Abstract
Abstract Traditional immunohistochemistry serves as a vital diagnostic assay as it allows for semiquantitatively probing the magnitude and spatial distribution of protein expression and posttranslational modifications. Unfortunately, an accurate portrayal of disease often requires several immunohistochemical stains, which are run on separate tissue sections. The application of transcriptomics analyses in cancer classification has led to the generation of multivariate index analyses that confer diagnostic, prognostic, and predictive signatures of disease. However this method is limited by the inability consider target expression in the context of tissue (stroma, epithelium) or cell (e.g. stem/progenitor cells) type. A unifying platform that would allow for the quantitative spatial analysis of multiplexed in situ target measurements promises to exploit the best features of both systems. To this end, we have developed a novel multiplexed fluorescence immunohistochemistry assay that is capable of quantitative analysis of >25 antigens in a single formalin-fixed paraffin-embedded tissue section, as well as image and data analysis capabilities for analyzing the multidimensional data. By inclusion of targets for cell and tissue compartments, including nuclei, cytoplasm and membrane, we have demonstrated the ability to investigate subcellular protein expression, phosphorylation and co-localization in individual cells. In the present study we have stained a cohort of 79 prostate cancer patients of varying Gleason grades for a total of 24 antigens, representing key mediators of cell signaling, tumor suppressors and oncogenes, tumor associated phenotypic targets, and structural proteins. The images are precisely registered and segmented into individual cells nuclear, cytoplasmic and membrane regions and the underlying expression of a given antigen is then quantitatively determined in each compartment by automated algorithms. With the advent of this new tool, we aim to define new prognostic molecular signatures in prostate cancer. Citation Information: Clin Cancer Res 2010;16(14 Suppl):A28.
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