Study On The Effect Of Thioridazine On Human Colon Cancer Stem Cells

It is known that CSCs constitute only a small percentage of tumor cells within atumolor mass containing heterogeneous population of tumolor cells within thetumolor microenvironment. CSCs are long-lived and resistant to chemo-radiationtherapy.They are reluctant to locate and give rise to uncontrolled amplification ofdifferentiated cell populations and contribute to tumor recurrence.Therefore, thedevelopment of strategies for targeted eradication of CSCs represent an important anddifficult point in cancer research.Colon cancer is the second leading cause of cancer related death in westerncountries, and the mortality rate for this cancer has increasd in our country recently. Itappears that tumor recurrence/relapse occurs although with good diagnosis, andbecomes resistant to chemotherapeutic agents. It is not easy to locate the lesion sitebecause of the slur of early symptom, the development of effective methods forcolon cancer is urgently needed. The finding of CSC might be a therapeutic target inthe treatment of colon cancer.Thioridazine was originally well known as an antipsychotic drug, has been reportedto induce apoptosis in various types of cancer cells, as a dopamine receptor antagonist,exerts its anti-leukemia CSC activity via antagonism of D2-family DRs differentiallyexpressed on neoplastic SCs.DR signaling. TRDZ is currently considered to helprelieve pain and stress and improve the quality of life of patients.According to the literature, thioridazine targets leukemia stem cells and gastriccancer stem cells and breast cancer stem cells and so on to inhibit tumor cellproliferation、induce apoptosis and exert the function of anti-tumor, however, fewresearches for colon cancer stem cells.In this study, we artificially separated EpCAM+CD44+sub-populations fromHCT116cell lines by immunomagnetic beads technique, and identified by induceddifferentiation, sphere formation, immunohistochemical expression of the markergene and so on. We then use cell proliferation assays(CCK8) and microscopicobservation to evaluate the ability of TRDZ to inhibit the growth of HCT116 cell-derived tumors. Apoptosis induction was estimated using AO/EB staining, JC-1staining, Annexin-V-FITC stainig and FACS analysis and so on. Results show thatEpCAM+CD44+HCT116single cell volumole which were clutrued in serumol-freemediumol formed tumor microspheres gradually, when cultured in DMEM/F12mediumol with10%fetal bovine serumol, the suspended cells gradually adherentgrowth; TRDZ inhibited the growth of the HCT116SC in a dose-dependent manner.When in the concentration of1μmol/L, there was a slight promotion in proliferation.Moreover, TRDZ inhibited colony formation abilitiy in a dose dependent; AO/EBresults indicate the higher concentration of thioridazine, the greater degree ofapoptosis; The result of Annexin-V-FITC stainig indicates the proportion of earlyapoptosis cells increased with the concentration of thioridazine; The mitochondrialmembrane potential decreased, and the higher concentration of thioridazine, thegreater percentage of apoptotic cells. Q-PCR detection results suggestanti-apoptosis mRNA bcl-2were high expressed in the normal group, andthe expression quantity during apoptosis in thioridazine-treated group showed adose-dependent decrease contrast to the expression of pro-apoptotic protein bax; Theexpression of cleaved caspase-3changed following the treatment of thioridazine.Collectively, thioridazine inhibit tumor cell proliferation, induce apoptosis; Ourfindings lays the experimental basis for developing new drugs targeting cancer stemcells and provides the foundation experiment for colon cancer treatment.