1 ) Method development
We develop novel methods and strategies for quantitative proteomics (discovery and targeted) and metabolomics (targeted) with a focus on robustness, precision, and throughput. This includes the system-wide quantification of relative changes of up to 10,000 proteins and phosphorylation sites across different samples, as well as the development of targeted assays for the precise and absolute quantification of protein concentrations or phosphorylation stoichiometries. Using stable isotope-labeled standard (SIS) peptides, we are able to absolutely quantify the endogenous concentrations of 274 proteins in human plasma from less than 10 碌L of blood.
1. The first comprehensive and quantitative analysis of human platelet protein composition allows the comparative analysis of structural and functional pathways. Burkhart, J.M., Vaudel, M., Gambaryan, S., Radau, S., Walter, U., Martens, L., Geiger, J., Sickmann, A., Zahedi, R.P., 2012.听Blood听120, e73-82.
2. Time-resolved characterization of cAMP/PKA-dependent signaling reveals that platelet inhibition is a concerted process involving multiple signaling pathways. Beck, F., Geiger, J., Gambaryan, S., Veit, J., Vaudel, M., Nollau, P., Kohlbacher, O., Martens, L., Walter, U., Sickmann, A., Zahedi, R.P., 2014.听叠濒辞辞诲听123, e1鈥揺10.
3. Multiple reaction monitoring-based, multiplexed, absolute quantitation of 45 proteins in human plasma. Kuzyk, M.A., Smith, D., Yang, J., Cross, T.J., Jackson, A.M., Hardie, D.B., Anderson, N.L., Borchers, C.H., 2009.听Mol. Cell. Proteomics听MCP 8, 1860鈥�1877.
4. Multi-site assessment of the precision and reproducibility of multiple reaction monitoring-based measurements of proteins in plasma. Addona, T.A., Abbatiello, S.E., Schilling, B., Skates, S.J., Mani, D.R., Bunk, D.M., Spiegelman, C.H., Zimmerman, L.J., Ham, A.-J.L., Keshishian, H., Hall, S.C., Allen, S., Blackman, R.K., Borchers, C.H., Buck, C., Cardasis, H.L., Cusack, M.P., Dodder, N.G., Gibson, B.W., Held, J.M., Hiltke, T., Jackson, A., Johansen, E.B., Kinsinger, C.R., Li, J., Mesri, M., Neubert, T.A., Niles, R.K., Pulsipher, T.C., Ransohoff, D., Rodriguez, H., Rudnick, P.A., Smith, D., Tabb, D.L., Tegeler, T.J., Variyath, A.M., Vega-Montoto, L.J., Wahlander, A., Waldemarson, S., Wang, M., Whiteaker, J.R., Zhao, L., Anderson, N.L., Fisher, S.J., Liebler, D.C., Paulovich, A.G., Regnier, F.E., Tempst, P., Carr, S.A., 2009.听Nat. Biotechnol.听27, 633鈥�641.
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2) MultiOMICS for precision medicine
As part of the Lady Davis Institute for Biomedical Research, we conduct fundamental research projects to improve the understanding, diagnosis and treatment of cancer in order to improve precision medicine. We apply multiOMICS approaches that allow the parallel quantification of the (phospho)proteome, the metabolome and the lipidome from minimal sample amounts. We apply quantitative (phospho)proteomics for the system-wide discovery of disease and treatment-specific changes and develop tailored strategies for the improved quantification of specific cancer markers and pathways using mass spectrometry.
1. Simultaneous Metabolite, Protein, Lipid Extraction (SIMPLEX): A Combinatorial Multimolecular Omics Approach for Systems Biology.听Coman, C., Solari, F.A., Hentschel, A., Sickmann, A., Zahedi, R.P., Ahrends, R., 2016听Mol. Cell. Proteomics MCP听15, 1453-1466.
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3) Proteogenomics
In precision medicine, genetic testing has become a main determinant for guiding cancer therapies, such as EGFR mutation status in non-small cell lung cancer. However, the response rates to these guided therapies are often modest. This may be due to discordance between the genome and the actual phenotype, which is reflected by the expression levels of the (mutated) cancer proteins and the activity of relevant signaling pathways. Therefore, we aim to complement genome data with system-wide information on protein expression and protein phosphorylation states from tissue biopsies, thus making it possible to identify causes for unanticipated therapy resistance and improving the selection of appropriate treatments.
