Mieke Buntinx

Layered double hydroxides exchanged with tungstate as biomimetic catalysts for mild oxidative bromination

The manufacture of a range of bulk and fine chemicals, including flame retardants, disinfectants and antibacterial and antiviral drugs, involves bromination. Conventional bromination methods typically use elemental bromine, a pollutant and a safety and health hazard. Attempts to develop alternative and more benign strategies have been inspired by haloperoxidase enzymes, which achieve selective halogenation at room temperature and nearly neutral pH by oxidizing inorganic halides with hydrogen peroxide,. The enzyme vanadium bromoperoxidase has attracted particular interest, in this regard, and several homogeneous inorganic catalysts mimicking its activity are available, although they are limited by the requirement for strongly acidic reaction media. A heterogenous mimic operating at neutral pH has also been reported, but shows only modest catalytic activity. Here we describe a tungstate-exchanged layered double hydroxide that catalyses oxidative bromination and bromide-assisted epoxidation reactions in a selective manner. We find that the catalyst is over 100 times more active than its homogeneous analogue. The low cost and heterogeneous character of this system, together with its ability to operate efficiently under mild conditions using bromides rather than elemental bromine, raise the prospect of being able to develop a clean and efficient industrial route to brominated chemicals and drugs and epoxide intermediates.

Cytokine-induced cell death in human oligodendroglial cell lines: I. Synergistic effects of IFN-γ and TNF-α on apoptosis

Multiple sclerosis is a chronic inflammatory disease of the central nervous system. Myelin and oligodendrocytes are considered the major targets of injury caused by a cell‐mediated immune response. There is circumstantial evidence that proinflammatory cytokines like tumor necrosis factor α (TNF‐α) and interferon γ (IFN‐γ) could have disease‐promoting roles in multiple sclerosis (MS). In the present study, the cytotoxic effects of IFN‐γ and TNF‐α on the human oligodendroglial cell lines human oligodendroglioma (HOG) and MO3.13 were analyzed. When the oligodendroglial cell lines were cultured in the presence of IFN‐γ or TNF‐α, apoptotic cell death was observed in both cell lines after >24 hr incubation. Apoptosis was evidenced by a decrease in cell viability, apoptotic changes in cell and nucleus morphology, and disruption of the membrane asymmetry. Our data show that TNF‐α and IFN‐γ induce apoptosis in a dose‐dependent fashion in both oligodendroglial cell lines and that their synergistic effect results in enhanced cell death. Understanding the regulation of cell death pathways in oligodendrocytes is critical for protecting myelin‐producing cells and their associated axons during injury in patients with MS.

Characterization of three human oligodendroglial cell lines as a model to study oligodendrocyte injury: morphology and oligodendrocyte-specific gene expression.

Oligodendrocytes, the myelin-forming cells of the central nervous system, are the target of pathogenic immune responses in multiple sclerosis. Primary cultures of human oligodendrocytes have been used to unravel the cellular and molecular mechanisms of immune-mediated injury of oligodendrocytes. However, these studies are hampered by the limited availability of viable human brain tissue. The present study was aimed at comparing the morphological and biochemical characteristics of the human oligodendroglial cell lines HOG, MO3.13 and KG-1C. We have determined oligodendrocyte-associated features of these lines and analyzed the degree to which they can be used as a model of human oligodendrocytes arrested at specific developmental stages. The oligodendroglial cell lines all exhibited markers of immature oligodendrocytes, such as CNPase and GalC, but not the astrocytic marker GFAP. Differentiation could be induced in HOG and MO3.13 cells, as was seen through a decrease in proliferation, an increase in process extension without formation of myelin sheets and up-regulation of genes associated with mature oligodendrocytes such as MBP and MOG. Microarray analysis revealed the expression of MAG, MOBP and OMG genes in HOG cells. The KG-1C cells displayed poor growth characteristics in the recommended conditions. In conclusion, our data show that the oligodendroglial cell lines HOG and MO3.13 can be used as a model of human oligodendrocytes ‘arrested’ in an immature developmental stage. Culturing in appropriate medium can induce further differentiation of these cells. These cell lines can therefore be applied as a model to study immune-mediated injury of oligodendrocytes in relation to disease.

