Members of the Ras protooncogene family are mutated in approximately 75% of
colon cancers. The
Raf kinases (Raf-1, b-Raf and a-Raf) directly interact with Ras and serve as mediators of mitogenic signals. Expression of the constitutively active alleles of Raf or Ras gene families results in
oncogenesis in a number of model systems. Previous studies emphasized the importance of Raf-1 and b-Raf in preventing apoptosis in addition to their roles in cell growth. In the present study, we examined whether inhibition of the Raf-1 or
b-Raf kinase decreases cell growth and increases apoptosis in
colon cancer cells. c-Raf and b-Raf were depleted in
colon cancer cell lines, such as HCT116, HT29 and Colo205, containing Ras or b-Raf mutations by RNA interference (RNAi). The results showed that
colon cancer cells with activating Ras mutations undergo apoptosis following Raf-1 inhibition, as determined by cell cycle analysis and the release of cytochrome c. Moreover, in b-Raf mutant
colon cancers, the inhibition of b-Raf as compared to Raf-1 is crucial for
cancer cell death. There is increasing evidence for both
MEK-independent Raf signaling and Raf-independent
MEK signaling. Thus, we investigated whether targeting multiple points of the
mitogen-activated protein kinase (MAPK) pathway with a
MEK inhibitor and Raf RNAi increases
cancer cell death. The results showed that combination
therapy, inhibiting Raf and
MEK kinases simultaneously, increased apoptosis in
colon cancer cells. Taken together, our data demonstrate that combination
therapy targeting the MAPK pathway at two distinct points,
Raf kinase and
MEK, has greater efficacy in increasing
cancer cell death and is likely to improve therapeutic outcomes for patients.