Susumu Muto

A New IκB Kinase β Inhibitor Prevents Human Breast Cancer Progression through Negative Regulation of Cell Cycle Transition

Constitutive nuclear factor-kappaB (NF-kappaB) activity plays a crucial role in the development and progression of lymphoma, leukemia, and some epithelial cancers. Given the contribution of NF-kappaB in carcinogenesis, a novel approach that interferes with its activity might have therapeutic potential against cancers that respond poorly to conventional treatments. Here, we have shown that a new IkappaB kinase beta inhibitor, IMD-0354, suppressed the growth of human breast cancer cells, MDA-MB-231, HMC1-8, and MCF-7, by arresting cell cycle and inducing apoptosis. In an electrophoretic mobility shift assay and a reporter assay, IMD-0354 abolished the NF-kappaB activity in MDA-MB-231 cells in a dose-dependent manner. In the cells incubated with IMD-0354, cell cycle arrested at the G0-G1 phase and apoptotic cells were increased. The expression of some cell cycle regulatory molecules and antiapoptotic molecules was suppressed in cells treated with IMD-0354. On the other hand, cyclin-dependent kinase suppressor p27Kip1 was up-regulated by the addition of IMD-0354. Daily administration of IMD-0354 inhibited tumor expansion in immunodeficient mice into which MDA-MB-231 cells were transplanted. These results indicate that NF-kappaB may contribute to cell proliferation through up-regulation of cell cycle progression; accordingly, inhibition of NF-kappaB activity might have a therapeutic ability in the treatment of human breast cancers.

Novel IkB kinase inhibitors for treatment of nuclear factor-kB-related diseases.

Introduction: NF-κB is a key regulator of inflammation and immunity in cancer development. The IκB kinase (IKK) is a multisubunit complex containing catalytic subunits termed IKK-α, -β and -γ. It is well known that many pro-inflammatory stimuli require the IKK-β subunit for NF-κB activation. Areas covered: NF-κB affects the progression of inflammation-related diseases, such as myocardial ischemia, bronchial asthma, arthritis, cancer and other diseases. We review the characteristics and effects of these inhibitors on inflammatory and other diseases. Expert opinion: Various synthesized IKK inhibitors have been developed and they will be used clinically in the near future.

Antiallergic and Anti-Inflammatory Effects of a Novel IκB Kinase β Inhibitor, IMD-0354, in a Mouse Model of Allergic Inflammation

Background: Nuclear factor (NF)-κB is a transcription factor known to regulate allergy-associated cytokine and chemokine production related to the induction of inflammation. IκB kinase β (IKKβ), which is responsible for activation of the NF-κB pathway, may be an ideal molecular target to inhibit this process. IMD-0354 [N-(3,5-bis-trifluoromethyl-phenyl)-5-chloro-2-hydroxy-benzamide] is an attractive novel IKKβ inhibitor that prevents the production of inflammatory cytokines in various diseases, although it is not known if IMD-0354 is effective against allergic inflammation. This study aimed to elucidate the antiallergic effects of a newly synthesized IKKβ inhibitor, IMD-0354, in a mouse model of allergic inflammation. Methods: We generated ovalbumin (OVA)-sensitized mice which were then challenged with OVA. IMD-0354 was administered intraperitoneally to therapeutic groups. Lung histopathology and the concentrations of cytokines and chemokines in bronchoalveolar lavage fluid (BALF) and supernatants of lung homogenates were determined. Results: Administration of IMD-0354 ameliorated airway hyperresponsiveness and reduced the numbers of bronchial eosinophils and mucus-producing cells in OVA-sensitized mice. The total numbers of cells and eosinophils in BALF were also reduced by treatment with IMD-0354. Treatment with IMD-0354 inhibited the production of Th2 cytokines such as interleukin (IL)-5 and IL-13 and eotaxin in the airways and/or lungs of OVA-sensitized mice, but it did not affect the restoration of Th1 cytokines such as IL-12 and interferon-γ under the same experimental conditions. IgE production was also inhibited by IMD-0354. Conclusion: A specific IKKβ inhibitor, IMD-0354, improved allergic airway inflammation and hyperresponsiveness in mice. IMD-0354 may have therapeutic potential for bronchial asthma.

