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.

本研究合成了两种具有增强水溶性的对映纯缩氨基硫脲-脯氨酸缀合物，即2-羟基-3-甲基-(S)-吡咯烷-2-羧酸酯-5-甲基苯甲醛缩氨基硫脲[l-Pro-STSC或(S)-H2L]与2-羟基-3-甲基-(R)-吡咯烷-2-羧酸酯-5-甲基苯甲醛缩氨基硫脲[d-Pro-STSC或(R)-H2L]，并通过元素分析、光谱手段（紫外-可见分光光度法、1H核磁共振谱与13C核磁共振谱）以及电喷雾电离质谱法对其进行了表征。本研究通过pH电位法、紫外-可见分光光度法、圆二色谱法、电子顺磁共振波谱法、1H核磁共振谱法以及荧光光谱法，探究了l-Pro-STSC的金属配位行为、化学计量比，以及其二价铁、三价铁、二价铜与二价锌配合物在30%（w/w）二甲基亚砜/水混合溶剂中的热力学稳定性。分别将二水合氯化铜（CuCl2·2H2O）与(S)-H2L、(R)-H2L反应，制备得到配合物[Cu[(S)-H2L]Cl]Cl与[Cu[(R)-H2L]Cl]Cl，并对其进行了全面表征。对[Cu[(R)-H2L]Cl]Cl的X射线衍射研究表明，其形成了平面正方形二价铜配合物，该配合物通过层间距为3.3 Å的分子堆积形成堆叠结构。针对两种手性配体及其对应的二价铜配合物的抗增殖活性，本研究在两种人类癌细胞系——SW480（结肠癌细胞）与CH1（卵巢癌细胞）中进行了测试。缩氨基硫脲-脯氨酸缀合物l-Pro-STSC与d-Pro-STSC仅表现出中等的细胞毒性效力：在CH1细胞中，二者的半数抑制浓度（IC50）分别为62 μM与75 μM，在SW480细胞中则均大于100 μM。然而，对应的二价铜配合物在CH1细胞中的细胞毒性效力分别提升了13倍与5倍；在SW480细胞中，其抗增殖活性的提升更为显著。在两种测试的细胞系中，l-Pro-STSC及其二价铜配合物的抗增殖活性均略高于带有d-Pro基团的对应化合物：其中[Cu(l-Pro-STSC)Cl]Cl在CH1与SW480细胞中的IC50值分别为4.6 μM与5.5 μM。