Identification of gel-separated tumor marker proteins by mass spectrometry

Abstract

Two-dimensional gel electrophoresis with subsequent analysis by mass spectrometry was applied to study differences in protein expression between benign and malignant solid tumors from human beast, lung and ovary cells. Cells from freshly resected clinical material were lysed and the extracts were subjected to isoelectric focusing with immobilized pH gradients followed by second-dimensional separation on 10-13% sodium dodecyl sulfate (SDS)/polyacrylamide gels. Polypeptides were identified using matrix-assisted laser desorption/ionization and electrospray ionization mass spectrometry after in-gel protein digestion. Some of the upregulated polypeptides in malignant cells are of potential importance as markers of tumor proliferation. Twenty such proteins were identified, ten constituting novel identifications and ten sequence verifications of previously gel-matched proteins. The proteins identified span a wide range of functions, but several cases of protein truncation were found. Truncated forms of cytokeratins 6D and 8, and of cathepsin D were identified. Truncated froms of these over-expressed proteins support the presence of proteolytic processing steps in tumor material. The protein processing and the difference between protein and mRNA abundancies in tumors of different malignancy and origin suggest that studies at the protein level are important for an understanding of tumor phenotypes.

Proteomic detection of changes in protein synthesis induced by lanthanum in BGC-823 human gastric cancer cells

“…The differential proteins were excised from twodimensional gel and digested with trypsin as previously described (Bergman et al 2000). Briefly, the spots were washed several times with 50% acetonitrile, which was then removed.…”

A database of protein expression in lung cancer

“…Specific protein differences between different types of cancer have been identified by other groups. In a recent study of breast, ovary and lung tumors, 20 differentially expressed proteins were identified [9] and in a prior study, 16 polypeptides were found to be associated with different histopathological features of lung cancer [10,11]. In a study of 25 adenocarcinomas of the lung, 12 small cell lung cancers, and 16 squamous cell tumors, by our group (manuscript submitted) an initial analysis of protein 2-D patterns uncovered a group of 52 protein spots that differed in average integrated intensity between the three groups.…”

A protein molecular weight map of ES2 clear cell ovarian carcinoma cells using a two-dimensional liquid separations/mass mapping technique

“…This capability becomes especially important for mapping changes that occur during cancer progression. In the transformation process changes in protein expression may result where proteins are expressed at elevated or reduced levels [17][18][19][20][21][22][23][24][25][26]. Alternatively, important mechanisms in the cancer process often involve structural changes such as sequence variations or post-translational modifications [26][27][28][29][30][31][32].…”