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The rationale for developing histone deacetylase (HDAC) inhibitors (HDACi) as anticancer agents was based on their ability to induce apoptosis and cell cycle arrest in cancer cells. However, while HDACi have been remarkably effective in the treatment of hematological malignancies, clinical studies with HDACi as single agents in solid cancers have been disappointing. Recent studies have shown that, in addition to inducing apoptosis in cancer cells, class I HDACi induce IκB kinase (IKK)-dependent expression of proinflammatory chemokines, such as interleukin-8 (IL8; CXCL8), resulting in the increased proliferation of tumor cells, and limiting the effectiveness of HDACi in solid tumors. Here, we discuss the mechanisms responsible for HDACi-induced CXCL8 expression, and opportunities for combination therapies targeting HDACs and IKK in solid tumors. [ABSTRACT FROM AUTHOR]

The advanced stages of cutaneous T cell lymphoma (CTCL) are characterized not only by decreased levels of pro-inflammatory cytokines, resulting in high susceptibility to infections, but also by high constitutive activity of NFκB, which promotes cell survival and resistance to apoptosis. The increased expression of the proto-oncogene Bcl3 belonging to IκB family is associated with the pathogenesis of the different types of human cancer, yet, the function and regulation of Bcl3 in CTCL have not been studied. Here, we show that Bcl3 is highly expressed in CTCL Hut-78 and HH cells. The suppression of Bcl3 levels decreases the expression of the pro-survival genes cIAP1 and cIAP2, reduces cell viability, and increases CTCL apoptosis. Interestingly, Bcl3 suppression concomitantly increases expression and the release of the pro-inflammatory cytokines IL-8 and IL-17 in CTCL cells. Chromatin immunoprecipitation studies show that Bcl3 regulates cIAP1, cIAP2, IL-8 and IL-17 gene expression through direct binding to their promoters. Bcl3 expression is regulated by bortezomib (BZ)-mediated proteasome inhibition, and BZ inhibits Bcl3 recruitment to its target promoters, resulting in decreased expression of cIAP1 and cIAP2, but increased expression of IL-8 and IL-17. The Bcl3 expression is regulated through NFκB subunit exchange on Bcl3 promoter. In untreated cells, the Bcl3 promoter is occupied predominantly by p65/p50 heterodimers, inducing Bcl3 expression; however, in BZ-treated cells, the p65/50 heterodimers are replaced by p52 subunits, resulting in Bcl3 transcriptional repression. These data provide the first insights into the function and regulation of Bcl3 in CTCL, and indicate that Bcl3 has an important pro-survival and immunosuppressive role in these cells. [ABSTRACT FROM AUTHOR]

Simple Summary: IFNγ is a multifunctional cytokine produced not only by activated lymphocytes but also in response to cancer immunotherapies; it has both antitumor and tumor-promoting effects. In ovarian cancer cells, the tumor-promoting effects of IFNγ are mediated by increased expression of the proto-oncogene Bcl3, the immune checkpoint PD-L1 and the proinflammatory cytokine IL-8. Recent studies have shown that IFNγ induces the expression of Bcl3, which then promotes the expression of PD-L1 and IL-8 in ovarian cancer cells, resulting in their increased proliferation and migration. In this review, we summarize the recent findings on the IFNγ tumor-promoting functions and on the mechanisms by which IFNγ induces Bcl3, PD-L1 and IL-8 expression in cancer cells, with a special focus on ovarian cancer. We also highlight the importance of a better understanding of these mechanisms to optimize the development of combinatorial approaches in cancer immunotherapies. IFNγ, a pleiotropic cytokine produced not only by activated lymphocytes but also in response to cancer immunotherapies, has both antitumor and tumor-promoting functions. In ovarian cancer (OC) cells, the tumor-promoting functions of IFNγ are mediated by IFNγ-induced expression of Bcl3, PD-L1 and IL-8/CXCL8, which have long been known to have critical cellular functions as a proto-oncogene, an immune checkpoint ligand and a chemoattractant, respectively. However, overwhelming evidence has demonstrated that these three genes have tumor-promoting roles far beyond their originally identified functions. These tumor-promoting mechanisms include increased cancer cell proliferation, invasion, angiogenesis, metastasis, resistance to chemotherapy and immune escape. Recent studies have shown that IFNγ-induced Bcl3, PD-L1 and IL-8 expression is regulated by the same JAK1/STAT1 signaling pathway: IFNγ induces the expression of Bcl3, which then promotes the expression of PD-L1 and IL-8 in OC cells, resulting in their increased proliferation and migration. In this review, we summarize the recent findings on how IFNγ affects the tumor microenvironment and promotes tumor progression, with a special focus on ovarian cancer and on Bcl3, PD-L1 and IL-8/CXCL8 signaling. We also discuss promising novel combinatorial strategies in clinical trials targeting Bcl3, PD-L1 and IL-8 to increase the effectiveness of cancer immunotherapies. [ABSTRACT FROM AUTHOR]

The article discusses research on the therapeutic use of dimethyl fumarate (DMF) in cutaneous T-cell lymphoma (CTCL). Topics discussed include therapeutic option DMF in CTCL treatment, the characteristic of malignant T cells, the restoration of apoptosis in CTCL cells using DMF, and the clinical trial for CTCL patients.

