Faculty Publications

Given high rates of smoking among cancer patients, smoking cessation treatment is crucial; yet limited data exist to guide integration of such trials into the oncologic context. In order to determine the feasibility of conducting smoking cessation clinical trials with cancer patients, screening and baseline data from a large randomized placebo-controlled pharmacotherapy trial were analyzed. Descriptive statistics and regression analyses were used to compare enrollees to decliners, describe program enrollees, and assess correlates of confidence in quitting smoking. Out of 14,514 screened patients, 263 (<2%) were eligible; 43 (16%) refused enrollment. Among the eligible patients, 220 (84%) enrolled. Enrollment barriers included smoking rate, medical history/contraindicated medication, lack of interest, and language. Compared to enrollees, decliners were more likely to have advanced cancer. The trial enrolled a sample of 67 (>30%) African Americans; participants had extensive smoking histories; many were highly nicotine dependent; and participants consumed about seven alcoholic beverages/week on average. Head and neck and breast cancer were the most common tumors. About 52 (25%) reported depressive symptoms. A higher level of confidence to quit smoking was related to lower depression and lower tumor stage. Integrating a smoking cessation clinical trial into the oncologic setting is challenging, yet feasible. Recruitment strategies are needed for patients with advanced disease and specific cancers. Once enrolled, addressing participant's depressive symptoms is critical for promoting cessation.

Dengue virus (DV) and West Nile virus (WNV) have become a global concern due to their widespread distribution and their ability to cause a variety of human diseases. Antiviral immune defenses involve NK cells. In the present study, we investigated the interaction between NK cells and these two flaviviruses. We show that the NK-activating receptor NKp44 is involved in virally mediated NK activation through direct interaction with the flavivirus envelope protein. Recombinant NKp44 directly binds to purified DV and WNV envelope proteins and specifically to domain III of WNV envelope protein; it also binds to WNV virus-like particles. These WNV-virus-like particles and WNV-domain III of WNV envelope protein directly bind NK cells expressing high levels of NKp44. Functionally, interaction of NK cells with infective and inactivated WNV results in NKp44-mediated NK degranulation. Finally, WNV infection of cells results in increased binding of rNKp44 that is specifically inhibited by anti-WNV serum. WNV-infected target cells induce IFN-gamma secretion and augmented lysis by NKp44-expressing primary NK cells that are blocked by anti-NKp44 Abs. Our findings show that triggering of NK cells by flavivirus is mediated by interaction of NKp44 with the flavivirus envelope protein. The Journal of Immunology, 2009, 183: 2610-2621.

In B lymphocytes, the B-cell adaptor for phosphatidylinositol 3-kinase (BCAP) facilitates signaling from the antigen receptor. Mice lacking BCAP have a predominantly immature pool of B cells with impaired immune function and increased susceptibility to apoptosis. Unexpectedly, we have found that natural killer (NK) cells from BCAP-deficient mice are more mature, more long-lived, more resistant to apoptosis, and exhibit enhanced functional activity compared with NK cells from wild-type mice. Surprisingly, these effects are evident despite a severe impairment of the immunoreceptor tyrosine-based activation motif-mediated Akt signaling pathway. The seemingly paradoxical phenotype reveals inherent differences in the signals controlling the final maturation of B cells and NK cells, which depend on positive and negative signals, respectively. Both enhanced interferon-gamma responses and augmented maturation of NK cells in BCAP-deficient mice are independent of available MHC class I ligands. Our data support a model in which blunting of BCAP-mediated activation signaling in developing NK cells promotes functionality, terminal maturation, and long-term survival.

Human NK cells lyse Ab-coated target cells through the process of Ab-dependent cellular cytotoxicity (ADCC). Improving ADCC responses is desirable because it is thought to be an important antitumor mechanism for some Abs. NK cell inhibitory receptors, such as killer cell Ig-like receptors, engage with MHC class I molecules on self-cells to block NK cell activation. Accordingly, we enhanced ADCC responses by blocking NK cell inhibitory receptors, thus perturbing induction of the self-recognition signal. In a cell line model of anti-lymphoma therapy, the combination of rituximab with an Ab that blocks inhibitory self-recognition yielded increased NK cell-mediated target cell lysis when compared with rituximab alone. To validate this proof-of-concept, we then used a more representative approach in which an individual's fresh primary NK cells encountered autologous, EBV-transformed B cells. In this system, rituximab and a combination of Abs that block NK cell inhibitory receptors yielded improved NK cell-mediated lysis over rituximab alone. The results show, for the first time, that disruption of inhibitory self-recognition can efficiently promote ADCC in a human model, applying an autologous system in which physiologic checkpoints are in place. This method provides an alternative approach to potentiate the therapeutic benefit of antitumor Abs that mediate ADCC.

The inhibitory killer cell Ig-like receptors (KIR) negatively regulate NK cell cytotoxicity by activating the Src homology 2 domain-containing protein tyrosine phosphatases 1 and 2 following ligation with MHC class I molecules expressed on normal cells. This requires tyrosine phosphorylation of KIR on ITIMs in the cytoplasmic domain. Surprisingly, we have-found that KIR3DL1 is strongly and constitutively phosphorylated on serine and weakly on threonine residues. In this study, we have mapped constitutive phosphorylation sites for casein kinases, protein kinase C, and an unidentified kinase on the KIR cytoplasmic domain. Three of these phosphorylation sites are highly conserved in human inhibitory KIR. Functional studies of the wild-type receptor and serine/threonine mutants indicated that phosphorylation of Ser(394) by protein kinase C slightly suppresses KIR3DL1 inhibitory function, and reduces receptor internalization and turnover. Our results provide evidence that serine/threonine phosphorylation is an important regulatory mechanism of KIR function.

