Analytical platform evaluation for quantification of ERG in prostate cancer using protein and mRNA detection methods

He, Jintang; Schepmoes, Athena A.; Shi, Tujin; Wu, Chaochao; Fillmore, Thomas L.; Gao, Yuqian; Smith, Richard D.; Qian, Weijun; Rodland, Karin D.; Liu, Tao; Camp, David G.; Rastogi, Anshu; Tan, Shyh-Han; Yan, Wusheng; Mohamed, Ahmed A.; Huang, Wei; Banerjee, Sreedatta; Kagan, Jacob; Srivastava, Sudhir; McLeod, David; Srivastava, Shiv; Petrovics, Gyorgy; Dobi, Albert; Srinivasan, Alagarsamy

doi:10.1186/s12967-015-0418-z

Analytical platform evaluation for quantification of ERG in prostate cancer using protein and mRNA detection methods

Journal Article · Thu Jan 01 00:00:00 EST 2015 · Journal of Translational Medicine

DOI:https://doi.org/10.1186/s12967-015-0418-z· OSTI ID:1184918

He, Jintang; Schepmoes, Athena A.; Shi, Tujin; Wu, Chaochao; Fillmore, Thomas L.; Gao, Yuqian; Smith, Richard D.; Qian, Weijun; Rodland, Karin D.; Liu, Tao; Camp, David G.; Rastogi, Anshu; Tan, Shyh-Han; Yan, Wusheng; Mohamed, Ahmed A.; Huang, Wei; Banerjee, Sreedatta; Kagan, Jacob; Srivastava, Sudhir; McLeod, David more »

Background: The established methods for detecting prostate cancer (CaP) are based on tests using PSA (blood), PCA3 (urine), and AMACR (tissue) as biomarkers in patient samples. The demonstration of ERG oncoprotein overexpression due to gene fusion in CaP has thus provided ERG as an additional biomarker. Based on this, we hypothesized that ERG protein quantification methods can be of use in the diagnosis of prostate cancer. Methods: Therefore, an antibody-free assay for ERG3 protein detection was developed based on PRISM (high-pressure high-resolution separations with intelligent selection and multiplexing)-SRM (selected reaction monitoring) mass spectrometry. We utilized TMPRSS2-ERG positive VCaP and TMPRSS2-ERG negative LNCaP cells to simulate three different sample types (cells, tissue, and post-DRE urine sediment). Results: Recombinant ERG3 protein spiked into LNCaP cell lysates could be detected at levels as low as 20 pg by PRISM-SRM analysis. The sensitivity of the PRISM-SRM assay was around approximately 10,000 VCaP cells in a mixed cell population model of VCaP and LNCaP cells. Interestingly, ERG protein could be detected in as few as 600 VCaP cells spiked into female urine. The sensitivity of the in-house enzyme-linked immunosorbent assay (ELISA) was similar to the PRISM-SRM assay, with detection of 30 pg of purified recombinant ERG3 protein and 10,000 VCaP cells. On the other hand, qRT-PCR exhibited a higher sensitivity, as TMPRSS2-ERG transcripts were detected in as few as 100 VCaP cells, in comparison to NanoString methodologies which detected ERG from 10,000 cells. Conclusions: Based on this data, we propose that the detection of both ERG transcriptional products with RNA-based assays, as well as protein products of ERG using PRISM-SRM assays, may be of clinical value in developing diagnostics and prognostics assays for prostate cancer given their sensitivity, specificity, and reproducibility.

View Accepted Manuscript (DOE)

Research Organization:: Pacific Northwest National Laboratory (PNNL), Richland, WA (US), Environmental Molecular Sciences Laboratory (EMSL)

Sponsoring Organization:: USDOE

Grant/Contract Number:: AC05-76RL01830

OSTI ID:: 1184918

Report Number(s):: PNNL-SA-107827; 48505; 400412000

Journal Information:: Journal of Translational Medicine, Journal Name: Journal of Translational Medicine Journal Issue: 1 Vol. 13; ISSN 1479-5876

Country of Publication:: United States

Language:: English

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Noninvasive Prenatal Diagnosis Significance of ERG Methylation as a Biomarker in Down’s Syndrome Liu, Xiangju; Xue, Ming Medical Science Monitor, Vol. 23 https://doi.org/10.12659/msm.898687	journal	January 2017
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Quantification of mutant SPOP proteins in prostate cancer using mass spectrometry-based targeted proteomics. Wang, Hui; Barbieri, Christopher E.; He, Jintang BioMed Central https://doi.org/10.7892/boris.110759	text	January 2017

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