AMP-activated protein kinase (AMPK) is an energy sensor and master regulator of metabolism. AMPK functions as a fuel gauge monitoring systemic and cellular energy status. Activation of AMPK occurs when the intracellular AMP/ATP ratio increases and leads to a metabolic switch from anabolism to catabolism.AMPKphosphorylates and inhibits acetyl-CoA carboxylase (ACC), which catalyzes carboxylation of acetyl-CoA to malonyl-CoA, the first and rate-limiting reaction in de novo synthesis of fatty acids.AMPKthus regulates homeostasis of acetyl- CoA, a key metabolite at the crossroads of metabolism, signaling, chromatin structure, and transcription. Nucleocytosolic concentration of acetyl-CoA affects histone acetylation and links metabolism and chromatin structure. Here we show that activation of AMPK with the widely used antidiabetic drug metformin or with the AMP mimetic 5-aminoimidazole- 4-carboxamide ribonucleotide increases the inhibitory phosphorylation of ACC and decreases the conversion of acetyl-CoA to malonyl-CoA, leading to increased protein acetylation and altered gene expression in prostate and ovarian cancer cells. Direct inhibition of ACC with allosteric inhibitor 5-(tetradecyloxy)- 2-furoic acid also increases acetylation of histones and non-histone proteins. Because AMPK activation requires liver kinase B1, metformin does not induce protein acetylation in liver kinase B1-deficient cells. Together, our data indicate that AMPK regulates the availability of nucleocytosolic acetyl-CoA for protein acetylation and that AMPK activators, such as metformin, have the capacity to increase protein acetylation and alter patterns of gene expression, further expanding the plethora of metformin's physiological effects. [ABSTRACT FROM AUTHOR]

Increased expression of the immunosuppressive cytokines, TGF-β1 and IL-10, is a hallmark of the advanced stages of cutaneous T cell lymphoma (CTCL), where it has been associated with suppressed immunity, increased susceptibility to infections, and diminished antitumor responses. Yet, little is known about the transcriptional regulation of TGF-β1 and IL-10 in CTCL, and about their function in regulating the CTCL cell responses. In this article, we show that TGF-β1 and IL-10 expression in CTCL cells is regulated by NF-κB and suppressed by bortezomib (BZ), which has shown promising results in the treatment of CTCL. However, although the TGF-β1 expression is IκBα dependent and is regulated by the canonical pathway, the IL-10 expression is IκBα independent, and its inhibition by BZ is associated with increased promoter recruitment of p52 that characterizes the noncanonical pathway. TGF-β1 suppression decreases CTCL cell viability and increases apoptosis, and adding exogenous TGF-β1 increases viability of BZ-treated CTCL cells, indicating TGF-β1 prosurvival function in CTCL cells. In addition, TGF-β1 suppression increases expression of the proinflammatory cytokines IL-8 and IL-17 in CTCL cells, suggesting that TGF-β1 also regulates the IL-8 and IL-17 expression. Importantly, our results demonstrate that BZ inhibits expression of the chemokine receptor CXCR4 in CTCL cells, resulting in their decreased migration, and that the CTCL cell migration is mediated by TGF-β1. These findings provide the first insights into the BZ-regulated TGF-β1 and IL-10 expression in CTCL cells, and indicate that TGF-β1 has a key role in regulating CTCL survival, inflammatory gene expression, and migration. [ABSTRACT FROM AUTHOR]

The increased expression of pro-inflammatory and pro-angiogenic chemokines contributes to ovarian cancer progression through the induction of tumor cell proliferation, survival, angiogenesis, and metastasis. The substantial potential of these chemokines to facilitate the progression and metastasis of ovarian cancer underscores the need for their stringent transcriptional regulation. In this Review, we highlight the key mechanisms that regulate the transcription of pro-inflammatory chemokines in ovarian cancer cells, and that have important roles in controlling ovarian cancer progression. We further discuss the potential mechanisms underlying the increased chemokine expression in drug resistance, along with our perspective for future studies. [ABSTRACT FROM AUTHOR]

Heme is an essential cofactor for enzymes of the electron transport chain (ETC) and ATP synthesis in mitochondrial oxidative phosphorylation (OXPHOS). Heme also binds to and destabilizes Bach1, a transcription regulator that controls expression of several groups of genes important for glycolysis, ETC, and metastasis of cancer cells. Heme synthesis can thus affect pathways through which cells generate energy and precursors for anabolism. In addition, increased heme synthesis may trigger oxidative stress. Since many cancers are characterized by a high glycolytic rate regardless of oxygen availability, targeting glycolysis, ETC, and OXPHOS have emerged as a potential therapeutic strategy. Here, we report that enhancing heme synthesis through exogenous supplementation of heme precursor 5-aminolevulinic acid (ALA) suppresses oxidative metabolism as well as glycolysis and significantly reduces proliferation of both ovarian and breast cancer cells. ALA supplementation also destabilizes Bach1 and inhibits migration of both cell types. Our data indicate that the underlying mechanisms differ in ovarian and breast cancer cells, but involve destabilization of Bach1, AMPK activation, and induction of oxidative stress. In addition, there appears to be an inverse correlation between the activity of oxidative metabolism and ALA sensitivity. Promoting heme synthesis by ALA supplementation may thus represent a promising new anti-cancer strategy, particularly in cancers that are sensitive to altered redox signaling, or in combination with strategies that target the antioxidant systems or metabolic weaknesses of cancer cells. [ABSTRACT FROM AUTHOR]

