Routine measurements of factor VIII activity and inhibitor titer in the presence of emicizumab utilizing anti-idiotype monoclonal antibodies

SummaryWe found a patient with mild hemophilia A who had no detectable factor VIII antigen (FVIII:Ag), as shown by two-site ELISA using inhibitor alloantibodies (TK). We then analyzed A2, A2/B, and C2 antigen of the patient's DDAVP-induced FVIII using several anti-FVIII monoclonal antibodies. Factor VIII activity (FVIII : C) was increased from 12 to 42 Uldl by the administration of DDAVP. The DDAVPinduced increases in the A2 and A2/B antigens were 40 and 36 Uldl, respectively. However, the increase in the C2 antigen was only 7.5 Uldl. SSCP analysis and subsequent sequencing demonstrated an Arg to Cys transition at codon 2159. The anti-FVII1:C titer of monoclonal antibody, NMC-VIII15 which recognized the C2 domain, against normal plasma was 450 Bethesda Ulmg of IgG. However, the titer against DDAVP-treated patient's plasma was only 15 Bethesda Ulmg. We also tested DDAVP-induced increase in the FVIII : Ag in another mild hemophilia A patient with the same mutation at Arg2159. Increase in his C2 antigen levels was only 19% of those in the A2 and A2/B antigen. We designate this abnormal FVIII as FVIII Ise. Our results show that a missense mutation at Arg2159 to Cys modifies the antigenicity of the C2 domain.

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Kinetic parameters of monoclonal antibodies ESH2, ESH4, ESH5, and ESH8 on coagulation factor VIII and their influence on factor VIII activity

Journal of Molecular Recognition ◽

10.1002/jmr.947 ◽

2009 ◽

Vol 22 (4) ◽

pp. 301-306 ◽

Cited By ~ 7

Author(s):

C. Egler ◽

T. Albert ◽

O. Brokemper ◽

M. Zabe-Kühn ◽

G. Mayer ◽

...

Keyword(s):

Monoclonal Antibodies ◽

Factor Viii ◽

Kinetic Parameters ◽

Coagulation Factor ◽

Coagulation Factor Viii ◽

Factor Viii Activity

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Measurement of factor VIII activity of B-domain deleted recombinant factor VIII

Seminars in Hematology ◽

10.1053/shem.2001.25889 ◽

2001 ◽

Vol 38 (2, Suppl 4) ◽

pp. 13-23 ◽

Cited By ~ 16

Author(s):

M. Mikaelsson ◽

U. Oswaldsson ◽

M. A. Jankowski

Keyword(s):

Factor Viii ◽

Recombinant Factor Viii ◽

Factor Viii Activity

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Thrombelastography during and after Elective Abdominal Surgery

Thrombosis and Haemostasis ◽

10.1055/s-0038-1646708 ◽

1978 ◽

Vol 39 (02) ◽

pp. 488-495 ◽

Cited By ~ 13

Author(s):

J M Butler

Keyword(s):

Abdominal Surgery ◽

Factor Viii ◽

Whole Blood ◽

Dominant Factor ◽

Fibrinogen Concentration ◽

Factor Viii Activity ◽

Elective Abdominal Surgery

SummaryThrombelastography has been performed on recalcified whole blood from 50 patients before, during and after elective abdominal surgery. The characteristic changes of the thrombelastographic indices r, k and mA are described.During operation r and k shortened, but no change in mA was observed. This response was in part associated with an increase in factor VIII activity. Following operation, while r time was somewhat shortened, much more marked changes in k and mA were evident. Increasing fibrinogen concentration was the dominant factor in determining the post-operative changes in the thrombelastograph.

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Heparin, Protamine and Polybrene

Thrombosis and Haemostasis ◽

10.1055/s-0038-1656300 ◽

1965 ◽

Vol 13 (02) ◽

pp. 550-560 ◽

Cited By ~ 2

Author(s):

Anthony Britten

Keyword(s):

Factor Viii ◽

Incubation Mixture ◽

Factor V ◽

Factor Viii Activity ◽

Rapid Loss

SummaryThe effects of incubating heparin, protamine or Polybrene with plasma were studied. All three drugs cause rapid loss of factor V from decalcified plasma, while Polybrene also accelerates the loss of factor VIII activity. These changes are related to temperature, the period of incubation and the dose of the drug used, and can be partially prevented by inclusion of neutralizing doses of the appropriate antagonist in the incubation mixture.The implications of these findings are discussed.

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Inhibition of human coagulation factor VIII by monoclonal antibodies. Mapping of functional epitopes with the use of recombinant factor VIII fragments

Biochemical Journal ◽

10.1042/bj2630187 ◽

1989 ◽

Vol 263 (1) ◽

pp. 187-194 ◽

Cited By ~ 41

Author(s):

A Leyte ◽

K Mertens ◽

B Distel ◽

R F Evers ◽

M J M De Keyzer-Nellen ◽

...

Keyword(s):

Monoclonal Antibodies ◽

Factor Viii ◽

Factor Xa ◽

Coagulation Factor ◽

Procoagulant Activity ◽

Coagulation Factor Viii ◽

Coagulation Cascade ◽

Restriction Enzyme Digestion ◽

Proteolytic Activation

The epitopes of four monoclonal antibodies against coagulation Factor VIII were mapped with the use of recombinant DNA techniques. Full-length Factor VIII cDNA and parts thereof were inserted into the vector pSP64, permitting transcription in vitro with the use of a promoter specific for SP6 RNA polymerase. Factor VIII DNA inserts were truncated from their 3′-ends by selective restriction-enzyme digestion and used as templates for ‘run-off’ mRNA synthesis. Translation in vitro with rabbit reticulocyte lysate provided defined radiolabelled Factor VIII fragments for immunoprecipitation studies. Two antibodies are shown to be directed against epitopes on the 90 kDa chain of Factor VIII, between residues 712 and 741. The 80 kDa chain appeared to contain the epitopes of the other two antibodies, within the sequences 1649-1778 and 1779-1840 respectively. The effect of antibody binding to these sequences was evaluated at two distinct levels within the coagulation cascade. Both Factor VIII procoagulant activity and Factor VIII cofactor function in Factor Xa generation were neutralized upon binding to the region 1779-1840. The antibodies recognizing the region 713-740 or 1649-1778, though interfering with Factor VIII procoagulant activity, did not inhibit in Factor Xa generation. These findings demonstrate that antibodies that virtually inhibit Factor VIII in coagulation in vitro are not necessarily directed against epitopes involved in Factor VIII cofactor function. Inhibition of procoagulant activity rather than of cofactor function itself may be explained by interference in proteolytic activation of Factor VIII. This hypothesis is in agreement with the localization of the epitopes in the proximity of thrombin-cleavage or Factor Xa-cleavage sites.

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