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Aims Tumours secrete proangiogenic factors to induce the ingrowth of blood vessels, the end targets of which are vascular endothelial cells (ECs). The MEOX2 homeoprotein inhibits nuclear factor-κB (NF-κB) signalling and EC activation in response to serum and proangiogenic factors. We hypothesize that MEOX2 interacts with components of this pathway in vascular ECs to modulate NF-κB activity and EC activation and that these interactions depend upon specific domains within the MEOX2 protein. Methods and results To test our hypothesis, we transduced ECs with MEOX2 expression constructs. MEOX2 protein localized to the nuclear fraction, as did IκBβ and p65. By co-immunoprecipitation, MEOX2 bound to both p65 and IκBβ. Immuno-fluorescence demonstrated that MEOX2 colocalizes in the nucleus with both p65 and IκBβ and that this colocalization requires the MEOX2 homeodomain and N-terminal domain. Finally, promoter assays revealed that MEOX2 expression has a biphasic effect on NF-κB-dependent promoters. At low levels, MEOX2 stimulates NF-κB activity, whereas at high levels, it represses, effects that also depend upon the homeodomain and the N-terminal domain. Conclusion Our results represent the first report of an interaction between a homeobox protein and IκBβ and suggest that MEOX2 modulates the activity of the RelA complex through direct interaction with its components. These observations implicate MEOX2 as a potentially important regulatory gene inhibiting not only the angiogenic response of ECs to proangiogenic factors, but also their response to chronic inflammatory stimulation that normally activates NF-κB, suggesting MEOX2 as a possible molecular target for the therapy of angiogenesis-dependent diseases such as cancer. [ABSTRACT FROM AUTHOR]

Abstract: While the classical pathway of NF-κB activation plays critical roles in a wide range of biological processes, the more recently described “non-canonical” NF-κB pathway has important but more restricted roles in both normal and pathological processes. The non-canonical NF-κB pathway, based on processing of the nf-κb2 gene product p100 to generate p52, appears to be involved in B-cell maturation and lymphoid development. Deregulated activation of this pathway has been observed in a variety of malignant and autoimmune diseases, thus inhibitors that specifically target p100 processing might be predicted to have potential roles as immunomodulators and in the therapy of malignant diseases. We review current understandings of NF-κB activation, particularly the mechanisms of p100 processing under both physiological and pathological conditions. [Copyright &y& Elsevier]

The aryl hydrocarbon (Ah) receptor mediates most of the toxic effects induced by 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) and related compounds, which are ubiquitous environmental contaminants causing toxic responses in human and wildlife. Nuclear factor kappa B (NF-κB) is a pleiotropic transcription factor that plays a pivotal role in a wide array of physiological and pathological responses including immune modulation, inflammatory responses and apoptosis. Many physiological functions adversely affected by TCDD are also known to be regulated by NF-κB, such as immune activation, maintenance of skin differentiation, control of cell proliferation and survival, as well as induction of xenobiotic metabolizing enzymes. In the past few years, evidence has emerged to show that the Ah receptor and NF-κB interact and transcriptionally modulate each other. This review discusses Ah receptor–NF-κB interactions and examines potential mechanistic explanations for toxic responses as a result of TCDD exposure and the suppression of cytochrome P450 1A1/1A2 by stress stimuli such as inflammation and infection. [Copyright &y& Elsevier]

Genomic instability is a hallmark of cancer development and progression, and characterizing the stresses that create and the mechanisms by which cells respond to genomic perturbations is essential. Here we demonstrate that antiapoptotic BCL-2 family proteins promoted tumor formation of transformed baby mouse kidney (BMK) epithelial cells by antagonizing BAX- and BAK-dependent apoptosis. Cell death in vivo correlated with hypoxia and induction of PUMA (p53 up-regulated modulator of apoptosis). Strikingly, carcinomas formed by transformed BMK cells in which apoptosis was blocked by aberrant BCL-2 family protein function displayed prevalent, highly polyploid, tumor giant cells. Examination of the transformed BMK cells in vivo revealed aberrant metaphases and ploidy changes in tumors as early as 9 d after implantation, which progressed in magnitude during the tumorigenic process. An in vitro ischemia system mimicked the tumor microenvironment, and gain of BCL-2 or loss of BAX and BAK was sufficient to confer resistance to apoptosis and to allow for accumulation of polyploid cells in vitro. These data suggest that in vivo, even in cells in which p53 function is compromised, apoptosis is an essential response to hypoxia and ischemia in the tumor microenvironment and that abrogation of this response allows the survival of cells with abnormal genomes and promotes tumorigenesis. [ABSTRACT FROM AUTHOR]

