IntroductionNatural killer (NK) cells are present in the bloodstream, spleen, bone marrow, and in nonlymphoid organs and represent one of the main effectors of the immunosurveillance against tumors, by exerting 2 major effector functions, cytolysis of target cells and production of cytokines and chemokines. 1,2 The activity of NK cells depends on the interplay between inhibitory receptors for major histocompatibility complex (MHC) class I molecules and activating receptors, which operate in concert to induce the elimination of tumor cells. 3,4 Among the activating receptors particularly relevant for tumor cell recognition and killing is NKG2D, the receptor for the MHC I-related molecules MICA/B, and ULBPs (UL16-binding proteins), belonging to the C-type lectin-like receptor family. 3,[5][6][7] The NKG2D activating receptor is expressed not only on NK cells, but also on ␥␦ T cells, CD8 ϩ T cells, and a subset of CD4 ϩ T cells. The expression of NKG2D ligands is largely confined to virus-infected, tumor, and stressed cells. 7 To promote escape of tumors from NKG2D-mediated immunosurveillance, NKG2D ligands undergo proteolytic shedding. Soluble NKG2D ligands (NKG2DLs) have been shown to down-regulate the cell surface NKG2D expression on NK cells, resulting in impaired killing of tumor cells. 8,9 Another activating receptor involved in NK-cell-mediated tumor cell killing is DNAX accessory molecule-1 (DNAM-1), a transmembrane glycoprotein constitutively expressed on the majority of T cells, NK cells, and macrophages; its ligands are Nectin-2 (Nec-2, CD112) and the poliovirus receptor (PVR, CD155), which belong to the nectin/nectin-like family. 10-12 DNAM-1 ligands have been initially described as adhesion molecules mainly regulating trans-endothelial migration 13 and only recently they have been found on a variety of tumor cells. 12,14 Both DNAM-1 and NKG2D cooperate in the induction of NK-cell killing against tumor cells of different histotypes, including those of hematopoietic origin. 14,15 Similarly to the NKG2D ligands, soluble isoforms of PVR have also been found in human serum and in the culture supernatant of tumor cell lines, and their role in tumor immunoevasion has been considered. 12 It has recently been demonstrated that agents that produce a genotoxic stress or DNA-replication inhibitors up-regulate NKG2D ligand expression through the activation of ATM (ataxia telangiectasia mutated) and ATR (ATM-and Rad3-related) protein kinases on human fibroblasts and on mouse tumor cell lines, and enhance their destruction by NK cells. 16,17 Increased ligand expression is regulated by the activation of the DNA damage response (DDR). This is a cellular program devoted to the maintenance of genome integrity through the inhibition of cell cycle and activation of the DNA repair systems, or by the induction of apoptosis or a protracted cell-cycle arrest known as cellular senescence. 17,18 No information is so far available on the regulation of DNAM-1 ligand expression through the DDR pathway.Submitted August 11, 2008; acc...

SummaryMany body surfaces harbor organ-specific γδ T cell compartments that contribute to tissue integrity. Thus, murine dendritic epidermal T cells (DETCs) uniquely expressing T cell receptor (TCR)-Vγ5 chains protect from cutaneous carcinogens. The DETC repertoire is shaped by Skint1, a butyrophilin-like (Btnl) gene expressed specifically by thymic epithelial cells and suprabasal keratinocytes. However, the generality of this mechanism has remained opaque, since neither Skint1 nor DETCs are evolutionarily conserved. Here, Btnl1 expressed by murine enterocytes is shown to shape the local TCR-Vγ7+ γδ compartment. Uninfluenced by microbial or food antigens, this activity evokes the developmental selection of TCRαβ+ repertoires. Indeed, Btnl1 and Btnl6 jointly induce TCR-dependent responses specifically in intestinal Vγ7+ cells. Likewise, human gut epithelial cells express BTNL3 and BTNL8 that jointly induce selective TCR-dependent responses of human colonic Vγ4+ cells. Hence, a conserved mechanism emerges whereby epithelia use organ-specific BTNL/Btnl genes to shape local T cell compartments.

