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Quantification of Intact and Truncated Stromal Cell-Derived Factor-1α in Circulation by Immunoaffinity Enrichment and Tandem Mass Spectrometry

  • Research Article
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Journal of The American Society for Mass Spectrometry

Abstract

Stromal cell-derived factor 1α (SDF-1α) or CXCL12 is a small pro-inflammatory chemoattractant cytokine and a substrate of dipeptidyl peptidase IV (DPP-IV). Proteolytic cleavage by DPP-IV inactivates SDF-1α and attenuates its interaction with CXCR4, its cell surface receptor. To enable investigation of suppression of such inactivation with pharmacologic inhibition of DPP-IV, we developed quantitative mass spectrometric methods that differentiate intact SDF-1α from its inactive form. Using top-down strategy in quantification, we demonstrated the unique advantage of keeping SDF-1α’s two disulfide bridges intact in the analysis. To achieve the optimal sensitivity required for quantification of intact and truncated SDF-1α at endogenous levels in blood, we coupled nano-flow tandem mass spectrometry with antibody-based affinity enrichment. The assay has a quantitative range of 20 pmol/L to 20 nmol/L in human plasma as well as in rhesus monkey plasma. With only slight modification, the same assay can be used to quantify SDF-1α in mice. Using two in vivo animal studies as examples, we demonstrated that it was critical to differentiate intact SDF-1α from its truncated form in the analysis of biomarkers for pharmacologic inhibition of DPP-IV activity. These novel methods enable translational research on suppression of SDF-1 inactivation with DPP-IV inhibition and can be applied to relevant clinical samples in the future to yield new insights on change of SDF-1α levels in disease settings and in response to therapeutic interventions.

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Acknowledgments

The authors thank Aimee L. Burton, Sophie Brennan, Irene Capodanno, and Xiaolan Shen for help with the mouse study. They are grateful for discussions with Joel Berger, David E. Kelley, and Nancy A. Thornberry.

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Author notes

  1. Nathan A. Yates

    Present address: Biomedical Mass Spectrometry Center for the Health Sciences, University of Pittsburgh, Pittsburgh, PA, USA

  2. Alessandro Pocai

    Present address: Cardiovascular and Metabolism, Janssen R&D, Spring House, PA, USA

Authors and Affiliations

  1. Pharmacokinetic Pharmacodynamics and Drug Metabolism, Merck and Co., Inc., Rahway, NJ, 07065, USA

    Weixun Wang, Bernard K. Choi, Anita Y. H. Lee, Sandra C. Souza, Nathan A. Yates & Lucinda H. Cohen

  2. Diabetes and Endocrinology, Merck and Co., Inc., Rahway, NJ, 07065, USA

    Wenyu Li, Zhege Lao, Timothy Kowalski & Alessandro Pocai

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  1. Weixun Wang
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Correspondence to Weixun Wang.

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Wang, W., Choi, B.K., Li, W. et al. Quantification of Intact and Truncated Stromal Cell-Derived Factor-1α in Circulation by Immunoaffinity Enrichment and Tandem Mass Spectrometry. J. Am. Soc. Mass Spectrom. 25, 614–625 (2014). https://doi.org/10.1007/s13361-013-0822-7

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  • DOI: https://doi.org/10.1007/s13361-013-0822-7

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