TUMOR IMAGING WITH TC-99M-LABELED HYDRAZINONICOTINAMIDE-FAB' CONJUGATES

This study compares the in vivo properties of direct Versus indirect T c-99m-labeling for two Fab' fragments from antibodies that recognize t umor-associated antigens. Methods: Fab' fragments of two IgG2a monoclo nal antibodies were either radiolabeled directly or via the linker bro moacetyl hydrazinonicotinamide hydrobromide (BAHNH) conjugated site sp ecifically at protein thiols. A thiol assay was used to determine the number of thiols in the Fab' and to monitor their consumption during c onjugation with BAHNH. Both preparations were labeled to > 95% incorpo ration of Tc-99m, with the isotope tracking the single 50 kD absorbanc e peak seen on size-exclusion HPLC. The labeled preparations were test ed in tumor-bearing and control mice, with dissections at 4 and 24 hr and gamma scintigraphy of the tumor-bearing mice. Results: The major d ifference between the two labeled preparations for either antibody fra gment was the greater accumulation of isotope in the tumor for the ind irectly labeled preparations, This increase ranged from 1.5- and 2.7-f old at 4 hr to 2.6- and 3.2-fold at 24 hr for the two antibodies, resp ectively. Since blood clearance was similar for the two labeling metho ds, the higher tumor accumulation with the indirectly labeled fragment s resulted in higher tumor to blood ratios. Tumors could be imaged wit h both antibodies with either type of labeling with greater clarity an d sensitivity at the 24 hr time point. Conclusion: While both labeling methods resulted in tumor detection through imaging, the images obtai ned with the indirectly labeled antibody fragments were more easily vi sualized due to the combination of higher radioisotope accumulation in the tumor and similar brood clearances compared to the direct labeled fragment.