Honokiol inhibits TNF-α-stimulated NF-κB activation and NF-κB-regulated gene expression through suppression of IKK activation

Honokiol, a small molecular weight lignan originally isolated from Magnolia officinalis, shows anti-angiogenic, anti-invasive and anti-proliferative activities in a variety of cancers. In this study, we investigated whether honokiol affects the transcription factor nuclear factor-kappa B (NF-κB) which controls a large number of genes involved in angiogenesis, metastasis and cell survival. We observed that the tumor necrosis factor-alpha (TNF-α)-induced NF-κB activation was blocked by honokiol in four different cancer cell lines as evidenced by EMSA. Honokiol did not directly affect the NF-κB-DNA binding. Immunoblot experiments demonstrated that honokiol inhibited the TNF-α-stimulated phosphorylation and degradation of the cytosolic NF-κB inhibitor IκBα. Furthermore, honokiol suppressed the intrinsic and TNF-α-stimulated upstream IκB kinases (IKKs) activities measured by a non-radioactive kinase assay using immunoprecipitated IKKs, suggesting a critical role of honokiol in abrogating the phosphorylation and degradation of IκBα. In a HeLa cell NF-κB-dependent luciferase reporter system, honokiol suppressed luciferase expression stimulated by TNF-α and by the transient transfection and expression of NIK (NF-κB-inducing kinase), wild type IKKβ, constitutively active IKKα and IKKβ, or the p65 subunit. Honokiol was also found to inhibit the nuclear translocation and phosphorylation of p65 subunit of NF-κB. RT-PCR results showed that honokiol suppressed NF-κB-regulated inflammatory and carcinogenic gene products including MMP-9, TNF-α, IL-8, ICAM-1 and MCP-1. In line with the observation that NF-κB activation may up-regulate anti-apoptotic genes, it was shown that honokiol enhanced TNF-α-induced apoptotic cell death. In summary, our results demonstrate that honokiol suppresses NF-κB activation and NF-κB-regulated gene expression through the inhibition of IKKs, which provides a possible mechanism for its anti-tumor actions.