N,N-bis(2-mercaptoethyl)methylamine: a new coligand for Tc-99m labeling of hydrazinonicotinamide peptides

Hydrazinonicotinamide (HYNIC) forms stable coordination complexes with Tc-99m when reacted with Tc(V)oxo species such as Tc-mannitol or other Tc-polyhydric complexes. However, radio-HPLC of Tc-For-MLFK-HYNIC] labeled via Tc-polyhydric ligands demonstrated multiple radiochemical species each with unique biodistribution patterns. This is likely due to the fact that Tc can bind to the hydrazino moiety, as well as polyhydric ligands, in a variety of coordination geometries. Tridentate ligands, such as bis(mercaptoethyl)methylamine (NS2), may constrain the possible coordination geometries and improve overall stability. To investigate this, we synthesized NS2, converted the Tc-mannitol-For-MLFK-HYNIC] to the corresponding NS2-containing complex Tc-NS2-For-MLFK-HYNIC], and compared its infection imaging and biodistribution properties with Tc-mannitol-For-MLFK-HYNIC]. Conversion to the NS2 complex was confirmed by HPLC which showed a single unique hydrophobic species with retention time greater than the Tc-mannitol-For-MLFK-HYNIC] complex. Imaging experiments with both preparations were performed in rabbits with E. coli infections in the left thigh. Tissue radioactivity measurements demonstrated that compared to Tc-mannitol-peptide, accumulation of Tc-NS2-peptide was lower in blood, heart, and normal muscle and higher in spleen, infected muscle, and pus (p < 0.01). These results indicate that the Tc-NS2-peptide complex is chemically more homogeneous and exhibits improved infection localization and biodistribution properties. In an effort to model the interactions of the metal-HYNIC core with NS2 and related ligand types, the reactions of ReCl3(NNC5H4NH)(NHNC5H4N)] and 99TcCl3(NNC5H4NH)(NHNC5H4N)], effective structural analogues for the M(NNC5H4NH(x))2] core, with NS2, C5H3N-2,6-(CH2SH)2, O(CH2CH2SH)2, and S(CH2CH2SH)2 were investigated and the compounds MCH3N(CH2CH2S)2](NNC5H4N)(NHNC5H4N] (M = 99Tc (5a), Re (5b)), ReC5H3N-2,6-(CH2S)2](NNC5H4N)(NHNC5H4N)].CH2Cl2.0.5MeOH (7), ReSCH2CH2)2O] (NNC5H4N)(NHNC5H4N)] (8), and Re(SCH2CH2)2S](NNC5H4NH)(NHNC5H4N)]Cl (9) were isolated. Similarly, the reaction of ReCl3(NNC5H4NH)(NHNC5H4N)] with the bidentate ligands pyridine-2-methanethiol and 3-(trimethlysilyl)pyridine-2-thiol led to the isolation of ReCl(C5H4N-2-CH2S) (NNC5H4N)(NHNC5H4N)] (10) and Re(2-SC5H3N-3-SiMe3)2 (NNC5H4N)(NHNC5H4N)] (11), respectively, while reaction with N-methylimidazole-2-thiol yielded the binuclear complex Re(OH)Cl(SC3H2N2CH3)2(NNC5H4N)2 (NHNC5H4N)2] (12). The analogous metal-(HYNIC-OH) precursor, ReCl3NNC5H3NH(CO2R)] NHNC5H3N(CO2R)]] (R = H, 13a; R = CH3, 13b) has been prepared and coupled to lysine to provide RCl3NNC5H3NH(CONHCH2CH2CH2CH2CH(NH2)CO2H)] NHNC5H3NH(CONHCH2CH2CH2CH2CH(NH2)CO2H)]].2HCl (14.2HCl), while the reaction of the methyl ester 13b with 2-mercaptopyridine yields Re(2-SC5H4N)2NNC5H3N(CO2Me)]NHNC5H3N(CO2Me)]] (15). While the chemical studies confirm the robustness of the M-HYNIC core (M = Tc, Re) and its persistence in ligand substitution reactions at adjacent coordination sites of the metal, the isolation of oligomeric structures and the insolubility of the peptide conjugates of 13, 14, and 15 underscore the difficulty of characterizing these materials on the macroscopic scale, an observation relevant to the persistent concerns with reagent purity and identity on the tracer level.