Treatment of gastric cancer cells with 5-fluorouracil/leucovorin and irinotecan induces distinct alterations in the mRNA expression of the apoptosis-related genes, including the novel gene BCL2L12.

Abstract

The BCL2 (bcl-2)family of genes is highly involved in the apoptotic mechanisms. We have recently discovered and cloned a novel member of the same family, BCL2L12. The aim of this study was to investigate the modulations in the BAX, BCL2 and BCL2L12 mRNA levels in gastric cancer cells, after their treatment with the anticancer drugs 5-fluorouracil and irinotecan as well as the antioxidant substance leucovorin. AGS gastric cancer cells were examined for their sensitivity to antineoplastic drugs and/or antioxidants using the MTT assay. Total RNA was extracted and reverse transcribed into cDNA. A highly sensitive quantitative real-time PCR method for the mRNA quantification was developed using the SYBR Green chemistry. GAPDH was used as a housekeeping gene. Relative quantification analysis was performed using the comparative threshold cycle method. Treatment of AGS cells with 5-fluorouracil (5 microM), leucovorin (10 microM), a combination of the latter and, eventually, irinotecan (1 microM) for 3 time periods (24, 48 and 72 h), resulted in significant modulations of the BCL2, BAX and BCL2L12 mRNA levels compared with the untreated cells. Our experimental data demonstrate that the molecular profile mainly of BCL2 and BCL2L12 genes may serve as a new potential molecular biomarker predicting treatment response in gastric cancer cells.