Validation of prometastatic proteins in low-grade breast cancer by targeted mass spectrometry approach based on selected reaction monitoring
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| Year of publication | 2012 |
| Type | Conference abstract |
| MU Faculty or unit | |
| Citation | |
| Description | Protein biomarkers, including those newly revealed for breast cancer metastasis, are expected to provide new improvements in prognosis, prediction, treatment and cancer progression monitoring. Untargeted proteomics on the basis of iTRAQ, TMT, superSILAC labelling or label-free quantification is able to identify potential protein targets by screening high number of proteins in rather limited sample sets. In a long process leading towards real clinical application, it is however absolutely essential to obtain statistically convincing data on biomarker selectivity and sensitivity by analyzing large sample cohorts. Besides well established immunochemical methods (imunohistochemistry or ELISA), targeted mass spectrometry approach based on selected reaction monitoring (SRM) can offer high specificity, sensitivity, accuracy and broad linear dynamic range of protein quantification in large sets of samples. Here, principles and features of this rapidly evolving approach based on monitoring of selected ion pairs originating from peptides (precursor ions and fragmentation product ions) in a triple quadrupole or qTOF mass spectrometer will be explained. Application of several available approaches for relative and absolute quantification in SRM mode will be discussed. We used SRM approach utilizing mTRAQ chemical tags for verification of perspective biomarkers identified in a large iTRAQ study focused on discovery of prometastatic targets in low grade breast cancer. 96 tissue lysates were measured in analytical triplicates using SRM method, targeting selected set of proteins. The preliminary results indicating similar trends as found in the untargeted proteomic study will be presented. Furthermore, additional data from immuno-SRM method development for determination of p53 mutations will be shown. This work was supported by the Czech Science Foundation No. P304/10/0868 and by the European Regional Development Fund and the State Budget of the Czech Republic (RECAMO CZ.1.05./2.1.00/03.0101). |
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