Differential Proteomic Analysis Of Uveal Melanoma Serologically Peptidome

AIM: Based on proteomic technology,to compare and analyze the serum peptidome profiling between uveal melanoma(UM)patients and healthy controls(C),between Pre-surgery(PS),1mo Post-operation(PO1)and 6mo Post-operation(PO6)of UM patients,and to establish a UM diagnosis model that has better sensitivity,specificity and grouping accuracy.METHODS: 1.8 patients with UM(age 21-55 y,mean 35.7y)and 10 healthy controls(age 24-54 y,mean 34.2y)were enrolled in Tongren Eye Center of Beijing Tongren Hospital.Venous blood samples were drawn.Randomly selected samples of PS2 and PO1-2 were analyzed with MB-hydrophobic interaction chromatography(MB-HIC C8),MB-immobilized metal ion affinity chromatography containing copper ions(MB-IMAC-Cu)and MB-weak-cation-exchange chromatography(MB-WCX).Matrix-assisted laser desorption/ionization time-of-flight(MALDI-TOF)mass spectrometery and Clin Prot Tools(CPT)bioinformatics software were used.2.18 patients with UM(age 21-68 y,mean 39.6y)were enrolled.Venous blood samples of PS,PO1 and PO6 were drawn and were all analyzed with MB-IMAC-Cu.3.Randomly selected samples of C3 and PO6-4 were analyzed to evaluate the within-run reproducibility.4.Andomly selected samples of PO1-9 were analyzed to evaluate the between-run reproducibility.5.Compared the serum peptidome profiling between PS and C to screen for differentially expressed peptides/proteins.6.Compared the serum peptidome profiling between PS,PO1 and PO6 of UM to screen for differentially expressed peptides/proteins.7.Uveal melanoma serum diagnostic model were built up,then verify the validity of the established model using several algorithms.RESULTS: 1.Most of the protein peaks were no more than 10 k Da.MB-IMAC Cu beads managed to capture the largest number of peptide peaks and highest peak amplitudes compared with the other two beads.2.The coefficients of variance(CVs)of PO6-4 serum samples were around 8.7%-30.9% with the mean coefficient of variance(MCV)15.2% while the CVs of C3 serum samples were around 4.5%-28.2% with MCV 18.1%,showing satisfactory within-run reproducibility.3.The CVs of PO1-9 serum samples were around 10.9%-31.8% with MCV 19.1%,showing satisfactory between-run reproducibility.4.Compared the serum peptidome profiling between PS and C,47 peptides were differentially expressed,of which 13 peptides were potential UM markers.Based on combined use of potential markers,peptides provide high sensitivity,specificity and accuracy rate to differentiate melanoma patients from healthy controls.5.Mean intensities(MIs)in PS and PO1 showed no statistical difference while significant differences were showed in PS and PO6 as well as in PO1 and PO6(P values <0.05).6.Potential markers were determined in UM patients.The sequence of markers were determined to be G.EGDFLAEGGGVR.G and G.EGDFLAEGGGVR.G and A.DSGEGDFLAEGGGVR.G.CONCLUSION: MB-IMAC Cu beads were used in this study because of its high-tech method in proteomics analysis.We utilized proteomic technique in UM patients and established a UM diagnosis model that had satisfactory sensitivity,specificity and grouping accuracy.Further researches are still necessary to explore its mechanism and significance.The combination of affinity bead separation and MALDI-TOF mass spectrometery shows a promising broad application prospect.