Institute of Clinical Pathology, Medical University of Vienna, Waehringer Guertel 18-20, A-1090 Vienna, Austria.
Br J Cancer. 2013 Feb 19;108(3):570-8. doi: 10.1038/bjc.2012.580. Epub 2013 Jan 8.
As metastasis is the prime cause of death from malignancies, there is vibrant interest to discover options for the management of the different mechanistic steps of tumour spreading. Some approved pharmaceuticals exhibit activities against diseases they have not been developed for. In order to discover such activities that might attenuate lymph node metastasis, we investigated 225 drugs, which are approved by the US Food and Drug Administration.
A three-dimensional cell co-culture assay was utilised measuring tumour cell-induced disintegrations of the lymphendothelial wall through which tumour emboli can intravasate as a limiting step in lymph node metastasis of ductal breast cancer. The disintegrated areas in the lymphendothelial cell (LEC) monolayers were induced by 12(S)-HETE, which is secreted by MCF-7 tumour cell spheroids, and are called 'circular chemorepellent induced defects' (CCIDs). The putative mechanisms by which active drugs prevented the formation of entry gates were investigated by western blotting, NF-κB activity assay and by the determination of 12(S)-HETE synthesis.
Acetohexamide, nifedipin, isoxsuprine and proadifen dose dependently inhibited the formation of CCIDs in LEC monolayers and inhibited markers of epithelial-to-mesenchymal-transition and migration. The migration of LECs is a prerequisite of CCID formation, and these drugs either repressed paxillin levels or the activities of myosin light chain 2, or myosin-binding subunit of myosin phosphatase. Isoxsuprine inhibited all three migration markers, and isoxsuprine and acetohexamide suppressed the synthesis of 12(S)-HETE, whereas proadifen and nifedipin inhibited NF-κB activation. Both the signalling pathways independently cause CCID formation.
The targeting of different mechanisms was most likely the reason for synergistic effects of different drug combinations on the inhibition of CCID formation. Furthermore, the treatment with drug combinations allowed also a several-fold reduction in drug concentrations. These results encourage further screening of approved drugs and their in vivo testing.
由于转移是恶性肿瘤死亡的主要原因，因此人们非常有兴趣发现针对肿瘤扩散不同机制步骤的治疗选择。一些已批准的药物对其未开发用于治疗的疾病表现出活性。为了发现可能减轻淋巴结转移的此类活性，我们研究了美国食品和药物管理局批准的 225 种药物。
利用三维细胞共培养测定法，测量肿瘤细胞诱导的淋巴管内皮壁破裂，肿瘤栓子通过该破裂部位进入淋巴管是乳腺癌淋巴结转移的一个限制步骤。由 MCF-7 肿瘤细胞球体分泌的 12(S)-HETE 诱导的淋巴管内皮细胞（LEC）单层中的破裂区域称为“圆形趋化抑制剂诱导缺陷”（CCID）。通过 Western blot、NF-κB 活性测定和 12(S)-HETE 合成的测定，研究了活性药物防止入口门形成的潜在机制。
醋甲唑胺、硝苯地平、异舒普林和普罗地芬可剂量依赖性地抑制 LEC 单层中 CCID 的形成，并抑制上皮间质转化和迁移的标志物。LEC 的迁移是 CCID 形成的前提，这些药物要么抑制桩蛋白水平，要么抑制肌球蛋白轻链 2 的活性，要么抑制肌球蛋白磷酸酶的肌球蛋白结合亚基。异舒普林抑制了所有三种迁移标志物，异舒普林和醋甲唑胺抑制了 12(S)-HETE 的合成，而普罗地芬和硝苯地平抑制了 NF-κB 的激活。这两种信号通路都可独立导致 CCID 的形成。
靶向不同机制很可能是不同药物组合对抑制 CCID 形成的协同作用的原因。此外，药物组合治疗还允许药物浓度降低几倍。这些结果鼓励进一步筛选已批准的药物并进行体内测试。
