Sequence-dependent Combination of 5-Fluorouracil and Targeted Agents in RAF-mutant Colorectal Cancer Cell Lines

Abstract

V-raf murine sarcoma viral oncogene homolog B (BRAF)-mutant colorectal cancer has limited therapeutic options for targeted agent and showed poor response to standard chemotherapy containing 5-fluorouracil (5-FU). We previously reported that 5-FU induced the expression of thymidine kinase 1 (TK1), a rate-limiting enzyme for thymidine salvage pathway, which is related with acquired resistance for 5-FU. As TK1 expression was dependent on growth-stimulatory signal such as Braf-MEK and PI3K-AKT pathway, we investigated whether the sequential treatment of 5-FU followed by targeted agent could induce synergy for cell viability and repress TK1 expression in compared with the treatment in reverse sequence. Treatment with 5-FU for 2 days followed by vemurafenib, a mutant BRAF inhibitor, for another 2 days induced significant increase of cell death in compared with the single treatment in colo205 and RKO cell lines. However, the combined treatment in reverse sequence showed the similar response to single treatment. Treatment of dactolisib (a PI3K/mTOR inhibitor) or MK-2206 (a AKT inhibitor) preceded by 5-FU induced cell death more than single agent. Induction of TK1 by 5-FU was abrogated by subsequent treatment with vemurafenib, dactolisib, or MK-2206, whereas 5-FU still induced TK1 expression in the cells pretreated with these targeted agents. In summary, the sequence-dependent treatment of 5-FU and targeted agents is effective through the extinction of 5-FU-induced TK1 activation, which could give a promising strategy for patients having BRAF-mutant colon cancers.