Proliferating cells coordinate histone and DNA synthesis to maintain correct stoichiometry for chromatin assembly. Histone mRNA levels must be repressed when DNA replication is inhibited to prevent toxicity and genome instability due to free non-chromatinized histone proteins. In mammalian cells, replication stress triggers degradation of histone mRNAs, but it is unclear if this mechanism is conserved from other species. The aim of this study was to identify the histone mRNA decay pathway in the yeast Saccharomyces cerevisiae and determine the mechanism by which DNA replication stress represses histone mRNAs. Using reverse transcription-quantitative PCR and chromatin immunoprecipitation-quantitative PCR, we show here that histone mRNAs can be degraded by both 5' → 3' and 3' → 5' pathways; however, replication stress does not trigger decay of histone mRNA in yeast. Rather, replication stress inhibits transcription of histone genes by removing the histone gene-specific transcription factors Spt10p and Spt21p from histone promoters, leading to disassembly of the preinitiation complexes and eviction of RNA Pol II from histone genes by a mechanism facilitated by checkpoint kinase Rad53p and histone chaperone Asf1p. In contrast, replication stress does not remove SCB-binding factor transcription complex, another activator of histone genes, from the histone promoters, suggesting that Spt10p and Spt21p have unique roles in the transcriptional downregulation of histone genes during replication stress. Together, our data show that, unlike in mammalian cells, replication stress in yeast does not trigger decay of histone mRNAs but inhibits histone transcription. [ABSTRACT FROM AUTHOR]

The DNA damage response (DDR) is an evolutionarily conserved process essential for cell survival. Previously, we found that decreased histone expression induces mitochondrial respiration, raising the question whether the DDR also stimulates respiration. Here, using oxygen consumption and ATP assays, RT-qPCR and ChIP-qPCR methods, and dNTP analyses, we show that DDR activation in the budding yeast Saccharomyces cerevisiae, either by genetic manipulation or by growth in the presence of genotoxic chemicals, induces respiration. We observed that this induction is conferred by reduced transcription of histone genes and globally decreased DNA nucleosome occupancy. This globally altered chromatin structure increased the expression of genes encoding enzymes of tricarboxylic acid cycle, electron transport chain, oxidative phosphorylation, elevated oxygen consumption, and ATP synthesis. The elevated ATP levels resulting from DDR-stimulated respiration drove enlargement of dNTP pools; cells with a defect in respiration failed to increase dNTP synthesis and exhibited reduced fitness in the presence of DNA damage. Together, our results reveal an unexpected connection between respiration and the DDR and indicate that the benefit of increased dNTP synthesis in the face of DNA damage outweighs possible cellular damage due to increased oxygen metabolism. [ABSTRACT FROM AUTHOR]

The proto-oncogene Bcl3 induces survival and proliferation in cancer cells; however, its function and regulation in ovarian cancer (OC) remain unknown. Here, we show that Bcl3 expression is increased in human OC tissues. Surprisingly, however, we found that in addition to promoting survival, proliferation, and migration of OC cells, Bcl3 promotes both constitutive and interferon-γ (IFN)-induced expression of the immune checkpoint molecule PD-L1. The Bcl3 expression in OC cells is further increased by IFN, resulting in increased PD-L1 transcription. The mechanism consists of an IFN-induced, Bcl3- and p300-dependent PD-L1 promoter occupancy by Lys-314/315 acetylated p65 NF-kB. Blocking PD-L1 by neutralizing antibody reduces proliferation of OC cells overexpressing Bcl3, suggesting that the pro-proliferative effect of Bcl3 in OC cells is partly mediated by PD-L1. Together, this work identifies PD-L1 as a novel target of Bcl3, and links Bcl3 to IFNγ signaling and PD-L1-mediated immune escape. [ABSTRACT FROM AUTHOR]

Metformin has been a frontline therapy for type 2 diabetes (T2D) for many years. Its effectiveness in T2D treatment is mostly attributed to its suppression of hepatic gluconeogenesis; however, the mechanistic aspects of metformin action remain elusive. In addition to its glucose-lowering effect, metformin possesses other pleiotropic health-promoting effects that include reduced cancer risk and tumorigenesis. Metformin inhibits the electron transport chain (ETC) and ATP synthesis; however, recent data reveal that metformin regulates AMP-activated protein kinase (AMPK) and the mechanistic target of rapamycin complex 1 (mTORC1) by multiple, mutually nonexclusive mechanisms that do not necessarily depend on the inhibition of ETC and the cellular ATP level. In this review, we discuss recent advances in elucidating the molecular mechanisms that are relevant for metformin use in cancer treatment. Highlights Metformin use is associated with lower cancer incidence in diabetic patients and is considered for cancer therapy of non-diabetic patients. Metformin inhibits ETC and elevates AMP/ATP ratio, resulting in AMPK activation and mTORC1 inhibition. In addition, at metformin concentrations that do not affect the ETC and AMP/ATP ratio, administration of metformin results in AMPK activation and mTORC1 inhibition by mechanisms not involving ETC. Metformin-induced inhibition of ATP synthesis affects the nuclear pore complex, which leads to nuclear exclusion of RagC. Without being able to pass through the nucleus, RagC cannot be potentiated for mTORC1 activation. [ABSTRACT FROM AUTHOR]

