ABSTRACT
Proteomic analysis of formalin-fixed paraffin-embedded (FFPE) tissue would enable retrospective biomarker investigations of this vast archive of pathologically characterized clinical samples that exist worldwide. These FFPE tissues are, however, refractory to proteomic investigations utilizing many state of the art methodologies largely due to the high level of covalently cross-linked proteins arising from formalin fixation. A novel tissue microdissection technique has been developed and combined with a method to extract soluble peptides directly from FFPE tissue for mass spectral analysis of prostate cancer (PCa) and benign prostate hyperplasia (BPH). Hundreds of proteins from PCa and BPH tissue were identified, including several known PCa markers such as prostate-specific antigen, prostatic acid phosphatase, and macrophage inhibitory cytokine-1. Quantitative proteomic profiling utilizing stable isotope labeling confirmed similar expression levels of prostate-specific antigen and prostatic acid phosphatase in BPH and PCa cells, whereas the expression of macrophage inhibitory cytokine-1 was found to be greater in PCa as compared with BPH cells.
Subject(s)
Formaldehyde , Gene Expression Regulation, Neoplastic , Neoplasm Proteins/analysis , Prostatic Neoplasms/metabolism , Prostatic Neoplasms/pathology , Proteomics , Tissue Fixation , Amino Acid Sequence , Cytokines/analysis , Cytokines/chemistry , Cytokines/metabolism , Gene Expression Profiling , Growth Differentiation Factor 15 , Humans , Immunohistochemistry , Male , Mass Spectrometry , Microdissection , Molecular Sequence Data , Neoplasm Proteins/chemistry , Phosphatidylethanolamine Binding Protein/analysis , Phosphatidylethanolamine Binding Protein/chemistry , Phosphatidylethanolamine Binding Protein/metabolism , Prostate/metabolism , Prostate/pathology , Prostate-Specific Antigen/analysis , Prostate-Specific Antigen/chemistry , Prostate-Specific Antigen/metabolism , Prostatic Hyperplasia/metabolism , Prostatic Hyperplasia/pathology , Protein Array Analysis , Proteome/analysis , Proteome/chemistry , Proteome/metabolismABSTRACT
Identification and quantitation of candidate biomarker proteins in large numbers of individual tissues is required to validate specific proteins, or panels of proteins, for clinical use as diagnostic, prognostic, toxicological, or therapeutic markers. Mass spectrometry (MS) provides an exciting analytical methodology for this purpose. Liquid Tissue MS protein preparation allows researchers to utilize the vast, already existing, collections offormalin-fixed paraffin-embedded (FFPE) tissues for the procurement of peptides and the analysis across a variety of MS platforms.