Viral and parasitic coinfections are known to lead to both enhanced disease progression and altered disease states. HTLV-1 and Strongyloides stercoralis are co-endemic throughout much of their worldwide ranges resulting in a significant incidence of coinfection. Independently, HTLV-1 induces a Th1 response and S. stercoralis infection induces a Th2 response. However, coinfection with the two pathogens has been associated with the development of S. stercoralis hyperinfection and an alteration of the Th1/Th2 balance. In this study, a model of HTLV-1 and S. stercoralis coinfection in CD34+ umbilical cord blood hematopoietic stem cell engrafted humanized mice was established. An increased level of mortality was observed in the HTLV-1 and coinfected animals when compared to the S. stercoralis infected group. The mortality was not correlated with proviral loads or total viral RNA. Analysis of cytokine profiles showed a distinct shift towards Th1 responses in HTLV-1 infected animals, a shift towards Th2 cytokines in S. stercoralis infected animals and elevated TNF-α responses in coinfected animals. HTLV-1 infected and coinfection groups showed a significant, yet non-clonal expansion of the CD4+CD25+ T-cell population. Numbers of worms in the coinfection group did not differ from those of the S. stercoralis infected group and no autoinfective larvae were found. However, infective larvae recovered from the coinfection group showed an enhancement in growth, as was seen in mice with S. stercoralis hyperinfection caused by treatment with steroids. Humanized mice coinfected with S. stercoralis and HTLV-1 demonstrate features associated with human infection with these pathogens and provide a unique opportunity to study the interaction between these two infections in vivo in the context of human immune cells. Author summary: Human infections with the virus HTLV-1 and the nematode Strongyloides stercoralis are co-endemic throughout much of their worldwide ranges resulting in a significant incidence of coinfection. The coincident infections result in S. stercoralis hyperinfection and an alteration of the balance of the immune responses to the virus and to the worm. In this study, a novel model of HTLV-1 and S. stercoralis coinfection was established in humanized mice. Mortality was increased in the HTLV-1 and coinfected animals as compared to the S. stercoralis infected group. Analysis of cytokine profiles showed a distinct shift towards Th1 responses in HTLV-1 infected animals, a shift towards Th2 cytokines in S. stercoralis infected animals and elevated TNF-α responses in coinfected animals. Numbers of worms in the coinfection group did not differ from those of the S. stercoralis infected group, however, infective larvae recovered from the coinfection group showed a significant enhancement in growth. Humanized mice coinfected with S. stercoralis and HTLV-1 demonstrate features associated with human infection with these pathogens and provide a unique opportunity to study the interaction between these two infections. [ABSTRACT FROM AUTHOR]

Presents a study on inhibition of HIV-1 replication by a Tat RNA-binding domain peptide analog. Methodology used to conduct study; Details on study conducted; Findings of study; Discussion on findings.

Regulation of expression of HTLV-1 gene products from integrated proviruses plays an important role in HTLV-1-associated disease pathogenesis. Previous studies have shown that T cell receptor (TCR)- and phorbol ester (PMA) stimulation of chronically infected CD4 T cells increases the expression of integrated HTLV-1 proviruses in latently infected cells, however the mechanism remains unknown. Analysis of HTLV-1 RNA and protein species following PMA treatment of the latently HTLV-1-infected, FS and SP T cell lines demonstrated rapid induction of tax/rex mRNA. This rapid increase in tax/rex mRNA was associated with markedly enhanced tax/rex mRNA stability while the stability of unspliced or singly spliced HTLV-1 RNAs did not increase. Tax/rex mRNA in the HTLV-1 constitutively expressing cell lines exhibited high basal stability even without PMA treatment. Our data support a model whereby T cell activation leads to increased HTLV-1 gene expression at least in part through increased tax/rex mRNA stability. [ABSTRACT FROM AUTHOR]

The article discusses a randomized phase-two study which determines whether changes in nuclear factor kappa B (NF-ƘB) activity corresponded with changes in the expression of NF-ƘB target genes. In the said study, the newly diagnosed patients with acute myelogenous leukemia (AML) eligible for standard induction chemotherapy were assigned to choline magnesium trisalicylate (CMT) or no CMT.

