SELDI protein profiling of dunning R-3327 derived cell lines: identification of molecular markers of prostate cancer progression.
BACKGROUND
We recently demonstrated the protein expression profiling of Dunning rat tumor cell lines of varying metastatic potential (G (0%), AT-1 ( approximately 20%), and MLL (100%)) using SELDI-TOF-MS. As a parallel effort, we have been pursuing the identification of the protein(s) comprising the individual discriminatory "peaks" and evaluating their utility as potential biomarkers for prostate cancer progression.


METHODS
To identify the observed SELDI-TOF-MS m/z (mass/charge) values with discriminatory expression between different sublines, we employed a combination of chemical pre-fractionation, liquid chromatography, gel electrophoresis and tandem mass spectroscopy. Identified proteins were then verified by immuno-assay and Western analysis.


RESULTS
A 17.5 K m/z SELDI-TOF-MS peak was found to retain discriminatory value in each of two separate study-sets with an increased expression in the metastatic MLL line. Sequence identification and subsequent immunoassays verified that Histone H2B is the observed 17.5 K m/z SELDI peak. SELDI-based immuno-assay and Western Blotting revealed that Histone H2B is specifically over-expressed in metastatic MLL lines.


CONCLUSIONS
SELDI-TOF MS analysis of the Dunning prostate cancer cell lines confirmed the consistent overexpression of a 17.5 K m/z peak in metastatic MLL subline. The 17.5 kDa protein from MLL has been isolated and identified as Histone H2B.
Malik
Gunjan
G
Center for Biomedical Proteomics, Virginia Prostate Center, Eastern Virginia Medical School, Norfolk, Virginia, USA.
Rojahn
Elizabeth
E
Ward
Michael D
MD
Gretzer
Mathew B
MB
Partin
Alan W
AW
Semmes
O John
OJ
Veltri
Robert W
RW
D000230:Adenocarcinoma / Q000737:chemistry / Q000378:metabolism* / Q000556:secondary; D000818:Animals; D014408:Biomarkers, Tumor / Q000032:analysis / Q000378:metabolism*; D045744:Cell Line, Tumor; D002851:Chromatography, High Pressure Liquid; D018450:Disease Progression; D004591:Electrophoresis, Polyacrylamide Gel; D008297:Male; D009362:Neoplasm Metastasis; D009363:Neoplasm Proteins / Q000032:analysis / Q000378:metabolism*; D011471:Prostatic Neoplasms / Q000737:chemistry / Q000378:metabolism* / Q000473:pathology; D040081:Protein Array Analysis; D040901:Proteomics; D051381:Rats; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization / Q000379:methods*; D053719:Tandem Mass Spectrometry
D014408:Biomarkers, Tumor; D009363:Neoplasm Proteins