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Thrombin generation assay using factor XIa to measure factors VIII and IX and their glycoPEGylated derivatives is robust and sensitive.
J Thromb Haemost. 2015 Nov; 13(11):2041-52.JT

Abstract

BACKGROUND

Conventional coagulation factor assays are associated with certain limitations, as they do not always reflect the clinical heterogeneity of bleeding in hemophilic patients or correctly reflect the individual patient response to treatment with bypassing agents or novel factor concentrates. The thrombin generation assay (TGA) is currently being assessed as a possible method for characterizing bleeding phenotypes in individuals with hemophilia.

OBJECTIVES

This study assessed the robustness and sensitivity of the TGA for measuring the activity of recombinant factor VIII (rFVIII), recombinant factor IX (rFIX) and their glycoPEGylated derivatives, N8-GP and N9-GP, in vitro.

METHODS

Factor-deficient plasma was spiked with 0.13-130 IU dL(-1) rFVIII or N8-GP (hemophilia A [HA] plasma), or rFIX or N9-GP (hemophilia B [HB] plasma). A calibrated automated thrombogram triggered with tissue factor (TF) or activated FXI (FXIa) was used to measure thrombin generation over time. Endogenous thrombin potential, peak thrombin, velocity index, lag time and time to peak thrombin were analyzed.

RESULTS

FXIa-triggered assays were not affected by glycoPEGylation and were sufficiently sensitive to differentiate between spiked samples mimicking severe and moderate HB and HA; TF-triggered assays were not sufficiently sensitive for this distinction in HA. Both FXIa-triggered and TF-triggered assays had an acceptable level of variability (≤ 20%), although TF-triggered assays were associated with greater variability.

CONCLUSIONS

FXIa-triggered TGA reactions produced more robust and sensitive results than TF-triggered TGA reactions, and have the potential for use in monitoring patients treated with glycoPEGylated or non-PEGylated coagulation factor concentrates. These promising results merit confirmation with clinical samples to correlate in vitro and in vivo data.

Authors+Show Affiliations

Hemophilia Biology, Novo Nordisk A/S, Måløv, Denmark.Hemophilia Biology, Novo Nordisk A/S, Måløv, Denmark.Hemophilia Biology, Novo Nordisk A/S, Måløv, Denmark.Hemophilia Biology, Novo Nordisk A/S, Måløv, Denmark.Hemophilia Biology, Novo Nordisk A/S, Måløv, Denmark.

Pub Type(s)

Comparative Study
Evaluation Study
Journal Article
Research Support, Non-U.S. Gov't

Language

eng

PubMed ID

26340413

Citation

Waters, E K., et al. "Thrombin Generation Assay Using Factor XIa to Measure Factors VIII and IX and Their glycoPEGylated Derivatives Is Robust and Sensitive." Journal of Thrombosis and Haemostasis : JTH, vol. 13, no. 11, 2015, pp. 2041-52.
Waters EK, Hilden I, Sørensen BB, et al. Thrombin generation assay using factor XIa to measure factors VIII and IX and their glycoPEGylated derivatives is robust and sensitive. J Thromb Haemost. 2015;13(11):2041-52.
Waters, E. K., Hilden, I., Sørensen, B. B., Ezban, M., & Holm, P. K. (2015). Thrombin generation assay using factor XIa to measure factors VIII and IX and their glycoPEGylated derivatives is robust and sensitive. Journal of Thrombosis and Haemostasis : JTH, 13(11), 2041-52. https://doi.org/10.1111/jth.13134
Waters EK, et al. Thrombin Generation Assay Using Factor XIa to Measure Factors VIII and IX and Their glycoPEGylated Derivatives Is Robust and Sensitive. J Thromb Haemost. 2015;13(11):2041-52. PubMed PMID: 26340413.
* Article titles in AMA citation format should be in sentence-case
TY - JOUR T1 - Thrombin generation assay using factor XIa to measure factors VIII and IX and their glycoPEGylated derivatives is robust and sensitive. AU - Waters,E K, AU - Hilden,I, AU - Sørensen,B B, AU - Ezban,M, AU - Holm,P K, Y1 - 2015/10/01/ PY - 2014/12/17/received PY - 2015/08/26/accepted PY - 2015/9/5/entrez PY - 2015/9/5/pubmed PY - 2016/9/17/medline KW - blood coagulation tests KW - factor VIII KW - factor XIa KW - hemophilia A KW - hemophilia B SP - 2041 EP - 52 JF - Journal of thrombosis and haemostasis : JTH JO - J Thromb Haemost VL - 13 IS - 11 N2 - BACKGROUND: Conventional coagulation factor assays are associated with certain limitations, as they do not always reflect the clinical heterogeneity of bleeding in hemophilic patients or correctly reflect the individual patient response to treatment with bypassing agents or novel factor concentrates. The thrombin generation assay (TGA) is currently being assessed as a possible method for characterizing bleeding phenotypes in individuals with hemophilia. OBJECTIVES: This study assessed the robustness and sensitivity of the TGA for measuring the activity of recombinant factor VIII (rFVIII), recombinant factor IX (rFIX) and their glycoPEGylated derivatives, N8-GP and N9-GP, in vitro. METHODS: Factor-deficient plasma was spiked with 0.13-130 IU dL(-1) rFVIII or N8-GP (hemophilia A [HA] plasma), or rFIX or N9-GP (hemophilia B [HB] plasma). A calibrated automated thrombogram triggered with tissue factor (TF) or activated FXI (FXIa) was used to measure thrombin generation over time. Endogenous thrombin potential, peak thrombin, velocity index, lag time and time to peak thrombin were analyzed. RESULTS: FXIa-triggered assays were not affected by glycoPEGylation and were sufficiently sensitive to differentiate between spiked samples mimicking severe and moderate HB and HA; TF-triggered assays were not sufficiently sensitive for this distinction in HA. Both FXIa-triggered and TF-triggered assays had an acceptable level of variability (≤ 20%), although TF-triggered assays were associated with greater variability. CONCLUSIONS: FXIa-triggered TGA reactions produced more robust and sensitive results than TF-triggered TGA reactions, and have the potential for use in monitoring patients treated with glycoPEGylated or non-PEGylated coagulation factor concentrates. These promising results merit confirmation with clinical samples to correlate in vitro and in vivo data. SN - 1538-7836 UR - https://www.unboundmedicine.com/medline/citation/26340413/Thrombin_generation_assay_using_factor_XIa_to_measure_factors_VIII_and_IX_and_their_glycoPEGylated_derivatives_is_robust_and_sensitive_ L2 - https://doi.org/10.1111/jth.13134 DB - PRIME DP - Unbound Medicine ER -