Mesenchymal stem cells (MSC) are present in most, if not all, tissues and are believed to contribute to tissue regeneration and the tissue immune microenvironment. Murine MSCs exert immunosuppressive effects through production of inducible nitric oxide synthase (iNOS), whereas human MSCs use indoleamine 2,3-dioxygenase (IDO). Thus, studies of MSC-mediated immunomodulation in mice may not be informative in the setting of human disease, although this critical difference has been mainly ignored. To address this issue, we established a novel humanized system to model human MSCs, using murine iNOS-/- MSCs that constitutively or inducibly express an ectopic human IDO gene. In this system, inducible IDO expression is driven by a mouse iNOS promoter that can be activated by inflammatory cytokine stimulation in a similar fashion as the human IDO promoter. These IDO-expressing humanized MSCs (MSC-IDO) were capable of suppressing T-lymphocyte proliferation in vitro. In melanoma and lymphoma tumor models, MSC-IDO promoted tumor growth in vivo, an effect that was reversed by the IDO inhibitor 1-methyl-tryptophan. We found that MSC-IDO dramatically reduced both tumor-infiltrating CD8+ Tcells andB cells. Our findings offer an important new line of evidence that interventional targeting of IDO activity could be used to restore tumor immunity in humans, by relieving IDO-mediated immune suppression of MSCs in the tumor microenvironment as well as in tumor cells themselves. [ABSTRACT FROM AUTHOR]

Disruption of adenylyl cyclase type 5 ( AC5) knockout ( KO) is a novel model for longevity. Because malignancy is a major cause of death and reduced lifespan in mice, the goal of this investigation was to examine the role of AC5 KO in protecting against cancer. There have been numerous discoveries in genetically engineered mice over the past several decades, but few have been translated to the bedside. One major reason is that it is difficult to alter a gene in patients, but rather a pharmacological approach is more appropriate. The current investigation employs a parallel construction to examine the extent to which inhibiting AC5, either in a genetic knockout ( KO) or by a specific pharmacological inhibitor protects against cancer. This study is unique, not only because a combined genetic and pharmacological approach is rare, but also there are no prior studies on the extent to which AC5 affects cancer. We found that AC5 KO delayed age-related tumor incidence significantly, as well as protecting against mammary tumor development in AC5 KO × MMTV- HER-2 neu mice, and B16F10 melanoma tumor growth, which can explain why AC5 KO is a model of longevity. In addition, a Food and Drug Administration approved antiviral agent, adenine 9-β-D-arabinofuranoside (Vidarabine or AraAde), which specifically inhibits AC5, reduces LP07 lung and B16F10 melanoma tumor growth in syngeneic mice. Thus, inhibition of AC5 is a previously unreported mechanism for prevention of cancers associated with aging and that can be targeted by an available pharmacologic inhibitor, with potential consequent extension of lifespan. [ABSTRACT FROM AUTHOR]

Mesenchymal stem cells (MSCs), also called multipotent mesenchymal stromal cells, exist in almost all tissues and are a key cell source for tissue repair and regeneration. Under pathological conditions, such as tissue injury, these cells are mobilized towards the site of damage. Tissue damage is usually accompanied by proinflammatory factors, produced by both innate and adaptive immune responses, to which MSCs are known to respond. Indeed, recent studies have shown that there are bidirectional interactions between MSCs and inflammatory cells, which determine the outcome of MSC-mediated tissue repair processes. Although many details of these interactions remain to be elucidated, we provide here a synthesis of the current status of this newly emerging and rapidly advancing field. [ABSTRACT FROM AUTHOR]

