The synthesized 2-(2-fluorophenyl)-6,7-methylenedioxyquinolin-4-one (CHM-1) promoted G2/M arrest through inhibition of CDK1 and induced apoptosis through the mitochondrial-dependent pathway in CT-26 murine colorectal adenocarcinoma cells.
BACKGROUND
In this study, we investigated the effects of 2-(2-fluorophenyl)-6,7-methylenedioxyquinolin-4-one (CHM-1) on cell viability, cell cycle arrest and apoptosis in CT-26 murine colorectal adenocarcinoma cells.


METHODS
For determining cell viability, the MTT assay was used. CHM-1 promoted G2/M arrest by PI staining and flow cytometric analysis. Apoptotic cells were evaluated by DAPI staining. We used CDK1 kinase assay, Western blot analysis and caspase activity assays for examining the CDK1 activity and proteins correlated with apoptosis and cell cycle arrest. The in vivo anti-tumor effects of CHM-1-P were evaluated in BALB/c mice inoculated with CT-26 cells orthotopic model.


RESULTS
CHM-1 induced CT-26 cell viability inhibition and morphologic changes in a dose-dependent and time-dependent manner and the approximate IC50 was 742.36 nM. CHM-1 induced significant G2/M arrest and apoptosis in CT-26 cells. CHM-1 inhibited the CDK1 activity and decreased CDK1, Cyclin A, Cyclin B protein levels. CHM-1 induced apoptosis in CT-26 cells and promoted increasing of cytosolic cytochrome c, AIF, Bax, BAD, cleavage of pro-caspase-9, and -3. The significant reduction of caspase-9 and -3 activity and increasing the viable CT-26 cells after pretreated with caspase-9 and -3 inhibitor indicated that CHM-1-induced apoptosis was mainly mediated a mitochondria-dependent pathway. CHM-1-P improved mice survival rate, and enlargement of the spleen and liver metastasis were significantly reduced in groups treated with either 10 mg/kg and 30 mg/kg of CHM-1-P and 5-FU in comparison to these of CT-26/BALB/c mice.


CONCLUSIONS
Taken together, CHM-1 acted against colorectal adenocarcinoma cells in vitro via G2/M arrest and apoptosis, and CHM-1-P inhibited tumor growth in vivo.
Chou
Li-Chen
LC
Graduate Institute of Pharmaceutical Chemistry, College of Pharmacy, China Medical University, No 91, Hsueh-Shih Road, Taichung 404, Taiwan.
Yang
Jai-Sing
JS
Huang
Li-Jiau
LJ
Wu
Hsi-Chin
HC
Lu
Chi-Cheng
CC
Chiang
Jo-Hua
JH
Chen
Kuan-Tin
KT
Kuo
Sheng-Chu
SC
Chung
Jing-Gung
JG
D000230:Adenocarcinoma / Q000188:drug therapy* / Q000378:metabolism / Q000473:pathology; D000818:Animals; D000970:Antineoplastic Agents / Q000494:pharmacology* / Q000627:therapeutic use; D017209:Apoptosis / Q000187:drug effects*; D016203:CDC2 Protein Kinase / Q000037:antagonists & inhibitors*; D053148:Caspase 3 / Q000378:metabolism; D053453:Caspase 9 / Q000378:metabolism; D002453:Cell Cycle / Q000187:drug effects*; D002455:Cell Division / Q000187:drug effects; D045744:Cell Line, Tumor; D049109:Cell Proliferation / Q000187:drug effects; D002470:Cell Survival / Q000187:drug effects*; D015179:Colorectal Neoplasms / Q000188:drug therapy* / Q000378:metabolism / Q000473:pathology; D019925:Cyclin A / Q000187:drug effects / Q000378:metabolism; D019926:Cyclin B / Q000187:drug effects / Q000378:metabolism; D004149:Dioxoles / Q000494:pharmacology* / Q000627:therapeutic use; D004305:Dose-Response Relationship, Drug; D005434:Flow Cytometry; D016195:G2 Phase / Q000187:drug effects; D006801:Humans; D008113:Liver Neoplasms / Q000556:secondary; D008540:Meiosis; D051379:Mice; D008807:Mice, Inbred BALB C; D008928:Mitochondria / Q000187:drug effects / Q000378:metabolism; D015363:Quinolones / Q000494:pharmacology* / Q000627:therapeutic use; D023041:Xenograft Model Antitumor Assays
C525063:2'-fluoro-6,7-methylenedioxy-2-phenyl-4-quinolone; D000970:Antineoplastic Agents; D019925:Cyclin A; D019926:Cyclin B; D004149:Dioxoles; D015363:Quinolones; D016203:CDC2 Protein Kinase; D053148:Caspase 3; D053453:Caspase 9
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