1. Drugging the catalytically inactive state of RET kinase in RET-rearranged tumors. Plenker, D., Riedel, M., Br盲gelmann, J., Dammert, M.A., Chauhan, R., Knowles, P.P., Lorenz, C., Keul, M., B眉hrmann, M., Pagel, O., Tischler, V., Scheel, A.H., Sch眉tte, D., Song, Y., Stark, J., Mrugalla, F., Alber, Y., Richters, A., Engel, J., Leenders, F., Heuckmann, J.M., Wolf, J., Diebold, J., Pall, G., Peifer, M., Aerts, M., Gevaert, K., Zahedi, R.P., Buettner, R., Shokat, K.M., McDonald, N.Q., Kast, S.M., Gautschi, O., Thomas, R.K., Sos, M.L., 2017.听Sci. Transl. Med.听9.
2. Alterations of the platelet proteome in type I Glanzmann thrombasthenia caused by different homozygous delG frameshift mutations in ITGA2B. Loroch, S., Trabold, K., Gambaryan, S., Rei脽, C., Schwierczek, K., Fleming, I., Sickmann, A., Behnisch, W., Zieger, B., Zahedi, R.P., Walter, U., Jurk, K., 2017.听Thromb. Haemost. 117, 556鈥�569.
3. Combined Quantification of the Global Proteome, Phosphoproteome, and Proteolytic Cleavage to Characterize Altered Platelet Functions in the Human Scott Syndrome. Solari, F.A., Mattheij, N.J.A., Burkhart, J.M., Swieringa, F., Collins, P.W., Cosemans, J.M.E.M., Sickmann, A., Heemskerk, J.W.M., Zahedi, R.P., 2016, Mol.听Cell. Proteomics MCP听15, 3154鈥�3169.
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Translational Research
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One of our key goals is the translation of targeted MS-based assays for proteins and small molecules into the clinic. This will allow for more sensitive and precise early detection of diseases, better treatment optimization, and an improved therapeutic monitoring.
We develop specific听Multiple Reaction Monitoring mass spectrometry (MRM-MS)听assays for quantifying small molecules in blood and plasma in close collaboration with clinicians at the Jewish General Hospital.
1. Multiple reaction monitoring-based, multiplexed, absolute quantitation of 45 proteins in human plasma. Kuzyk, M.A., Smith, D., Yang, J., Cross, T.J., Jackson, A.M., Hardie, D.B., Anderson, N.L., Borchers, C.H., 2009.听Mol. Cell. Proteomics听MCP 8, 1860鈥�1877.
2. MRM-based multiplexed quantitation of 67 putative cardiovascular disease biomarkers in human plasma. Domanski, D., Percy, A.J., Yang, J., Chambers, A.G., Hill, J.S., Freue, G.V.C., Borchers, C.H., 2012.听Proteomics听12, 1222鈥�1243.听
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We furthermore develop听immuno-MALDI (iMALDI)听assays for the highly sensitive, specific and automated quantification of proteins and their phosphorylation states from cells, fresh-frozen tissues and FFPE tissues. The workflow includes the extraction of proteins, enzymatic digestion, the spike-in of stable isotope-labeled reference standard (SIS) peptides, and the combined immunocapture of the endogenous peptide and its SIS counterpart. The signal is measured by MALDI-MS, allowing the absolute quantification of proteins. As the workflow requires no LC system and can be fully automated, it represents a promising technology to precisely quantify proteins from minimal sample amounts such as needle biopsies with the robustness required for a real clinical setting.
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1. How iMALDI can improve clinical diagnostics. Popp, R., Basik, M., Spatz, A., Batist, G., Zahedi, R.P., Borchers, C.H., 2018.
Analyst听Clin. Chem. 64, 1271鈥�1272.
143, 2197鈥�2203.
2. Immuno-MALDI-TOF-MS in the Clinic. Zahedi, R.P., Parker, C.E., Borchers, C.H., 2018.