Proteomic validation of protease drug targets: pharmacoproteomics of matrix metalloproteinase inhibitor drugs using isotope-coded affinity tag labelling and tandem mass spectrometry.
We illustrate the use of quantitative proteomics, namely isotope-coded affinity tag labelling and tandem mass spectrometry, to assess the targets and effects of the blockade of matrix metalloproteinases by an inhibitor drug in a breast cancer cell culture system. Treatment of MT1-MMP-transfected MDA-MB-231 cells with AG3340 (Prinomastat) directly affected the processing a multitude of matrix metalloproteinase substrates, and indirectly altered the expression of an array of other proteins with diverse functions. Therefore, broad spectrum blockade of MMPs has wide-ranging biological consequences. In this human breast cancer cell line, secreted substrates accumulated uncleaved in the conditioned medium and plasma membrane protein substrates were retained on the cell surface, due to reduced processing and shedding of these proteins (cell surface receptors, growth factors and bioactive molecules) to the medium in the presence of the matrix metalloproteinase inhibitor. Hence, proteomic investigation of drug-perturbed cellular proteomes can identify new protease substrates and at the same time provides valuable information for target validation, drug efficacy and potential side effects prior to commitment to clinical trials.
Butler
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Centre for Blood Research, Life Sciences Institute, University of British Columbia, Vancouver, V6T 1Z3, Canada.
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D000970:Antineoplastic Agents / Q000009:adverse effects / Q000494:pharmacology*; D001943:Breast Neoplasms / Q000188:drug therapy / Q000201:enzymology; D045744:Cell Line, Tumor; D015195:Drug Design; D004353:Drug Evaluation, Preclinical / Q000379:methods; D016326:Extracellular Matrix Proteins; D005786:Gene Expression Regulation / Q000187:drug effects; D006801:Humans; D007553:Isotope Labeling; D061965:Matrix Metalloproteinase Inhibitors; D020782:Matrix Metalloproteinases / Q000378:metabolism; D009930:Organic Chemicals / Q000494:pharmacology; D010447:Peptide Hydrolases / Q000378:metabolism; D011480:Protease Inhibitors / Q000009:adverse effects / Q000494:pharmacology*; D011499:Protein Processing, Post-Translational / Q000187:drug effects; D021381:Protein Transport / Q000187:drug effects; D040901:Proteomics / Q000379:methods*; D013379:Substrate Specificity; D049490:Systems Biology / Q000379:methods; D053719:Tandem Mass Spectrometry
D000970:Antineoplastic Agents; D016326:Extracellular Matrix Proteins; D061965:Matrix Metalloproteinase Inhibitors; D009930:Organic Chemicals; D011480:Protease Inhibitors; C113282:prinomastat; D010447:Peptide Hydrolases; D020782:Matrix Metalloproteinases