Triple negative breast cancer (TNBC) cells express increased levels of the pro-inflammatory and pro-angiogenic chemokine interleukin-8 (IL-8, CXCL8), which promotes their proliferation and migration. Because TNBC patients are unresponsive to current targeted therapies, new therapeutic strategies are urgently needed. While proteasome inhibition by bortezomib (BZ) or carfilzomib (CZ) has been effective in treating hematological malignancies, it has been less effective in solid tumors, including TNBC, but the mechanisms are incompletely understood. Here we report that proteasome inhibition significantly increases expression of IL-8, and its receptors CXCR1 and CXCR2, in TNBC cells. Suppression or neutralization of the BZ-induced IL-8 potentiates the BZ cytotoxic and anti-proliferative effect in TNBC cells. The IL-8 expression induced by proteasome inhibition in TNBC cells is mediated by IκB kinase (IKK), increased nuclear accumulation of p65 NFκB, and by IKK-dependent p65 recruitment to IL-8 promoter. Importantly, inhibition of IKK activity significantly decreases proliferation, migration, and invasion of BZ-treated TNBC cells. These data provide the first evidence demonstrating that proteasome inhibition increases the IL-8 signaling in TNBC cells, and suggesting that IKK inhibitors may increase effectiveness of proteasome inhibitors in treating TNBC. [ABSTRACT FROM AUTHOR]

Overexpression of the pro-angiogenic chemokine IL-8 (CXCL8) is associated with a poor prognosis in several solid tumors, including epithelial ovarian cancer (EOC). Even though histone deacetylase (HDAC) inhibition has shown remarkable antitumor activity in hematological malignancies, it has been less effective in solid tumors, including EOC. Here we report results that may explain the decreased efficiency of HDAC inhibition in EOC, based on our data demonstrating that HDAC inhibition specifically induces expression of IL-8/CXCL8 in SKOV3, CAOV3, andOVCAR3cells. Suppression or neutralization of vorinostat-induced IL-8/CXCL8 potentiates the vorinostat inhibitory effect on cell viability and proliferation. The IL-8/CXCL8 expression induced by vorinostat in EOC cells is dependent on IκB kinase (IKK) activity and associated with a gene-specific recruitment of IKKβ and IKK-dependent recruitment of p65 NFκB to the IL-8/CXCL8 promoter. In addition, HDACinhibition induces acetylation of p65 and histone H3 and their IL-8/CXCL8 promoter occupancy. In vivo results demonstrate that combining vorinostat and the IKK inhibitor Bay 117085 significantly reduces tumor growth in nude mice compared with control untreated mice or either drug alone. Mice in the combination group had the lowest IL-8/CXCL8 tumor levels and the lowest tumor expression of the murine neutrophil [7/4] antigen, indicating reduced neutrophil infiltration. Together, our results demonstrate that HDAC inhibition specifically induces IL-8/CXCL8 expression in EOC cells and that the mechanism involves IKK, suggesting that using IKK inhibitors may increase the effectiveness of HDAC inhibitors when treating ovarian cancer and other solid tumors characterized by increased IL-8/CXCL8 expression. [ABSTRACT FROM AUTHOR]

Previously, we have shown that N,N-dimethylacetamide (DMA) prevents inflammation-induced preterm birth in a murine model, inhibits lipopolysaccharide (LPS)-induced increases in placental proinflammatory cytokines and upregulates the antiinflammatory cytokine interleukin-10 (IL-10). However, DMA's mechanism of action remains to be elucidated. In the current study, we investigate how DMA produces its antiinflammatory effect. Using in vitro and ex vivo models, we show that DMA suppresses secretion of proinflammatory cytokines in lipopolysaccharide (LPS)-induced RAW 264.7 cells, TNFα-challenged JEG-3 cells and LPS-stimulated human placental explants. DMA significantly attenuated secretion of TNFα, IL-6, IL-10 and granulocyte macrophage colony stimulating factor (GM-CSF) from LPS-stimulated RAW 264.7 cells; IL-6 secretion from TNFα-stimulated JEG-3 cells; and TNFα, IL-6, IL-10, GM-CSF and Interleukin-8 (IL-8) from LPS-stimulated human placental explants. We further investigated whether DMA's effect on cytokine expression involves the nuclear factor kappa-light-chain-enhancer of activated B cells (NF-κB) and mitogen-activated protein kinase (MAPK) pathways. DMA (10 mM) significantly inhibited degradation of nuclear factor of kappa light polypeptide gene enhancer in B cell inhibitor a (IκBα) in LPS-stimulated RAW 264.7 cells, but there was no significant change in the expression of phosphorylated or native forms of downstream proteins in the MAPK pathway. In addition, DMA significantly attenuated luciferase activity in cells co-transfected with NF-κB-Luc reporter plasmid, but not with AP-1-Luc or CEBP-Luc reporters. Overall, our findings suggest that the antiinflammatory activity of DMA is mediated by inhibition of the NF-κB pathway via decreased IκBa degradation. [ABSTRACT FROM AUTHOR]

