title: Prevalidation of potential protein biomarkers in toxicology using iTRAQ reagent technology.
abstract: Today, toxicoproteomics still relies mainly on 2-DE followed by MS for detection and identification of proteins, which might characterize a certain state of disease, indicate toxicity or even predict carcinogenicity. We utilized the classical 2-DE/MS approach for the evaluation of early protein biomarkers which are predictive for chemically induced hepatocarcinogenesis in rats. We were able to identify statistically significantly deregulated proteins in N-nitrosomorpholine exposed rat liver tissue. Based on literature data, biological relevance in the early molecular process of hepatocarcinogenicity could be suggested for most of these potential biomarkers. However, in order to ensure reliable results and to create the prerequisites necessary for integration in routine toxicology studies in the future, these protein expression patterns need to be prevalidated using independent technology platforms. In the current study, we evaluated the usefulness of iTRAQ reagent technology (Applied Biosystems, Framingham, USA), a recently introduced MS-based protein quantitation method, for verification of the 2-DE/MS biomarkers. In summary, the regulation of 26 2-DE/MS derived protein biomarkers could be verified. Proteins like HSP 90-beta, annexin A5, ketohexokinase, N-hydroxyarylamine sulfotransferase, ornithine aminotransferase, and adenosine kinase showed highly comparable fold changes using both proteomic quantitation strategies. In addition, iTRAQ analysis delivered further potential biomarkers with biological relevance to the processes of hepatocarcinogenicity: e.g. placental form of glutathione S-transferase (GST-P), carbonic anhydrase, and aflatoxin B1 aldehyde reductase. Our results show both the usefulness of iTRAQ reagent technology for biomarker prevalidation as well as for identification of further potential marker proteins, which are indicative for liver hepatocarcinogenicity.
authors: Applied Biosystems, Mass Spectrometry and Proteomics, Darmstadt, Germany.
authors: Matthias
authors: M
authors: Glückmann
authors: Kerstin
authors: K
authors: Fella
authors: Dietmar
authors: D
authors: Waidelich
authors: Dietrich
authors: D
authors: Merkel
authors: Volker
authors: V
authors: Kruft
authors: Peter-Jürgen
authors: PJ
authors: Kramer
authors: Yvonne
authors: Y
authors: Walter
authors: Jürgen
authors: J
authors: Hellmann
authors: Michael
authors: M
authors: Karas
authors: Michaela
authors: M
authors: Kröger
mesh_terms: D000818:Animals; D015415:Biomarkers / Q000737:chemistry / Q000378:metabolism*; D002273:Carcinogens / Q000494:pharmacology; D015180:Electrophoresis, Gel, Two-Dimensional; D008099:Liver / Q000166:cytology / Q000187:drug effects / Q000378:metabolism / Q000473:pathology; D008297:Male; D013058:Mass Spectrometry / Q000379:methods; D008969:Molecular Sequence Data; D009602:Nitrosamines / Q000494:pharmacology; D020543:Proteome / Q000032:analysis*; D051381:Rats; D017208:Rats, Wistar; D015203:Reproducibility of Results; D014116:Toxicology / Q000295:instrumentation / Q000379:methods
chemical_list: D015415:Biomarkers; D002273:Carcinogens; D009602:Nitrosamines; D020543:Proteome; C002741:N-nitrosomorpholine