Quantification of mutant SPOP proteins in prostate cancer using mass spectrometry-based targeted proteomics

Wang, Hui; Barbieri, Christopher E.; He, Jintang; Gao, Yuqian; Shi, Tujin; Wu, Chaochao; Schepmoes, Athena A.; Fillmore, Thomas L.; Chae, Sung-Suk; Huang, Dennis; Mosquera, Juan Miguel; Qian, Wei-Jun; Smith, Richard D.; Srivastava, Sudhir; Kagan, Jacob; Camp, David G.; Rodland, Karin D.; Rubin, Mark A.; Liu, Tao

doi:10.1186/s12967-017-1276-7

Title: Quantification of mutant SPOP proteins in prostate cancer using mass spectrometry-based targeted proteomics

Journal Article · Tue Aug 15 00:00:00 EDT 2017 · Journal of Translational Medicine

DOI:https://doi.org/10.1186/s12967-017-1276-7· OSTI ID:1406773

Wang, Hui ^[1]; Barbieri, Christopher E. ^[2]; He, Jintang ^[1]; Gao, Yuqian ^[1]; Shi, Tujin ^[1]; Wu, Chaochao ^[1]; Schepmoes, Athena A. ^[1]; Fillmore, Thomas L. ^[1]; Chae, Sung-Suk ^[2]; Huang, Dennis ^[2]; Mosquera, Juan Miguel ^[2]; Qian, Wei-Jun ^[1];

^[1]; Srivastava, Sudhir ^[3]; Kagan, Jacob ^[3]; Camp, David G. ^[1];

^[1]; Rubin, Mark A. ^[2];

^[1]

Pacific Northwest National Lab. (PNNL), Richland, WA (United States)
Weill Cornell Medical College, New York, NY (United States). Inst. of Precision Medicine
National Cancer Institute, Bethesda, MD (United States). Division of Cancer Prevention, Cancer Biomarkers Research Group

Speckle-type POZ protein (SPOP) is an E3 ubiquitin ligase adaptor protein that functions as a potential tumor suppressor, and SPOP mutations have been identified in ~10% of human prostate cancers. However, it remains unclear if mutant SPOP proteins can be utilized as biomarkers for early detection, diagnosis, prognosis or targeted therapy of prostate cancer. Moreover, the SPOP mutation sites are distributed in a relatively short region where multiple lysine residues, posing significant challenges for bottom-up proteomics analysis of the SPOP mutations. To address this issue, PRISM (high-pressure, high-resolution separations coupled with intelligent selection and multiplexing)-SRM (selected reaction monitoring) mass spectrometry assays have been developed for quantifying wild-type SPOP protein and 11 prostate cancer-derived SPOP mutations. Despite inherent limitations due to amino acid sequence constraints, all the PRISM-SRM assays developed using Arg-C digestion showed a linear dynamic range of at least two orders of magnitude, with limits of quantification range from 0.1 to 1 fmol/µg of total protein in the cell lysate. Applying these SRM assays to analyze HEK293T cells with and without expression of the three most frequent SPOP mutations in prostate cancer (Y87N, F102C or F133V) led to confident detection of all three SPOP mutations in corresponding positive cell lines but not in the negative cell lines. Expression of the F133V mutation and wild-type SPOP was at much lower levels compared to that of F102C and Y87N mutations; however, at present it is unknown if this also affects the activity of the SPOP protein. In summary, PRISM-SRM enables multiplexed, isoform-specific detection of mutant SPOP proteins in cell lysates, which holds great potential in biomarker development for prostate cancer.

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

Sponsoring Organization:: USDOE; National Institutes of Health (NIH)

Grant/Contract Number:: AC05-76RL01830; ACNl-5006-001; P41-RR018522

OSTI ID:: 1406773

Alternate ID(s):: OSTI ID: 1757986

Report Number(s):: PNNL-SA-125993; 49639; 453040220

Journal Information:: Journal of Translational Medicine, Vol. 15, Issue 1; ISSN 1479-5876

Publisher:: BioMed Central Ltd.Copyright Statement

Country of Publication:: United States

Language:: English

Citation Metrics:

Cited by: 6 works

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Related Subjects

60 APPLIED LIFE SCIENCES
59 BASIC BIOLOGICAL SCIENCES
mass spectometry
targeted proteomics
SPOP M
prostate cancer
biomarker
PRISM-SRM

