A novel tetraazamacrocycle bearing a thiol pendant arm for labeling biomolecules with radiolanthanides

Abstract

The novel tetraazamacrocycle 10-(2-sulfanylethyl)-1,4,7,10-tetraazacyclododecane-1,4,7-triacetic acid (H₄DO3ASH) was synthesized and characterized by multinuclear NMR spectroscopy, 2D NMR techniques and mass spectrometry. The protonation constants of H₄DO3ASH were determined by potentiometry at 25 °C in 0.1 M KCl ionic strength, and the protonation sequence was assigned based on ¹H- and ¹³C-NMR titrations. The stability constants of the DO3ASH complexes with Ce³⁺, Sm³⁺ and Ho³⁺ have been determined by potentiometry and UV-Vis spectroscopy. They are very similar, comprising a narrow range (logK_ML = 21.0–22.0). UV-Vis spectrophotometric data on Ce³⁺–DO3ASH and relaxivity measurements on the Gd³⁺–DO3ASH complex suggest that the thiol group does not coordinate to the metal, even in its deprotonated form. For labeling with radioactive lanthanides(III), various conditions were tested and both complexes, ¹⁵³Sm/¹⁶⁶Ho–DO3ASH, were obtained in quantitative yield (> 98%) at pH = 6. At room temperature, formation kinetics were faster for the ¹⁵³Sm than for the ¹⁶⁶Ho complex (5 vs. 60 min, respectively, needed for complete labeling). The stability of these hydrophilic complexes (¹⁵³Sm, logD = −2.1; ¹⁶⁶Ho, logD = −1.6) has been studied in different buffers, in human serum and in the presence of excess of cysteine and glutathione. ¹⁵³Sm–DO3ASH has shown a high stability under these conditions and a relatively low protein binding (2.1%), while ¹⁶⁶Ho–DO3ASH was less stable, including in the presence of cysteine and glutathione, and had a slightly higher protein binding (6.7%). In vivo studies have been performed only for the more stable ¹⁵³Sm–DO3ASH complex and its biological profile and in vivo stability has been compared to that of ¹⁵³Sm–DO3A in the same animal model. The biodistribution profile presents a similar trend with rapid total excretion from the whole animal body, mainly via the urinary pathway. The most striking difference found is related to a slightly slower clearance of ¹⁵³Sm–DO3ASH from organs like blood, bone and muscle as compared to ¹⁵³Sm–DO3A. Additionally, the fraction of ¹⁵³Sm–DO3ASH taken by the hepatobiliar tract is also modestly higher than that of ¹⁵³Sm–DO3A.