Recombinant coagulation factor VIIa labelled with the fac‐99 mTc(CO)3‐core: synthesis and in vitro evaluation of a putative new radiopharmaceutical for imaging in acute bleeding lesion

Coagulation in blood is initiated when coagulation factor VII (FVII) binds to exposed TF and is activated to FVIIa, and the TF/FVIIa complex may therefore provide a marker of vascular injury potentially applicable in diagnostic imaging of acute gastrointestinal (GI) bleeding. Methods: Recombinant FVIIa (rFVIIa) was radiolabeled with technetium‐99 m in a direct labeling reaction using the ‘carbonyl approach’ using the IsoLink^® carbonyl labeling agent. The properties of ^{99 m}Tc(CO)₃‐rFVIIa complex was analyzed by TCA precipitation, HPLC and FVIIa functional integrity was tested in in vitro assays. Results: Labeling of rFVIIa was possible without tagging with a chelater. Incorporation of radioactivity depended strongly on rFVIIa concentration and temperature. More than 95% incorporation was achieved after 30 min at 45°C with 0.76 mg/ml rFVIIa. ^{99 m}Tc(CO)₃‐rFVIIa was obtained in 46% radiochemical yield and in >95% radiochemical purity. Pull down experiments showed that the biological activity (binding to tissue factor and to anti‐FVII antibody) of the radiolabelled product remained intact in the formulation mixture as well as in human serum. By computer modeling analysis, two candidate sites for stabilizing the ^{99 m}Tc(CO)‐ligand structure in FVIIa were identified. Conclusion: Radiolabelled rFVIIa derivatives may represent a novel tool for the diagnosis of acute gastrointestinal bleeding lesions. Copyright © 2010 John Wiley & Sons, Ltd.