Objective: Through establishing several types of 5-fluorouracil (5-FU) resistant human colon cancer cell lines, to investigate the biological characteristics of the 5-FU resistant colon cancer cells and the mechanism for drug resistance.
Methods: Using human colon cancer HT-29, LoVo and SW480 cells, the 5-FU resistant colon cancer cell lines HT-29/5-FU, LoVo/5-FU and SW480/5-FU were established by repeated exposure to excessive and increasing concentrations of 5-FU. In the established drug-resistant cell lines and their parent cells after treatment with different concentrations of 5-FU, the sensitivities to 5-FU, the cell cycle distributions, the mRNA and protein expressions of drug resistance-related molecules [P-glycoprotein (P-gp), multidrug resistance-associated protein 1 (MRP1), and ATP-binding cassette superfamily G member 2 (ABCG2)] and phosphatase and tensin homolog deleted on chromosome 10 (PTEN) and protein kinase B (Akt) were determined by MTT assay, flow cytometry, qRT-PCR and Western blot respectively, and the Akt activities were measured by Akt kinase assay kit.
Results: Compared with their corresponding parent cell line, the IC50 values to 5-FU in HT-29/5-FU, LoVo/5-FU and SW480/5-FU were all significantly increased (all P<0.05), with the drug resistance indexes of 7.213, 5.849 and 15.940, respectively. The numbers of cells in G0/G1 phase in both drug-resistant cell lines and their parent cell lines were increased with the increase of the treatment concentration of 5-FU (all P<0.05), but the number of cells in G0/G1 phase in each drug-resistant cell line was significantly less than that in their respective corresponding parent cell line under the same concentration of 5-FU (all P<0.05). Compared with their corresponding parent cell line, the mRNA and protein expressions of P-gp, MRP1, ABCG2 and Akt were significantly increased and the mRNA and protein expressions of PTEN were significantly decreased in all the drug-resistant cell lines (all P<0.05), and the Akt activities were significantly enhanced (P<0.05).
Conclusion: The 5-FU resistant colon cancer cell lines are successfully established. The mechanism for drug-resistance may probably be associated with the activation of PI3K/Akt pathway resulted from PTEN down-regulation.