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Updated: May 2, 2026
Phosphopeptide Enrichment Coupled with Label-free Quantitative Mass Spectrometry to Investigate the Phosphoproteome in Prostate Cancer
Published on: August 2, 2018
Application of molecular technologies for phosphoproteomic analysis of clinical samples
M Pierobon1, J Wulfkuhle1, L Liotta1
1Center for Applied Proteomics and Molecular Medicine, George Mason University, Manassas, VA, USA.
Abstract:
The integration of small kinase inhibitors and monoclonal antibodies into oncological practice has opened a new paradigm for treating cancer patients. As proteins are the direct targets of the new generations of targeted therapeutics, many of which are kinase/enzymatic inhibitors, there is an increasing interest in developing technologies capable of monitoring post-translational changes of the human proteome for the identification of new predictive, prognostic and therapeutic biomarkers. It is well known that the vast majority of the activation/deactivation of these drug targets is driven by phosphorylation. This review provides a description of the main proteomic platforms (planar and bead array, reverse phase protein microarray, phosphoflow, AQUA and mass spectrometry) that have successfully been used for measuring changes in phosphorylation level of drug targets and downstream substrates using clinical specimens. Major emphasis was given to the strengths and weaknesses of the different platforms and to the major barriers that are associated with the analysis of the phosphoproteome. Finally, a number of examples of application of the above-mentioned technologies in the clinical setting are reported.
Insights
New targeted cancer therapies focus on proteins, driving interest in monitoring phosphorylation changes. This review details proteomic platforms for identifying biomarkers in clinical specimens.
Area of Science:
- Oncology
- Proteomics
- Biomarker Discovery
Background:
- Targeted cancer therapeutics, including kinase inhibitors and monoclonal antibodies, directly target proteins.
- Protein phosphorylation is a key mechanism regulating the activity of these drug targets.
- There is a growing need for technologies to monitor proteome-wide post-translational modifications for biomarker identification.
Purpose of the Study:
- To review proteomic platforms for measuring phosphorylation changes in clinical specimens.
- To discuss the strengths, weaknesses, and challenges of phosphoproteome analysis.
- To provide examples of these technologies in clinical applications.
Main Methods:
- Review of established proteomic platforms: planar and bead arrays, reverse phase protein microarrays, phosphoflow, AQUA, and mass spectrometry.
- Analysis of platform capabilities for measuring drug target and downstream substrate phosphorylation levels.
- Examination of barriers and limitations in phosphoproteome analysis.
Main Results:
- Several proteomic platforms are effective for quantifying phosphorylation levels in clinical samples.
- Each platform possesses distinct advantages and disadvantages regarding sensitivity, throughput, and cost.
- Significant challenges remain in phosphoproteome analysis, including sample preparation and data interpretation.
Conclusions:
- Proteomic technologies are crucial for identifying predictive, prognostic, and therapeutic biomarkers in oncology.
- Understanding the phosphoproteome aids in optimizing targeted cancer therapies.
- Further technological advancements are needed to overcome current limitations in clinical phosphoproteomics.
