MOLECULAR CHARACTERIZATION OF ESTABLISHED HUMAN COLON CARCINOMA CELL LINES (HCT-8) MADE RESISTANT TO 5-FLUOROURACIL BY DIFFERENT SELECTION SCHEDULES

To provide some insight into molecular mechanisms of 5 fluorouracil (5-FU) clinical resistance in colorectal cancer, we hypothesized that different in vitro exposure schedules of human colorectal cancer cell lines mimicking clinical infusion or bolus regimens could lead to differential gene expression. Resistant HCT-8 colon cancer cell lines (HCT-8/FUI/15R and HCT-8/FUB/2R) were selected from parental sensitive HCT-8 cells by long-term and short-term exposure schedules, respectively. Expression levels of the 437 genes evaluated by the Atlas Select cDNA Expression Human Tumor Array were not substantially different between HCT-8/FUB/2R and HCT-8 cell lines except for three genes downregulated in the resistant subline. Several genes were differentially expressed in HCT-8/FUI/15R cells compared to the parental cell line: 43 genes, including three chemoresistance-related genes, were upregulated, and three genes were downregulated. HCT-8/FUB/2R cells were substantially more resistant to 5-FU in comparison to HCT-8/FUI/15R cells after both 4- and 72-h exposures. No substantial differences were observed among resistant and parental cells in sensitivity to SN-38, the active metabolite of irinotecan, and oxaliplatin. Analysis of the mRNA levels of thymidylate synthase, thymidine phosphorylase, and bcl-2 genes evaluated by reverse transcription and real time PCR (RT-PCR) assay showed comparable results in resistant sublines and sensitive parental cells, whereas expression of the dihydropyrimidine dehydrogenase gene was markedly increased in both resistant cell lines compared to parental cells