title: Polo-like kinase 1, a new therapeutic target in hepatocellular carcinoma.
abstract: OBJECTIVE
  To investigate the role of polo-like kinase 1 (PLK1) as a therapeutic target for hepatocellular carcinoma (HCC).
  
  
  METHODS
  PLK1 gene expression was evaluated in HCC tissue and HCC cell lines. Gene knockdown with short-interfering RNA (siRNA) was used to study PLK1 gene and protein expression using real-time reverse transcription polymerase chain reaction (RT-PCR) and Western blotting, and cell proliferation using 3-(4,5-dimethylthiazol-2-yl)-5-(3-carboxymethoxyphenyl)-2(4-sulfophenyl)-2H-tetrazolium (MTS) and bromodeoxyuridine (BrdU) assays. Apoptosis was evaluated using the terminal deoxynucleotidyl transferase dUTP nick end labeling (TUNEL) assay, and caspase-inhibition assay. Huh-7 cells were transplanted into nude mice and co-cultured with PLK1 siRNA or control siRNA, and tumor progression was compared with controls.
  
  
  RESULTS
  RT-PCR showed that PLK1 was overexpressed 12-fold in tumor samples compared with controls, and also was overexpressed in Huh-7 cells. siRNA against PLK1 showed a reduction in PLK1 gene and protein expression of up to 96% in Huh-7 cells, and a reduction in cell proliferation by 68% and 92% in MTS and BrdU cell proliferation assays, respectively. There was a 3-fold increase in apoptosis events, and TUNEL staining and caspase-3 assays suggested that this was caspase-independent. The pan-caspase inhibitor Z-VAD-FMK was unable to rescue the apoptotic cells. Immnofluorescence co-localized endonuclease-G to fragmented chromosomes, implicating it in apoptosis. Huh-7 cells transplanted subcutaneously into nude mice showed tumor regression in siPLK1-treated mice, but not in controls.
  
  
  CONCLUSIONS
  Knockdown of PLK1 overexpression in HCC was shown to be a potential therapeutic target, leading to apoptosis through the endonuclease-G pathway.
authors: Institute of Molecular and Cell Biology, Agency for Science, Technology and Research, Singapore 138673, Singapore.
authors: Wei Chuen
authors: WC
authors: Mok
authors: Shanthi
authors: S
authors: Wasser
authors: Theresa
authors: T
authors: Tan
authors: Seng Gee
authors: SG
authors: Lim
mesh_terms: D000328:Adult; D000368:Aged; D000369:Aged, 80 and over; D000590:Amino Acid Chloromethyl Ketones / Q000494:pharmacology; D000818:Animals; D017209:Apoptosis; D015153:Blotting, Western; D006528:Carcinoma, Hepatocellular / Q000201:enzymology* / Q000235:genetics / Q000473:pathology / Q000628:therapy*; D053148:Caspase 3 / Q000378:metabolism; D061945:Caspase Inhibitors / Q000494:pharmacology; D018797:Cell Cycle Proteins / Q000235:genetics / Q000378:metabolism*; D049109:Cell Proliferation; D004706:Endodeoxyribonucleases / Q000378:metabolism; D005260:Female; D005455:Fluorescent Antibody Technique; D055785:Gene Knockdown Techniques; D015316:Genetic Therapy; D056945:Hep G2 Cells; D006801:Humans; D008113:Liver Neoplasms / Q000201:enzymology* / Q000235:genetics / Q000473:pathology / Q000628:therapy*; D008297:Male; D051379:Mice; D008819:Mice, Nude; D008875:Middle Aged; D017346:Protein Serine-Threonine Kinases / Q000235:genetics / Q000378:metabolism*; D011518:Proto-Oncogene Proteins / Q000235:genetics / Q000378:metabolism*; D034622:RNA Interference; D060888:Real-Time Polymerase Chain Reaction; D020133:Reverse Transcriptase Polymerase Chain Reaction; D014162:Transfection; D047368:Tumor Burden; D023041:Xenograft Model Antitumor Assays; D000097869:Polo-Like Kinase 1
chemical_list: D000590:Amino Acid Chloromethyl Ketones; D053148:Caspase 3; D061945:Caspase Inhibitors; D018797:Cell Cycle Proteins; D004706:Endodeoxyribonucleases; D017346:Protein Serine-Threonine Kinases; D011518:Proto-Oncogene Proteins; D000097869:Polo-Like Kinase 1; C096713:benzyloxycarbonylvalyl-alanyl-aspartyl fluoromethyl ketone; C052391:endonuclease G; C505407:CASP3 protein, human