A single-domain antibody that blocks factor VIIa activity in the absence but not presence of tissue factor.
J Thromb Haemost
; 17(12): 2035-2046, 2019 12.
Article
in En
| MEDLINE
| ID: mdl-31423724
ABSTRACT
BACKGROUND:
Activated factor VII (FVIIa) is pertinent to the initiation of blood coagulation. Proteolytic and amidolytic activity of FVIIa are greatly enhanced by its cofactor, tissue factor (TF).OBJECTIVE:
We aimed to generate a single-domain antibody (sdAb) that recognizes free FVIIa rather than TF-bound FVIIa.METHODS:
A llama-derived phage library was used to screen for anti-FVIIa sdAbs.RESULTS:
One sdAb, KB-FVIIa-004, bound to FVIIa, but not to its precursor FVII or to homologous proteins (prothrombin, factor X, or their activated derivatives). FVIIa amidolytic activity was inhibited by KB-FVIIa-004 (Ki = 28-45 nM) in a competitive manner. KB-FVIIa-004 also inhibited FVIIa-mediated FX activation (Ki = 26 nM). In contrast, KB-FVIIa-004 was inefficient in prolonging the clotting time of the prothrombin time-test, which was prolonged by a maximum of 10 s at high sdAb concentrations (10 µM). Furthermore, FVIIa/TF amidolytic activity or FVIIa/TF-mediated FX activation remained unaffected up to a 50-fold to 1000-fold molar excess of KB-FVIIa-004. These data suggest that KB-FVIIa-004 loses its inhibitory activity in the presence of TF. A KB-FVIIa-004/albumin fusion-protein (004-HSA) was generated for in vivo testing. By using 004-HSA, we observed that this sdAb blocked the therapeutic capacity of FVIIa to correct bleeding in FVIII-deficient mice.DISCUSSION:
This observation is compatible with the view that FVIIa functions independently of TF under these conditions. In conclusion, we have generated a sdAb that specifically blocks TF-independent activity of FVIIa. This antibody can be used to gain insight into the roles of TF-bound and TF-free FVIIa.Key words
Full text:
1
Collection:
01-internacional
Database:
MEDLINE
Main subject:
Blood Coagulation
/
Thromboplastin
/
Factor VIIa
/
Single-Domain Antibodies
/
Anticoagulants
Type of study:
Prognostic_studies
Limits:
Animals
/
Female
/
Humans
/
Male
Language:
En
Journal:
J Thromb Haemost
Year:
2019
Document type:
Article