l- and d‑Proline Thiosemicarbazone Conjugates: Coordination Behavior in Solution and the Effect of Copper(II) Coordination on Their Antiproliferative Activity

Two enantiomerically pure thiosemicarbazone–proline conjugates with enhanced aqueous solubility, namely, 2-hydroxy-3-methyl-(S)-pyrrolidine-2-carboxylate-5-methylbenzaldehyde thiosemicarbazone [l-Pro-STSC or (S)-H2L] and 2-hydroxy-3-methyl-(R)-pyrrolidine-2-carboxylate-5-methylbenzaldehyde thiosemicarbazone [d-Pro-STSC or (R)-H2L] have been synthesized and characterized by elemental analysis, spectroscopic methods (UV–vis and 1H and 13C NMR), and electrospray ionization mass spectrometry. The metal complexation behavior of l-Pro-STSC, stoichiometry, and thermodynamic stability of iron­(II), iron­(III), copper­(II), and zinc­(II) complexes in 30% (w/w) dimethyl sulfoxide/H2O solvent mixture have been studied by pH-potentiometric, UV–vis-spectrophotometric, circular dichroism, electron paramagnetic resonance, 1H NMR spectroscopic, and spectrofluorimetric measurements. By the reaction of CuCl2·2H2O with (S)-H2L and (R)-H2L, respectively, the complexes [Cu­[(S)-H2L]­Cl]Cl and [Cu­[(R)-H2L]­Cl]Cl have been prepared and comprehensively characterized. An X-ray diffraction study of [Cu­[(R)-H2L]­Cl]Cl showed the formation of a square-planar copper­(II) complex, which builds up stacks with interplanar separation of 3.3 Å. The antiproliferative activity of two chiral ligands and their corresponding copper­(II) complexes has been tested in two human cancer cell lines, namely, SW480 (colon carcinoma) and CH1 (ovarian carcinoma). The thiosemicarbazone–proline conjugates l- and d-Pro-STSC show only moderate cytotoxic potency with IC50 values of 62 and 75 μM, respectively, in CH1 cells and >100 μM in SW480 cells. However, the corresponding copper­(II) complexes are 13 and 5 times more potent in CH1 cells, based on a comparison of IC50 values, and in SW480 cells the increase in the antiproliferative activity is even higher. In both tested cell lines, l-Pro-STSC as well as its copper­(II) complex show slightly stronger antiproliferative activity than the compounds with a d-Pro moiety, yielding IC50 values of 4.6 and 5.5 μM for [Cu­(l-Pro-STSC)­Cl]Cl in CH1 and SW480 cells, respectively.