The role of calcium-sensitive potassium channels (KCa3.1) in non-small cell lung cancer progression

目前的项目旨在破译钙敏感钾通道KCa3.1(由KCNN4基因编码)在非小细胞肺癌(NSCLC)进展中的作用机制。在我们之前的工作中，我们已经发现KCa3.1通道启动子的低甲基化和通道蛋白的过度表达是患者预后不良的强烈预测因素。肺癌是最常见的非小细胞肺癌(NSCLC)亚型，是世界范围内癌症相关死亡的主要原因，5年生存率仅为10%。大多数患者死于疾病的转移性传播。因此，了解肿瘤转移的机制具有重要的病理生理意义。所谓的转移级联包括循环中的肿瘤细胞与内皮细胞的黏附以及它们随后在转移部位的跨内皮细胞迁移。虽然离子通道在肿瘤细胞凋亡、增殖和迁移中的作用已经得到了很好的研究，但它们在转移级联的不同步骤中的作用在很大程度上仍然是未知的。这里我们重点研究钙敏感的钾通道KCa3.1。该通道通过控制NSCLC细胞的增殖、凋亡和迁移而促进其侵袭性表型。然而，到目前为止，对于KCa3.1通道驱动转移从而决定患者预后的机制的了解非常有限。在本项目中，我们将研究KCa3.1通道在非小细胞肺癌转移级联的外渗过程中的作用。KCa3.1通道在非小细胞肺癌和内皮细胞中均有表达。我们推测KCa3.1通道正在调节NSCLC细胞与内皮细胞的黏附和它们的跨内皮迁移，NSCLC和内皮细胞的KCa3.1通道都参与了这一过程。我们将使用活细胞和3D成像技术、原子力显微镜、微流体和小鼠转移模型的组合。KCa3.1通道依赖机制将通过补充的药理学、siRNA和/或基因敲除方法来确定。我们想要证明KCa3.1通道(I)以一种ROS依赖的方式调节相关黏附分子的表达和功能，(Ii)确定NSCLC细胞与内皮细胞的初始黏附和其向血管内迁移的能力之间的微调平衡，以及(Iii)使内皮细胞能够采用允许NSCLC细胞跨内皮迁移的力学特性。综上所述，我们的项目将提供进一步的证据，证明离子通道长期以来被认为是其他疾病(如高血压)的重要和容易下药的靶点，正在成为肿瘤学中一类新的治疗靶点。

The current project aims at deciphering mechanisms by which the calcium-sensitive potassium channel KCa3.1 (encoded by the KCNN4 gene) contributes to the progression of non-small cell lung cancer (NSCLC). We had identified in our previous work the hypomethylation of the KCa3.1 channel promoter and hence overexpression of the channel protein as strong predictors of a poor patient prognosis. Lung cancer, with non-small cell lung carcinoma (NSCLC) being the most frequent subtype, is a leading cause of cancer-related death worldwide with a 5 year survival rate of only ~10 %. Most patients die because of the metastatic spread of the disease. Thus, understanding mechanisms of metastasis is of great pathophysiological significance. The socalled metastatic cascade involves amongst others the adhesion of circulating tumor cells to the endothelium and their subsequent transendothelial migration at the site of metastasis. While the contribution of ion channels to tumor cell apoptosis, proliferation and migration has been well studied, their role in distinct steps of the metastatic cascade is still largely elusive. Here we focus on the calcium-sensitive potassium channel KCa3.1. The channel contributes to the aggressive phenotype of NSCLC cells by controlling their proliferation, apoptosis and migration. However, so far there is only very limited knowledge about the mechanisms by which KCa3.1 channels are driving metastasis and thereby determine patient prognosis. In the present project we will investigate the role of KCa3.1 channels in the extravasation step of the metastatic cascade in NSCLC. KCa3.1 channels are expressed both in NSCLC and in endothelial cells. We hypothesize that KCa3.1 channels are regulating the adhesion of NSCLC cells to the endothelium and their transendothelial migration with KCa3.1 channels from both, NSCLC and endothelial cells, being involved in this process. We will employ a combination of live-cell as well as 3D imaging techniques, atomic force microscopy, microfluidics and a murine model of metastasis. KCa3.1 channel-dependent mechanisms will be determined with a complimentary pharmacological, siRNA and/or knock-out approach. We want to show that KCa3.1 channels (i) regulate the expression and function of the relevant adhesion molecules in a ROS-dependent manner, (ii) ascertain the fine-tuned balance between initial adhesion of NSCLC cells to endothelial cells and their ability to migrate intravascularly to the site of transmigration, and (iii) enable endothelial cells to adopt mechanical properties permissive for transendothelial migration of NSCLC cells. Taken together, our project will provide further evidence that ion channels, long known as important and easily drugable targets in other pathologies such as hypertension, are emerging as a novel class of therapeutic targets in oncology.