IKK Inhibitor Research Articles

Inhibition of IL-6 Release in Vitro by in Vivo Administration of an IKK Inhibitor in Mice with Lung Fibrosis Induced by Poly I:C

Inhibition of IL-6 Release in Vitro by in Vivo Administration of an IKK Inhibitor in Mice with Lung Fibrosis Induced by Poly I:C

ZNF395 Is an Activator of a Subset of IFN-Stimulated Genes.

Activation of the interferon (IFN) pathway in response to infection with pathogens results in the induction of IFN-stimulated genes (ISGs) including proinflammatory cytokines, which mount the proper antiviral immune response. However, aberrant expression of these genes is pathogenic to the host. In addition to IFN-induced transcription factors non-IFN-regulated factors contribute to the transcriptional control of ISGs. Here, we show by genome wide expression analysis, siRNA-mediated suppression and Doxycycline-induced overexpression that the cellular transcription factor ZNF395 activates a subset of ISGs including the chemokines CXCL10 and CXCL11 in keratinocytes. We found that ZNF395 acts independently of IFN but enhances the IFN-induced expression of CXCL10 and CXCL11. Luciferase reporter assays revealed a requirement of intact NFκB-binding sites for ZNF395 to stimulate the CXCL10 promoter. The transcriptional activation of CXCL10 and CXCL11 by ZNF395 was abolished after inhibition of IKK by BMS-345541, which increased the stability of ZNF395. ZNF395 encodes at least two motifs that mediate the enhanced degradation of ZNF395 in response to IKK activation. Thus, IKK is required for ZNF395-mediated activation of transcription and enhances its turn-over to keep the activity of ZNF395 low. Our results support a previously unrecognized role of ZNF395 in the innate immune response and inflammation.

Targeting the Canonical Nuclear Factor-κB Pathway with a High-Potency IKK2 Inhibitor Improves Outcomes in a Mouse Model of Idiopathic Pneumonia Syndrome

Targeting the Canonical Nuclear Factor-κB Pathway with a High-Potency IKK2 Inhibitor Improves Outcomes in a Mouse Model of Idiopathic Pneumonia Syndrome

Apoptosis and surfactant protein-C expression inhibition induced by lipopolysaccharide in AEC II cell may associate with NF-κB pathway.

Lipopolysaccharide (LPS), a Gram-negative bacterial outer membrane component, is one of the major causes of septic shock. Herein we investigate LPS-induced apoptosis of rat alveolar epithelial type II cells (AEC II) and the effects of LPS on surfactant protein-C (SP-C) expression in AEC II, along with the possible molecular mechanisms. LPS exposure impaired cell viability and increased apoptosis of AEC II significantly in concentration-dependent manner embodied in increased caspase-3 expression and the activity of caspase-3. Simultaneously, our results also indicated that LPS inhibited surfactant protein-C (SP-C) expression in AEC II. Mechanistic studies revealed that LPS treatment significantly increased the expression of NF-κB p50, NF-κB p65 and IKKβ proteins as well as induced IκB-α phosphorylation. Moreover, pretreatment with IKK inhibitor IKK-16 or NF-κB inhibitor PDTC ameliorated LPS-caused alterations in cleaved caspase-3 expression, the activity of caspase-3 and SP-C expression. Taken together, these results demonstrate that LPS can induce apoptosis of AEC II and decrease SP-C expression partly through activating the NF-κB pathway.

Aspirin Inhibits IKK-β-mediated Prostate Cancer Cell Invasion by Targeting Matrix Metalloproteinase-9 and Urokinase-Type Plasminogen Activator