IκB kinase β inhibitor IMD-0354 suppresses airway remodelling in a Dermatophagoides pteronyssinus-sensitized mouse model of chronic asthma

Background Nuclear factor (NF)‐κB is a transcription factor that regulates cytokine and chemokine production in various inflammatory diseases, including bronchial asthma. IκB kinase (IKK) β is important for NF‐κB activation in inflammatory conditions, and is possibly related to airway remodelling. Thus, inhibition of the IKKβ–NF‐κB pathway may be an ideal strategy for the management of airway remodelling.

Effectiveness of IκB kinase inhibitors in murine colitis-associated tumorigenesis

PurposeNuclear factor κB (NF-κB) activation is involved in various inflammatory illnesses, for example inflammatory bowel disease, and is thought to be a key factor in the tumor-promoting mechanism of inflammation-associated tumorigenesis. This study investigated whether inhibitors of IKKβ, which is a critical kinase for NF-κB activation, reduce colitis-associated tumorigenesis in mice.MethodsWe used a mouse model of the disease whereby administration of azoxymethane (AOM) followed by repeated dextran sulfate sodium (DSS) ingestion causes severe colonic inflammation and the subsequent development of multiple tumors. Effects of IKKβ inhibitors, NBD peptide, and IMD-0354 were examined.ResultsIn a colitis-associated cancer model, treatment with the IKKβ inhibitors NBD peptide and IMD-0354 significantly reduced the number of tumors compared with the untreated group. The tumors were also significantly smaller in the inhibitor-treated mice than in the untreated mice. Macrophage and neutrophil infiltration decreased with the inhibitor treatment. NF-κB activation and the expression of Cox-2 and iNOS were observed in tumor tissues, and the inhibitors ameliorated their expression. These inhibitors blocked NF-κB activation and the expression of proinflammatory cytokines mediated by the culture supernatant of inflamed colon in murine primary macrophages. In-vitro and in-vivo experiments showed that these drugs, especially NBD peptide, could also inhibit the proliferation of colonic epithelial cells.ConclusionThese results imply that IKKβ-targeted NF-κB blockade is an attractive therapeutic approach for the prevention of colitis-associated tumors.

Pathophysiological roles of nuclear factor kappaB (NF-kB) in pulmonary arterial hypertension: effects of synthetic selective NF-kB inhibitor IMD-0354

AIMS Proliferation of pulmonary arterial smooth muscle cells (PASMCs) is one histological sign of pulmonary arterial hypertension (PAH). We hypothesized that a signalling cascade from fibroblast growth factor 2 (FGF₂) to plasminogen activator inhibitor 1 (PAI-1) and monocyte chemotactic protein-1 (MCP-1) via nuclear transcription factor nuclear factor kappaB (NF-kB) play a critical role in progression of PAH, and tested this hypothesis both in vivo and in vitro using a synthetic selective NF-kB inhibitor, N-(3,5-Bis-trifluoromethyl-phenyl)-5-chloro-2-hydroxy-benzamide (IMD-0354). METHODS AND RESULTS Monocrotaline (MCT) was injected into 75 Sprague-Dawley rats. Starting at day 14 after MCT injection, we administered IMD-0354 (MCT + IMD group) or vehicle (MCT group) daily. At day 32, 65% of the MCT + IMD group were alive compared with 0% of the MCT group. IMD-0354 prevented increase of right ventricular pressure, and suppressed proliferation and induced apoptosis of PASMCs. mRNA transcript levels of FGF₂, PAI-1, and tissue plasminogen activator (t-PA) were lower in MCT + IMD compared with MCT. In in vitro experiments, IMD-0354 inhibited p65 translocation to the nucleus promoted by FGF₂ in PASMCs. Furthermore, the time courses of extracellular signal-regulated kinase (Erk) 1/2, MCP-1, and PAI-1 stimulated with FGF₂ were each markedly shortened by IMD-0354. CONCLUSIONS We speculate that the positive-feedback loop (Erk1/2-NF-kB-MCP-1-Erk1/2) is associated with progression of PAH by causing FGF₂-induced inflammation in MCT rats. IMD-0354 has potential as a new therapeutic tool for PAH.