Quantification of mutant SPOP proteins in prostate cancer using mass spectrometry-based targeted proteomics Wang, Hui; Barbieri, Christopher; He, Jintang https://doi.org/10.6084/M9.FIGSHARE.C.3854275 The current record is supplemented by this collection	collection	January 2017
Quantification of mutant SPOP proteins in prostate cancer using mass spectrometry-based targeted proteomics Wang, Hui; Barbieri, Christopher; He, Jintang https://doi.org/10.6084/m9.figshare.c.3854275.v1 The current record is supplemented by this collection	collection	January 2017
MOESM1 of Quantification of mutant SPOP proteins in prostate cancer using mass spectrometry-based targeted proteomics Wang, Hui; Barbieri, Christopher; He, Jintang https://doi.org/10.6084/m9.figshare.c.3854275_d1 The current record is supplemented by this dataset	dataset	January 2017
MOESM1 of Quantification of mutant SPOP proteins in prostate cancer using mass spectrometry-based targeted proteomics Wang, Hui; Barbieri, Christopher; He, Jintang https://doi.org/10.6084/m9.figshare.c.3854275_d1.v1 The current record is supplemented by this dataset	dataset	January 2017
MOESM2 of Quantification of mutant SPOP proteins in prostate cancer using mass spectrometry-based targeted proteomics Wang, Hui; Barbieri, Christopher; He, Jintang https://doi.org/10.6084/M9.FIGSHARE.C.3854275_D2 The current record is supplemented by this audiovisual	audiovisual	January 2017
MOESM2 of Quantification of mutant SPOP proteins in prostate cancer using mass spectrometry-based targeted proteomics Wang, Hui; Barbieri, Christopher; He, Jintang https://doi.org/10.6084/m9.figshare.c.3854275_d2.v1 The current record is supplemented by this audiovisual	audiovisual	January 2017
MOESM3 of Quantification of mutant SPOP proteins in prostate cancer using mass spectrometry-based targeted proteomics Wang, Hui; Barbieri, Christopher; He, Jintang https://doi.org/10.6084/M9.FIGSHARE.C.3854275_D3 The current record is supplemented by this audiovisual	audiovisual	January 2017
MOESM3 of Quantification of mutant SPOP proteins in prostate cancer using mass spectrometry-based targeted proteomics Wang, Hui; Barbieri, Christopher; He, Jintang https://doi.org/10.6084/m9.figshare.c.3854275_d3.v1 The current record is supplemented by this audiovisual	audiovisual	January 2017
MOESM4 of Quantification of mutant SPOP proteins in prostate cancer using mass spectrometry-based targeted proteomics Wang, Hui; Barbieri, Christopher; He, Jintang https://doi.org/10.6084/m9.figshare.c.3854275_d4 The current record is supplemented by this dataset	dataset	January 2017
MOESM4 of Quantification of mutant SPOP proteins in prostate cancer using mass spectrometry-based targeted proteomics Wang, Hui; Barbieri, Christopher; He, Jintang https://doi.org/10.6084/m9.figshare.c.3854275_d4.v1 The current record is supplemented by this dataset	dataset	January 2017
MOESM5 of Quantification of mutant SPOP proteins in prostate cancer using mass spectrometry-based targeted proteomics Wang, Hui; Barbieri, Christopher; He, Jintang https://doi.org/10.6084/M9.FIGSHARE.C.3854275_D5 The current record is supplemented by this audiovisual	audiovisual	January 2017
MOESM5 of Quantification of mutant SPOP proteins in prostate cancer using mass spectrometry-based targeted proteomics Wang, Hui; Barbieri, Christopher; He, Jintang https://doi.org/10.6084/m9.figshare.c.3854275_d5.v1 The current record is supplemented by this audiovisual	audiovisual	January 2017
MOESM6 of Quantification of mutant SPOP proteins in prostate cancer using mass spectrometry-based targeted proteomics Wang, Hui; Barbieri, Christopher; He, Jintang https://doi.org/10.6084/M9.FIGSHARE.C.3854275_D6 The current record is supplemented by this audiovisual	audiovisual	January 2017
MOESM6 of Quantification of mutant SPOP proteins in prostate cancer using mass spectrometry-based targeted proteomics Wang, Hui; Barbieri, Christopher; He, Jintang https://doi.org/10.6084/m9.figshare.c.3854275_d6.v1 The current record is supplemented by this audiovisual	audiovisual	January 2017
MOESM7 of Quantification of mutant SPOP proteins in prostate cancer using mass spectrometry-based targeted proteomics Wang, Hui; Barbieri, Christopher; He, Jintang https://doi.org/10.6084/m9.figshare.c.3854275_d7 The current record is supplemented by this audiovisual	audiovisual	January 2017
MOESM7 of Quantification of mutant SPOP proteins in prostate cancer using mass spectrometry-based targeted proteomics Wang, Hui; Barbieri, Christopher; He, Jintang https://doi.org/10.6084/m9.figshare.c.3854275_d7.v1 The current record is supplemented by this audiovisual	audiovisual	January 2017
MOESM8 of Quantification of mutant SPOP proteins in prostate cancer using mass spectrometry-based targeted proteomics Wang, Hui; Barbieri, Christopher; He, Jintang https://doi.org/10.6084/M9.FIGSHARE.C.3854275_D8 The current record is supplemented by this audiovisual	audiovisual	January 2017
MOESM8 of Quantification of mutant SPOP proteins in prostate cancer using mass spectrometry-based targeted proteomics Wang, Hui; Barbieri, Christopher; He, Jintang https://doi.org/10.6084/m9.figshare.c.3854275_d8.v1 The current record is supplemented by this audiovisual	audiovisual	January 2017
MOESM9 of Quantification of mutant SPOP proteins in prostate cancer using mass spectrometry-based targeted proteomics Wang, Hui; Barbieri, Christopher; He, Jintang https://doi.org/10.6084/M9.FIGSHARE.C.3854275_D9 The current record is supplemented by this audiovisual	audiovisual	January 2017
MOESM9 of Quantification of mutant SPOP proteins in prostate cancer using mass spectrometry-based targeted proteomics Wang, Hui; Barbieri, Christopher; He, Jintang https://doi.org/10.6084/m9.figshare.c.3854275_d9.v1 The current record is supplemented by this audiovisual	audiovisual	January 2017