Abstract
Renal cell carcinoma accounts for about 3% of adult malignancies and 85% of neoplasms arising from the kidney. To identify potential progression markers for kidney cancer we examined non-neoplastic and neoplastic kidney tissue from three groups of patients, which represent different tumor stages (pT1, pT2, pT3) by a fluorescence two-dimensional difference gel electrophoresis (2D-DIGE) approach combined with MALDI-ToF-MS/MS. Delta2D software package was used for gel image based quantification and statistical analysis. Thereby, a comprehensive Principal Component Analysis (PCA) could be performed and allowed a robust quality control of the experiment as well as a classification of the analyzed samples, which correlated with the predicted stages from the pathological examination. Additionally for selected candidate proteins we detected a correlation to the tumor grading as revealed by immunohistochemistry. On the 2D protein map 176 spots out of 989 were detected as at least 2-fold differentially expressed. These spots were analyzed by MALDI-ToF-MS/MS and 187 different proteins were identified. The functional clustering of the identified proteins revealed ten groups. Within these groups we found 86 enzymes, 63 proteins of unknown function, 14 transporter, 8 peptidases and 7 kinases. From the systems biology approach we could map many of these proteins in major pathways involved in remodelling of cytoskeleton, mitochondrial dysfunctions and changes in lipid metabolism. Due to complexity of the highly interconnected pathway network, further expression and functional validation of these proteins might provide new insights in kidney cancer progression to design novel diagnostic and therapeutic strategies.
Highlights
Renal cell carcinoma is the most common cancer of the kidney and accounts for about 209 000 new cases per year worldwide and 000 deaths [1]
We investigated the stage-related protein alterations in kidney cancer from defined pathological characterized samples using 2D-DIGE [17] and MALDI-ToF-MS/MS for protein identification
We analyzed the proteome of kidney cancer pool specimens from 27 cancer patients and 9 corresponding control tissues (Table 1) by 2D-DIGE in a pI range of 3.0–10.0 and molecular weight range between 10 kDa and approx
Summary
Renal cell carcinoma is the most common cancer of the kidney and accounts for about 209 000 new cases per year worldwide and 000 deaths [1]. Rapid progress has been made to understand tumorigenesis in renal cell carcinomas and other tumor entities, the primary events leading to dysfunction of the control of proliferation remain unclear. Studies of genes involved in cell cycle control and even array analysis studying the global gene expression pattern of neoplastic cells in comparison with non-neoplastic cells led to the identification of differentially expressed genes [2]. Using the rendered gene expression pattern several research groups identified differentially expressed genes, which could be used to gels a total amount of 150 mg protein and for preparative gels a total of 700 mg protein were filled to a volume of 450 ml with rehydration buffer (8 M urea, 2 M thiourea, 2% CHAPS, 15 mM dithiothreitol and 0.5% IPG-buffer pH 3-10 NL [GE Healthcare, Munich, Germany]).
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