Expression of the immune checkpoint programmed death ligand-1 (PD-L1) is increased in ovarian cancer (OC) and correlates with poor prognosis. Interferon-γ (IFNγ) induces PD-L1 expression in OC cells, resulting in their increased proliferation and tumor growth, but the mechanisms that regulate the PD-L1 expression in OC remain unclear. Here, we show that the IFNγ-induced PD-L1 expression in OC cells is associated with increased levels of STAT1, Tyr-701 pSTAT1 and Ser-727 pSTAT1. Suppression of JAK1 and STAT1 significantly decreases the IFNγ-induced PD-L1 expression in OC cells, and STAT1 overexpression increases the IFNγ-induced PD-L1 expression. In addition, IFNγ induces expression of the transcription factor interferon regulatory factor 1 (IRF1) and IRF1 suppression attenuates the IFNγ-induced gene and protein levels of PD-L1. Chromatin immunoprecipitation results show that IFNγ induces PD-L1 promoter acetylation and recruitment of STAT1, Ser-727 pSTAT1 and IRF1 in OC cells. Together, these findings demonstrate that the IFNγ-induced PD-L1 expression in OC cells is regulated by JAK1, STAT1, and IRF1 signaling, and suggest that targeting the JAK1/ STAT1/IRF1 pathway may provide a leverage to regulate the PD-L1 levels in ovarian cancer. • PDL1 levels are increased in ovarian cancer (OC) and correlate with poor prognosis. • IFNγ induces PDL1 expression in OC, but the mechanisms remain unknown. • The IFNγ induced PDL1 in OC cells is mediated by JAK1, STAT1 and IRF1. • Targeting JAK1/ STAT1/IRF1 may provide a leverage to regulate PD-L1 levels in OC. [ABSTRACT FROM AUTHOR]

Bortezomib (BZ) is the first clinically approved proteasome inhibitor that has shown remarkable anticancer activity in patients with hematological malignancies. However, many patients relapse and develop resistance; yet, the molecular mechanisms of BZ resistance are not fully understood. We have recently shown that in solid tumors, BZ unexpectedly increases expression of the pro-inflammatory and pro-angiogenic chemokine interleukin-8 (IL-8), while it inhibits expression of other NFκB-regulated genes. Since monocytes and macrophages are major producers of IL-8, the goal of this study was to test the hypothesis that BZ increases the IL-8 expression in human monocytes and macrophages. Here, we show that BZ dramatically increases the IL-8 expression in lipopolysaccharide (LPS)-stimulated U937 macrophages as well as in unstimulated U937 monocytes and peripheral blood mononuclear cells, while it inhibits expression of IL-6, IL-1 and tumor necrosis factor-α. In addition, our results show that the underlying mechanisms involve p38 mitogen-activated protein kinase, which is required for the BZ-induced IL-8 expression. Together, these data suggest that the BZ-increased IL-8 expression in monocytes and macrophages may represent one of the mechanisms responsible for the BZ resistance and indicate that targeting the p38-mediated IL-8 expression could enhance the BZ effectiveness in cancer treatment. [ABSTRACT FROM AUTHOR]

R3152 --> 667.11 --> The immune checkpoint programmed death ligand‐1 (PD‐L1) is highly expressed in cancer cells, including ovarian cancer (OC) cells. High PD‐L1 expression in ovarian cancer correlates with poor prognosis, but the mechanisms that regulate the PD‐L1 expression in OC remain poorly understood. We have recently shown that IFNγ induces the PD‐L1 expression in OC cells, resulting in their increased proliferation and invasion. Here, we show that the IFNγ‐induced PD‐L1 expression in OC cells is associated with increased levels of STAT1, Tyr‐701 pSTAT1 and Ser‐727 pSTAT1 in OC cells. JAK1 and STAT1 suppression significantly decreases the IFNγ‐induced PD‐L1 mRNA and protein levels in OC cells. In addition, IRF1 suppression significantly decreases the IFNγ‐induced gene and protein levels of PD‐L1, indicating that the IFNγ‐induced PD‐L1 expression in OC cells is dependent on IRF1 signaling. Chromatin immunoprecipitation data demonstrate that IFNγ induces occupancy of STAT1, Ser‐727 pSTAT1 and IRF1 at STAT1 and IRF1 binding sites in human PD‐L1 promoter. Together, these results show that the IFNγ‐induced PD‐L1 expression in OC cells is regulated by IRF1/JAK1/STAT1 signaling, and suggest that targeting the IRF1/JAK1/STAT1 pathway may provide leverage to regulate the PD‐L1 levels in ovarian cancer. [ABSTRACT FROM AUTHOR]

R3153 --> 666.5 --> The proto‐oncogene Bcl3 is an atypical member of IkB family, which regulates expression of a number of NFkB target genes, including anti‐apoptotic genes, cytokines, and immune checkpoints. Bcl3 expression is increased in many types of human cancers and is associated with increased tumor growth and malignant potential. Interferon‐γ (IFNγ) is a pleiotropic cytokine that has been used in cancer treatment; in addition, its expression is induced in response to radiation therapy and immune checkpoint blockade during cancer treatment. We have recently shown that IFNγ induces Bcl3 expression in ovarian cancer cells, resulting in their increased proliferation, migration, and invasion. In this study, we have investigated the mechanisms of how IFNγ induces the Bcl3 expression in ovarian cancer cells. Our results demonstrate that the IFNγ‐induced Bcl3 expression is dependent on JAK1 and STAT1 signaling, as well as on p65 NFkB. In addition, we have identified a potential interferon regulatory factor 1 (IRF1) binding site in human Bcl3 promoter and found that the IFNγ‐induced Bcl3 expression in ovarian cancer cells is associated with an increased occupancy of IRF1, Ser‐727 phosphorylated STAT1 and acetylated histone H3 at the IRF1 binding site in Bcl3 promoter. These findings identify Bcl3 as a novel target of IFNγ/JAK1/STAT1 signaling and suggest that targeting the JAK1/STAT1 pathway may suppress the IFNγ‐induced Bcl3 expression in ovarian cancer. [ABSTRACT FROM AUTHOR]

