Specific MALDI Imaging and Profiling for Biomarker Hunting and Validation: Fragment of the 11S Proteasome Activator Complex, Reg Alpha Fragment, Is a New Potential Ovary Cancer Biomarker

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Abstract
MALDI imaging mass spectrometry represents a new analytical tool to directly provide the spatial distribution and relative abundance of proteins in tissue. Twenty-five ovary carcinomas (stages III and IV) and 23 benign ovaries were directly analyzed using MALDI-TOF MS. The biomarker with the major prevalence (80%) has been fully identified using MALDI MS and nanoESI MS and MS/MS after separation by RP-HPLC and trypsin enzymatic digestion. This marker with an m/z of 9744 corresponds to 84 amino acid residues from the 11S proteasome activator complex, named PA28 or Reg-alpha. Validation of this marker has been performed using MALDI imaging, classical immunocytochemistry with an antibody raised against the C-terminal part of the protein, specific MALDI imaging, and Western blot analysis. The validation, using immunocytochemistry, confirmed the epithelial localization of this fragment with nucleus localization in benign epithelial cells and a cytoplasmic localization in carcinoma cells. This indicates that this antibody could be used to discriminate the borderline tumor cases. At this point, a multicentric study needs to be conducted in order to clearly establish the potential of this biomarker. Taken together these studies reflect that direct tissue analysis and specific MALDI imaging strategies facilitate biomarker hunting and validation which can be named pathological proteomics. Keywords: MALDI direct analysis • MALDI imaging • Ovarian cancer • biomarker hunting • biomarker validation • Proteasome 11S