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John W Crabb, Bo Hu, Jack S Crabb, Arun D Singh; Protein Candidate Biomarkers of Uveal Melanoma Metastasis. Invest. Ophthalmol. Vis. Sci. 2015;56(7 ):5339.
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Proteomic analysis of metastasized and non-metastasized uveal melanoma primary tumors was pursued for insights into mechanisms and biomarkers of uveal melanoma metastasis.
Fifteen uveal melanoma specimens collected from enucleated eyes at the Cole Eye Institute, Cleveland Clinic were subjected to global quantitative proteomic analysis using LC MS/MS iTRAQ technology. Bruch’s membrane choroid complex protein extracts from nine normal human postmortem eyes were pooled and utilized as a reference control specimen. Tumor and control protein was extracted in SDS and digested with trypsin. Peptides were labeled with iTRAQ tags, fractionated by cation exchange chromatography, and analyzed by LC MS/MS. Protein identification utilized the Mascot search engine and the human Uni-Prot/Swiss-Protein database with the false discovery rate £ 1%. iTRAQ tags were quantified by the weighted average method (Matrix Science, 2.4.1). Differentially expressed proteins exhibited group differences with p £ 0.05 (t-test) in a sample set composed of five metastasized and five non-metastasized ocular tumors. Logistic regression modeling and principal component analysis was used to test the quantitative capability of differentially expressed proteins to classify the metastatic status of five independent ocular tumor specimens
Over 1700 proteins were quantified with two or more peptides from 15 uveal melanoma primary tumors. Thirty-six differentially expressed proteins were identified from quantitative comparison of proteins in 5 metastatic and 5 non-metastatic ocular tumors. Among differentially expressed proteins, the levels of collagen alpha-3 (VI) and heat shock protein beta-1 correctly classified metastasis in five independent tumor samples. The metastatic status of one independent tumor specimen was also classified correctly by 13 other differentially expressed proteins and by principal component analysis.
While additional uveal melanoma specimens must be studied to confirm the identification of differentially expressed proteins and to establish their discriminatory capabilities between metastatic and non-metastatic tumors, the current results suggest collagen alpha-3 (VI) and heat shock protein beta-1 as candidate biomarkers of metastasis.
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