Proinflammatory and pro-angiogenic chemokine interleukin-8 (IL-8, CXCL8) contributes to ovarian cancer progression through its induction of tumor cell proliferation, survival, angiogenesis, and metastasis. Proteasome inhibition by bortezomib, which has been used as a frontline therapy in multiple myeloma, has shown only limited effectiveness in ovarian cancer and other solid tumors. However, the responsible mechanisms remain elusive. Here, we show that proteasome inhibition dramatically increases the IL-8 expression and release in ovarian cancer cells. The responsible mechanism involves an increased nuclear accumulation of IκB kinase β (IKKβ) and an increased recruitment of the nuclear IKKβ, p65-phosphorylated at Ser536, and the transcription factor early growth response-1 (EGR-1) to the endogenous IL-8 promoter. Coimmunoprecipitation studies identified the nuclear EGR-1 associated with IKKβ and with p65, with preferential binding to S536P-p65. Both IKKβ activity and EGR-1 expression are required for the increased IL-8 expression induced by proteasome inhibition in ovarian cancer cells. Interestingly, in multiple myeloma cells the IL-8 release is not increased by bortezomib. Together, these data indicate that the increased IL-8 release may represent one of the underlying mechanisms responsible for the decreased effectiveness of proteasome inhibition in ovarian cancer treatment and identify IKKβ and EGR-1 as potential new targets in ovarian cancer combination therapies. [ABSTRACT FROM AUTHOR]

Expression of the proinflammatory and proangiogenic chemokine IL-8, which is regulated at the transcriptional level by NF-κB, is constitutively increased in androgen-independent metastatic prostate cancer and correlates with poor prognosis. Inhibition of NF-κB–dependent transcription was used as an anticancer strategy for the development of the first clinically approved 26S proteasome inhibitor, bortezomib (BZ). Even though BZ has shown remarkable antitumor activity in hematological malignancies, it has been less effective in prostate cancer and other solid tumors; however, the mechanisms have not been fully understood. In this article, we report that proteasome inhibition by BZ unexpectedly increases IL-8 expression in androgen-independent prostate cancer PC3 and DU145 cells, whereas expression of other NF-κB–regulated genes is inhibited or unchanged. The BZ-increased IL-8 expression is associated with increased in vitro p65 NF-κB DNA binding activity and p65 recruitment to the endogenous IL-8 promoter. In addition, proteasome inhibition induces a nuclear accumulation of IκB kinase (IKK)α, and inhibition of IKKα enzymatic activity significantly attenuates the BZ-induced p65 recruitment to IL-8 promoter and IL-8 expression, demonstrating that the induced IL-8 expression is mediated, at least partly, by IKKα. Together, these data provide the first evidence, to our knowledge, for the gene-specific increase of IL-8 expression by proteasome inhibition in prostate cancer cells and suggest that targeting both IKKα and the proteasome may increase BZ effectiveness in treatment of androgen-independent prostate cancer. [ABSTRACT FROM AUTHOR]

Introduction: Increasing evidence now supports the association between the fetal inflammatory response syndrome (FIRS) with the pathogenesis of preterm labor, intraventricular hemorrhage and bronchopulmonary dysplasia. Polymorphonuclear leukocyte (PMNs) and mononuclear cell (MONOs) infiltration of the placenta is associated with these disorders. The aim of this study was to reveal cell-specific differences in gene expression and cytokine release in response to endotoxin that would elucidate inflammatory control mechanisms in the newly born. Methods: PMNs and MONOs were separately isolated from the same cord blood sample. A genome-wide microarray screened for gene expression and related pathways at 4 h of LPS stimulation (n = 5). RT-qPCR and ELISA were performed for selected cytokines at 4 h and 18 h of LPS stimulation. Results: Compared to PMNs, MONOs had a greater diversity and more robust gene expression that included pro- inflammatory (PI) cytokines, chemokines and growth factors at 4 h. Only MONOs had genes changing expression (all up regulated including interleukin-10) that were clustered in the JAK/STAT pathway. Pre-incubation with IL-10 antibody, for LPS-stimulated MONOs, led to up regulated PI and IL-10 gene expression and release of PI cytokines after 4 h. Discussion: The present study suggests a dominant role of MONO gene expression in control of the fetal inflammatory response syndrome at 4 hrs of LPS stimulation. LPS-stimulated MONOs but not PMNs of the newborn have the ability to inhibit PI cytokine gene expression by latent IL-10 release. [ABSTRACT FROM AUTHOR]

Interferon-γ (IFNγ) is a pleiotropic cytokine that has a crucial role in immune response and tumor immunity. Because of its anti-tumor effects, IFNγ has been used in cancer treatment. However, IFNγ also has tumor-promoting functions that are less well understood. Here, we show that IFNγ induces expression of the pro-inflammatory and pro-angiogenic chemokine interleukin-8 (IL-8, CXCL8) in ovarian cancer (OC) cells. The IFNγ-induced IL-8 expression is dependent on JAK1, STAT1, and p65 NFκB, and is associated with an increased occupancy of K314/315 acetylated p65 NFκB and Ser-727 phosphorylated STAT1 at the IL-8 promoter. Neutralization of IL-8 using anti-IL-8 antibody reduces IFNγ-induced migration of OC cells, and their invasion ability in 3D spheroids. Together, these findings identify IL-8 as a novel target induced by IFNγ/JAK1/STAT1/p65 NFκB signaling, and indicate that the IFNγ-induced IL-8 contributes to IFNγ pro-tumorigenic effects in ovarian cancer cells. • IFNγ induces IL-8 expression in ovarian cancer cells. • IFNγ induced IL-8 expression is mediated by JAK1/STAT1 and NFκB signaling. • IL-8 neutralization decreases migration and invasiveness of IFNγ-treated cells. • IL-8 is a novel target induced by IFNγ signaling in solid cancer cells. • IL-8 neutralization might increase IFNγ effectiveness in cancer treatment. [ABSTRACT FROM AUTHOR]

