Influence of the denticity of ligand systems on the in vitro and in vivo behavior of 99mTc(I)-tricarbonyl complexes:: A hint for the future functionalization of biomolecules

Functionalization of biologically relevant molecules for the labeling with the novel fac-[Tc-99m(OH2)(3)(CO)(3)](+) precursor has gained considerable attention recently. Therefore, we tested seven different tridentate (histidine L-1, iminodiacetic acid L-2, N-2-picolylamineacetic acid L-3, N,N-2-picolylaminediacetic acid L-4) and bidentate (histamine L-5, a-picolinic acid L-6, 2,4-dipicolinic acid L-7) ligand systems, with the potential to be bifunctionalized and attached to a biomolecule. The ligands allowed mild radiolabeling conditions with fac-[Tc-99m(OH2)(3)(CO)(3)](+) (30 min, 75 degrees C). The ligand concentrations necessary to obtain yields of >95% of the corresponding organometallic complexes 1-7 ranged from 10(-6) to 10(-4) M. Complexes of the general formula fac-[(TcL)-Tc-99m(CO)(3)] (L = tridentate ligand) and fac-[Tc-99m(OH2)L'(CO)(3)] (L' = bidentate ligand), respectively, were produced. Challenge studies with cysteine and histidine revealed significant displacement of the ligands in complexes 5-7 but only little exchange with complexes 1-4 after 24 h at 37 degrees C in PBS buffer. However, no decomposition to (TCO4-)-T-99m was observed under these conditions. All complexes showed a hydrophilic character (log P-o/w values ranging from -2.12 to 0.32). Time-dependent FPLC analyses of compounds 1-7 incubated in human plasma at 37 degrees C showed again no decomposition to (TCO4-)-T-99m after 24 h at 37 degrees C. However, the complexes with bidentate ligands (5-7) became almost completely protein bound after 60 min, whereas the complexes with tridentate coordinated ligands (1-4) showed no reaction with serum proteins. The compounds were tested for their in vivo stability and the biodistribution characteristics in BALB/c mice. The complexes with tridentate coordinated ligand systems (1-4) revealed generally a good and fast clearance from all organs and tissues. On the other hand, the complexes with only bidentate coordinated ligands (5-7) showed a significantly higher retention of activity in the liver, the kidneys, and the blood pool. Detailed radiometric analyses of murine plasma samples, 30 min p.i. of complex fac-[(TcL1)-Tc-99m(CO)(3)], 1, revealed almost no reaction of the radioactive complex with the plasma proteins. By contrast, in plasma samples of mice, which were injected with complex fac-[Tc-99m(OH2)L-5(CO)(3)](+), 5, the entire radioactivity coeluded with the proteins. On the basis of these in vitro and in vivo experiments, it appears that functionalization of biomolecules with tridentate-chelating ligand systems is preferable for the labeling with fac-[99mTc(OH2)(3)(CO)(3)](+), since this will presumably result in radioactive bioconjugates with better pharmacokinetic profiles.