Abstract: 【Abstract】ObjectiveTo construct a colon cancer chemotherapyresistant cell line COLO, and study its characteristics and its relationship with tumor stem cells. MethodsWe constructed two 5fluorouraci (5FU)resistant colon cancer cell line COLO/5FU1 and COLO/5FU2, which were resistant to 0.10 μmol/ml and 0.20 μmol/ml 5FU respectively through gradiently increased drug concentration. The characteristics of 5FUresistant cell lines were compared with parental colon cancer cell line COLO related to proliferation, colony forming ability, migration and invasion, sphere forming ability, expression of stemness genes and cross drugresistance. ResultsIn the cell viability assay, 4 days after regular training, the absorbancy of colon cancer 5FUresistant cell lines COLO/5FU2, COLO/5FU1 and parental colon cancer cell line COLO were 0.61±0.13, 0.54±0.07 and 0.41±0.09 respectively, with significant difference (F=63.43, P=0.033). With the increased concentration of 5FU, 5FUresistant cell lines presented increasing clonality. The cloning efficiency of COLO/5FU2, COLO/5FU1 and parental colon cancer cell line COLO were (87.6±12.7)%, (65.3±9.7)% and (38.5±7.6)% respectively, with significant difference (F=33.64, P=0.017). In each high power field of vision, the cell numbers of migration through the basement membrane of COLO/5FU2, COLO/5FU1 and parental colon cancer cell line COLO were 482±39, 434±45 and 373±38 respectively; and the cell numbers of invasion through the basement membrane were 174±42, 112±31 and 87±29 respectively, with significant differences (F=109.61, P=0.009; F=67.31, P=0.032). Compared with parental colon cancer cell line COLO, 5FUresistant cell lines had higher expression of stemness genes (F=47.31, P=0.042). 5FUresistant cell lines were crossresistant to other chemotherapeutic drugs such as mitoxantrone. For example, after incubation for 96 hours, inhibition rate of mitoxantrone to parent colon cancer cell line COLO was higher significantly than COLO/5FU1 and COLO/5FU2 (0.749±0.042, 0.423±0.024, 0.342±0.018), with significant difference (F=12.61, P=0.028). The microsphere forming rates of COLO/5FU2, COLO/5FU1 and parental colon cancer cell line COLO were (8.90±0.97)%, (6.20±0.75)% and (3.90±0.32)% respectively, with significant difference (F=164.32, P=0.006). ConclusionColon cancer drugresistant cell line COLO possess tumor stem celllike characteristics, which are enriched in cancer stem cells.

Key words: Colonic neoplasms, Drug resistance, neoplasms, Neoplastic stem cells, Drugresistant cell line