The effects of pharmacological PAI-1 inhibition on thrombus formation and neointima formation after arterial injury

Objective: Plasminogen activator inhibitor (PAI)-1 plays a role in neointimal formation after percutaneous coronary intervention (PCI), the effect of overexpression or lack of PAI-1 is controversial. Murine arterial injury models develop neointimal hyperplasia similar to that observed in clinical coronary arterial restenosis after PCI. Methods and results: To clarify the role of PAI-1 in thrombus formation and neointimal formation after arterial injury, we used a specific PAI-1 inhibitor (IMD-1622) in a rat aorta–vein shunt model and a mouse arterial injury model. While the non-treated shunt model showed massive thrombus formation, IMD-1622 administration suppressed this. Injured arteries with vehicles showed significant neointimal formation with enhancement of adhesion molecules, fibrinogen accumulation and cell proliferation on day 28 after injury. However, intimal thickening and expression of these factors were suppressed in PAI-1 recipients. Conclusion: A specific PAI-1 inhibitor prevents thrombus formation and arterial neointimal formation after arterial injury. Thus, PAI-1 plays a critical role in arterial remodeling after mechanical injury. PAI-1 regulation may be useful to prevent thrombus and neointimal formation after PCI.

An IκB kinase 2 inhibitor IMD-0354 suppresses the survival of adult T-cell leukemia cells

Adult T‐cell leukemia (ATL) is a fatal T‐cell malignancy associated with human T‐cell leukemia virus type I infection. The aberrant expression of nuclear factor‐κB (NF‐κB) is considered to contribute to the malignant phenotype and chemo‐resistance of ATL cells. Because of the poor prognosis of ATL, the development of new therapeutic strategies is direly needed. In the present study, we show that an IκB kinase 2 (IKK2) inhibitor, IMD‐0354, efficiently inhibits the survival of CD4+ CD25+  primary ATL cells and prevents the growth of or induces apoptosis of patient‐derived ATL cell lines. Assays of transcription with integrated forms of reporter genes revealed that IMD‐0354 suppresses NF‐κB‐dependent transcriptional activity. Moreover, the daily administration of IMD‐0354 prevents the growth of tumors in mice inoculated with ATL cells. Our results suggest that targeting IKK2 with a small molecule inhibitor, such as IMD‐0354, is an attractive strategy for the treatment of ATL. (Cancer Sci 2012; 103: 100–106)

Activation of IκB Kinase β and NF-κB Is Essential for Helicobacter pylori-Induced Chronic Gastritis in Mongolian Gerbils

ABSTRACT The Mongolian gerbil model of Helicobacter pylori infection resembles human gastritis. In this study, we investigated the role of NF-κB activation in H. pylori-infected gerbils. Activated macrophages were significantly increased in H. pylori-infected gastric mucosa and were identified as being important cells with potent activation of NF-κB, which plays an important part in producing proinflammatory cytokines. Macrophage depletion by the administration of clodronate resulted in milder inflammation in gerbils infected with H. pylori. In macrophages, the inhibition of IκB kinase β (IKKβ), which is a critical kinase for NF-κB activation, resulted in lower proinflammatory cytokine expression caused by heat-killed H. pylori cells. Furthermore, treatment with IKKβ inhibitor resulted in milder inflammation in gerbils with H. pylori gastritis. Collectively, our data suggest that H. pylori-mediated gastric inflammation critically depends on the efficient recruitment and activation of macrophages, with sufficient NF-κB activation.

Effects of specific chemical suppressors of plasminogen activator inhibitor-1 in cardiovascular diseases.

Introduction: Plasminogen activator inhibitor-1 (PAI-1) is critical in thrombus formation and inflammation. Although these are essential pathological features of cardiovascular diseases, the effects of PAI-1 inhibition against the development of cardiovascular remodeling have not been well studied. Areas covered: The review explores the therapeutic value of PAI-1 in the progression of various cardiovascular diseases. To date, the authors have reported that a novel PAI-1 inhibitor suppressed the development of experimental autoimmune myocarditis, vascular remodeling after arterial injury, and heart transplant rejection using rodent models. Pathologically, the PAI-1 inhibitor improved histological remodeling of myocardium and arteries with suppression of inflammation and thrombus formation. Expert opinion: PAI-1 inhibitors appear to exhibit potent effects on the prevention of adverse tissue remodeling. However, PAI-1 is a multifunctional protein and more research is needed to further elucidate the association between PAI-1 expression and cardiovascular disease.