Development of an ELISA to detect the secreted prostate cancer biomarker AGR2 in voided urine.

Abstract

BACKGROUND:

Comparative transcriptomics between sorted cells identified AGR2 as one of the highest up-regulated genes in cancer. Overexpression in primary tumors was verified by tissue microarray analysis. AGR2 encodes a 19-kDa secreted protein that might be found in urine.

METHODS:

Monoclonal antibodies were generated against AGR2. One antibody pair, P1G4 (IgG1) to capture and P3A5 (IgG2a) to detect, showed good performance characteristics in a sandwich ELISA. This assay could detect AGR2 at sub ng/ml quantities.

RESULTS:

AGR2 was detected in tissue digestion media of tumor specimens and culture media of AGR2-secreting prostate cancer cell lines. Additional testings involved frozen section immunohistochemistry, immunoprecipitation, and Western blot analysis. Voided urine samples were collected from pre-operative cancer patients, and urinary protein was desalted and concentrated by filtration. The amount of AGR2 detected was scored as pg/100 µg total protein, and then converted to pg/ml urine. The developed ELISA detected AGR2 protein, ranging from 3.6 to 181 pg/ml, in an initial cohort of samples. AGR2 was not detected in the urine of non-cancer and a bladder cancer patient.

CONCLUSIONS:

For prostate cancer, an AGR2 urine test could be used for diagnosis. The data, although derived from a small number of samples assayed, showed that developing such a test for clinical application is viable because AGR2 is specific to cancer cells, and apparently secreted into urine.

• Authors: Elizabeth A Wayner, Sue-Ing Quek, Rumana Ahmad, Melissa E Ho, Michelle A Loprieno, Yong Zhou, William J Ellis, Lawrence D True, Alvin Y Liu
• Journal: The Prostate (Prostate) Vol. 72 Issue 9 Pg. 1023-34 (Jun 15 2012) ISSN: 1097-0045 [Electronic] United States
• PMID: 22072305 (Publication Type: Comparative Study, Journal Article, Research Support, N.I.H., Extramural)
• Copyright: Copyright © 2011 Wiley Periodicals, Inc.

Chemical References

• AGR2 protein, human
• Biomarkers, Tumor
• Mucoproteins
• Oncogene Proteins
• Proteins

Topics

• Amino Acid Sequence
• Animals
• Biomarkers, Tumor (chemistry, metabolism, urine)
• Enzyme-Linked Immunosorbent Assay (methods)
• Humans
• Male
• Mice
• Molecular Sequence Data
• Molecular Weight
• Mucoproteins
• Oncogene Proteins
• Prostatic Neoplasms (chemistry, metabolism, urine)
• Proteins (chemistry, genetics, metabolism)