title: Antibody-independent targeted quantification of TMPRSS2-ERG fusion protein products in prostate cancer.
abstract: Fusions between the transmembrane protease serine 2 (TMPRSS2) and ETS related gene (ERG) represent one of the most specific biomarkers that define a distinct molecular subtype of prostate cancer. Studies of TMPRSS2-ERG gene fusions have seldom been performed at the protein level, primarily due to the lack of high-quality antibodies suitable for quantitative studies. Herein, we applied a recently developed PRISM (high-pressure high-resolution separations with intelligent selection and multiplexing)-SRM (selected reaction monitoring) strategy for quantifying ERG protein in prostate cancer cell lines and tumors. The highly sensitive PRISM-SRM assays provided confident detection of 6 unique ERG peptides in both TMPRSS2-ERG positive cell lines and tissues, but not in cell lines or tissues lacking the TMPRSS2-ERG rearrangement, clearly indicating that ERG protein expression is significantly increased in the presence of the TMPRSS2-ERG gene fusion. Significantly, our results provide evidence that two distinct ERG protein isoforms are simultaneously expressed in TMPRSS2-ERG positive samples as evidenced by the concomitant detection of two mutually exclusive peptides in two patient tumors and in the VCaP prostate cancer cell line. Three peptides, shared across almost all fusion protein products, were determined to be the most abundant peptides, providing "signature" peptides for detection of ERG over-expression resulting from TMPRSS2-ERG gene fusion. The PRISM-SRM assays provide valuable tools for studying TMPRSS2-ERG gene fusion protein products in prostate cancer.
authors.affiliation: Biological Sciences Division, Pacific Northwest National Laboratory, Richland, WA, USA.
authors.forename: Jintang
authors.initials: J
authors.lastname: He
authors.affiliation: Biological Sciences Division, Pacific Northwest National Laboratory, Richland, WA, USA.
authors.forename: Xuefei
authors.initials: X
authors.lastname: Sun
authors.affiliation: Biological Sciences Division, Pacific Northwest National Laboratory, Richland, WA, USA.
authors.forename: Tujin
authors.initials: T
authors.lastname: Shi
authors.affiliation: Biological Sciences Division, Pacific Northwest National Laboratory, Richland, WA, USA.
authors.forename: Athena A
authors.initials: AA
authors.lastname: Schepmoes
authors.affiliation: Environmental Molecular Sciences Laboratory, Pacific Northwest National Laboratory, Richland, WA, USA.
authors.forename: Thomas L
authors.initials: TL
authors.lastname: Fillmore
authors.affiliation: Biological Sciences Division, Pacific Northwest National Laboratory, Richland, WA, USA.
authors.forename: Vladislav A
authors.initials: VA
authors.lastname: Petyuk
authors.affiliation: Biological Sciences Division, Pacific Northwest National Laboratory, Richland, WA, USA.
authors.forename: Fang
authors.initials: F
authors.lastname: Xie
authors.affiliation: Environmental Molecular Sciences Laboratory, Pacific Northwest National Laboratory, Richland, WA, USA.
authors.forename: Rui
authors.initials: R
authors.lastname: Zhao
authors.affiliation: Biological Sciences Division, Pacific Northwest National Laboratory, Richland, WA, USA.
authors.forename: Marina A
authors.initials: MA
authors.lastname: Gritsenko
authors.affiliation: Biological Sciences Division, Pacific Northwest National Laboratory, Richland, WA, USA.
authors.forename: Feng
authors.initials: F
authors.lastname: Yang
authors.affiliation: Department of Pathology and Laboratory Medicine, Weill Cornell Medical College, New York, NY, USA.
authors.forename: Naoki
authors.initials: N
authors.lastname: Kitabayashi
authors.affiliation: Department of Pathology and Laboratory Medicine, Weill Cornell Medical College, New York, NY, USA.
authors.forename: Sung-Suk
authors.initials: SS
authors.lastname: Chae
authors.affiliation: Department of Pathology and Laboratory Medicine, Weill Cornell Medical College, New York, NY, USA.
authors.forename: Mark A
authors.initials: MA
authors.lastname: Rubin
authors.affiliation: Department of Urology, University of Michigan Medical School, Ann Arbor, MI, USA; Michigan Center for Translational Pathology, University of Michigan Medical School, Ann Arbor, MI, USA.
authors.forename: Javed
authors.initials: J
authors.lastname: Siddiqui
authors.affiliation: Department of Urology, University of Michigan Medical School, Ann Arbor, MI, USA.
authors.forename: John T
authors.initials: JT
authors.lastname: Wei
authors.affiliation: Michigan Center for Translational Pathology, University of Michigan Medical School, Ann Arbor, MI, USA.
authors.forename: Arul M
authors.initials: AM
authors.lastname: Chinnaiyan
authors.affiliation: Biological Sciences Division, Pacific Northwest National Laboratory, Richland, WA, USA.
authors.forename: Wei-Jun
authors.initials: WJ
authors.lastname: Qian
authors.affiliation: Biological Sciences Division, Pacific Northwest National Laboratory, Richland, WA, USA.
authors.forename: Richard D
authors.initials: RD
authors.lastname: Smith
authors.affiliation: Division of Cancer Prevention, National Cancer Institute, Rockville, MD, USA.
authors.forename: Jacob
authors.initials: J
authors.lastname: Kagan
authors.affiliation: Division of Cancer Prevention, National Cancer Institute, Rockville, MD, USA.
authors.forename: Sudhir
authors.initials: S
authors.lastname: Srivastava
authors.affiliation: Biological Sciences Division, Pacific Northwest National Laboratory, Richland, WA, USA.
authors.forename: Karin D
authors.initials: KD
authors.lastname: Rodland
authors.affiliation: Biological Sciences Division, Pacific Northwest National Laboratory, Richland, WA, USA. Electronic address: tao.liu@pnnl.gov.
authors.forename: Tao
authors.initials: T
authors.lastname: Liu
authors.affiliation: Biological Sciences Division, Pacific Northwest National Laboratory, Richland, WA, USA. Electronic address: dave.camp@pnnl.gov.
authors.forename: David G
authors.initials: DG
authors.lastname: Camp
mesh_terms: D000595:Amino Acid Sequence; D045744:Cell Line, Tumor; D015321:Gene Rearrangement; D006801:Humans; D008297:Male; D008969:Molecular Sequence Data; D015514:Oncogene Proteins, Fusion / Q000032:analysis* / Q000235:genetics; D010455:Peptides / Q000032:analysis / Q000235:genetics; D011467:Prostate / Q000378:metabolism / Q000473:pathology*; D011471:Prostatic Neoplasms / Q000235:genetics / Q000473:pathology*
chemical_list: D015514:Oncogene Proteins, Fusion; D010455:Peptides; C515553:TMPRSS2-ERG fusion protein, human
keywords: ERG protein isoform; PRISM-SRM; Prostate cancer; TMPRSS2-ERG gene fusion; Targeted quantification