Leukemia inhibitory factor is produced by myelin-reactive T cells from multiple sclerosis patients and protects against tumor necrosis factor-α-induced oligodendrocyte apoptosis

In multiple sclerosis (MS), damage to oligodendrocytes is believed to be caused by an aberrant immune response initiated by autoreactive T cells. Increasing evidence indicates that these T cells are not exclusively detrimental but might also exert protective effects. We report for the first time that myelin‐reactive T‐cell clones from eight MS patients (6/19) and five healthy controls (4/11) produce leukemia inhibitory factor (LIF), a member of the neuropoietic family of neurotrophins. In addition, T‐cell clones specific for tetanus toxoid, CD4+ and CD8+ T cells, and monocytes, but not B cells, secreted LIF. LIF‐producing T lymphocytes and macrophages were also identified immunohistochemically in both active and chronic‐active MS lesions. We further demonstrated dose‐dependent protective effects of LIF on tumor necrosis factor‐α‐induced apoptosis of oligodendrocytes. In conclusion, our data demonstrate that peripheral and CNS‐infiltrating T cells from MS patients produce LIF, a protective factor for oligodendrocytes. This study emphasizes that secretion of LIF may contribute to the neuroprotective effects of autoreactive T cells.

Cytokine-induced cell death in human oligodendroglial cell lines. II: Alterations in gene expression induced by interferon-γ and tumor necrosis factor-α

Cytokines, such as interferon‐γ (IFN‐γ) and tumor necrosis factor‐α (TNF‐α), can initiate dual effects resulting in either cell growth or cell death. In this study, the human oligodendroglial cell lines HOG and MO3.13 were used as a model to study the molecular mechanisms of cytokine‐induced cell death in human oligodendrocytes. We have previously shown that TNF‐α and IFN‐γ induce apoptosis in both oligodendroglial cell lines within 72 hr. In the present study, the cell death pathways operating within these cells were further investigated at the gene expression level. Both cell lines express a broad repertoire of caspases and apoptosis‐related genes. Some of these genes are specifically up‐regulated by cytokine treatment; e.g., caspase‐1 is up‐regulated by IFN‐γ. In addition to direct cytotoxic effects, IFN‐γ and TNF‐α also enhance the expression of Fas, TNFR1, and MHC class I molecules in both cell lines. This suggests that cytokines can make oligodendrocytes more vulnerable to different cell death pathways in an inflammatory environment. cDNA microarray analysis of the HOG cell line revealed that TNF‐α induces genes that regulate apoptosis, survival, inflammation, cell metabolism, and cell signaling. The data suggest that oligodendroglial cells activate both death and survival pathways upon cytokine challenges. However, the survival pathways seem to be unable to compete with the death signal after more than 24 hr of cytokine treatment. These results may contribute to the development of therapeutic strategies aimed at interfering with cytokine‐induced cell death of oligodendrocytes in patients with multiple sclerosis.

Transition metal anion exchanged layered double hydroxides as a bioinspired model of vanadium bromoperoxidase

Abstract Tungstate-exchanged layered double hydroxides (LDHs) and V-bromoperoxidase enzymes perform oxidative brominations in a highly similar way: first H 2 O 2 binds on the metal to form a peroxometal complex; next the peroxometal oxidizes Br − to “Br + ”; this electrophilic “Br + ” halogenates an organic compound, or oxidizes a second H 2 O 2 molecule to form excited state singlet oxygen. Full evidence for this similarity is given, based on spectroscopic observation of peroxotungstate and 1 O 2 and on identification of the organic bromination products. In comparison with the homogeneous oxometallates, or with heterogeneous Ti-catalysts, the bioinspired LDH–WO 4 2− catalyst displays much higher rates of Br − oxidation; this rate enhancement is explained. The activity of LDH–WO 4 2− can be enhanced by changing the elemental composition of the octahedral layer of the LDH structure. Since LDH–WO 4 2− is stable toward leaching and high H 2 O 2 concentrations, it is a practical catalyst for oxidative bromination.

