Proteome Studies And Biomarker Discovery Associated With Hepatocellular Carcinoma Metastasis

Proteome studies and biomarker discovery associated with hepatocellular carcinoma metastasis Shi-Jian Ding (Biochemistry and Molecular Biology) Directed by Professor Qi-Chang XiaAbstract Hepatocellular carcinoma (HCC) is one of the most common andaggressive malignant tumors with especially high prevalence in China, the extremelypoor prognosis of HCC is largely the result of a high rate of recurrence andmetastasis. To better understand the mechanism underlying the HCC metastasis andto search potential markers for HCC prognosis, differential proteome analysis on twoHCC cell strains with high and low metastatic potentials, MHCC97-H andMHCC97-L, was conducted using various proteomics approaches. Bytwo-dimensional gel electrophoresis (2-DE), image analysis and mass spectrometry,the expression levels of 60 protein spots were found quantitatively or qualitativelychanged between MHCC97-H and MHCC97-L and among them 14 protein spotswere further identified using in-gel tryptic digestion, peptide mass fingerprinting andtandem mass spectrometry. During this study, a 2-DE database was also constructedand is available at http://www.proteomics.ac.cn/2D/. At present the database contains68 identified protein spots including both differentially-expressed and housekeepingproteins. To provide comprehensive and complementary proteome coverage, twotypes of isotope-coded affinity tags (ICAT), conventional H-ICAT and new 2cleavable C-ICAT, were employed to quantitative analysis of the protein expression 13between MHCC97-H and MHCC97-L cells. Database search using SEQUEST andXPRESS showed that 123 proteins were unambiguously identified and theexpressions of 43 proteins were showed more than 2 fold changes in these two cellstrains. Some low abundant proteins and high molecular weight proteins were alsoobserved which were poorly tractable by 2-DE analysis. These results provide thebasis for searching potential markers for HCC prognosis and give some clues toelucidate the mechanism of HCC metastasis. To further explore the clinicalsignificance of the proteomic finding, cytokeratin 19 was screened from comparativeproteomic analysis of MHCC97-H and MHCC97-L cells and was found to correlatewith HCC metastasis in more progressively high metastatic HCC cells, HCC tissuebiopsies and HCC metastasis nude mice model using Western blot, IIAbstractimmunohistochemistry and ELISA analysis. Cytokeratin 19 may be a useful markerfor predicting tumor metastasis and a therapeutic target for the treatment of HCCpatients with metastases.