Background/Aims: Aspirin has been demonstrated to possess potent chemopreventive and anticancer effects on prostate cancer. However, the more detailed molecular mechanisms of aspirin to suppress prostate cancer cell invasion have not been clearly elucidated. Methods: Transwell assays were performed to evaluate the effects of aspirin on cell invasion. Matrix metalloproteinases (MMPs) and serine proteinases activities in cell media were examined by gelatin zymography and ELISA. In addition, inhibitor of κB (IκB) kinase-β (IKK-β) phosphorylation and IKK-β kinase activity were measured to assess the effects of aspirin on IKK-β activation. Results: We found that aspirin suppressed the invasion and attachment in human prostate cancer cells. Aspirin treatment significantly resulted in reduction of matrix metalloproteinase-9 (MMP-9) and upregulation of tissue inhibitors of metalloproteinase-1 (TIMP-1) activity, which are the proteolytic enzymes contributing to the degradation of extracellular matrix and basement membrane in cell invasion and metastasis. Our data further showed that aspirin was able to inhibit both urokinase-type plasminogen activator (uPA) and plasminogen activator inhibitor-1 (PAI-1) expression in the cells. In addition, aspirin treatment caused a strong decrease in nuclear factor-kappa B (NF-κB) activation, inhibitor of κB (IκB)-α phosphorylation together with translocation of NF-κB p65 to nucleus and IκB kinase (IKK)- β activation. Moreover, the inhibitory effects of aspirin on cell invasion were reversed by IKK-β overexpression, while the IKK inhibitor sensitizes the anti-invasive effect of aspirin in prostate cancer cells. Conclusion: The present research concluded that aspirin suppressed prostate cancer cell invasion by reducing MMP-9 activity and uPA expression through decreasing of IKK-β-mediated NF-κB activation, indicating that the ability of aspirin to inhibit cell invasion might be useful in the chemoprevention of metastatic prostate cancer.

Inhibition of IKKβ by celastrol and its analogues – an in silico and in vitro approach

Context: Alzheimer’s disease (AD) is the most common form of dementia affecting the aged population and neuroinflammation is one of the most observed AD pathologies. NF-κB is the central regulator of inflammation and inhibitor κB kinase (IKK) is the converging point in NF-κB activation. Celastrol is a natural triterpene used as a treatment for inflammatory conditions. Objective: This study determines the neuroprotective and inhibitory effect of celastrol on amyloid beta1-42 (Aβ1-42) induced cytotoxicity and IKKβ activity, respectively. Materials and methods: Retinoic acid differentiated IMR-32 cells were treated with celastrol (1 μM) before treatment with Aβ1-42 (IC30 10 μM) for 24 h. The cytotoxicity and IKK phosphorylation were measured by MTT and western blotting analysis, respectively. We screened 36 celastrol analogues for the IKKβ inhibition by molecular docking and evaluated their drug like properties to delineate the neuroprotective effects. Results: Celastrol (1 μM) inhibited Aβ1-42 (10 μM) induced IκBα phosphorylation and protected IMR-32 cells from cell death. Celastrol and 25 analogues showed strong binding affinity with IKKβ as evidenced by strong hydrogen-bonding interactions with critical active site residues. All the 25 analogues displayed strong anti-inflammatory properties but only 11 analogues showed drug-likeness. Collectively, molecule 15 has highest binding affinity, CNS activity and more drug likeness than parent compound celastrol. Discussion and conclusion: The decreased expression of pIκBα in celastrol pretreated cells affirms the functional representation of inhibited IKKβ activity in these cells. The neuroprotective potentials of celastrol and its analogues may be related to IKK inhibition.

IL-17A exacerbates diabetic retinopathy by impairing Müller cell function via Act1 signaling.

