99m Tc(N)-DBODC(5) is the lead compound of a new series of monocationic 99m Tc(N)-based potential myocardial imaging agents that exhibit original biodistribution properties. This study was addressed to elucidate the mechanisms of distribution, retention, and elimination of this promising 99m Tc(N)-agent. Methods: The sex-related in vitro and in vivo stability and the subcellular distribution of 99m Tc(N)-DBODC(5) were investigated. Studies were performed by considering binding to the serum proteins; stability in rat serum, human serum, and rat liver homogenates; and the chemical integrity of the complex after extraction from rat tissues such as heart, liver, and kidney, as well as from intestinal fluids and urine. The effect of cyclosporin A on the in vivo pharmacokinetic properties of 99m Tc(N)-DBODC(5) was also evaluated. Subcellular distribution of 99m Tc(N)-DBODC(5) in ex vivo rat heart was determined by standard differential centrifugation techniques. Results: No significant in vitro serum protein binding and no notable biotransformation of the native compound into different species by the in vitro action of the serum and liver enzymes was evidenced. In vivo experiments showed that sex affects the pharmacokinetic profile of the 99m Tc(N)-complexes including metabolism and excretion. Chromatographic profiles of 99m Tc(N)-radioactivity extracted from tissues and fluids of female rats were always coincident with the control. Conversely, a small percentage of metabolized species was detected by high-performance liquid chromatography in liver extracts of male rats. Furthermore, administration of cyclosporin A caused a significant reduction of lung, liver, and kidney washout along with a considerable variation in activity distribution in the intestinal tract in both male and female rats, thus indicating a possible implication of Pgp transporters in determining the biologic behavior of 99m Tc(N)-DBODC(5). However, this phenomenon was more pronounced in females. Subcellular distribution studies showed that 86.3% 6 7.4% of 99m Tc(N)-DBODC(5) was localized into mitochondrial fraction as a result of the interaction with the negative membrane potential. Conclusion: Evidence showing that the new 99m Tc(N)-myocardial tracers behave as multidrug resistance-associated protein P-glycoprotein substrates, combined with their selective mitochondrial accumulation, strongly supports the possibility that diagnostic application of 99m Tc(N)-DBODC(5) can be extended to tumor imaging and noninvasive multidrug resistance studies. (Fig. 1) (DBODC 5 bis-(N-ethoxyethyl)dithiocarbamato; PNP5 5 bis-(dimethoxypropylphosphinoethyl)ethoxyethylamine, PNP3 5 bis-(dimethoxypropylphosphinoethyl)methoxyethylamine) exhibited the most interesting biodistribution properties (3). In particular, animal studies of [ 99m Tc(N)(DBODC)(PNP5)] 1 (abbreviated 99m Tc(N)-DBODC(5)) revealed high and persistent myocardial uptake and rapid blood, lung, and liver clearance yielding high-quality images of the heart as early as 20 to 30 min after injection (4,5).S...

(99m)Tc(N)-DBODC5 is a cationic mixed compound under clinical investigation as potential myocardial imaging agent. In spite of this, analogously to the other cationic (99m)Tc-agents, presents a relatively low first-pass extraction. Thus, modification of (99m)Tc(N)-DBODC(5) direct to increase its first-pass extraction keeping unaltered the favorable imaging properties would be desirable. This work describes the synthesis and biological evaluation of a series of novel cationic (99m)Tc-nitrido complexes, of general formula [(99m)TcN(DTC-Ln)(PNP)](+) (DTC-Ln= alicyclic dithiocarbamates; PNP = diphosphinoamine), as potential radiotracers for myocardial perfusion imaging. The synthesis of cationic (99m)Tc-(N)-complexes were accomplished in two steps. Biodistribution studies were performed in rats and compared with the distribution profiles of (99m)Tc(N)-DBODC5 and (99m)Tc-Sestamibi. The metabolisms of the most promising compounds were evaluated by HPLC methods. Biological studies revealed that most of the complexes have a high initial and persistent heart uptake with rapid clearance from nontarget tissues. Among tested compounds, 2 and 12 showed improved heart uptake with respect to the gold standard (99m)Tc-complexes with favorable heart-to-liver and slightly lower heart-to-lung ratios. Chromatographic profiles of (99m)Tc(N)-radioactivity extracted from tissues and fluids were coincident with the native compound evidencing remarkable in vivo stability of these agents. This study shows that the incorporation of alicyclic dithiocarbamate in the [(99m)Tc(N)(PNP)](+) building block yields to a significant increase of the heart uptake at early injection point suggesting that the first-pass extraction fraction of these novel complexes may be increased with respect to the other cationic (99m)Tc-agents keeping almost unaltered the favorable target/nontarget ratios.

A general procedure is presented for the preparation of a new class of nitrido asymmetrical Tc-99m complexes containing two different bidentate ligands bound to the same [Tc(N)]2+ core that could be used to design either essential or target specific imaging agents. This procedure is based on the chemical properties of a new monosubstituted [Tc(N)(R2PS)Cl(PPh3)] species composed of a TcN multiple bond and an ancillary phosphine thiol ligand (R2PSH). This intermediate readily reacts with bidentate mononegative ligands (S--Y) containing soft pi-donor coordinating atoms to give neutral pentacoordinate asymmetrical complexes of the type [Tc(N)(R2PS)(S--Y)]. The ability of several bidentate ligands containing different combination of heteroatoms (S, N, O) to form complexes with the [Tc(N)(R2PS)]+ building block was investigated. It was found that mononegative dithiocarbamate (DTC) or cysteine carboxyl derivate ligands promptly react with the monosubstituted species to form the final mixed compound in high yield. Preliminary biodistribution data in rats of some representative [Tc(N)(R2PS)(DTC)] compounds revealed an interesting initial brain uptake (in the range 0.20 +/- 0.01% ID/g and 0.91 +/- 0.06% ID/g), indicating their ability to cross in and out of the intact BBB. In these complexes the dithiocarbamate, or more generally the bidentate ligand (S--Y), can be designed to carry a functional group or a bioactive molecule, which could be involved in a trapping mechanism to increase brain retention for longer time intervals. These results could be conveniently utilized to devise a new procedure for the production of a novel class of brain perfusion and/or brain receptor imaging agents.

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