This result was consistent with the measurement of trypan and CCK-8. safety and showed preliminary evidence of clinical benefit for cancer patients. The molecules 41BB and CD3 are commonly used as stimulators in the CAR structure, and their expression in NK cells can promote the activation of NK cells, leading to the enhanced perforin- and granzyme-mediated lysis of tumor cells. This study showed that genetically modified NK-92 cells combined with antibody-mediated ADCC using rituximab and trastuzumab monoclonal antibodies lysed tumor cells more efficient than the NK-92 cell lines. It also showed that the anti-tumor activity of chimeric stimulator molecules of the CAR-modified CD16 receptor was stronger than that of CD16 (allotype V158). These studies provide a rationale for the use of genetically modified NK-92 cells in combination with IgG1 anti-tumor monoclonal antibodies. We also provide a rationale for the chimeric modified CD16 receptor that can improve the anti-tumor effect of NK92 cells via ADCC. Supplementary Information The online version of this article (10.1007/s10616-021-00476-1) contains supplementary Mouse monoclonal to STAT3 material, which is available to authorized users. Keywords: Natural killer cells, NK-92, CD16, CAR, ADCC, Chimeric receptor Introduction Natural killer (NK) cells play a pivotal role in mediating innate immunity, which defends against virus-infected and malignant cells (Weiner et al. 2009; Paust et al. 2010; Gras Navarro et al. 2015). Unlike cytotoxic T lymphocytes, NK cells target tumors, and virus-infected cells circumvent major histocompatibility complex (MHC) (Gladow et al. 2004). NK cells are activated without stimulation (Carson et al. 2001). However, a variety of cytokines, including interleukin (IL)-2 and IL12, can enhance NK cell cytotoxic activity. In addition to these mechanisms, NK cells play a role in anti-tumor immunity alone or in combination with specific antibodies via antibody-dependent cell-mediated cytotoxicity (ADCC). For adoptive immunotherapy, NK cells provide a effector cell type with the capacity of recognizing antigen-positive and MHC-I-negative or MHC-I-low LDE225 (NVP-LDE225, Sonidegib) tumor cells (Yokoyama and Kim 2006). However, NK cells can malfunction due to the lack of adhesion molecules presented on the cell surface, for example, the HLA-G molecule, which can bind?to immunoglobulin-like receptors (KIRs), or the production of molecules by lymphoblasts, such as MICA and MICB that binding to NK cell activating receptors, such as NKG2D (Fernandez-Messina et al. 2012; Yang et al. 2013). Allogeneic NK cells are used for manipulation unless the T cell is depleted to prevent graft versus host disease (GVHD). By contrast, the activated NK cell line NK-92, which continuously grows and is IL-2 dependent, can easily be expanded in vitro and is highly cytotoxic against a variety of malignant cells. Homologous NK cells from patients are often poor, but the safety of NK92 cells has been studied in phase I trials (Scott et al. 2001). Preclinical studies showed that NK-92 cells do not form tumors when transplanted into severe combined immunodeficiency (SCID) or athymic mice. Several clinical studies show repeated infusions of irradiated NK-92 cells were harmless. Clinical responses were observed in patients with melanoma, lung cancer, Merkel cell carcinoma, lymphoma, and kidney cancer (Klingemann et al. 2016). Despite the allogeneic nature of NK-92 cells, the formation of anti-human leukocyte antigen (HLA) antibodies was observed in less than half of the patients. Moreover, the pharmacodynamics of NK-92 clearance did LDE225 (NVP-LDE225, Sonidegib) not differ upon repeated doses. NK-92 cells do not express the CD16 Fc LDE225 (NVP-LDE225, Sonidegib) receptor, which is necessary for the NK-mediated ADCC lysis of tumor cells. Many patients benefit from adoptive immunotherapy of some mAbs, such LDE225 (NVP-LDE225, Sonidegib) as the rituximab (Rituxan) and the trastuzumab (Herceptin), and cell-mediated immunity is recognized as one of the mechanisms responsible for their clinical efficiency (Koene et al. 1997; Cartron et al. 2002). In addition to lysing malignant and virus-infected cells, ADCC, which is mediated by the CD16 (FccRIIIa) receptor present on the NK cells surface, can primarily be activated in NK cells. Therefore some clinical studies employing the IgG1 isotype mAbs trastuzumab (Herceptin) or rituximab (Rituxan) showed that breast and lymphoma NSG mice, respectively, whose NK-92 cells express CD16 have improved overall survival compared to NK-92 cell lines (Bibeau et al. 2009). Similarly, after transduction with the FcIIIRa/FcRI (referred to as CD16/) receptor fusion gene, CD4+ and CD8+ cytotoxic T lymphocytes stably expressed the CD16/ receptor on their surface and mediated ADCC. Thus, associating a therapeutic mAb and CD16/-transduced.