We investigated the effects of caloric restriction (CR) on growth of tumors and metastases in the 4T1 mammary tumor model and found that CR, compared with normal diet, reduced the growth of mammary tumors and metastases and the total number of metastases that originated both spontaneously from the primary tumor and also experimentally from i.v. injection of the tumor cells. CR also decreased proliferation and angiogenesis and increased apoptosis in tumors. CR reduced levels of insulin, leptin, insulin-like growth factor 1, insulin-like growth factor binding protein 3 and increased adiponectin in tumors. We also demonstrated that tumors from CR mice possessed lower levels of transforming growth factor-β, lower intratumor deposition of collagen IV and reduced invasiveness due to a decrease in tumor secretion of active matrix metalloproteinase 9. Our results suggest that CR-induced metabolic and signaling changes affect the stroma and the tumor cells resulting in a microenvironment that prevents proliferation of breast tumors and their metastases. [ABSTRACT FROM AUTHOR]

It is a long-standing observation that inflammatory responses and infections decrease drug metabolism capacity in human and experimental animals. Cytochrome P-450 3A4 cyp304 is responsible for the metabolism of over 50% of current prescription drugs, and cyp3a4 expression is transcriptionally regulated by pregnane X receptor (PXR), which is a ligand-dependent transcription factor. In this study, we report that NF-κB activation by lipopolysaccharide and tumor necrosis factor-a plays a pivotal role in the suppression of cyp3a4 through interactions of NF-κB with the PXR-retinoid X receptor (RXR) complex. Inhibition of NF-κB by NF-κB-specific suppressor SRIκBα reversed the suppressive effects of lipopolysaccharide and tumor necrosis factor-α. Furthermore, we showed that NF-κB p65 disrupted the association of the PXR-RXRα complex with DNA sequences as determined by electrophoretic mobility shift assay and chromatin immunoprecipitation assays. NF-κB p65 directly interacted with the DNA-binding domain of RXRα and may prevent its binding to the consensus DNA sequences, thus inhibiting the transactivation by the PXR-RXRα complex. This mechanism of suppression by NF-κB activation may be extended to other nuclear receptor-regulated systems where RXRα is a dimerization partner. [ABSTRACT FROM AUTHOR]

Mantle cell lymphoma (MCL) is a distinct form of non-Hodgkin''s lymphoma (NHL) derived from CD5+ B cells. MCL cells overexpress cyclin D1 as a consequence of translocation of the gene into the immunoglobulin heavy-chain gene locus. MCL is an aggressive form of NHL with frequent relapses after standard-dose chemotherapy. In this context, a variety of novel therapies for patients with MCL have been investigated. In this study, we use an expanded panel of attenuated adenoviruses to study adenovirus-mediated cytotoxicity of MCL cells. Our results demonstrate: 1) adenovirus infection of MCL cells despite the absence of receptor/coreceptor molecules known to be important for adenovirus infection of other cells types; 2) cytotoxicity of MCL cells after infection with specific adenovirus mutants; 3) a high degree of cytotoxicity after infection of some patient samples with viruses lacking the E1B 19k “antiapoptotic” gene; and 4) cytotoxicity after infection with viruses containing mutations in E1A pRb or p300 binding. The extent of cytotoxicity with the panel of viruses demonstrated interpatient variability, but 100% cytotoxicity, as determined by molecular analysis, was detected in some samples. These studies provide the foundation for: 1) the development of adenoviruses as cytotoxic agents for MCL and 2) analyses of key regulatory pathways operative in MCL cells. [Copyright &y& Elsevier]

Abstract: bcl-3 is a protooncogene which undergoes chromosomal translocation in a subset of chronic B-cell lymphocytic leukemia cells. Bcl-3 is a unique IκB family protein that regulates transcription of a number of NF-κB target genes through interactions with NF-κB dimers. Based on previous studies, suggesting that Bcl-3 interacts with the Fyn tyrosine kinase in platelets, we investigated possible interactions of Bcl-3 with Lck, a related tyrosine kinase important in lymphoid cells. Protein–protein interactions between Bcl-3 and the Lck tyrosine kinase were identified both in vitro and in vivo. Lck enhanced Bcl-3-mediated activation of a p52/Bcl-3-responsive promoter in reporter gene assays independent of its tyrosine kinase activity, but requiring the Lck SH3 protein interaction domain. These studies suggest that Bcl-3 might participate in oncogenic pathways involving Lck. [Copyright &y& Elsevier]