Abstract: Proteasome inhibitors are known to suppress the proteasome-mediated degradation of IκBα in stimulated cells. This results in the cytoplasmic retention of NFκB and its reduced nuclear transcriptional activity. In this study, we show that in the metastatic prostate cancer cells, the proteasome inhibitors exhibit a novel, previously unrecognized effect: they increase the cellular levels of IκBα, which then translocates to the nucleus, associates with the nuclear p65 NFκB, thus inhibiting the constitutive NFκB DNA binding activity and inducing apoptosis. The proteasome inhibition-induced nuclear translocation of IκBα is dependent on de novo protein synthesis, occurs also in other cell types, and does not require IκBα phosphorylation on Ser-32. Since NFκB activity is constitutively increased in many human cancers as well as in inflammatory disorders, the proteasome inhibition-induced nuclear translocation of IκBα could thus provide a new therapeutic strategy aimed at the specific inhibition of NFκB activity by the nuclear IκBα. [Copyright &y& Elsevier]

Background: Neutrophils followed by monocytic cells are recruited into the lung during the early development of bronchopulmonary dysplasia (BPD). Objectives: We determined: (1) the capacity of polymorphonuclear leukocytes (PMNs) and peripheral blood monocytic cells (PBMCs) of the newborn to produce and release the anti-inflammatory cytokine, interleukin (IL)-10, after stimulation by lipopolysaccharide (LPS) or tumor necrosis factor (TNF), and (2) the levels of exogenous IL-10 and/or dexamethasone (DEX) needed to inhibit the release of the pro-inflammatory chemokine IL-8 from stimulated cells. Methods: PMNs and PBMCs were isolated from cord blood of healthy term infants. RT-PCR and ELISA were used to detect mRNA and cytokine levels from culture media, respectively. Results: We found that PMNs did not produce IL-10 mRNA or release IL-10 but did produce IL-8 mRNA by 1 h. PBMCs did produce IL-10 mRNA after 4 h (with IL-8 mRNA expression by 1 h). LPS-stimulated PBMCs released IL-10 to a maximum of 1,038 pg/ml/5 million cells (56 femtomolar). Equimolar doses of exogenous IL-10 or DEX produced up to 83% inhibition of IL-8 from PMNs. Exogenous IL-10 was more potent than DEX, on an equimolar basis, with regard to IL-8 release from PBMCs (90 vs. 33% respectively at a 10 nanomolar level). No inhibition of IL-8 release by IL-10 or DEX was observed at 100 femtomolar level. IL-10 and DEX did not have an additive inhibitory effect on IL-8 release. Conclusions: We conclude that for the newborn: (1) PBMCs produce IL-10 far below the level needed to inhibit a submaximal release of IL-8 from PMNs or PBMCs, and (2) exogenous IL-10 was equipotent or more potent than therapeutic levels of DEX on inhibition of IL-8 from these cells. Further studies are needed to determine if exogenous IL-10 may be useful in the treatment of BPD or other inflammatory disorders of the newborn. Copyright © 2007 S. Karger AG, Basel [ABSTRACT FROM AUTHOR]

Increased activation of the transcription factor NFκB in the neutrophils has been associated with the pathogenesis of sepsis, acute lung injury (AU). bronchopulmonary dysplasia (BPD). and other neutrophil-mediated inflammatory disorders. Despite re- cent progress in analyzing early NFκB activation in human neutrophils, activation of NFκB in persistently stimulated neutrophils has not been previously studied. Because it is the persistent NFκB activation that is thought to be involved in the host response to sepsis and the pathogenesis of ALl and BPD. we hypothesized that continuously stimulated human neutrophils may exhibit a late phase of NFκB activity. The goal of this study was to analyze the NFκB activation and expression of 1κB and NFκB proteins during neutrophil stimulation with inflammatory signals for prolonged times. We demonstrate that neutrophil stimulation with lipopolysaccharide (LPS) and tumor necrosis factor-α (TNFα) induces, in addition to the early activation at 30-60 mm, a previously unrecognized late phase of NFκB activation, In LPS-stimulated neutrophils. this NFκB activity typically had a biphasic character, whereas TNFα-stimulated neutrophils exhibited a continuous NFκB activity peaking around 9 h after stimulation. In contrast to the early NFκB activation that inversely correlates to the nuclear levels of IκBα, however, in continuously stimulated neutrophils, NFκB is persistently activated despite considerable levels of IκBα present in the nucleus. Our data suggest that NFκB is persistently activated in human neutrophils during neutrophil-mediated inflammatory disorders, and this persistent NFκB activity may represent one of the underlying mechanisms for the continuous production of proinflammatory mediators. [ABSTRACT FROM AUTHOR]

