Molecular Pharmacology and Pathology Program, Department of Pathology and Bosch Institute, University of Sydney, Sydney, New South Wales, Australia.
Mol Pharmacol. 2013 Dec;84(6):911-24. doi: 10.1124/mol.113.088393. Epub 2013 Oct 1.
The chelator di-2-pyridylketone 4,4-dimethyl-3-thiosemicarbazone (Dp44mT) shows potent and selective anticancer and antimetastatic activity. However, the mechanism by which it is initially transported into cells to induce cytotoxicity is unknown. Hence, the current investigation examined the cellular uptake of ¹⁴C-Dp44mT relative to two structurally related ligands, namely the aroylhydrazone ¹⁴C-pyridoxal isonicotinoyl hydrazone (¹⁴C-PIH) and the thiosemicarbazone (¹⁴C-2-benzoylpyridine 4-ethyl-3-thiosemicarbazone (¹⁴C-Bp4eT). In marked contrast to the cellular uptake of ¹⁴C-PIH and ¹⁴C-Bp4eT, which were linear as a function of concentration, ¹⁴C-Dp44mT uptake was saturable using SK-N-MC neuroepithelioma cells (Bmax, 4.28 × 10⁷ molecules of chelator/cell; and Kd, 2.45 μM). Together with the fact that ¹⁴C-Dp44mT uptake was temperature-dependent and significantly (P < 0.01) decreased by competing unlabeled Dp44mT, these observations indicated a saturable transport mechanism consistent with carrier/receptor-mediated transport. Other unlabeled ligands that shared the saturated N4 structural moiety with Dp44mT significantly (P < 0.01) inhibited ¹⁴C-Dp44mT uptake, illustrating its importance for carrier/receptor recognition. Nevertheless, unlabeled Dp44mT most markedly decreased (¹⁴C-Dp44mT uptake, demonstrating that the putative carrier/receptor shows high selectivity for Dp44mT. Interestingly, in contrast to ¹⁴C-Dp44mT, uptake of its Fe complex [Fe(¹⁴C-Dp44mT)₂] was not saturable as a function of concentration and was much greater than the ligand alone, indicating an alternate mode of transport. Studies examining the tissue distribution of ¹⁴C-Dp44mT injected intravenously into a mouse tumor model demonstrated the ¹⁴C label was primarily identified in the excretory system. Collectively, these findings examining the mechanism of Dp44mT uptake and its distribution and excretion have clinical implications for its bioavailability and uptake in vivo.
螯合剂二吡啶酮 4,4-二甲基-3-硫代缩氨基脲（Dp44mT）具有很强的选择性抗肿瘤和抗转移活性。然而，其最初被细胞摄取以诱导细胞毒性的机制尚不清楚。因此，目前的研究检测了相对于两种结构相关配体，即芳基酰肼 ¹⁴C-吡啶醛异烟酰基腙（¹⁴C-PIH）和硫代缩氨基脲（¹⁴C-2-苯甲酰基吡啶 4-乙基-3-硫代缩氨基脲（¹⁴C-Bp4eT）的¹⁴C-Dp44mT 的细胞摄取。与 ¹⁴C-PIH 和 ¹⁴C-Bp4eT 的细胞摄取呈浓度依赖性的线性形成鲜明对比的是，使用 SK-N-MC 神经上皮细胞瘤细胞，¹⁴C-Dp44mT 的摄取是饱和的（Bmax，4.28×107 个螯合剂/细胞；Kd，2.45 μM）。¹⁴C-Dp44mT 的摄取与温度有关，并且明显（P < 0.01）被未标记的 Dp44mT 竞争降低，这些观察结果表明存在一种饱和的转运机制，与载体/受体介导的转运一致。其他与 Dp44mT 共享饱和 N4 结构部分的未标记配体显著（P < 0.01）抑制了¹⁴C-Dp44mT 的摄取，说明了其对载体/受体识别的重要性。然而，未标记的 Dp44mT 最显著地降低了（¹⁴C-Dp44mT 的摄取，表明假定的载体/受体对 Dp44mT 具有很高的选择性。有趣的是，与 ¹⁴C-Dp44mT 相反，其 Fe 配合物[Fe（¹⁴C-Dp44mT）₂]的摄取不是浓度依赖性的饱和，并且远远大于单独的配体，表明存在另一种转运方式。研究检查了静脉注射到小鼠肿瘤模型中的¹⁴C-Dp44mT 的组织分布，表明¹⁴C 标记主要在排泄系统中被识别。总的来说，这些研究 Dp44mT 摄取机制及其分布和排泄的研究结果对其在体内的生物利用度和摄取具有临床意义。
