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

Geldanamycin is a benzoquinone ansamycin with multiple pharmacologic properties. 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 gene expression and nuclear factor (NF)-kappaB activation. Geldanamycin inhibited tumor necrosis factor (TNF)-alpha -mediated IL-8 gene expression in A549 human respiratory epithelial cells as measured by enzyme-linked immunosorbent assay and Northern blot analyses. In cells transiently transfected with an IL-8 promoter-luciferase reporter plasmid, geldanamycin inhibited TNF-a-mediated luciferase activity. Geldanamycin inhibited TNF-a-mediated NF-kappaB activation as measured by electromobility shift assays and transient transfections with a NF-kappaB-dependent luciferase reporter plasmid. In contrast, 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. Geldanamycin added directly to nuclear extracts of TNF-a-treated cells reduced the formation of the NF-kappaB/DNA complex. These results demonstrate that geldanamycin inhibits TNF-a-mediated IL-8 gene expression in A549 cells by inhibiting activation of the IL-8 promoter. The mechanism of inhibition involves inhibition of NF-KB activation, which is independent of I kappaB alpha degradation or p65 nuclear translocation. Geldanamycin appears to directly inhibit the ability of NF-KB to bind DNA. The observed in vitro effects could account, in part, for the anti-inflammatory properties of geldanamycin observed in vivo.