A small-molecule metastasis inhibitor, norcantharidin, downregulates matrix metalloproteinase-9 expression by inhibiting Sp1 transcriptional activity in colorectal cancer cells.
Norcantharidin (NCTD) is a small-molecule metastasis inhibitor without renal toxicity derived from a renal toxic compound cantharidin, which is found in blister beetles (Mylabris phalerata Pall.), commonly used in traditional Chinese medicine. The anti-metastatic capacity of NCTD is apparently through the downexpression of matrix metalloproteinase-9 (MMP-9) activity. The aim of this study was to clarify the transcriptional regulation of MMP-9 gene by NCTD in colorectal cancer CT-26 cells. NCTD not only downregulated MMP-9 mRNA and protein expression, but also inhibited gelatinase activity in a concentration- and time-dependent manner. In CT26 cells with transfection of cis-element reporter plasmids, NCTD treatment decreased reporter luciferase activity from a Sp1 construct, augmented with a NF-kappaB construct, but this did not occur with an AP-1 construct. Further transfecting with constructs containing wild-type or various mutant MMP-9 promoters in CT26 cells indicated that Sp1, but not the others, was required for NCTD-inhibition of MMP-9 promoter transactivation. More evidence by electrophoretic mobility shift assay demonstrated that NCTD inhibited the DNA-binding activity of Sp1. In addition, the increase effect of NF-kappaB-luciferase activity by NCTD may include the upexpression of nuclear STAT1 and result in competitive suppression of NF-kappaB-binding activity in MMP-9 promoter. In conclusion, the metastasis inhibitor NCTD downregulates MMP-9 expression by inhibiting Sp1 transcriptional activity in colorectal cancer CT26 cells.
Chen
Yu-Jen
YJ
Department of Radiation Oncology, Mackay Memorial Hospital, Taipei 104, Taiwan.
Chang
Wei-Min
WM
Liu
Yi-Wen
YW
Lee
Chia-Yun
CY
Jang
Yi-Hua
YH
Kuo
Cheng-Deng
CD
Liao
Hui-Fen
HF
D001483:Base Sequence; D015153:Blotting, Western; D019086:Bridged Bicyclo Compounds, Heterocyclic / Q000494:pharmacology*; D045744:Cell Line, Tumor; D015179:Colorectal Neoplasms / Q000201:enzymology* / Q000473:pathology; D017931:DNA Primers; D015536:Down-Regulation / Q000187:drug effects*; D024202:Electrophoretic Mobility Shift Assay; D006801:Humans; D020780:Matrix Metalloproteinase 9 / Q000378:metabolism*; D009362:Neoplasm Metastasis / Q000517:prevention & control*; D020133:Reverse Transcriptase Polymerase Chain Reaction; D016329:Sp1 Transcription Factor / Q000037:antagonists & inhibitors*; D014158:Transcription, Genetic / Q000187:drug effects*
D019086:Bridged Bicyclo Compounds, Heterocyclic; D017931:DNA Primers; D016329:Sp1 Transcription Factor; C069741:norcantharidin; D020780:Matrix Metalloproteinase 9