title: Heteroarylketones inhibit astroglial interleukin-6 expression via a STAT3/NF-κB signaling pathway.
abstract: BACKGROUND
  Elevated brain levels of the pleiotropic cytokine interleukin-6, which is mainly secreted from activated local astrocytes, contribute to pathological events including neuroinflammation and neurodegeneration. Thus, inhibition of pathological IL-6 expression provides a rationale strategy for targeting the onset or further progression of neurological disorders including Alzheimer's disease, multiple sclerosis, Parkinson's disease and traumatic brain injury. The purpose of this study was to identify and to characterize new potent inhibitors of astrocytic IL-6 expression for further therapeutic development of novel anti-inflammatory and neuroprotective drugs.
  
  
  METHODS
  Oncostatin M (OSM)-treated human glioma U343 cells were used as model for induction of astrocytic IL-6 expression. This model was characterized by immunoblotting, siRNA technique, ELISA and qRT-PCR and used to screen low molecular weight compound libraries for IL-6-lowering effects. To validate bioactive compounds identified from library screens, bacterial lipopolysaccharide was used to induce IL-6 expression in cultivated primary astrocytes and in mice in vivo. To dissect underlying molecular mechanisms, protein extracts from OSM-treated U343 cells were analyzed by phospho-specific immunoblotting and immunocytochemistry as well as by co-immunoprecipitation.
  
  
  RESULTS
  OSM-treatment (100 ng/ml; 24 h) led to 30-fold increase of IL-6 secretion from U343 cells. The temporal profile of IL-6 mRNA induction displayed a biphasic induction pattern with peak synthesis at 1 h (6.5-fold) and 16 h (5.5-fold) post stimulation. IL-6 protein release did not show that biphasic pattern and was detected as early as 3 h post stimulation reaching a maximum at 24 h. The screen of compound libraries identified a set of heteroarylketones (HAKs) as potent inhibitors of IL-6 secretion. HAK compounds affected the second peak in IL-6 mRNA synthesis, whereas the first peak was insensitive to HAK treatment. HAK compounds also suppressed lipopolysaccharide-induced IL-6 expression in primary murine astrocytes as well as in brain and plasma samples from lipopolysaccharide-treated mice. Finally, HAK compounds were demonstrated to specifically suppress the OSM-induced phosphorylation of STAT3 at serine 727 and the physical interaction of pSTAT3S727 with p65.
  
  
  CONCLUSIONS
  Heteroarylketone compounds are potent inhibitors of IL-6 expression in vitro and in vivo and may represent a new class of potent anti-inflammatory and neuroprotective drugs.
authors.affiliation: Probiodrug AG, Weinbergweg 22, Halle/Saale, 06120, Germany.
authors.forename: Ingo
authors.initials: I
authors.lastname: Schulz
authors.forename: Claudia
authors.initials: C
authors.lastname: Engel
authors.forename: André J
authors.initials: AJ
authors.lastname: Niestroj
authors.forename: Ulrike
authors.initials: U
authors.lastname: Zeitschel
authors.forename: Katja
authors.initials: K
authors.lastname: Menge
authors.forename: Astrid
authors.initials: A
authors.lastname: Kehlen
authors.forename: Antje
authors.initials: A
authors.lastname: Meyer
authors.forename: Steffen
authors.initials: S
authors.lastname: Rossner
authors.forename: Hans-Ulrich
authors.initials: HU
authors.lastname: Demuth
mesh_terms: D000818:Animals; D000970:Antineoplastic Agents / Q000494:pharmacology; D001253:Astrocytes / Q000166:cytology / Q000187:drug effects* / Q000378:metabolism*; D045744:Cell Line, Tumor; D002478:Cells, Cultured; D006801:Humans; D015850:Interleukin-6 / Q000235:genetics / Q000378:metabolism*; D007659:Ketones / Q000737:chemistry / Q000494:pharmacology*; D051379:Mice; D008810:Mice, Inbred C57BL; D048052:Mitogen-Activated Protein Kinase 3 / Q000235:genetics / Q000378:metabolism; D015394:Molecular Structure; D016328:NF-kappa B / Q000235:genetics / Q000378:metabolism*; D053683:Oncostatin M / Q000494:pharmacology; D011897:Random Allocation; D051381:Rats; D050796:STAT3 Transcription Factor / Q000235:genetics / Q000378:metabolism*; D015398:Signal Transduction / Q000187:drug effects*
chemical_list: D000970:Antineoplastic Agents; D015850:Interleukin-6; D007659:Ketones; D016328:NF-kappa B; D050796:STAT3 Transcription Factor; D053683:Oncostatin M; D048052:Mitogen-Activated Protein Kinase 3