IntroductionIn addition to their role in providing antitumor and antiviral immunity, 1 natural killer (NK) cells are also able to regulate the T-cell arm of the adaptive immune response by secreting different cytokines and chemokines. 2 Moreover, several studies have provided evidence of cognate cell-cell interactions between NK cells and various leukocyte types, including dendritic cells (DCs) and B and T lymphocytes. [3][4][5] Although NK cells have been thought to mainly promote adaptive immune responses, recent in vivo studies suggest that they can also restrain T cell-mediated immune responses. Therefore, the depletion of NK cells results in enhanced T-cell proliferation and effector functions during murine cytomegalovirus infection 6 and in an antitumor response against lymphoma cells. 7 Conversely, several studies have indicated that the depletion of NK cells is associated with increased severity of autoimmune diseases. In fact, NK cell-depleted mice develop a more severe form of experimental autoimmune encephalomyelitis, 8 and NK-cell-mediated downregulation of autoreactive cytotoxic T lymphocytes has been shown to have a protective role in type 1 diabetes. 9 These findings suggest that NK cells may be crucial for terminating T cell-mediated responses and for preventing inappropriate T-cell activation and effector functions leading to the development of autoimmune diseases.NK cell-mediated attenuation of T-cell responses can involve several mechanisms, including the production of inhibitory cytokines (eg, TGF-␤ and IL-10) 10,11 and killing of DCs and/or activated T cells. 4,12,13 In regard to the NK cell-mediated killing of T cells, IL-2-activated mouse and human NK cells recognize and lyse T-cell blasts in a perforin-dependent manner through the activating receptor NKG2D. 4,12 Interestingly, the results of our previous study indicated that Ag stimulation of human T cells was sufficient to induce the surface expression of the NKG2D ligands (NKG2DLs) MHC class I-related chain A (MICA), MICB, and UL16-binding proteins 1-3 (ULBP1-3). 4,14 To date, little is known about the existence of additional receptor-ligand interactions that might contribute to the NK cell-mediated recognition of T lymphocytes.DNAX accessory molecule-1 (DNAM-1) is an activating receptor belonging to the Ig superfamily that is constitutively expressed by most NK cells, T cells, macrophages, and DCs. 15,16 DNAM-1 interacts with lymphocyte function-associated antigen 1 (LFA-1), and this association is required for its functional activity on both NK and cytotoxic T cells. 17 Ligands for DNAM-1 (DNAM1Ls) include Nectin-2 and poliovirus receptor (PVR), which belong to the Nectin/Nectin-like family of adhesion molecules. 18 DNAM1Ls are often expressed by tumor cells and can activate or enhance tumor cell lysis in vitro. 15,18 Recent studies have reported that they can also be expressed by monocytes, DCs, and phytohemagglutinin (PHA)-stimulated CD4 ϩ T lymphocytes. 19,20 Little information is available about the molecular mechanisms regulating...

Increasing evidence indicates that cancer cell stress induced by chemotherapeutic agents promote antitumor immune responses and contribute to their full clinical efficacy. In this article, we identify the signaling events underlying chemotherapy-induced NKG2D and DNAM-1 ligand expression on multiple myeloma (MM) cells. Our findings indicate that sublethal doses of doxorubicin and melphalan initiate a DNA damage response (DDR) controlling ligand upregulation on MM cell lines and patient-derived malignant plasma cells in Chk1/2-dependent and p53-independent manner. Drug-induced MICA and PVR gene expression are transcriptionally regulated and involve DDR-dependent E2F1 transcription factor activity. We also describe the involvement of changes in the redox state in the control of DDR-dependent upregulation of ligand surface expression and gene transcriptional activity by using the antioxidant agent N-acetyl-l-cysteine. Finally, in accordance with much evidence indicating that DDR and oxidative stress are major determinants of cellular senescence, we found that redox-dependent DDR activation upon chemotherapeutic treatment is critical for MM cell entry in premature senescence and is required for the preferential ligand upregulation on senescent cells, which are preferentially killed by NK cells and trigger potent IFN-γ production. We propose immunogenic senescence as a mechanism that promotes the clearance of drug-treated tumor cells by innate effector lymphocytes, including NK cells.

