Oncogenic Ras increases sensitivity of colon cancer cells to 5-FU-induced apoptosis.
Despite the fact that objective response rates to 5-FU are as low as 20%, 5-FU remains the most commonly used drug for the treatment of colorectal cancer. The lack of understanding of resistance to 5-FU, therefore, remains a significant impediment in maximizing its efficacy. We used intestinal epithelial cells with an inducible K-RasV12 to demonstrate that expression of oncogenic Ras promotes cell death upon 5-FU treatment. Accordingly, transient expression of the mutant RasV12, but not the WT Ras, enhanced 5-FU-induced apoptosis in 293T cells. Consistent with these data, we showed that targeted deletion of the mutant Ras allele in the HCT116 colon cancer cell line protected cells from 5-FU-induced apoptosis. Using isogenic colon cancer cell lines that differ only by the presence of the mutant Ras allele, HCT116 and Hke-3 cells, we demonstrated that signaling by oncogenic Ras promotes both accumulation of p53 and its phosphorylation on serine15 in response to 5-FU, a situation that favors apoptosis over growth arrest. However, despite the differential induction of p53 in HCT116 and Hke-3 cells, the expression of Puma, a gene with an important role in p53-dependent apoptosis, was not affected by Ras signaling. In contrast, we showed that Ras interferes with 5-FU-induced expression of gelsolin, a protein with known antiapoptotic activity. We ascertained the role of gelsolin in 5-FU-induced apoptosis by demonstrating that silencing of gelsolin expression through RNAi sensitized cells to 5-FU-induced apoptosis and that re-expression of gelsolin in cells harboring mutant Ras protected cells from 5-FU-induced apoptosis. These data therefore demonstrate that Ras mutations increase sensitivity to 5-FU-induced apoptosis at least in part through the negative regulation of gelsolin expression. Our data indicate that Ras mutations promote apoptosis in response to 5-FU treatment and imply that tumors with Ras mutations and/or reduced expression of gelsolin may show enhanced apoptosis in response to 5-FU also in vivo.
Klampfer
Lidija
L
Albert Einstein Cancer Center, Montefiore Medical Center, Department of Oncology, Bronx, NY 10467, USA. lklampf@aecom.yu.edu
Swaby
Laurie-Anne
LA
Huang
Jie
J
Sasazuki
Takehiko
T
Shirasawa
Senji
S
Augenlicht
Leonard
L
D017209:Apoptosis / Q000187:drug effects*; D053148:Caspase 3; D020169:Caspases / Q000378:metabolism; D045744:Cell Line, Tumor; D003110:Colonic Neoplasms / Q000473:pathology*; D004305:Dose-Response Relationship, Drug; D005472:Fluorouracil / Q000633:toxicity*; D018260:Gelsolin / Q000494:pharmacology; D006801:Humans; D007700:Kinetics
D018260:Gelsolin; C505407:CASP3 protein, human; D053148:Caspase 3; D020169:Caspases; D005472:Fluorouracil