Targeted quantitation of CVD-linked plasma proteins for biomarker verification and validation

Abstract

Despite significant advances in treatment, cardiovascular disease (CVD) remains one of the leading causes of morbidity and mortality in developed and developing countries. Judicious monitoring of common risk factors has been unable to control this global epidemic, necessitating novel biomarkers for improved screening and earlier disease detection and management. Although numerous plasma proteins have been associated with CVD, only a few of these potential biomarkers have been validated for clinical use. Here we review the quantitative proteomic methods used to verify and validate new biomarker candidates in human plasma. These methods center on a bottom-up approach involving multiple or selected reaction monitoring, for targeted detection, with stable isotope-labeled standards, for peptide normalization. Also included are a discussion of future strategies for improved CVD protein biomarker verification and validation, recommendations for method translation to the clinic, and future projections for protein biomarker research.

Keywords::

• cardiovascular disease
• multiple reaction monitoring
• plasma
• protein
• targeted quantitation

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Erratum

Financial & competing interests disclosure

The authors acknowledge support and funding from Genome Canada, Genome BC, the Western Economic Diversification of Canada, and the Networks of Centres of Excellence Centre of Excellence for Commercialization and Research Prevention of Organ Failure Centre of Excellence.

C Borchers is Chief Scientific Officer of MRM Proteomics INC and principal investigator on one grant related to this work (Genome Canada STIC grant funding) and receives support from four additional sources of funding not related to this work: NSERC Discovery Grant, Genome Canada LSARP grant, Genome Canada STIC grant for metabolomics 2011-2013, Genome Canada STIC grant for metabolomics 2013-2015. The authors have no other relevant affiliations or financial involvement with any organization or entity with a financial interest in or financial conflict with the subject matter or materials discussed in the manuscript. This includes employment, consultancies, honoraria, stock ownership or options, expert testimony, grants or patents received or pending, or royalties.

No writing assistance was utilized in the production of this manuscript.

Key issues

• • Few approved protein biomarkers for cardiovascular disease currently exist, but more are required for improved diagnostic accuracy and enhanced personalized medicine.

• • Targeted absolute quantitative proteomic techniques involving MRM-MS (multiple reaction monitoring-mass spectrometry) and stable isotope-labeled standard (SIS) peptides are ideal for candidate biomarker verification and validation in pre-clinical trials.

• • The MRM with SIS peptide/protein approach is rapid and robust, while offering high multiplexing and specificity, with the potential for high sensitivity.

• • Clinical validation of candidate protein biomarkers can be conducted with the MRM-MS with SIS peptide approach or with ELISAs.

• • Instrumentation for biofluid biomarker monitoring already exists in clinical laboratories for metabolomics and can be readily configured for protein-based analyses.

• • For clinical implementation, regular QC of the method and platform must be performed, the method must be usable by a non-expert, and user-friendly software with minimal intervention or subjectivity must be developed.

Notes