are often dysfunctional and low in number in cervical cancer patients 19,23,24 . In order to achieve a more potent and effective cytotoxic effect of NK cells in patients with cervical cancer it is therefore critical to have adequate numbers of functional effector NK cells. In regard to generating large numbers of NK cells for therapeutic purposes, NK cells expanded from PBMC and other sources have limited expansion capacity as compared to cord blood derived NK cells 51 . Adoptive transfer of large numbers of cytotoxic UCB-NK could be a viable treatment option, because UCB-NK have a highly activated phenotype with more than 75% stable expression rates of NKG2D, DNAM-1, NKp30, NKp44 and NKp46 in all mature UCB-NK and lack inhibitory KIRs resulting in HLA independent cytolytic efficacy; additional advantages of UCB-NK over PBNK are fewer impurities (such as T and B cells) detected upon full NK maturation, thereby reducing chances of GVHD upon adoptive transfer 31,38 . In this study, UCB-NK were not tested in combination with CET due to their low surface expression of CD16a in vitro , however, UCB-NK further mature upon adoptive