Integrated pipeline for mass spectrometry-based discovery and confirmation of biomarkers demonstrated in a mouse model of breast cancer.
Despite their potential to impact diagnosis and treatment of cancer, few protein biomarkers are in clinical use. Biomarker discovery is plagued with difficulties ranging from technological (inability to globally interrogate proteomes) to biological (genetic and environmental differences among patients and their tumors). We urgently need paradigms for biomarker discovery. To minimize biological variation and facilitate testing of proteomic approaches, we employed a mouse model of breast cancer. Specifically, we performed LC-MS/MS of tumor and normal mammary tissue from a conditional HER2/Neu-driven mouse model of breast cancer, identifying 6758 peptides representing >700 proteins. We developed a novel statistical approach (SASPECT) for prioritizing proteins differentially represented in LC-MS/MS datasets and identified proteins over- or under-represented in tumors. Using a combination of antibody-based approaches and multiple reaction monitoring-mass spectrometry (MRM-MS), we confirmed the overproduction of multiple proteins at the tissue level, identified fibulin-2 as a plasma biomarker, and extensively characterized osteopontin as a plasma biomarker capable of early disease detection in the mouse. Our results show that a staged pipeline employing shotgun-based comparative proteomics for biomarker discovery and multiple reaction monitoring for confirmation of biomarker candidates is capable of finding novel tissue and plasma biomarkers in a mouse model of breast cancer. Furthermore, the approach can be extended to find biomarkers relevant to human disease.
Whiteaker
Jeffrey R
JR
Fred Hutchinson Cancer Research Center, Seattle, Washington 98109, USA.
Zhang
Heidi
H
Zhao
Lei
L
Wang
Pei
P
Kelly-Spratt
Karen S
KS
Ivey
Richard G
RG
Piening
Brian D
BD
Feng
Li-Chia
LC
Kasarda
Erik
E
Gurley
Kay E
KE
Eng
Jimmy K
JK
Chodosh
Lewis A
LA
Kemp
Christopher J
CJ
McIntosh
Martin W
MW
Paulovich
Amanda G
AG
D000465:Algorithms; D000818:Animals; D014408:Biomarkers, Tumor / Q000096:biosynthesis* / Q000097:blood; D002135:Calcium-Binding Proteins / Q000096:biosynthesis / Q000097:blood; D002853:Chromatography, Liquid; D016208:Databases, Factual; D004797:Enzyme-Linked Immunosorbent Assay; D016326:Extracellular Matrix Proteins / Q000096:biosynthesis / Q000097:blood; D005260:Female; D008325:Mammary Neoplasms, Experimental / Q000378:metabolism*; D051379:Mice; D015233:Models, Statistical; D053495:Osteopontin / Q000096:biosynthesis / Q000097:blood; D020543:Proteome / Q000096:biosynthesis*; D040901:Proteomics; D021241:Spectrometry, Mass, Electrospray Ionization; D053719:Tandem Mass Spectrometry
D014408:Biomarkers, Tumor; D002135:Calcium-Binding Proteins; D016326:Extracellular Matrix Proteins; D020543:Proteome; C506896:Spp1 protein, mouse; C084612:fibulin 2; D053495:Osteopontin