Induction of Resistance to the Multi-Targeted Antifolate MTA (LY231514) in Widr Human Colon Cancer Cells

Abstract

The enzyme thymidylate synthase (TS) catalyses the methylation of 2′-deoxyuridine-5′monophosphate (dUMP) to 2′-deoxythymidine-5′-monophosphate (dTMP), an essential precursor during DNA synthesis (1). 5,10-Methylene tetrahydrofolate (CH2-THF) is the limiting methyl donor during this reaction. TS, is usually elevated in tumors (2) and is therefore an interesting target for anticancer agents such as the antifolate multi-targeted antifolate (MTA, ALIMTA, PEMETREXED)(3,4), which inhibits activity of TS by competition with the binding site of CH2-THF of TS. MTA is currently been developed as an anticancer agent for treatment of colorectal, non-small-cell lung cancer and mesothelioma (4). Resistance to this agent might develop similarly to other antifolates, and might be due to elevated activity or mutations of the target enzyme TS, impaired polyglutamation of antifolates, decreased transport into the cells, but also apoptosis regulating proteins may be involved (5,6). For this purpose we induced resistance to the antifolate MTA in the colon cancer cell line WiDr by exposure to 50 mM MTA for 4 hours every week, resulting in the WiDr-4LY cell line, and by continuous exposure for 72 hr to 20 mM MTA every week, resulting in WiDr-cLY. The resistant WiDr variants were studied on the level of TS and folyl polyglutamate synthetase (FPGS).