The development of genetically modified K562 cells as artificial antigen presenting cells for NKT cells (P4310)

Abstract CD1d-restricted type-I Natural Killer T cells (NKTs) have been shown to mediate anti-tumor immune responses in mouse models and are associated with improved outcome in several types of cancer. However, the therapeutic application of NKTs has been limited by low numbers and functional defects of these cells in patients with cancer. To provide a means for effective ex-vivo NKT-cell expansion for cell and gene therapy applications we explored native and engineered properties of K562 cells to function as artificial antigen presenting cells (aAPC). K562 cells express an activating allele of HLA-C and propagate NK cells which compete with NKTs in the same culture. Therefore, we used zinc finger nucleases to delete the endogenous HLA-C gene in K562 cells and transduced parental and HLA-Cnull K562 cells with CD1d cDNA followed by single cell sorting and clonal expansion. The clones exhibited a range of CD1d expression. They were pulsed with a NKT ligand, αGalactosylceramide and tested as aAPC for NKTs using CFSE proliferation assay. We found that aAPC clones with an intermediate level of CD1d expression induced the highest rate of NKT-cell proliferation. No difference was observed between K562/CD1d and HLA-CnullK562/CD1d clones when purified NKTs were used as responders. In contrast to K562/CD1d, HLA-CnullK562/CD1d clones did not expand NK cells when added to primary PBMC. Therefore, these HLAnullCD1d+ aAPC can be used to selectively expand primary NKTs for adoptive cell therapy.