Diabetic retinopathy (DR), one of the most serious complications of diabetes, has been associated with inflammatory processes. We have recently reported that interleukin (IL)-17A, a proinflammatory cytokine, is increased in the plasma of diabetic patients. Further investigation is required to clarify the role of IL-17A in DR. Ins2Akita (Akita) diabetic mice and high-glucose (HG)-treated primary Müller cells were used to mimic DR-like pathology. Diabetes induced retinal expression of IL-17A and IL-17 receptor A (IL-17RA) in Müller cells in contrast to ganglion cells. Further evidence demonstrated that retinal Müller cells cultured in vitro increased IL-17A and IL-17RA expression as well as IL-17A secretion in the HG condition. In both the HG-treated Müller cells and Akita mouse retina, the Act1/TRAF6/IKK/NF-κB signaling pathway was activated. IL-17A further enhanced inflammatory signaling activation, whereas Act1 knockdown or IKK inhibition blocked the downstream signaling activation by IL-17A. HG- and diabetes-induced Müller cell activation and dysfunction, as determined by increased glial fibrillary acidic protein, vascular endothelial growth factor and glutamate levels and decreased glutamine synthetase and excitatory amino acid transporter-1 expression, were exacerbated by IL-17A; however, they were alleviated by Act1 knockdown or IKK inhibition. In addition, IL-17A intravitreal injection aggravated diabetes-induced retinal vascular leukostasis, vascular leakage and ganglion cell apoptosis, whereas Act1 silencing or anti-IL-17A monoclonal antibody ameliorated the retinal vascular damage and neuronal cell apoptosis. These findings establish that IL-17A exacerbates DR-like pathology by the promotion of Müller cell functional impairment via Act1 signaling.

Biphasic regulation of chondrocytes by Rela through induction of anti-apoptotic and catabolic target genes.

In vitro studies have shown that Rela/p65, a key subunit mediating NF-κB signalling, is involved in chondrogenic differentiation, cell survival and catabolic enzyme production. Here, we analyse in vivo functions of Rela in embryonic limbs and adult articular cartilage, and find that Rela protects chondrocytes from apoptosis through induction of anti-apoptotic genes including Pik3r1. During skeletal development, homozygous knockout of Rela leads to impaired growth through enhanced chondrocyte apoptosis, whereas heterozygous knockout of Rela does not alter growth. In articular cartilage, homozygous knockout of Rela at 7 weeks leads to marked acceleration of osteoarthritis through enhanced chondrocyte apoptosis, whereas heterozygous knockout of Rela results in suppression of osteoarthritis development through inhibition of catabolic gene expression. Haploinsufficiency or a low dose of an IKK inhibitor suppresses catabolic gene expression, but does not alter anti-apoptotic gene expression. The biphasic regulation of chondrocytes by Rela contributes to understanding the pathophysiology of osteoarthritis.

IinQ attenuates systemic inflammatory responses via selectively impairing the Myddosome complex formation upon TLR4 ligation

A specific small-molecule inhibitor of the TLR4 signaling complex upstream of the IKK would likely provide therapeutic benefit for NF-κB-mediated inflammatory disease. We previously identified brazilin as a selective upstream IKK inhibitor targeting the Myddosome complex. In this study, using a cell-based ubiquitination assay for IRAK1 and a chemical library comprising a series of structural analogues of brazilin, a novel small molecule, 2-hydroxy-5,6-dihydroisoindolo[1,2-a]isoquinoline-3,8-dione (IinQ), was identified as a selective and potent inhibitor of IRAK1-dependent NF-κB activation upon TLR4 ligation. In RAW264.7 macrophages, IinQ drastically suppressed activation of upstream IKK signaling events including membrane-bound IRAK1 ubiquitination and IKK phosphorylation by the TLR4 ligand, resulting in reduced expression of proinflammatory mediators including IL-6, TNF-α, and nitric oxide. Interestingly, IinQ did not suppress NF-κB activation via the TLR3 ligand, DNA damaging agents, or a protein kinase C activator, indicating IinQ is specific for TLR4 signaling. Analysis of upstream signaling events further confirmed that IinQ disrupts the MyD88-IRAK1-TRAF6 complex formation induced by LPS treatment, without affecting TLR4 oligomerization. Moreover, intravenous administration of IinQ significantly reduced lethality and attenuated systemic inflammatory responses in an in vivo mouse model of endotoxin shock following LPS challenge. Thus, IinQ represents a novel class of brazilin analogues with improved potency and specificity toward disruption of Myddosome complex formation in TLR4 signaling, indicating that IinQ may be a promising therapeutic candidate for the treatment of systemic inflammatory diseases.

New Insights into the Pro-Inflammatory Activities of Ang1 on Neutrophils: Induction of MIP-1β Synthesis and Release.