New arylthioindoles (ATIs) were obtained by replacing the 2-alkoxycarbonyl group with a bioisosteric 5-membered heterocycle nucleus. The new ATIs 5, 8, and 10 inhibited tubulin polymerization, reduced cell growth of a panel of human transformed cell lines, and showed higher metabolic stability than the reference ester 3. These compounds induced mitotic arrest and apoptosis at a similar level as combretastatin A-4 and vinblastine and triggered caspase-3 expression in a significant fraction of cells in both p53-proficient and p53-defective cell lines. Importantly, ATIs 5, 8, and 10 were more effective than vinorelbine, vinblastine, and paclitaxel as growth inhibitors of the P-glycoprotein-overexpressing cell line NCI/ADR-RES. Compound 5 was shown to have medium metabolic stability in both human and mouse liver microsomes, in contrast to the rapidly degraded reference ester 3, and a pharmacokinetic profile in the mouse characterized by a low systemic clearance and excellent oral bioavailability.

Modulation of the host immune system represents a promising therapeutic approach against cancer, including multiple myeloma. Recent findings indicate that the NK group 2D (NKG2D)- and DNAX accessory molecule-1 (DNAM-1)-activating receptors play a prominent role in tumor recognition and elimination by cytotoxic lymphocytes, suggesting that the levels of NKG2D and DNAM-1 ligand expression on tumor cells may be a critical factor to improve the immune response against cancer. In this study, we tested the effect of 17-allylaminogeldanamycin and radicicol, drugs targeting the heat shock protein-90 (HSP-90) chaperone protein and displaying antimyeloma activity, on the expression of NKG2D and DNAM-1 ligands in human myeloma cell lines. We demonstrate that HSP-90 inhibitors are able to up-regulate both MHC class I chain-related (MIC) A and MICB protein surface and mRNA expression in human myeloma cell lines, without any significant effect on the basal expression of the DNAM-1 ligand poliovirus receptor CD155, or induction of nectin-2 and UL16-binding proteins. Activation of the transcription factor heat shock factor-1 by HSP-90 inhibitors is essential for the up-regulation of MICA/MICB expression and knockdown of heat shock factor-1 using small hairpin RNA interference blocks this effect. Moreover, in vitro and in vivo binding of heat shock factor-1 to MICA and MICB promoters indicates that it may enhance NKG2D ligand expression at the transcriptional level. Finally, exposure to HSP-90 inhibitors renders myeloma cells more efficient to activate NK cell degranulation and a blocking Ab specific for NKG2D significantly reduces this effect. Thus, these results provide evidence that targeting NKG2D ligands expression may be an additional mechanism supporting the antimyeloma activity of HSP-90 inhibitors and suggest their possible immunotherapeutic value.

New arylthioindole derivatives having different cyclic substituents at position 2 of the indole were synthesized as anticancer agents. Several compounds inhibited tubulin polymerization at submicromolar concentration and inhibited cell growth at low nanomolar concentrations. Compounds 18 and 57 were superior to the previously synthesized 5. Compound 18 was exceptionally potent as an inhibitor of cell growth: it showed IC50 = 1.0 nM in MCF-7 cells, and it was uniformly active in the whole panel of cancer cells and superior to colchicine and combretastatin A-4. Compounds 18, 20, 55, and 57 were notably more potent than vinorelbine, vinblastine, and paclitaxel in the NCI/ADR-RES and Messa/Dx5 cell lines, which overexpress P-glycoprotein. Compounds 18 and 57 showed initial vascular disrupting effects in a tumor model of liver rhabdomyosarcomas at 15 mg/kg intravenous dosage. Derivative 18 showed water solubility and higher metabolic stability than 5 in human liver microsomes.

Our data indicate that chemerin is up-regulated during decidualization and might contribute to NK cell accumulation and vascular remodeling during early pregnancy.