Polo-like kinase 1,a new therapeutic target in hepatocellular carcinoma

AIM： 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-dim ethylthiazol-2-yl）-5-（3-carboxymethoxyphenyl）-2（4-sulf ophenyl）-2H-tetrazolium （MTS） and bromodeoxyuridine （BrdU） assays. Apoptosis was evaluated using the terminal deoxynucleotidyl transferase dUTP nick end label- ing （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 progres- sion was compared with controls. RESULTS： RT-PCR showed that PLK1 was overexpre- ssed 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 siPLKl-treated mice, but not in controls. CONCLUSION： Knockdown of PLK1 overexpression in HCC was shown to be a potential therapeutic target, leading to apoptosis through the endonuclease-G path- way.

AIM: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-dim ethylthiazol-2-yl)-5-(3-carboxymethoxyphenyl)-2(4-sulf ophenyl)-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.CONCLUSION:Knockdown of PLK1 overexpression in HCC was shown to be a potential therapeutic target,leading to apoptosis through the endonuclease-G pathway.