We reported the expression of angiopoietin Tie2 receptor on human neutrophils and the capacity of angiopoietins (Ang1 and Ang2) to induce pro-inflammatory activities, such as platelet-activating factor synthesis, β2-integrin activation and neutrophil migration. Recently, we observed differential effects between both angiopoietins, namely, the capacity of Ang1, but not Ang2, to promote rapid interleukin-8 synthesis and release, as well as neutrophil viability. Herein, we addressed whether Ang1 and/or Ang2 could modulate the synthesis and release of macrophage inflammatory protein-1β (MIP-1β) by neutrophils. Neutrophils were isolated from blood of healthy volunteers; intracellular and extracellular MIP-1β protein concentrations were assessed by ELISA. After 24 hours, the basal intracellular and extracellular MIP-1β protein concentrations were ≈500 and 100 pg/106 neutrophils, respectively. Treatment with Ang1 (10 nM) increased neutrophil intracellular and extracellular MIP-1β concentrations by 310 and 388% respectively. Pretreatment with PI3K (LY294002), p38 MAPK (SB203580) and MEK (U0126) inhibitors completely inhibited Ang1-mediated increase of MIP-1β intracellular and extracellular protein levels. Pretreatment with NF-κB complex inhibitors, namely Bay11-7085 and IKK inhibitor VII or with a transcription inhibitor (actinomycin D) and protein synthesis inhibitor (cycloheximide), did also abrogate Ang1-mediated increase of MIP-1β intracellular and extracellular protein levels. We validated by RT-qPCR analyses the effect of Ang1 on the induction of MIP-1β mRNA levels. Our study is the first one to report Ang1 capacity to induce MIP-1β gene expression, protein synthesis and release from neutrophils, and that these effects are mediated by PI3K, p38 MAPK and MEK activation and downstream NF-κB activation.

Lymphocyte-specific deletion of IKK2 or NEMO mediates an increase in intrarenal Th17 cells and accelerates renal damage in an ischemia-reperfusion injury mouse model.

Acute kidney injury (AKI) is associated with poor patient outcome and a global burden for end-stage renal disease. Ischemia-reperfusion injury (IRI) is one of the major causes of AKI, and experimental work has revealed many details of the inflammatory response in the kidney, such as activation of the NF-κB pathway. Here, we investigated whether deletion of the NF-κB kinases IKK2 or NEMO in lymphocytes or systemic inhibition of IKK2 would cause different kidney inflammatory responses after IRI induction. Serum creatinine, blood urea nitrogen (BUN) level, and renal tubular injury score were significantly increased in CD4creIKK2f/f (CD4xIKK2Δ) and CD4creNEMOf/f (CD4xNEMOΔ) mice compared with CD4cre mice after IRI induction. The frequency of Th17 cells infiltrating the kidneys of CD4xIKK2Δ or CD4xNEMOΔ mice was also significantly increased at all time points. CCL20, an important chemokine in Th17 cell recruitment, was significantly increased at early time points after the induction of IRI. IL-1β, TNF-α, and CCL2 were also significantly increased in different patterns. A specific IKK2 inhibitor, KINK-1, reduced BUN and serum creatinine compared with nontreated mice after IRI induction, but the frequency of kidney Th17 cells was also significantly increased. In conclusion, although systemic IKK2 inhibition improved kidney function, lymphocyte-specific deletion of IKK2 or NEMO aggravated kidney injury after IRI, and, in both conditions, the percentage of Th17 cells was increased. Our findings demonstrate the critical role of the NF-κB pathway in Th17 activation, which advises caution when using systemic IKK2 inhibitors in patients with kidney injury, since they might impair the T cell response and aggravate renal disease.