Abstract: Human T cell leukemia virus 1 (HTLV-1) gene expression is regulated by both the viral Tax protein and by cellular transcriptional factors. We have previously shown that immune activation stimuli such as phorbol esters (PMA) and phytohemagglutinin (PHA) cooperate with HTLV-1 Tax expression to enhance HTLV-1 gene expression in infected T cells through increased activity of the HTLV-1 LTR. We now extend these studies to demonstrate roles for the T cell receptor complex, Lck, and Ras molecules in the coactivation of the HTLV-1 LTR by Tax and T cell activation stimuli. We also observe coactivation of Tax-responsive cellular promoter elements containing NF-κB and serum response factor (SRF) binding sites by Tax and T cell activation stimuli. These results suggest a model whereby T cell receptor stimulation and Tax expression coactivate HTLV-1 gene expression and cellular gene expression, enhancing activation of latent HTLV-1 and expression of cellular genes involved in disease pathogenesis. [Copyright &y& Elsevier]

RNA degradation via the ribonuclease H (RNase H) activity of human immunodeficiency virus type I (HIV-1) reverse transcriptase (RT) is a critical component of the reverse transcription process. In this connection, mutations of RT that inactivate RNase H activity result in noninfectious virus particles. Thus, interfering with the RNase H activity of RT represents a potential vehicle for the inhibition of HIV-1 replication. Here, we demonstrate an approach for inhibiting the RNase H activity of HIV-1 RT by targeting its RNADNA hybrid substrates. Specifically, we show that the binding of the 4,5-disubstituted 2-deoxystreptamine aminoglycosides, neomycin, paromomycin, and ribostamycin, to two different chimeric RNA-DNA duplexes, which mimic two distinct intermediates in the reverse transcription process, inhibits specific RT-mediated RNase H cleavage, with this inhibition being competitive in nature. UV melting and isothermal titration calorimetry studies reveal a correlation between the relative binding affinities of the three drugs for each of the chimeric RNA-DNA host duplexes and the relative extents to which the drugs inhibit RT-mediated RNase H cleavage of the duplexes. Significantly, this correlation also extends to the relative efficacies with which the drugs inhibit HIV- 1 replication. In the aggregate, our results highlight a potential strategy for AIDS chemotherapy that should not be compromised by the unusual genetic diversity of HIV- 1. [ABSTRACT FROM AUTHOR]

Describes the adenovirus infection of primary malignant lymphoid cells. Induction of host cell pro-apoptotic responses; Mechanism of cell cycle regulation; Activation of regulatory pathways in malignant cells.

C-terminal truncations of the NFKB2 p100 gene product have been observed in a number of cases of human cutaneous T cell lymphomas, as well as human B-cell lymphomas and myelomas. The contribution of these alterations to lymphomagenesis is not understood; however, truncation at amino acid 666 to generate 80–85 kD proteins in the HUT78 cell line is associated with addition of a short (serine-alanine-serine) fusion at the 3′ end of p80HT, as well as with increased expression of NFKB2 mRNA. We therefore examined the effects of p80HT on the regulation of NFKB2 expression, as well as the properties of a series of other tumor-associated, and site directed mutations of NFKB2. While p80HT had not itself acquired novel transcriptional activation properties with respect to the NFKB2 P1 or P2 promoters or the IL-6 κB promoter, p80HT had lost the potent inhibitory (IκB-like) activity associated with the wild-type, p100 gene product. Loss of the inhibitory property depended on the SAS residues in the fusion protein, direct truncation at aa666 was fully inhibitory, as was a substitution of three alanines for the SAS residues. The presence of as few as two C-terminal ankyrin motifs was sufficient for inhibition of NF-κB-mediated transcriptional activation. Assays of a series of additional lymphoma-associated NF-κB-2 truncation suggested that the C-terminal truncation associated with these proteins was also associated with a loss of the IκB-like activities of p100 NF-κB-2, for at least some NF-κB target promoters. Thus, the loss of IκB-like activity of lymphoma-associated NFKB2 mutations may play an important role in the genesis of a subset of human lymphomas. Oncogene (2000) 19, 1334–1345. [ABSTRACT FROM AUTHOR]

