Department of Pharmacology, Creighton University School of Medicine, Omaha, Nebraska 68178, USA.
J Pharmacol Exp Ther. 2010 May;333(2):393-403. doi: 10.1124/jpet.109.164814. Epub 2010 Jan 28.
Signaling through G protein-coupled receptors (GPCRs) promotes breast cancer metastasis. G proteins convey GPCR signals by dissociating into Galpha and Gbetagamma subunits. The aim of the present study was to determine whether blockade of Gbetagamma signaling suppresses breast cancer cell migration and invasion, which are critical components of metastasis. Conditioned media (CM) of NIH-3T3 fibroblasts are widely used as chemoattractants in in vitro cancer metastasis studies. Expression of a Gbetagamma scavenger peptide attenuated NIH-3T3 CM-induced migration and invasion of both metastatic breast cancer MDA-MB-231 and MDA-MB-436 cells by 40 to 50% without effects on cell viability. Migration and invasion of cells in response to NIH-3T3 CM were also blocked by 8-(4,5,6-trihydroxy-3-oxo-3H-xanthen-9-yl)-1-naph-thalene-carboxylic acid) (M119K), a Gbetagamma inhibitor, with maximum inhibition exceeding 80% and half-maximal inhibitory concentration (IC50) values of 1 to 2 microM. M119K also attenuated Rac-dependent formation of lamellipodia, a key structure required for metastasis. Constitutively active Rac1 rescued Gbetagamma blockade-mediated inhibition of breast cancer cell migration, whereas dominant negative Rac1 inhibited cell migration similar to Gbetagamma blockade. Furthermore, M119K suppressed Gi protein-coupled CXC chemokine receptor 4 (CXCR4)-dependent MDA-MB-231 cell migration by 80% with an IC50 value of 1 microM, whereas tyrosine kinase receptor-dependent cell migration was significantly less inhibited. However, CXCR4-dependent inhibition of adenylyl cyclase, a Gialpha-mediated response in MDA-MB-231 cells, was not blocked by M119K but was blocked by pertussis toxin, which selectively inactivates Gialpha. This report is the first to directly demonstrate the role of Gbetagamma in cancer cell migration and invasion and suggests that targeting Gbetagamma signaling pathways may provide a novel strategy for suppressing breast cancer metastasis.
G 蛋白偶联受体 (GPCR) 的信号转导促进乳腺癌转移。G 蛋白通过与 Galpha 和 Gbetagamma 亚基解离来传递 GPCR 信号。本研究的目的是确定 Gbetagamma 信号阻断是否抑制乳腺癌细胞迁移和侵袭，这是转移的关键组成部分。NIH-3T3 成纤维细胞的条件培养基 (CM) 广泛用作体外癌症转移研究中的趋化剂。Gbetagamma 清除肽的表达减弱了转移性乳腺癌 MDA-MB-231 和 MDA-MB-436 细胞对 NIH-3T3 CM 的迁移和侵袭的诱导作用，抑制率为 40%至 50%，而对细胞活力没有影响。NIH-3T3 CM 诱导的细胞迁移和侵袭也被 Gbetagamma 抑制剂 8-(4,5,6-三羟基-3-氧代-3H-呫吨-9-基)-1-萘甲酸（M119K）阻断，最大抑制率超过 80%，半最大抑制浓度 (IC50) 值为 1 至 2 μM。M119K 还减弱了 Rac 依赖性的片状伪足形成，这是转移所必需的关键结构。组成型激活的 Rac1 挽救了 Gbetagamma 阻断介导的乳腺癌细胞迁移抑制，而显性负 Rac1 抑制细胞迁移类似于 Gbetagamma 阻断。此外，M119K 以 1 μM 的 IC50 值抑制 80%由 Gi 蛋白偶联 CXC 趋化因子受体 4 (CXCR4) 依赖性 MDA-MB-231 细胞迁移，而酪氨酸激酶受体依赖性细胞迁移受抑制程度显著降低。然而，M119K 并未阻断 CXCR4 依赖性腺苷酸环化酶抑制，这是 MDA-MB-231 细胞中 Gialpha 介导的反应，而该反应被百日咳毒素阻断，百日咳毒素选择性失活 Gialpha。本报告首次直接证明了 Gbetagamma 在癌细胞迁移和侵袭中的作用，并表明靶向 Gbetagamma 信号通路可能为抑制乳腺癌转移提供一种新策略。
