Previous reports have demonstrated evidence for a negative feedback loop between Erk and Raf. Cells resistant to MEK inhibitor induced cell cycle arrest have an intact feedback loop. Abrogation of the negative feedback loop by MEK inhibitors results in accumulation of phospho-MEK and activation of Raf proteins (Cancer Res. 68(15):6145-53, 2008). The data suggest that resistance to MEK inhibitors may be due in part to enhanced cell survival signaling through Raf-dependant, Erk-independent pathways. Our present study examined the impact of MAPK signaling on the sensitivity of human cancer cells to Sorafenib. The effects of MEK inhibitors, Sorafenib and the combination of both agents were evaluated in a broad spectrum of cancer cell lines, primary human cancer cells derived from CLL patients, as well as in xenografts. Synergistic cytotoxicity was observed with co-administration of Sorafenib and MEK inhibitors, including RDEA119, U0126, or PD184352, in solid tumor cell lines, such as Hepatoma (Hep3B, HUH7, HepG2), NSCLC (A549), Colon (HCT116), Glioma (D37), pancreatic (BxPC3), osteosarcoma (U2-OS), and leukemia cell lines (K562) as well as primary cancer cells derived from CLL patients. Cell death seen with the sorafenib/MEK inhibitor combination is typical of apoptosis, ie, cytochrome c release, as well as caspase and PARP cleavage. In addition, knock-down of Bim by shRNA substantially attenuated Sorafanib/MEK inhibitor-mediated cell death As expected, MEK inhibitor treatment decreased levels of phospho-Erk1/2 but resulted Raf activation and increased phosphoMEK, consistent with abrogation of the negative feed back loop. Sorafenib blocked the accumulation of phospho-MEK mediated by MEK inhibitors, and the co-exposure of cells to Sorafenib/MEK inhibitors enhanced Bim(EL) expression. Moreover, either knock-down of Erk1 and Erk2, or Raf-1 and B-Raf by selective RNA interference markedly promoted Sorafenib-, or U0126-induced apoptosis, respectively. These findings provide evidence that modulation of Raf activity by feedback inhibition between Erk and Raf participates in resistance to MEK inhibitor induced cytotoxicity, and Bim plays an important role in Sorafenib/MEKinhbitor-induced cell death. Additionally, these findings suggest that sequential targeting of multiple proteins in the MAPK pathway may be necessary for maximal anti-tumor activity. Our results support further clinical testing of Sorafenib in combination with a MEK inhibitor.

Citation Information: In: Proc Am Assoc Cancer Res; 2009 Apr 18-22; Denver, CO. Philadelphia (PA): AACR; 2009. Abstract nr 3692.

100th AACR Annual Meeting-- Apr 18-22, 2009; Denver, CO