Programmed cell death-2 (PDCD2) protein is enriched in embryonic, hematopoietic, and neural stem cells, however, its function in stem/progenitor cell differentiation is unclear. We investigated the effects of PDCD2 knockdown on the development and differentiation of hematopoietic progenitor cells (HPC). CD34+ cells derived from normal human bone marrow and K562 leukemic cells were effectively transduced with short-hairpin RNA to knockdown PDCD2. Colony-forming assays were used to investigate the effects of PDCD2 loss on HPC clonogenic potential and on 12-O-tetradecanoyl-phorbol-13-acetate–and arabinofuranosylcytosine–induced terminal differentiation. In CD34+ clonogenic progenitors, PDCD2 knockdown decreased the total number of colony-forming units, increased the number of colony-forming units–granulocyte-erythroid-macrophage-megakaryocyte and burst-forming unit–erythroid primitive colonies, and decreased the number of burst-forming unit–erythroid mature colonies. Similar results were observed in K562 cells, suggesting that PDCD2 is important for HPC differentiation and/or survival, and for erythroid lineage commitment. Furthermore, 12-O-tetradecanoyl-phorbol-13-acetate–induced megakaryocytic differentiation and proliferation of K562 cells was not affected by PDCD2 knockdown. In contrast, arabinofuranosylcytosine-induced erythroid differentiation of K562 cells was significantly reduced with PDCD2 knockdown, with no effect on cell proliferation. The effects of PDCD2 knockdown were attributed to a cell cycle arrest at G0/G1, along with increased messenger RNA expression of early progenitor factors c-MYB and GATA-2, and decreased expression of erythroid factors GATA-1, EpoR, and γ -globin. We conclude that PDCD2 loss of function(s) impedes erythroid differentiation by inducing cell cycle arrest and increasing expression of early hematopoietic progenitor factors. These findings suggest that PDCD2 has a novel regulatory role in human hematopoiesis and is essential for erythroid development. [Copyright &y& Elsevier]

An abstract of the research paper titled,"Coordination of the canonical and noncanonical IKK/NF-kB signaling pathways in the Human T-cell leukemia virus type 1 (HTLV-1) Taxmediated tumorigenesis," is presented.

An abstract of the research paper titled " Activation of HTLV-1 Expression in Chronically-Infected CD4+ T cells: Mechanisms and Implications for Pathogenesis," by Alison Swaims, Hsin-Ching Lin, Peter Simon, Celine Granier, Yingyu Zhang, Arthur Roberts and Satish Devadas is presented.

Tumors secrete proangiogenic factors to induce the ingrowth of blood vessels from the stroma. These peptides bind to cell surface receptors on vascular endothelial cells (ECs), triggering signaling cascades that activate and repress batteries of downstream genes responsible for the angiogenic phenotype. To determine if microRNAs (miRNAs) affect regulation of the EC phenotype by GAX, a homeobox gene and negative transcriptional regulator of the angiogenic phenotype, we tested the effect of miR-221 on GAX expression. miR-221 strongly upregulated GAX, suggesting that miR-221 downregulates a repressor of GAX. We next expressed miR-221 in ECs and identified ZEB2, a modulator of the epithelial-mesenchymal transition, as being strongly downregulated by miR-221. Using miR-221 expression constructs and an inhibitor, we determined that ZEB2 is upregulated by serum and downregulates GAX, while the expression of miR-221 upregulates GAX and downregulates ZEB2. A mutant miR-221 fails to downregulate ZEB2 or upregulate GAX. Finally, using chromatin immunoprecipitation, we identified two ZEB2 binding sites that modulate the ability of ZEB2 to downregulate GAX promoter activity. We conclude that miR-221 upregulates GAX primarily through its ability to downregulate the expression of ZEB2. These observations suggest a strategy for inhibiting angiogenesis by either recapitulating miR-221 expression or inhibiting ZEB2 activation. [ABSTRACT FROM AUTHOR]