Uniformed Services University of the Health Sciences (USU)-Center for Prostate Disease Research (CPDR) will serve as National Cancer Institute (NCI)-Early Detection Research Network (EDRN)-Biomarker Reference Laboratory (BRL). CPDR will provide support for current NCI-EDRN efforts focused on the development of a multiplex quantitative mass spectrometry-based detection of biomarkers by using normal and cancer cells and adjacent stroma cells enriched by laser capture microdissection (LCM). Our focus on LCM based CaP biomarker discovery and validation in relation to key observations in CaP molecular genetics research. Enrichment of prostate cancer epithelial cells by Laser Capture Microdissection (LCM) for development of a SRM-MS based assay for detection of truncated ERG protein. -The initial detection of the biomarkers will focus on human prostate tissue with the ERG oncogene rearrangements. The CPDR will enrich the fraction of prostate cancer cells by microdissection of cancer epithelial cells and matched benign epithelial cells from 23 prostatectomy specimens. - Detection of aberrant ERG rearrangements and expression in prostatectomy specimens by IHC and RT-PCR assay: Prostatectomy specimens will be pre-screened for ERG protein expression by IHC using ERG-MAb and for ERG fusion transcripts by RT-PCR. - Enrichment of benign and cancer epithelial cells from whole mount prostatectomy sections: For pilot experiments, three prostatectomy specimens will be used for LCM of 100,000 ERG positive cancer cells and matched benign epithelial cells. This number of cells should be sufficient to optimize detection of the truncated ERG by SRM-MS at the Pacific Northwest National Laboratory (PNNL), DOE. As ERG expression is relatively homogenous in individual tumor foci, entire ERG positive focus (based on ERG IHC of consecutive tissue section) can be dissected from FFPE-whole mounted prostate sections. Additional 20 pairs of cancer and benign epithelial cells ((50,000 cells/specimen;10 ERG positive and 10 ERG negative) will be obtained which will be sent for blinded examination at PNNL. Enrichment of prostate cancer epithelial cells by Laser Capture Microdissection (LCM) for development of a SRM-MS based assay for detection of truncated ETV1 protein. The purpose is identical to the previous one and the steps and task requirements will be identical. This study will be performed in collaboration with PNNL and other proteomic EDRN centers. Since the frequency of ETV1 is much lower (>5%), assistance from other EDRN investigators will also be sought in obtaining such specimens.
