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Detection and differentiation of active and inactive isoforms of coagulation factors II, VII, IX, and X in prothrombin complex concentrate by mass spectrometry.
Wilhelm, C; Kiessig, S T; Mandago, M; Wittke, S.
Afiliação
  • Wilhelm C; University of Applied Sciences Bremerhaven, An der Karlstadt 8, 27568 Bremerhaven, Germany.
  • Kiessig ST; PreviPharma Consulting GmbH, CUBEX41, Universitätsmedizin Mannheim, Haus 41, Theodor-Kutzer-Ufer 1-3, 68167 Mannheim, Germany.
  • Mandago M; PreviPharma Consulting GmbH, CUBEX41, Universitätsmedizin Mannheim, Haus 41, Theodor-Kutzer-Ufer 1-3, 68167 Mannheim, Germany.
  • Wittke S; University of Applied Sciences Bremerhaven, An der Karlstadt 8, 27568 Bremerhaven, Germany. Electronic address: swittke@hs-bremerhaven.de.
J Pharm Biomed Anal ; 209: 114475, 2022 Feb 05.
Article em En | MEDLINE | ID: mdl-34839053
ABSTRACT

PURPOSE:

Prothrombin complex concentrates (PCCs) are plasma products containing a mixture of four inactive/proactive coagulation factors. The activated forms of human coagulation factors, like Thrombin (FIIa), Convertin (FVIIa), activated Christmas factor (FIXa) and the activated Stuart-Prower factor (FXa), are impurities in PCCs. Until now no valid assay exists to differentiate the non activated proform (inactive) from active coagulation factor isoforms in PCCs in one measurement. Therefore, the aim of this study was to establish a mass spectrometry (LC-MS/MS)-based assay to address this issue in the ready to use medicinal product.

METHODS:

Bottom-up proteomics combining double digestion (Glu-C & Lys-C) and LC-MS/MS, was used to differentiate the inactive and active forms of the coagulation factors Prothrombin (FII), Proconvertin (FVII), Christmas factor (FIX) and the Stuart-Prower-factor (FX) in PCCs. RESULTS AND

CONCLUSIONS:

A targeted pseudo-multiple reaction monitoring (pMRM-LC-MS/MS)-assay was developed for the specific detection of four different coagulation factors in PCCs. Proteotypic peptides for the inactive/active isoforms (zymogen) of the four coagulation factors were identified and validated by the investigation of six investigational and one commercially available PCCs. In conclusion, the semi-quantitative determination and the distinction between the active and the inactive isoform of the respective coagulation factors were possible in one liquid chromatography tandem mass spectrometry (LC-MS/MS) run.
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Texto completo: 1 Coleções: 01-internacional Base de dados: MEDLINE Assunto principal: Fator IX / Protrombina Tipo de estudo: Diagnostic_studies / Prognostic_studies Limite: Humans Idioma: En Ano de publicação: 2022 Tipo de documento: Article

Texto completo: 1 Coleções: 01-internacional Base de dados: MEDLINE Assunto principal: Fator IX / Protrombina Tipo de estudo: Diagnostic_studies / Prognostic_studies Limite: Humans Idioma: En Ano de publicação: 2022 Tipo de documento: Article