No abstract

The p38 mitogen-activated protein kinase (MAPK) participates in intracellular signaling cascades resulting in inflammatory responses. Therefore, inhibition of the p38 MAPK pathway may form the basis of a new strategy for treatment of inflammatory diseases. However, p38 MAPK activation during systemic inflammation in humans has not yet been shown, and its functional significance in vivo remains unclear. Hence, we exposed 24 healthy male subjects to an i.v. dose of LPS (4 ng/kg), preceded 3 h earlier by orally administered 600 or 50 mg BIRB 796 BS (an in vitro p38 MAPK inhibitor) or placebo. Both doses of BIRB 796 BS significantly inhibited LPS-induced p38 MAPK activation in the leukocyte fraction of the volunteers. Cytokine production (TNF-α, IL-6, IL-10, and IL-1R antagonist) was strongly inhibited by both low and high dose p38 MAPK inhibitor. In addition, p38 MAPK inhibition diminished leukocyte responses, including neutrophilia, release of elastase-α1-antitrypsin complexes, and up-regulation of CD11b with down-regulation of L-selectin. Finally, blocking p38 MAPK decreased C-reactive protein release. These data identify p38 MAPK as a principal mediator of the inflammatory response to LPS in humans. Furthermore, the anti-inflammatory potential of an oral p38 MAPK inhibitor in humans in vivo suggests that p38 MAPK inhibitors may provide a new therapeutic option in the treatment of inflammatory diseases.

Violacein, a pigment isolated from Chromobacterium violaceum in the Amazon River, presents diverse biologic properties and attracts interest as a consequence of its antileukemic activity. Elucidation of the molecular mechanism mediating this activity will provide further relevant information for understanding its effects on the cellular physiology of untransformed cells and for considering its possible clinical application. Here, we show that violacein causes apoptosis in HL60 leukemic cells but is ineffective in this respect in other types of leukemia cells or in normal human lymphocytes and monocytes. Violacein cytotoxicity in HL60 cells was preceded by activation of caspase 8, transcription of nuclear factor B (NF-B) target genes, and p38 mitogenactivated protein (MAP) kinase activation. Thus, violacein effects resemble tumor necrosis factor ␣ (TNF-␣) signal transduction in these cells. Accordingly, infliximab, an antibody that antagonizes TNF-␣-induced signaling abolished the biologic activity of violacein. Moreover, violacein directly activated TNF receptor 1 signaling, because a violacein-dependent association of TNF receptor-associated factor 2 (TRAF2) to this TNF receptor was observed in coimmunoprecipitation experiments. Hence, violacein represents the first member of a novel class of cytotoxic drugs mediating apoptosis of HL60 cells by way of the specific activation of TNF receptor 1. IntroductionViolacein ( Figure 1A), a purple-colored pigment produced by one of the strains of Chromobacterium violaceum found in the Amazon River, Brazil, is an indole derivative characterized as 3-(1,2-dihydro-5-(5-hydroxy-1H-indol-3-yl)-2-oxo-3H-pyrrol-3-ilydene)-1,3-dihydro-2H-indol-2-one. 1 The biosynthesis and biologic properties of this pigment have been extensively studied; in particular, its antitumoral, antibacterial, antiulcerogenic, antileishmanial, and antiviral activities are of interest. [1][2][3][4][5][6] Its activity on acute myeloblastic leukemia (HL60) cells is of special interest because the compound effectively induces cell death in these cells.The development of violacein as a possible novel therapeutic avenue for the treatment of leukemia, however, depends on the establishment of the molecular basis for this cytotoxicity. Evidently, a possible future application of violacein as a therapeutic agent critically depends on its ability to target leukemia cells more effectively than normal blood cells. Hence, research comparing the cytotoxicity of violacein in transformed cells and the corresponding untransformed ones is urgently called for. In addition, identification of the molecular details mediating violacein effects on leukemic cells will provide insight into the possible benefit of this compound in comparison to existing therapeutics.The above-mentioned considerations prompted us to investigate the effects of violacein in HL60 cells. This cell line is generally accepted as a valid model for studying myeloid leukemia biology. 7,8 Melo et al 6 showed earlier that HL60 cells react to violacein with bo...

Hybrid drug 1 (NO-ASA) continues to attract intense research from chemists and biologists alike. It consists of ASA and a -ONO2 group connected through a spacer and is in preclinical development as an antitumor drug. We report that, contrary to current beliefs, neither ASA nor NO contributes to this antitumor effect. Rather, an unsubstituted QM was identified as the sole cytotoxic agent. QM forms from 1 after carboxylic ester hydrolysis and, in accordance with the HSAB theory, selectively reacts with cellular GSH, which in turn triggers cell death. Remarkably, a derivative lacking ASA and the -ONO2 group is 10 times more effective than 1. Thus, our data provide a conclusive molecular mechanism for the antitumor activity of 1. Equally importantly, we show for the first time that a "presumed invisible" linker in a hybrid drug is not so invisible after all and is in fact solely responsible for the biological effect.