Reproducible quantification of cancer-associated proteins in body fluids using targeted proteomics.
The rigorous testing of hypotheses on suitable sample cohorts is a major limitation in translational research. This is particularly the case for the validation of protein biomarkers; the lack of accurate, reproducible, and sensitive assays for most proteins has precluded the systematic assessment of hundreds of potential marker proteins described in the literature. Here, we describe a high-throughput method for the development and refinement of selected reaction monitoring (SRM) assays for human proteins. The method was applied to generate such assays for more than 1000 cancer-associated proteins, which are functionally related to candidate cancer driver mutations. We used the assays to determine the detectability of the target proteins in two clinically relevant samples: plasma and urine. One hundred eighty-two proteins were detected in depleted plasma, spanning five orders of magnitude in abundance and reaching below a concentration of 10 ng/ml. The narrower concentration range of proteins in urine allowed the detection of 408 proteins. Moreover, we demonstrate that these SRM assays allow reproducible quantification by monitoring 34 biomarker candidates across 83 patient plasma samples. Through public access to the entire assay library, researchers will be able to target their cancer-associated proteins of interest in any sample type using the detectability information in plasma and urine as a guide. The generated expandable reference map of SRM assays for cancer-associated proteins will be a valuable resource for accelerating and planning biomarker verification studies.
Department of Biology, Institute of Molecular Systems Biology, ETH Zurich, 8093 Zurich, Switzerland. huettenhain@imsb.biol.ethz.ch
Ruth
R
Hüttenhain
Martin
M
Soste
Nathalie
N
Selevsek
Hannes
H
Röst
Atul
A
Sethi
Christine
C
Carapito
Terry
T
Farrah
Eric W
EW
Deutsch
Ulrike
U
Kusebauch
Robert L
RL
Moritz
Emma
E
Niméus-Malmström
Oliver
O
Rinner
Ruedi
R
Aebersold
D014408:Biomarkers, Tumor / Q000097:blood / Q000652:urine; D001826:Body Fluids / Q000378:metabolism*; D015331:Cohort Studies; D005260:Female; D052138:Genes, Neoplasm / Q000235:genetics; D006801:Humans; D009154:Mutation / Q000235:genetics; D009363:Neoplasm Proteins / Q000097:blood* / Q000652:urine*; D009369:Neoplasms / Q000097:blood* / Q000235:genetics / Q000378:metabolism / Q000652:urine*; D010051:Ovarian Neoplasms / Q000097:blood; D010455:Peptides / Q000378:metabolism; D060066:Protein Interaction Maps; D040901:Proteomics / Q000379:methods*; D015203:Reproducibility of Results
D014408:Biomarkers, Tumor; D009363:Neoplasm Proteins; D010455:Peptides