Classical NF-κB pathway is responsible for APOBEC3B expression in cancer cells

APOBEC3B (A3B) is a DNA cytosine deaminase and catalyzes cytosine deamination, resulting in mutations in genomic DNA. A3B is aberrantly expressed in a variety of cancers and considered to be a source of genomic mutations that contribute to cancer progression and metastasis. However, the mechanisms through which A3B expression is dysregulated in cancer cells are not fully elucidated. Here we report that the classical NF-κB pathway plays a crucial role in the transcriptional regulation of A3B in various cancer cells, including lymphoid malignancies. PMA, a strong activator of PKC, induces A3B at both mRNA and protein levels in cancer cell lines, and specific inhibitors of both PKC and IKK downregulate A3B expression. Using luciferase reporter and EMSA assays, we identify 3 NF-κΒ binding sites in the A3B promoter and reveal that NF-κB p65/p50 and p65/c-Rel heterodimers are important for A3B transcription. These results suggest that the classical NF-κB pathway is responsible for activation of A3B mRNA expression and further imply that inhibition of PKC and IKK might augment cancer treatment by reducing cancer progression and metastasis through downregulation of A3B expression.

TNF activation of NF-κB is essential for development of single-positive thymocytes.

NF-κB activation has been implicated at multiple stages of thymic development of T cells, during which it is thought to mediate developmental signals originating from the T cell receptor (TCR). However, the Card11-Bcl10-Malt1 (CBM) complex that is essential for TCR activation of NF-κB in peripheral T cells is not required for thymocyte development. It has remained unclear whether the TCR activates NF-κB independent of the CBM complex in thymocyte development or whether another NF-κB activating receptor is involved. In the present study, we generated mice in which T cells lacked expression of both catalytic subunits of the inhibitor of κB kinase (IKK) complex, IKK1 and IKK2, to investigate this question. Although early stages of T cell development were unperturbed, maturation of CD4 and CD8 single-positive (SP) thymocytes was blocked in mice lacking IKK1/2 in the T cell lineage. We found that IKK1/2-deficient thymocytes were specifically sensitized to TNF-induced cell death in vitro. Furthermore, the block in thymocyte development in IKK1/2-deficient mice could be rescued by blocking TNF with anti-TNF mAb or by ablation of TNFRI expression. These experiments reveal an essential role for TNF activation of NF-κB to promote the survival and development of single positive T cells in the thymus.

IKK inhibitor suppresses epithelial-mesenchymal transition and induces cell death in prostate cancer.

IκB kinase (IKK)/nuclear factor κB (NF-κB) pathway activation is a key event in the acquisition of invasive and metastatic capacities in prostate cancer. A potent small-molecule compound, BMS-345541, was identified as a highly selective IKKα and IKKβ inhibitor to inhibit kinase activity. This study explored the effect of IKK inhibitor on epithelial-mesenchymal transition (EMT), apoptosis and metastasis in prostate cancer. Here, we demonstrate the role of IKK inhibitor reducing proliferation and inducing apoptosis in PC-3 cells. Furthermore, BMS345541 inhibited IκBα phosphorylation and nuclear level of NF-κB/p65 in PC-3 cells. We also observed downregulation of the N-cadherin, Snail, Slug and Twist protein in a dose-dependent manner. BMS‑345541 induced upregulation of the epithelial marker E-cadherin and phosphorylated NDRG1 at protein level. Moreover, BMS‑345541 reduced invasion and metastasis of PC-3 cells invitro. In conclusion, IKK has a key role in both EMT and apoptosis of prostate cancer. IKK inhibitor can reverse EMT and induce cell death in PCa cells. IKK was identified as a potential target structure for future therapeutic intervention in PCa.

Conditional knockout of polarity complex (atypical) PKCι reveals an anti-inflammatory function mediated by NF-κB.

The conserved proteins of the polarity complex made up of atypical PKC (aPKC, isoforms ι and ζ), Par6, and Par3 determine asymmetry in several cell types, from Caenorhabditis elegans oocytes to vertebrate epithelia and neurons. We previously showed that aPKC is down-regulated in intestinal epithelia under inflammatory stimulation. Further, expression of constitutively active PKCι decreases NF-κB activity in an epithelial cell line, the opposite of the effect reported in other cells. Here we tested the hypothesis that aPKC has a dual function in epithelia, inhibiting the NF-κB pathway in addition to having a role in apicobasal polarity. We achieved full aPKC down-regulation in small intestine villi and colon surface epithelium using a conditional epithelium-specific knockout mouse. The results show that aPKC is dispensable for polarity after cell differentiation, except for known targets, including ROCK and ezrin, claudin-4 expression, and barrier permeability. The aPKC defect resulted in increased NF-κB activity, which could be rescued by IKK and ROCK inhibitors. It also increased expression of proinflammatory cytokines. In contrast, expression of anti-inflammatory IL-10 decreased. We conclude that epithelial aPKC acts upstream of multiple mechanisms that participate in the inflammatory response in the intestine, including, but not restricted to, NF-κB.