Abstract: Tumor necrosis factor-α (TNFα) is a potent pro-inflammatory cytokine that plays a major role in the pathogenesis of acute and chronic inflammatory disorders such as septic shock and arthritis, respectively. Leukocytes stimulated with inflammatory signals such as lipopolysaccharide (LPS) are the predominant producers of TNFα, and thus control of TNFα release from stimulated leukocytes represents a potential therapeutic target. Here, we report that leptomycin B (LMB), a specific inhibitor of CRM1-dependent nuclear protein export, inhibits TNFα release from LPS-stimulated human peripheral blood neutrophils and mononuclear cells. In addition, immunofluorescence confocal microscopy and immunoblotting analysis indicate that TNFα is localized in the nucleus of human neutrophils and mononuclear cells. This study demonstrates that the cellular release of TNFα from stimulated leukocytes is mediated by the CRM1-dependent nuclear export mechanism. Inhibition of CRM1-dependent cellular release of TNFα could thus provide a novel therapeutic approach for disorders involving excessive TNFα release. [Copyright &y& Elsevier]

Prolonged exposure to hyperoxia induces pulmonary epithelial cell death and acute lung injury. Although both apoptotic and nonapoptotic morphologies are observed in hyperoxic animal lungs, nonapoptotic cell death had only been recorded in transformed lung epithelium cultured in hyperoxia. To test whether the nonapoptotic characteristics in hyperoxic animal lungs are direct effects of hyperoxia, the mode of cell death was determined both morphologically and biochemically in human primary lung epithelium exposed to 95% O2. In contrast to characteristics observed in apoptotic cells, hyperoxia induced swelling of nuclei and an increase in cell size, with no evidence for any augmentation in the levels of either caspase-3 activity or annexin V incorporation. These data suggest that hyperoxia can directly induce nonapoptotic cell death in primary lung epithelium. Although hyperoxia-induced nonapoptotic cell death was associated with NF-κB activation, it is unknown whether NF-κB activation plays any causal role in nonapoptotic cell death. This study shows that inhibition of NF-κB activation can accelerate hyperoxia-induced epithelial cell death in both primary and transformed lung epithelium. Corresponding to the reduced cell survival in hyperoxia, the levels of MnSOD were also low in NF-κB-deficient cells. These results demonstrate that NF-κB protects lung epithelial cells from hyperoxia-induced nonapoptotic cell death. [Copyright &y& Elsevier]

Human neutrophils differ from other cells by containing high amount of IκBα in the nucleus, and this increased nuclear IκBα accumulation is associated with the inhibition of NFκB activity and increased apoptosis. However, the mechanisms regulating NFκB activation and IκBα degradation in human neutrophils are little understood. The objective of this study was to provide a further insight into the mechanisms regulating NFκB activity and IκBα degradation in human neutrophils. We show that okadaic acid (OA), an inhibitor of protein phosphatases PP1 and PP2A, induces sustained activation of NFκB and degradation of the nuclear IκBα, and increases interleukin-8 expression in the neutrophils. Furthermore, inhibitors of protein kinase C-δ (PKCδ) and IκB kinase (IKK) inhibit the OA-induced activation of NFκB. Collectively, our results indicate that in human neutrophils, the sustained activation of NFκB is regulated by a continuous phosphorylation and degradation of the nuclear IκBα. [Copyright &y& Elsevier]

• Reduced nucleocytosolic acetyl-CoA synthesis downregulates histone transcription • Decreased acetyl-CoA levels affect PIC assembly at histone promoters • Decreased acetyl-CoA reduces histone acetylation at histone promoters • Decreased histone levels induce respiration and ATP synthesis • Decreased acetyl-CoA synthesis is compensated for by elevated ATP synthesis [ABSTRACT FROM AUTHOR]

Adenosine monophosphate (AMP)-activated protein kinase (AMPK) serves as an energy sensor and master regulator of metabolism. In general, AMPK inhibits anabolism to minimize energy consumption and activates catabolism to increase ATP production. One of the mechanisms employed by AMPK to regulate metabolism is protein acetylation. AMPK regulates protein acetylation by at least five distinct mechanisms. First, AMPK phosphorylates and inhibits acetyl-CoA carboxylase (ACC) and thus regulates acetyl-CoA homeostasis. Since acetyl-CoA is a substrate for all lysine acetyltransferases (KATs), AMPK affects the activity of KATs by regulating the cellular level of acetyl-CoA. Second, AMPK activates histone deacetylases (HDACs) sirtuins by increasing the cellular concentration of NAD+, a cofactor of sirtuins. Third, AMPK inhibits class I and II HDACs by upregulating hepatic synthesis of α-hydroxybutyrate, a natural inhibitor of HDACs. Fourth, AMPK induces translocation of HDACs 4 and 5 from the nucleus to the cytoplasm and thus increases histone acetylation in the nucleus. Fifth, AMPK directly phosphorylates and downregulates p300 KAT. On the other hand, protein acetylation regulates AMPK activity. Sirtuin SIRT1-mediated deacetylation of liver kinase B1 (LKB1), an upstream kinase of AMPK, activates LKB1 and AMPK. AMPK phosphorylates and inactivates ACC, thus increasing acetyl-CoA level and promoting LKB1 acetylation and inhibition. In yeast cells, acetylation of Sip2p, one of the regulatory β-subunits of the SNF1 complex, results in inhibition of SNF1. This results in activation of ACC and reduced cellular level of acetyl-CoA, which promotes deacetylation of Sip2p and activation of SNF1. Thus, in both yeast and mammalian cells, AMPK/SNF1 regulate protein acetylation and are themselves regulated by protein acetylation. [ABSTRACT FROM AUTHOR]