NK cells play critical roles in immune responses against tumors or virus infections by generating type 1 cytokine and cytotoxicity responses. In contrast, during type 2 dominant immune responses, such as allergic diseases, activities of NK cells are often impaired. These type 2 immune-mediated diseases have been reported to be closely associated with local production of PGD(2). PGD(2) is an eicosanoid primarily synthesized by mast cells and alveolar macrophages, and it functions through two major receptors, D prostanoid receptor (DP) and chemoattractant receptor-like molecule on the Th2 cell. Within the immune system, PGD(2) binding to DP generally leads to suppression of cellular functions. In the current study, we show that: 1) DP is expressed in human NK cells as detected by mRNA analysis and Western blot; 2) PGD(2) inhibits cytotoxicity, chemotaxis, and type 1 cytokine production of human NK cells via signaling through DP; 3) PGD(2) signaling via DP elevates intracell!

Tolerance of natural killer (NK) cells toward normal cells is mediated through their expression of inhibitory receptors that detect the normal expression of self in the form of class I major histocompatibility complex (MHC-I) molecules on target cells. These MHC-I-binding inhibitory receptors recruit tyrosine phosphatases, which are believed to counteract activating receptor-stimulated tyrosine kinases. The perpetual balance between signals derived from inhibitory and activating receptors controls NK cell responsiveness and provides an interesting paradigm of signaling cross talk. This review summarizes our knowledge of the intracellular mechanisms by which cell surface receptors influence biological responses by NK cells. Special emphasis focuses on the dynamic signaling events at the NK immune synapse and the unique signaling characteristics of specific receptors, such as NKG2D, 2B4, and KIR2DL4.

The invention provides a natural killer cell, NK-92, modified to express an Fc receptor on the surface of the cell, such as CD 16 (FcgRIII-A), or other Fcy or Fc receptors. The modified NK-92 cell can be further modified to concurrently express an assocd. accessory signaling protein, such as FceRI-g,TCR-z, or to concurrently express interleukin-2 (IL-2) or other cytokines. Addnl. methods are disclosed for various assays, assessments, and therapeutic treatments with the modified NK-92 cells. [on SciFinder (R)]

The major histocompatibility complex nonrestricted cytotoxic leukemic T cell line T acute lymphoblastic leukemia (TALL)-104 is being pursued. Pill as a therapeutic agent for cancer. However, the receptors and effector mechanisms responsible for its broad tumoricidal function remain undefined. Here, we examined the roles played by natural cytotoxicity receptors (NCR), killer cell immunoglobulin-like receptors, cytolytic granule components, and tumor necrosis factor (TNF) family members in tumor recognition and lysis by TALL-104 cells. The perforin-granzyme pathway, TNF-related apoptosis-inducing ligand (TRAIL), and Fas were each involved in the lysis of particular tumor targets by TALL-104. Furthermore, phorbol 12-myristate 13-acetate/ionomycin treatment induced surface expression of Fas-L and TRAIL. In addition, supernatants from CD3-stimulated TALL-104 cultures exhibited antiproliferative activity, which was blocked 50-90% by anti-TNF-alpha monoclonal antibody (mAb). Althou gh negative for the NCR natural killer (NK)p44, this cell line was found to express NKp46. An anti-NKp46 antibody strongly blocked TALL-104-mediated lysis of certain targets and directly induced cytokine production, granule release, and redirected lysis responses. Anti-NKG2D and anti-2B4 also stimulated redirected c),totoxicity by TALL-104. By contrast, anti-NKG2A mAb, did not stain the cells or inhibit killing responses. Alternatively, KIR3DL2 was detected on TALL-104, mid expression of its reported ligand, human leukocyte antigen (HLA)-A, on target cells provided protection from cytotoxicity. Thus, NKp46, NKG2D, and 2B4 are activating receptors, and KIR3DL2 is an inhibitory receptor on TALL-104. The data demonstrate the ability of TALL-104 cells to recognize a wide variety of tumors with NK cell receptors mid kill them with a broad arsenal of cytolytic effector mechanisms, including cytolytic granules and TNF family ligands.

BACKGROUND: The inherently complex signaling networks of tumors result from genetic and epigenetic alterations that occur during cancer initiation and progression. METHODS: In an attempt to identify early molecular changes associated with dominantly inherited predisposition to "two-hit" renal tumors, the expression profiles of primary cultures of phenotypically normal renal epithelial cells from individuals bearing a germline mutation in either the von Hippel-Lindau (VHL) or the tuberous sclerosis complex (TSC) gene were compared to that of renal epithelial cells from control nonmutation carriers by microarray analysis. RESULTS: Reliability of the microarray data from pooled samples was confirmed by real-time RT-PCR. Principal Component Analysis revealed substantial differences in the gene expression profiles of the renal epithelial cells from VHL and TSC mutation carriers. In several instances, the microarray data confirm our present knowledge of the cellular pathways affected by biallelic VHL and TSC mutations. CONCLUSIONS: These findings demonstrate that heterozygosity for a mutant tumor suppressor gene may alter the expression profiles of phenotypically normal epithelial cells in a gene-specific manner. Detectable effects of "one-hit" represent early molecular changes in tumorigenesis that may serve as targets for chemopreventive intervention.