Monoclonal anti-human factor VII antibodies. Detection in plasma of a second protein antigenically and genetically related to factor VII.

Abstract

Several murine monoclonal anti-human Factor VII antibodies were produced using hybridoma technology. Two noncompetitive monoclonal antibodies were used to examine by Western blotting the Factor VII cross-reactive material (CRM) in normal human plasma and three commercially available congenitally Factor VIIdeficient plasmas, and to construct a facile "sandwich" immunoassay for plasma Factor VII. A second, previously undescribed, form of Factor VII CRM was detected in human plasma, which on Western blotting stained with an apparent intensity 5-8% that of Factor VII. This glycoprotein, tentatively called VII*, has a molecular weight 4,500 D less than Factor VII, lacks detectable Factor VII functional activity, does not bind to barium citrate, and is not recognized by a monoclonal antibody that recognizes Factor VII but not a-chymotrypsin-treated Factor VII. VII* was not proteolytically produced from Factor VII during in vitro coagulation or after infusion of human Factor VII into rabbits. As determined by Western blotting, the human hepatoma cell line, HepG2, cultured in the presence of vitamin K, secreted relatively greater levels of VII* in proportion to VII (75%) than that found in human plasma. Warfarin treatment of HepG2 cells decreased the quantity of VII secreted by 77%, whereas it only inhibited the secretion of VII* by 14%. Immunologic studies of the plasmas from a patient on chronic warfarin therapy and an individual given a short course of high dose warfarin therapy corroborated the in vitro synthetic studies obtained with HepG2 cells. The data are consistent with the production of VII* by posttranslational, proteolytic, modification of VII, that, at least in the HepG2 cells studied, occurs intracellularly. However, other mechanisms for the production of VII*, in particular, alternative RNA splicing of the transcript from a single gene, cannot be excluded.

Mechanism underlying factor VII deficiency in Jewish populations with the Ala244Val mutation

“…A purified murine monoclonal antibody (MC1476), which recognizes an epitope in the amino terminal region of factor VII (Broze, 1985), was used in immunoprecipitation experiments. To immunoprecipitate potential degradation products of factor VII, we used a commercially available rabbit polyclonal antibody to factor VII (Celsus, Cincinnati, Ohio).…”

The P303T mutation in the human factor VII (FVII) gene alters the conformational state of the enzyme and causes a severe functional deficiency

“…Recombinant FVIIWT and FVII303T were isolated from cell supernatants and concentrated using either a previously characterized anti-FVII monoclonal antibody, reacting with the Gla domain (Broze et al, 1985), or an anti-FVII monoclonal antibody that recognizes an epitope on the light chain of FVII in the presence of calcium ions (Clone no. HVII-1; Sigma-Aldrich, St Louis, MO, USA), covalently coupled to activated agarose (Affi-Gel 10; Bio-Rad, Santa Barbara, CA, USA).…”

Activation of Cancer Cell Migration and Invasion by Ectopic Synthesis of Coagulation Factor VII

“…By Northern blotting of human tissue mRNA, FVII gene expression was only detected in the liver (20), confirming a selective hepatic synthesis under normal conditions. However, fVII is expressed in some hepatocellular carcinoma cells (24,25), and this ectopic fVII synthesis has been proposed to trigger liver cancer-specific invasion activity mediated by binding of fVIIa to TF pathway inhibitor-2 (26). Among cancers other than hepatocellular carcinoma, TF/fVIIa complex formation has been observed at the invasive edge of bladder cancer (27), ovarian cancer (28), and laryngeal carcinoma tissues (29).…”