IκB Kinase Inhibitor Attenuates Sepsis-Induced Cardiac Dysfunction in CKD.

Patients with CKD requiring dialysis have a higher risk of sepsis and a 100-fold higher mortality rate than the general population with sepsis. The severity of cardiac dysfunction predicts mortality in patients with sepsis. Here, we investigated the effect of preexisting CKD on cardiac function in mice with sepsis and whether inhibition of IκB kinase (IKK) reduces the cardiac dysfunction in CKD sepsis. Male C57BL/6 mice underwent 5/6 nephrectomy, and 8 weeks later, they were subjected to LPS (2 mg/kg) or sepsis by cecal ligation and puncture (CLP). Compared with sham operation, nephrectomy resulted in significant increases in urea and creatinine levels, a small (P<0.05) reduction in ejection fraction (echocardiography), and increases in the cardiac levels of phosphorylated IκBα, Akt, and extracellular signal-regulated kinase 1/2; nuclear translocation of the NF-κB subunit p65; and inducible nitric oxide synthase (iNOS) expression. When subjected to LPS or CLP, compared with sham-operated controls, CKD mice exhibited exacerbation of cardiac dysfunction and lung inflammation, greater increases in levels of plasma cytokines (TNF-α, IL-1β, IL-6, and IL-10), and greater increases in the cardiac levels of phosphorylated IKKα/β and IκBα, nuclear translocation of p65, and iNOS expression. Treatment of CKD mice with an IKK inhibitor (IKK 16; 1 mg/kg) 1 hour after CLP or LPS administration attenuated these effects. Thus, preexisting CKD aggravates the cardiac dysfunction caused by sepsis or endotoxemia in mice; this effect may be caused by increased cardiac NF-κB activation and iNOS expression.

Wedelolactone mitigates UVB induced oxidative stress, inflammation and early tumor promotion events in murine skin: plausible role of NFkB pathway

UVB (Ultra-violet B) radiation is one of the major etiological factors in various dermal pathology viz. dermatitis, actinic folliculitis, solar urticaria, psoriasis and cancer among many others. UVB causes toxic manifestation in tissues by inciting inflammatory and tumor promoting events. We have designed this study to assess the anti-inflammatory and anti-tumor promotion effect of Wedelolactone (WDL) a specific IKK inhibitor. Results indicate significant restoration of anti-oxidative enzymes due to WDL treatments. We also found that WDL was effective in mitigating inflammatory markers consisting of MPO (myeloperoxidase), Mast cells trafficking, Langerhans cells suppression and COX 2 expression up regulation due to UVB exposure. We also deduce that WDL presented a promising intervention in attenuating early tumor promotion events caused by UVB exposure as indicated by the results of ODC (Ornithine Decarboxylase), Thymidine assay, Vimentin and VEGF (Vascular-endothelial growth factor) expression. This study was able to provide substantial cues for the therapeutic ability of Wedelolactone against inflammatory and tumor promoting events in murine skin depicting plausible role of NFkB pathway.

Noncanonical WNT-5B signaling induces inflammatory responses in human lung fibroblasts.

