Received for publication January 15, 2008.
Revised May 5, 2008.
Accepted for publication May 5, 2008.

Activation of NF-kB pathway by simvastatin and RhoA silencing increases doxorubicin cytotoxicity in human colon cancer HT29 cells

Abstract

Doxorubicin efficacy in cancer therapy is hampered by the dose-dependent side effects, which may be overcome by reducing the drug's dose and increasing its efficacy. In the present work we suggest that the activation of the nuclear factor-kappa B (NF-kB) pathway and of nitric oxide (NO) synthase increases the doxorubicin efficacy in human colon cancer HT29 cells. To induce NF-kB, we took into account the effect of doxorubicin itself and of the 3-hydroxy-3-methylglutaryl coenzyme A reductase inhibitor simvastatin; as NF-kB inhibitors, we chose the sesquiterpene lactones parthenolide and artemisinin. Simvastatin increased the NF-kB activity and NO synthesis, elicited the tyrosine nitration of the multidrug resistance-related protein 3, and enhanced the doxorubicin intracellular accumulation and cytotoxicity. Simvastatin potentiated the effect of doxorubicin on the NF-kB pathway and the inducible NO synthase expression. The effects of simvastatin were due to the inhibition of the small G-protein RhoA and of its effector Rho kinase. Parthenolide and artemisinin prevented all the statin effects, by inducing RhoA/Rho kinase activation. On the opposite they did not reduce the NF-kB translocation and doxorubicin intracellular content when RhoA was silenced by small interfering RNA (siRNA). Interestingly, RhoA siRNA was sufficient to increase NF-kB translocation, NO synthase activity, doxorubicin accumulation and cytotoxicity also in non-stimulated cells. Our results suggest that artemisinin, a widely used antimalarial drug, may impair the response to doxorubicin in colon cancer cells; on the contrary, simvastatin and RhoA siRNA may represent future therapeutic approaches to improve doxorubicin efficacy, reducing the risk of doxorubicin-dependent adverse effects.

Key words: Nitric oxide, Nitric oxide synthases, NFkappaB, MDR/p-Glycoprotein, RNA/siRNA, Resistance