NK-92 cells transduced with retroviral vectors encoding marker genes and class I MHC suppression genes for improved use in adoptive cellular immunotherapy

Abstract

2592 Background: Human NK-92 cells have a natural killer phenotype and exhibit high cytotoxicity towards targets lacking class I MHC expression. They are currently being tested for use in adoptive cellular immunotherapy and bone marrow purging to augment autologous stem cell transplant. One problem encountered in initial clinical trials employing adoptive cellular immunotherapy involves the generation of host cytotoxic T lymphocyte (CTL) responses to the transplanted cells limiting their effectiveness in vivo. Methods: LXSN based retroviruses encoding green fluorescent protein (GFP), and two separate class I MHC suppression genes (Herpes simplex virus (HSV) ICP47 and Human Cytomegalovirus (HCMV) US11) were constructed and used to transduce NK-92 cells. Stable populations of transgenic NK-92 cells were selected either in media containing a neomycin homologue or by fluorescent cell sorting for GFP encoding vectors. Populations were then analyzed for class I MHC expression or GFP fluorescence using anti-class I MHC antibodies and flow cytometry. Results: NK-92 cells transduced with ICP47 and US11 encoding retroviruses expressed 52% and 32% less class I MHC respectively. The levels of cell surface class I MHC expression observed remained consistent over 8 months in culture. Current work focuses on determining whether the decrease in class I MHC expression noted is significant enough to abrogate a host CTL response. Modified NK-92 cells expressing GFP were transduced with additional retroviruses encoding the class I suppression genes and populations of fluorescent cells expressing reduced cell surface class I MHC were isolated. In these dual transduced populations, class I MHC expression was reduced by 53% and 83% for ICP47 and US11 respectively. The modified NK-92 cells were tested against a panel of class I MHC positive and negative target cell lines and only minimal alterations of NK-92 cytotoxicity were observed. Conclusions: These results demonstrate the potential to improve cells used for adoptive cellular immunotherapy by the incorporation of multiple transgenes encoding beneficial traits. No significant financial relationships to disclose.