Cargando…
Characterization of IXINITY® (Trenonacog Alfa), a Recombinant Factor IX with Primary Sequence Corresponding to the Threonine-148 Polymorph
The goal of these studies was to extensively characterize the first recombinant FIX therapeutic corresponding to the threonine-148 (Thr-148) polymorph, IXINITY (trenonacog alfa [coagulation factor IX (recombinant)]). Gel electrophoresis, circular dichroism, and gel filtration were used to determine...
Autores principales: | , , , , |
---|---|
Formato: | Online Artículo Texto |
Lenguaje: | English |
Publicado: |
Hindawi Publishing Corporation
2016
|
Materias: | |
Acceso en línea: | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4779506/ https://www.ncbi.nlm.nih.gov/pubmed/26997955 http://dx.doi.org/10.1155/2016/7678901 |
_version_ | 1782419628098584576 |
---|---|
author | Monroe, Dougald M. Jenny, Richard J. Van Cott, Kevin E. Buhay, Shelly Saward, Laura L. |
author_facet | Monroe, Dougald M. Jenny, Richard J. Van Cott, Kevin E. Buhay, Shelly Saward, Laura L. |
author_sort | Monroe, Dougald M. |
collection | PubMed |
description | The goal of these studies was to extensively characterize the first recombinant FIX therapeutic corresponding to the threonine-148 (Thr-148) polymorph, IXINITY (trenonacog alfa [coagulation factor IX (recombinant)]). Gel electrophoresis, circular dichroism, and gel filtration were used to determine purity and confirm structure. Chromatographic and mass spectrometry techniques were used to identify and quantify posttranslational modifications. Activity was assessed as the ability to activate factor X (FX) both with and without factor VIIIa (FVIIIa) and in a standard clotting assay. All results were consistent across multiple lots. Trenonacog alfa migrated as a single band on Coomassie-stained gels; activity assays were normal and showed <0.002 IU of activated factor IX (FIXa) per IU of FIX. The molecule has >97% γ-carboxylation and underwent the appropriate structural change upon binding calcium ions. Trenonacog alfa was activated normally with factor XIa (FXIa); once activated it bound to FVIIIa and FXa. When activated to FIXa, it was inhibited efficiently by antithrombin. Glycosylation patterns were similar to plasma-derived FIX with sialic acid content consistent with the literature reports of good pharmacokinetic performance. These studies have shown that trenonacog alfa is a highly pure product with a primary sequence and posttranslational modifications consistent with the common Thr-148 polymorphism of plasma-derived FIX. |
format | Online Article Text |
id | pubmed-4779506 |
institution | National Center for Biotechnology Information |
language | English |
publishDate | 2016 |
publisher | Hindawi Publishing Corporation |
record_format | MEDLINE/PubMed |
spelling | pubmed-47795062016-03-20 Characterization of IXINITY® (Trenonacog Alfa), a Recombinant Factor IX with Primary Sequence Corresponding to the Threonine-148 Polymorph Monroe, Dougald M. Jenny, Richard J. Van Cott, Kevin E. Buhay, Shelly Saward, Laura L. Adv Hematol Research Article The goal of these studies was to extensively characterize the first recombinant FIX therapeutic corresponding to the threonine-148 (Thr-148) polymorph, IXINITY (trenonacog alfa [coagulation factor IX (recombinant)]). Gel electrophoresis, circular dichroism, and gel filtration were used to determine purity and confirm structure. Chromatographic and mass spectrometry techniques were used to identify and quantify posttranslational modifications. Activity was assessed as the ability to activate factor X (FX) both with and without factor VIIIa (FVIIIa) and in a standard clotting assay. All results were consistent across multiple lots. Trenonacog alfa migrated as a single band on Coomassie-stained gels; activity assays were normal and showed <0.002 IU of activated factor IX (FIXa) per IU of FIX. The molecule has >97% γ-carboxylation and underwent the appropriate structural change upon binding calcium ions. Trenonacog alfa was activated normally with factor XIa (FXIa); once activated it bound to FVIIIa and FXa. When activated to FIXa, it was inhibited efficiently by antithrombin. Glycosylation patterns were similar to plasma-derived FIX with sialic acid content consistent with the literature reports of good pharmacokinetic performance. These studies have shown that trenonacog alfa is a highly pure product with a primary sequence and posttranslational modifications consistent with the common Thr-148 polymorphism of plasma-derived FIX. Hindawi Publishing Corporation 2016 2016-02-21 /pmc/articles/PMC4779506/ /pubmed/26997955 http://dx.doi.org/10.1155/2016/7678901 Text en Copyright © 2016 Dougald M. Monroe et al. https://creativecommons.org/licenses/by/4.0/ This is an open access article distributed under the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited. |
spellingShingle | Research Article Monroe, Dougald M. Jenny, Richard J. Van Cott, Kevin E. Buhay, Shelly Saward, Laura L. Characterization of IXINITY® (Trenonacog Alfa), a Recombinant Factor IX with Primary Sequence Corresponding to the Threonine-148 Polymorph |
title | Characterization of IXINITY® (Trenonacog Alfa), a Recombinant Factor IX with Primary Sequence Corresponding to the Threonine-148 Polymorph |
title_full | Characterization of IXINITY® (Trenonacog Alfa), a Recombinant Factor IX with Primary Sequence Corresponding to the Threonine-148 Polymorph |
title_fullStr | Characterization of IXINITY® (Trenonacog Alfa), a Recombinant Factor IX with Primary Sequence Corresponding to the Threonine-148 Polymorph |
title_full_unstemmed | Characterization of IXINITY® (Trenonacog Alfa), a Recombinant Factor IX with Primary Sequence Corresponding to the Threonine-148 Polymorph |
title_short | Characterization of IXINITY® (Trenonacog Alfa), a Recombinant Factor IX with Primary Sequence Corresponding to the Threonine-148 Polymorph |
title_sort | characterization of ixinity® (trenonacog alfa), a recombinant factor ix with primary sequence corresponding to the threonine-148 polymorph |
topic | Research Article |
url | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4779506/ https://www.ncbi.nlm.nih.gov/pubmed/26997955 http://dx.doi.org/10.1155/2016/7678901 |
work_keys_str_mv | AT monroedougaldm characterizationofixinitytrenonacogalfaarecombinantfactorixwithprimarysequencecorrespondingtothethreonine148polymorph AT jennyrichardj characterizationofixinitytrenonacogalfaarecombinantfactorixwithprimarysequencecorrespondingtothethreonine148polymorph AT vancottkevine characterizationofixinitytrenonacogalfaarecombinantfactorixwithprimarysequencecorrespondingtothethreonine148polymorph AT buhayshelly characterizationofixinitytrenonacogalfaarecombinantfactorixwithprimarysequencecorrespondingtothethreonine148polymorph AT sawardlaural characterizationofixinitytrenonacogalfaarecombinantfactorixwithprimarysequencecorrespondingtothethreonine148polymorph |