Study On The Drug Resistance Of Cancer Stem Cells Identified From Colorectal Cancer Cells

Colon cancer is a common malignant tumor of the digestive tract,whose incidence rate is the third of the malignant tumor of the digestive tract in our country ,inferior to the stomach cancer and the esophageal cancer. The chemotherapy is one of the main therapies of colon cancer,but the emergence of the multidrug resistance in tumor cells during the chemotherapy allows patients to a variety of chemotherapeutic agents tolerance, eventually resulting in chemotherapy failure.The emergence of multidrug resistance is a main reason that influnces the chemotherapy effect of colon cancer.Recently,with the thorough research of malignant tumor, scientists have proposed a new theory—cancer stem cell hypothesis.The cancer stem cell hypothesis considers: cancer stem cells(CSCs) are a small population in tumor tissues that have stem cell characteristics,these cells have the capacity of self-renewal and nondirectional differentiation potential.Cancer stem cells are considered to be the points of origin of tumor formation and proliferation and the basis for the emergence of multidrug resistance,tumor formation, growth and metastasis.So far cancer stem cells have been idetified from a variety of tumor tissues,such as leukemia,brain tumors, breast cancer,lung cancer and prostatic carcinoma,which gives further proof to the cancer stem cell hypothesis.The proposing of cancer stem cell hypothesis provides new way for the research on the mechanism of tumor formation,growth,and studies based on the isolation of cancer stem cells bring new hope for the improving of cancer therapies.In this study,colorectal cancer stem cells with the phaenotype of EpCAM~+CD44~+ will be identified from human colorectal cancer cell lines Colo205 and HCT-116 and the chemotherapeutic agents sensitivity be detected,so as to give some experimental basis of the possibility of colorectal cancer stem cells to be a therapeutic target for the treatment of colorectal cancer.Methods: (1)Isolation of colorectal cancer stem cells:Use inderect magnetic activated cell sorting to isolate colorectal cancer stem cells with the phaenotype of EpCAM~+CD44~+ from human colorectal cancer cell lines Colo205 and HCT-116 in logarithmic growth;(2)Identification of colorectal cancer stem cells:Use cell proliferation assay, cell differentiation assay,soft agar colony-formation assay and tumorigenicity assay in NOD/SCID mice to identify the EpCAM~+CD44~+ colorectal cancer stem cells isolated by magnetic activated cell sorting; (3) Chemotherapeutics sensitivity texting:Use MTT assay with the commonly used three chemotherapy agents mitomycin,diamminedichloroplatinum and 5-fluorouracil for colorectal cancer treatment to compare the sensitivity of chemotherapeutic agents in EpCAM~+CD44~+ colorectal cancer stem cells with colorectal cancer cells.Results: (1)Colorectal cancer stem cells with the phaenotype of EpCAM~+CD44~+ from human colorectal cancer cell lines Colo205 and HCT-116 have been isolated,and the EpCAM~+CD44~+ colorectal cancer stem cells can survive and grow in serum free medium and form cancer stem cell spheres in suspension;(2)The proliferation speed of EpCAM~+CD44~+ colorectal cancer stem cells in vitro is slower than that of colon cancer cells;(3)The cancer stem cell spheres formed in serum free medium can turn into adherent monolayer cancer cells in serum-containing medium;(4)The colony-formation ability in soft agar of EpCAM~+CD44~+ colorectal cancer stem cells is higher than that of colon cancer cells;(5)The tumorigenicity ability in NOD/SCID mouse of EpCAM~+CD44~+ colorectal cancer stem cells is higher than that of colon cancer cells;(6)The sensitivity with mitomycin,diamminedichloro-platinum and 5-fluorouracil in EpCAM~+CD44~+ colorectal cancer stem cells is lower than that of colon cancer cells.Conclusions:(1)The existence of colorectal cancer stem cells with the phaenotype of EpCAM~+CD44~+ in human colorectal cancer cell lines Colo205 and HCT-116;(2)The EpCAM~+CD44~+ colorectal cancer stem cells possess drug resistance to mitomycin,diamminedichloroplatinum and 5-fluorouracil.