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Martin L, Zouhiri N, Audran M, et al. A validated, sensitive electrophoretic method for the detection of activin receptor type II-Fc fusion proteins in human blood. Drug Test Anal. 2018;doi: 10.1002/dta.2376.
Martin, L., Zouhiri, N., Audran, M., & Marchand, A. (2018). A validated, sensitive electrophoretic method for the detection of activin receptor type II-Fc fusion proteins in human blood. Drug testing and analysis, . https://doi.org/10.1002/dta.2376
Martin, L, et al. "A validated, sensitive electrophoretic method for the detection of activin receptor type II-Fc fusion proteins in human blood." Drug testing and analysis vol. (2018). doi: https://doi.org/10.1002/dta.2376
Martin L, Zouhiri N, Audran M, Marchand A. A validated, sensitive electrophoretic method for the detection of activin receptor type II-Fc fusion proteins in human blood. Drug Test Anal. 2018 Mar 02; doi: 10.1002/dta.2376. Epub 2018 Mar 02. PMID: 29499588.
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Drug Test Anal. 2018 Mar 02; doi: 10.1002/dta.2376. Epub 2018 Mar 02.

A validated, sensitive electrophoretic method for the detection of activin receptor type II-Fc fusion proteins in human blood.

Drug testing and analysis

L Martin, N Zouhiri, M Audran, A Marchand

Affiliations

  1. Analysis Department, Agence Française de Lutte contre le Dopage (AFLD), Châtenay-Malabry, France.

PMID: 29499588 DOI: 10.1002/dta.2376

Abstract

New therapeutic proteins that trap circulating members of the transforming growth factor (TGF) beta superfamily (activins and growth differentiation factors) show promising effects on erythropoiesis and muscular growth. They are dimeric recombinant fusion proteins composed of the extracellular domain of a human activin receptor (ActRIIA or IIB) linked to the Fc part of human IgG1. Sotatercept (ActRIIA-Fc) and Luspatercept (a modified ActRIIB-Fc) in particular are now in phase 2/3 of clinical trials against anemia and included in the prohibited list established by the World Anti-Doping Agency. To prevent a potential misuse by athletes in the near future, a robust and sensitive method of detection is needed. We validated an approach adapted from an electrophoretic method used for the detection of recombinant erythropoietins that allowed detection of various ActRIIA-Fc and ActRIIB-Fc proteins, including variants produced in different cell types, after a single immuno-extraction step. After separation by sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE), an initial testing procedure performed by single-blotting can indicate the presence of an ActRII-Fc (indifferently type IIA or IIB). A confirmation performed by double-blotting using different antibodies for detection allows a more precise identification of the type of ActRII-Fc (IIA, IIB). Starting from a few hundred microliters of serum or plasma, this method is specific, sensitive, and easy to perform. It could easily be adopted by anti-doping laboratories.

Copyright © 2018 John Wiley & Sons, Ltd.

Keywords: activin receptor type II-Fc fusion proteins; electrophoretic method; erythropoiesis stimulating agent; immuno-purification; plasma; serum

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