Geldanamycin inhibits NF-kappa B activation and interleukin-8 gene expression in cultured human respiratory epithelium

Geldanamycin is a benzoquinone ansamycin with multiple pharmacologic proper ties. Recent data demonstrated that geldanamycin conferred protection in an animal model of inflammation-associated acute lung injury. In the current study, we investigated the effects of geldanamycin on interleukin (IL)-8 ge ne expression and nuclear factor (NF)-kappaB activation. Geldanamycin inhib ited tumor necrosis factor (TNF)-alpha -mediated IL-8 gene expression in A5 49 human respiratory epithelial cells as measured by enzyme-linked immunoso rbent assay and Northern blot analyses. In cells transiently transfected wi th an IL-8 promoter-luciferase reporter plasmid, geldanamycin inhibited TNF -a-mediated luciferase activity. Geldanamycin inhibited TNF-a-mediated NF-k appaB activation as measured by electromobility shift assays and transient transfections with a NF-kappaB-dependent luciferase reporter plasmid. In co ntrast, geldanamycin did not affect TNF-a-mediated degradation of the NF-KB inhibitory protein I kappaB alpha and did not block nuclear translocation of the NF-kappaB p65 subunit as measured by Western blot analyses. Geldanam ycin added directly to nuclear extracts of TNF-a-treated cells reduced the formation of the NF-kappaB/DNA complex. These results demonstrate that geld anamycin inhibits TNF-a-mediated IL-8 gene expression in A549 cells by inhi biting activation of the IL-8 promoter. The mechanism of inhibition involve s inhibition of NF-KB activation, which is independent of I kappaB alpha de gradation or p65 nuclear translocation. Geldanamycin appears to directly in hibit the ability of NF-KB to bind DNA. The observed in vitro effects could account, in part, for the anti-inflammatory properties of geldanamycin obs erved in vivo.