Synthesis and characterization of 5,6,7,8-tetrahydroquinoline C5a receptor antagonists

A novel series of substituted 2-aryl-5-amino-5,6,7,8-tetrahydroquinoline C5a receptor antagonists is reported. Synthetic routes were developed that allow the substituents on the tetrahydroquinoline core to be efficiently varied, facilitating determination of structure-activity relationships. Members of the series display high binding affinity for the C5a receptor and are potent functional antagonists.

Pharmacological profile of JNJ-27141491 [(S)-3-[3,4-difluorophenyl)-propyl]-5-isoxazol-5-yl-2-thioxo-2,3-dihydro-1H-imidazole-4-carboxyl acid methyl ester], as a noncompetitive and orally active antagonist of the human chemokine receptor CCR2.

The interaction between CC chemokine receptor 2 (CCR2) with monocyte chemoattractant proteins, such as MCP-1, regulates the activation and recruitment of inflammatory leukocytes. In this study, we characterized (S)-3-[3,4-difluoro-phenyl)-propyl]-5-isoxazol-5-yl-2-thioxo-2,3-dihydro-1H-imidazole-4-carboxyl acid methyl ester (JNJ-27141491) as a noncompetitive and orally active functional antagonist of human (h)CCR2. JNJ-27141491 strongly suppressed hCCR2-mediated in vitro functions, such as MCP-1-induced guanosine 5′-O-(3-[35S]thio)triphosphate binding; MCP-1, -3, and -4-induced Ca2+ mobilization; and leukocyte chemotaxis toward MCP-1 (IC50 = 7–97 nM), whereas it had little or no effect on the function of other chemokine receptors tested. The inhibition of CCR2 function was both insurmountable and reversible, consistent with a noncompetitive mode of action. JNJ-27141491 blocked the binding of 125I-MCP-1 to human monocytes (IC50 = 0.4 μM), but it failed to affect MCP-1 binding to mouse, rat, and dog cells (IC50 > 10 μM). Therefore, transgenic mice, in which the mouse (m)CCR2 gene was replaced by the human counterpart, were generated for in vivo testing. In these mice, oral administration of JNJ-27141491 dose-dependently [5–40 mg/kg q.d. (once daily) or b.i.d.] inhibited monocyte and neutrophil recruitment to the alveolar space 48 h after intratracheal mMCP-1/lipopolysaccharide instillation. Furthermore, treatment with JNJ-27141491 (20 mg/kg q.d.) significantly delayed the onset and temporarily reduced neurological signs in an experimental autoimmune encephalomyelitis model of multiple sclerosis. Taken together, these results identify JNJ-27141491 as a noncompetitive, functional antagonist of hCCR2, capable of exerting oral anti-inflammatory activity in transgenic hCCR2-expressing mice.

Synthesis and structure-activity relationship of benzetimide derivatives as human CXCR3 antagonists.

The synthesis and evaluation of benzetimide derivatives showing potent CXCR3 antagonism are described. Optimization of the screening hits led to the identification of more potent CXCR3 antagonists devoid of anti-cholinergic activity and identification of the key pharmacophore moieties of the series.

Design and optimization of aniline-substituted tetrahydroquinoline C5a receptor antagonists

A series of aniline-substituted tetrahydroquinoline C5a receptor antagonists were discovered. A functionality requirement of ortho substitution on the aniline was revealed. Secondary anilines, in general, outperformed tertiary analogs in inhibition of C5a-induced calcium mobilization. Further enhancement of activity was realized in the presence of an ortho hydroxyalkyl side chain. The functional IC(50) of selected analogs was optimized to the single-digit nanomolar level.