title: Biomarker discovery and identification in laser microdissected head and neck squamous cell carcinoma with ProteinChip technology, two-dimensional gel electrophoresis, tandem mass spectrometry, and immunohistochemistry.
abstract: Head and neck cancer is a frequent malignancy with a complex, and up to now not clear etiology. Therefore, despite of improvements in diagnosis and therapy, the survival rate with head and neck squamous-cell carcinomas is poor. For a better understanding of the molecular mechanisms behind the process of tumorigenesis and tumor progression, we have analyzed changes of protein expression between microdissected normal pharyngeal epithelium and tumor tissue by ProteinChip technology. For this, cryostat sections from head and neck tumors (n = 57) and adjacent mucosa (n = 44) were laser-microdissected and analyzed on ProteinChip arrays. The derived mass spectrometry profiles exhibited numerous statistical differences. One peak significantly higher expressed in the tumor (p = 0.000029) was isolated by two-dimensional gel electrophoresis and identified as annexin V by in-gel proteolytic digestion, peptide mapping, tandem mass spectrometry analysis, and immuno-deplete assay. The relevance of this single marker protein was further evaluated by immunohistochemistry. Annexin-positive tissue areas were re-analyzed on ProteinChip arrays to confirm the identity of this protein. In this study, we could show that biomarker in head and neck cancer can be found, identified, and assessed by combination of ProteinChip technology, two-dimensional gel electrophoresis, and immunohistochemistry. In our experience, however, such studies only make sense if a relatively pure microdissected tumor tissue is used. Only then minute changes in protein expression between normal pharyngeal epithelium and tumor tissue can be detected, and it will become possible to educe a tumor-associated protein pattern that might be used as a marker for tumorigenesis and progression.
authors.affiliation: Core Unit Chip Application (CUCA), Institute of Human Genetics and Anthropology, Friedrich-Schiller-University, 07740 Jena, Germany.
authors.forename: Christian
authors.initials: C
authors.lastname: Melle
authors.forename: Gunther
authors.initials: G
authors.lastname: Ernst
authors.forename: Bettina
authors.initials: B
authors.lastname: Schimmel
authors.forename: Annett
authors.initials: A
authors.lastname: Bleul
authors.forename: Sven
authors.initials: S
authors.lastname: Koscielny
authors.forename: Andreas
authors.initials: A
authors.lastname: Wiesner
authors.forename: Ralf
authors.initials: R
authors.lastname: Bogumil
authors.forename: Ursula
authors.initials: U
authors.lastname: Moller
authors.forename: Dirk
authors.initials: D
authors.lastname: Osterloh
authors.forename: Karl-Jurgen
authors.initials: KJ
authors.lastname: Halbhuber
authors.forename: Ferdinand
authors.initials: F
authors.lastname: von Eggeling
mesh_terms: D017304:Annexin A5 / Q000378:metabolism; D014408:Biomarkers, Tumor / Q000378:metabolism*; D002294:Carcinoma, Squamous Cell / Q000378:metabolism* / Q000473:pathology / Q000601:surgery; D015180:Electrophoresis, Gel, Two-Dimensional; D006258:Head and Neck Neoplasms / Q000378:metabolism* / Q000473:pathology / Q000601:surgery; D006801:Humans; D007150:Immunohistochemistry; D007834:Lasers; D013058:Mass Spectrometry; D042282:Microdissection; D040081:Protein Array Analysis; D040901:Proteomics / Q000379:methods*
chemical_list: D017304:Annexin A5; D014408:Biomarkers, Tumor