A strategy based on mass spectrometry proteomics is used to discover biomarkers for prostate and bladder cancer. The markers targeted are proteins secreted by the cancer cells and/or cancer-associated cells. Tumor tissue specimens are obtained from surgically resected organs after pathology. The specimens are digested by collagenase into single cells in a serum-free media. The digestion media, termed COL, is made cell-free by centrifugation, and it contains proteins made by the cells. A matched non-cancer specimen is processed similarly for comparative analysis. Differentially expressed protein species in the cancer and non-cancer COL pairs are screened by a method termed glycopeptide capture followed by tandem mass spectrometry for identification. Since virtually all secreted proteins are post-translationally modified by the addition of carbohydrate groups this method selectively analyzes these proteins. Quantification is achieved by isotopic labeling of the peptides, i.e. light isotopes for cancer and heavy isotopes for non-cancer. Candidate markers will be validated by Western blotting of the COL samples and immunohistochemistry if specific antibodies are available, or by in situ nucleic acid probe hybridization. This marker discovery process is illustrated by TIMP1, found in our initial experiments. The feasibility of proteomic analysis of urine samples to detect markers will be done. Preliminary experiments have generated a human urinary proteome of 239 protein species. Urine samples will be obtained from cancer patients and age-matched volunteers. The goal is to develop an assay for cancer associated protein markers that can be detected in voided urine for the diagnosis of prostate and bladder cancer. ? ? ?

Agency
National Institute of Health (NIH)
Institute
National Cancer Institute (NCI)
Type
Research Project--Cooperative Agreements (U01)
Project #
5U01CA111244-02
Application #
6951555
Study Section
Special Emphasis Panel (ZCA1-SRRB-E (O1))
Program Officer
Kagan, Jacob
Project Start
2004-09-22
Project End
2009-07-31
Budget Start
2005-09-01
Budget End
2006-07-31
Support Year
2
Fiscal Year
2005
Total Cost
$458,951
Indirect Cost
Name
University of Washington
Department
Urology
Type
Schools of Medicine
DUNS #
605799469
City
Seattle
State
WA
Country
United States
Zip Code
98195
Shi, Tujin; Quek, Sue-Ing; Gao, Yuqian et al. (2017) Multiplexed targeted mass spectrometry assays for prostate cancer-associated urinary proteins. Oncotarget 8:101887-101898
Ahmad, Rumana; Nicora, Carrie D; Shukla, Anil K et al. (2016) An efficient method for native protein purification in the selected range from prostate cancer tissue digests. Chin Clin Oncol 5:78
Vitello, Elizabeth A; Quek, Sue-Ing; Kincaid, Heather et al. (2016) Cancer-secreted AGR2 induces programmed cell death in normal cells. Oncotarget 7:49425-49434
Ho, Melissa E; Quek, Sue-Ing; True, Lawrence D et al. (2016) Bladder cancer cells secrete while normal bladder cells express but do not secrete AGR2. Oncotarget 7:15747-56
Alavi, Mohammed; Mah, Vei; Maresh, Erin L et al. (2015) High expression of AGR2 in lung cancer is predictive of poor survival. BMC Cancer 15:655
Quek, Sue-Ing; Wong, Olivia M; Chen, Adeline et al. (2015) Processing of voided urine for prostate cancer RNA biomarker analysis. Prostate 75:1886-95
Shi, Tujin; Gao, Yuqian; Quek, Sue Ing et al. (2014) A highly sensitive targeted mass spectrometric assay for quantification of AGR2 protein in human urine and serum. J Proteome Res 13:875-82
Shi, Tujin; Sun, Xuefei; Gao, Yuqian et al. (2013) Targeted quantification of low ng/mL level proteins in human serum without immunoaffinity depletion. J Proteome Res 12:3353-61
Shi, Tujin; Fillmore, Thomas L; Gao, Yuqian et al. (2013) Long-gradient separations coupled with selected reaction monitoring for highly sensitive, large scale targeted protein quantification in a single analysis. Anal Chem 85:9196-203
Ho, Melissa E; Quek, Sue-Ing; True, Lawrence D et al. (2013) Prostate cancer cell phenotypes based on AGR2 and CD10 expression. Mod Pathol 26:849-59

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