ABSTRACT
The present study demonstrated the anti-tumor effects of the quinoline derivative [5-(3-chloro-oxo-4-phenyl-cyclobutyl)-quinoli-8-yl-oxy] acetic acid hydrazide (CQAH) against colorectal carcinoma. Substantial apoptotic effects of CQAH on HCT116 and LoVo human colon cancer cell lines were observed. Apoptosis was identified based on cell morphological characteristics, including cell shrinkage and chromatin condensation as well as Annexin V/propidium iodide double staining followed by flow cytometric analysis and detection of apoptosis-associated proteins by western blot analysis. CQAH induced caspase-3 and PARP cleavage, reduced the expression of the anti-apoptotic proteins myeloid cell leukemia-1 and B-cell lymphoma (Bcl) extra large protein and elevated the expression of the pro-apoptotic protein Bcl-2 homologous antagonist killer. In addition, pharmacological inhibition of c-Jun N-terminal kinase (JNK), but not extracellular signal-regulated kinase or p38, significantly reduced CQAH-mediated cell death as well as cleavage of caspase-3 and PARP. Co-treatment of CQAH with the commercial chemotherapeutics 5-fluorouracil and camptothecin-11 significantly improved their efficacies. Comparison of the apoptotic effects of CQAH with those of two illustrated structure-activity associations for this compound type, indicating that substitution at position-4 of the azetidine phenyl ring is pivotal for inducing apoptosis. In conclusion, the results of the present study indicated CQAH and its analogues are potent candidate drugs for the treatment of colon carcinoma.
Subject(s)
Antineoplastic Agents/pharmacology , Apoptosis/drug effects , Quinolones/pharmacology , Antineoplastic Agents/chemistry , Camptothecin/analogs & derivatives , Camptothecin/pharmacology , Caspase 3/metabolism , Cell Line, Tumor , Colonic Neoplasms/pathology , Extracellular Signal-Regulated MAP Kinases/metabolism , Fluorouracil/pharmacology , HCT116 Cells , Humans , Irinotecan , JNK Mitogen-Activated Protein Kinases/metabolism , Myeloid Cell Leukemia Sequence 1 Protein/metabolism , Poly(ADP-ribose) Polymerases/metabolism , Proto-Oncogene Proteins c-bcl-2/metabolism , Quinolones/chemistry , p38 Mitogen-Activated Protein Kinases/metabolismABSTRACT
The present study aimed to investigate the effects of cinobufacini injection on the proliferation and expression of topoisomerases in human HepG-2 hepatocarcinoma cells. The cells were divided into a control group and an experimental group, in which 0.105, 0.21, 0.42 mg/l cinobufacini was injected. Cell proliferation was assessed using a 3-(4,5-dimethylthiazol-2-yl) -2,5-diphenyltetrazolium bromide assay, levels of apoptosis were detected using annexin V/propidium iodide staining and cell cycles were analyzed using flow cytometric analysis. The mRNA and protein expression levels of topoisomerase (TOPO) I and TOPO II were determined by reverse transcription-quantitative polymerase chain reaction and western blotting, respectively. Cinobufacini injection significantly inhibited the proliferation of the HepG-2 cells (P<0.05), induced apoptosis (P<0.05) in a dose- and time-dependent manner, induced tumor cell arrest at the S phase in a dose-dependent manner, and downregulated the mRNA and protein expression levels of TOPO I and TOPO II (P<0.05) in a dose-dependent manner. Therefore, cinobufacini was found to inhibit human HepG-2 hepatocellular carcinoma cell proliferation, and downregulation of the expression levels of TOPO I and TOPO II may contribute to the effect on proliferation observed in the HepG2 cells following cinobbufacini injection.
Subject(s)
Carcinoma, Hepatocellular/genetics , DNA Topoisomerases, Type II/biosynthesis , DNA Topoisomerases, Type I/biosynthesis , Liver Neoplasms/genetics , Amphibian Venoms/administration & dosage , Carcinoma, Hepatocellular/drug therapy , Carcinoma, Hepatocellular/pathology , Cell Proliferation/drug effects , DNA Topoisomerases, Type I/genetics , DNA Topoisomerases, Type II/genetics , Dose-Response Relationship, Drug , Enzyme Inhibitors/administration & dosage , Gene Expression Regulation, Neoplastic/drug effects , Hep G2 Cells , Humans , Liver Neoplasms/drug therapy , Liver Neoplasms/pathology , RNA, Messenger/biosynthesisABSTRACT
AIM: To identify genes differentially expressed in mouse hepatocarcinoma ascites cell line with low potential of lymphogenous metastasis. METHODS: A subtracted cDNA library of mouse hepatocarcinoma cell line with low potential of lympho-genous metastasis Hca-P and its synogenetic cell line Hca-F with high metastatic potential was constructed by suppression subtracted hybridization (SSH) method. The screened clones of the subtracted library were sequenced and GenBank homology search was performed. RESULTS: Fifteen differentially expressed cDNA fragments of Hca-P were obtained which revealed 8 known genes, 4 expressed sequence tags (ESTs) and 3 cDNAs showed no homology. CONCLUSION: Tumor metastasis is an incident involving multiple genes. SSH is a useful technique to detect differentially expressed genes and an effective method to clone novel genes.