COPD is a progressive chronic lung disease characterized by pulmonary inflammation. Several recent studies indicate aberrant expression of WNT ligands and Frizzled receptors in the disease. For example, WNT-5A/B ligand expression was recently found to be increased in lung fibroblasts of COPD patients. However, possible effects of WNT-5A and WNT-5B on inflammation have not been investigated yet. In this study, we assessed the regulation of inflammatory cytokine release in response to WNT-5A/B signaling in human lung fibroblasts. Primary human fetal lung fibroblasts (MRC-5), and primary lung fibroblasts from COPD patients and non-COPD controls were treated with recombinant WNT-5A or WNT-5B to assess IL-6 and CXCL8 cytokine secretion and gene expression levels. Following WNT-5B, and to a lesser extent WNT-5A stimulation, fibroblasts showed increased IL-6 and CXCL8 cytokine secretion and mRNA expression. WNT-5B-mediated IL-6 and CXCL8 release was higher in fibroblasts from COPD patients than in non-COPD controls. In MRC-5 fibroblasts, WNT-5B-induced CXCL8 release was mediated primarily via the Frizzled-2 receptor and TAK1 signaling, whereas canonical β-catenin signaling was not involved. In further support of noncanonical signaling, we showed activation of JNK, p38, and p65 NF-κB by WNT-5B. Furthermore, inhibition of JNK and p38 prevented WNT-5B-induced IL-6 and CXCL8 secretion, whereas IKK inhibition prevented CXCL8 secretion only, indicating distinct pathways for WNT-5B-induced IL-6 and CXCL8 release. WNT-5B induces IL-6 and CXCL8 secretion in pulmonary fibroblasts. In summary, WNT-5B mediates this via Frizzled-2 and TAK1. As WNT-5 signaling is increased in COPD, this WNT-5-induced inflammatory response could represent a therapeutic target.

New IKK inhibitors: Synthesis of new imidazo[1,2-a]quinoxaline derivatives using microwave assistance and biological evaluation as IKK inhibitors

The inhibition of the NF-κB-dependent pathways by IKK inhibitors plays an important role in immunity, inflammation, and cancer. New imidazoquinoxalines tricyclic derivatives are prepared using microwave assistance and their biological activities as IKK inhibitors are described. Compounds 6a present a potent inhibition activity and selectivity for IKK2. Docking studies in the IKK2 binding site allowed identification of residues most likely to interact with theses inhibitors and explain their potent IKK2 inhibition activity and selectivity.

Plasma L5 levels are elevated in ischemic stroke patients and enhance platelet aggregation

AbstractL5, the most electronegative and atherogenic subfraction of low-density lipoprotein (LDL), induces platelet activation. We hypothesized that plasma L5 levels are increased in acute ischemic stroke patients and examined whether lectin-like oxidized LDL receptor-1 (LOX-1), the receptor for L5 on endothelial cells and platelets, plays a critical role in stroke. Because amyloid β (Aβ) stimulates platelet aggregation, we studied whether L5 and Aβ function synergistically to induce prothrombotic pathways leading to stroke. Levels of plasma L5, serum Aβ, and platelet LOX-1 expression were significantly higher in acute ischemic stroke patients than in controls without metabolic syndrome (P < .01). In mice subjected to focal cerebral ischemia, L5 treatment resulted in larger infarction volumes than did phosphate-buffered saline treatment. Deficiency or neutralizing of LOX-1 reduced infarct volume up to threefold after focal cerebral ischemia in mice, illustrating the importance of LOX-1 in stroke injury. In human platelets, L5 but not L1 (the least electronegative LDL subfraction) induced Aβ release via IκB kinase 2 (IKK2). Furthermore, L5+Aβ synergistically induced glycoprotein IIb/IIIa receptor activation; phosphorylation of IKK2, IκBα, p65, and c-Jun N-terminal kinase 1; and platelet aggregation. These effects were blocked by inhibiting IKK2, LOX-1, or nuclear factor–κB (NF-κB). Injecting L5+Aβ shortened tail-bleeding time by 50% (n = 12; P < .05 vs L1-injected mice), which was prevented by the IKK2 inhibitor. Our findings suggest that, through LOX-1, atherogenic L5 potentiates Aβ-mediated platelet activation, platelet aggregation, and hemostasis via IKK2/NF-κB signaling. L5 elevation may be a risk factor for cerebral atherothrombosis, and downregulating LOX-1 and inhibiting IKK2 may be novel antithrombotic strategies.