Factors Regulating the Cytotoxic Activity of the Human Natural Killer Cell Line, NK-92

Abstract

NK-92, a highly cytotoxic, interleukin-2 (IL-2)-dependent human natural killer (NK) cell line, has been of interest for basic and translational research. We report on a comprehensive analysis of NK-92 for factors implicated in NK cytotoxicity to elucidate factors underlying NK-92's high cytolytic activity and target range. Thus, we hope to develop a method to identify patients best suited to NK-92 immunotherapy. In addition, as a model system, we hope to increase understanding of the basis for the elevated activity exhibited by activated NK (ANK) cells. NK-92 exhibits an unusual receptor expression profile, expressing a relatively large number of activating (NKp30, NKp46, 2B4, NKGD, E, CD28) receptors. Conversely, it expresses few inhibitory receptors (NKGA/B, low levels of KIR2DL4, ILT-2), lacking most of the killer inhibitory receptors (KIRs) clonally expressed on normal NK cells. In addition, NK-92 expresses high levels of molecules involved in the perforin-granzyme cytolytic pathway as well as additional cytotoxic effector molecules including tumor necrosis factor (TNF)-superfamily members FasL, TRAIL, TWEAK, TNF-alpha, indicating the ability to kill via alternative mechanisms. NK-92 also expresses other molecules implicated immune effector cell regulation (CD80, CD86, CD40L, TRANCE) whose relevance in NK killing is unclear. This study provides initial data to develop a method to identify NK-92 susceptible cells (cells expressing ligands for NK-92 activating receptors ie CD48 for 2B4 and CD80/86 for CD28). Furthermore, this work suggests mechanisms that may contribute to ANK cell activity, including modulation of receptor expression to favor activation, up-regulation of cytotoxic effector molecules, and acquisition of new cytolytic pathways.

NK resistance of tumor cells from multiple myeloma and chronic lymphocytic leukemia patients: implication of HLA-G

“…In contrast to KIR2DL4 surface expression, which is almost undetectable on NK-92, 17,28 these cells express high levels of ILT2. For NK-92 interaction with HLA-G, ILT2 is the relevant receptor.…”

“…NK-sensitive erythroleukemia cell line K562, myeloma cell lines U266, RPMI 8226, IM9, the M8 melanoma cell line and its stable transfectant M8-HLA-G1 were used as target for NK-mediated cytotoxicity assay and maintained in culture in RPMI 1640 supplemented with 10% fetal bovine serum (FBS). NK-92, a highly cytotoxic human NK cell line, with characteristics of activated NK cells 27,28 was used as an effector cell population and maintained in X-VIVO 10 supplemented with 5% human AB serum and 100 U ml -1 interleukin-2 (Chiron, Emervyille, CA, USA). All cell lines were obtained from ATCC (Manassa, VA, USA).…”

“…Moreover, NK activity is often altered in these patients. [33][34][35] We tested the sensitivity of patient-derived tumor cells to NK cells from unrelated healthy donors as well as NK-92, a highly cytotoxic human NK cell line with characteristics of activated NK cells 27,28 and which represents a model system to investigate susceptibility to NK-mediated killing and mechanisms of tumor escape.…”

A Peptide Against the N-Terminus of Myristoylated Alanine-Rich C Kinase Substrate Inhibits Degranulation of Human LeukocytesIn Vitro

“…tein in eosinophils (2), lysozyme in monocytes/macrophages (3,4), and granzyme in natural killer (NK) cells and cytotoxic lymphocytes (5)(6)(7)(8). These mediators are released at sites of injury and contribute to inflammation and repair in the lung and elsewhere.…”

“…Due to difficulties in isolating sufficient amounts of other leukocyte types from blood, we investigated the release of membrane-bound inflammatory mediators from commercially available human leukocyte cell lines. The human promyelocytic cell line HL-60 clone 15 was used to assess secretion of EPO (19)(20)(21)(22), the monocytic leukemia cell line U937 was used to assess secretion of lysozyme (3,4,23), and the lymphocyte NK cell line NK-92 was used to assess the release of granzyme (6)(7)(8). In all cases, the cells were preincubated with a range of concentrations of a synthetic peptide identical to the 24-amino-acid MARCKS N-terminus (myristoylated N-terminal sequence peptide [MANS]), or a missense control peptide (random N-terminal sequence peptide [RNS]), which consists of the same 24 amino acids arranged in random order.…”