Matesic Diane F, Ali Amna, Sidorova Tatyana S, Burns Timothy J
Department of Pharmaceutical Sciences, College of Pharmacy and Health Sciences, Mercer University Atlanta, Georgia 30341.
Curr Bioact Compd. 2013;9(3):255-262. doi: 10.2174/157340720903140119155322.
Cell-cell communication through gap junctions is aberrant or absent in a majority of human cancer cells, compared to cells in corresponding normal tissues. This and other evidence has led to the hypothesis that gap junction channels, comprised of connexin proteins, are important in growth control and cancer progression. The major goal of this ongoing study was to identify bioactive compounds that specifically upregulate gap junction channel-mediated cell-cell communication as potential anti-tumor therapies. Control of cell-cell communication is linked to growth regulatory intracellular signaling pathways; we therefore further aimed to identify signaling pathways modulated by these compounds in order to assess their potential as targeted anti-tumor therapies. Compounds were screened for their ability to upregulate gap junction-mediated cell-cell communication by using a fluorescent dye transfer assay to measure cell-cell communication between tumor promoter-treated astroglial cells or -transformed epithelial cells. Western blotting using connexin-specific and phosphorylation site-specific antibodies was used to monitor phosphorylation changes in signaling pathway proteins. Our results identified three compounds that upregulate gap junction-mediated cell-cell communication in our screening assays, chaetoglobosin K(ChK), 4-phenyl-3-butenoic acid (PBA) and the methyl ester of PBA (PBA-Me). Further analyses demonstrated that in tumorigenic cells, ChK downregulates phosphorylation of Akt kinase, an enzyme in the PI3-kinase signaling pathway that is found to be upregulated in a number of human cancers, on a key activation site. However, ChK did not inhibit PI-3 kinase as did the classic PI-3 kinase inhibitor, Wortmannin. PBA and PBA-Me were found to upregulate phosphorylation of p38 MAPK on a key activation site in tumorigenic cells, which is downregulated in several human cancer cell types. ChK and PBA also decreased activation of SAPK/JNK, another kinase found to be upregulated in a number of human cancers. These studies highlight the potential of monitoring gap junction intercellular communication for identifying experimental anti-tumor compounds.
与相应正常组织中的细胞相比，大多数人类癌细胞中通过间隙连接进行的细胞间通讯异常或缺失。这一现象及其他证据促使人们提出假说，即由连接蛋白组成的间隙连接通道在生长控制和癌症进展中起重要作用。这项正在进行的研究的主要目标是鉴定能够特异性上调间隙连接通道介导细胞间通讯的生物活性化合物，作为潜在的抗肿瘤疗法。细胞间通讯的控制与生长调节性细胞内信号通路相关；因此，我们进一步旨在鉴定这些化合物所调节的信号通路，以评估它们作为靶向抗肿瘤疗法的潜力。通过使用荧光染料转移测定法来测量肿瘤启动子处理的星形胶质细胞或转化上皮细胞之间的细胞间通讯，筛选化合物上调间隙连接介导的细胞间通讯的能力。使用连接蛋白特异性和磷酸化位点特异性抗体进行蛋白质印迹，以监测信号通路蛋白的磷酸化变化。我们的结果在筛选试验中鉴定出三种上调间隙连接介导的细胞间通讯的化合物，即球毛壳菌素K（ChK）、4-苯基-3-丁烯酸（PBA）和PBA的甲酯（PBA-Me）。进一步分析表明，在致瘤细胞中，ChK下调Akt激酶的磷酸化，Akt激酶是PI3激酶信号通路中的一种酶，在许多人类癌症中被发现上调，位于一个关键的激活位点。然而，ChK并不像经典的PI-3激酶抑制剂渥曼青霉素那样抑制PI-3激酶。发现PBA和PBA-Me在致瘤细胞的一个关键激活位点上调p38丝裂原活化蛋白激酶（p38 MAPK）的磷酸化，而在几种人类癌细胞类型中该位点的磷酸化是下调的。ChK和PBA还降低了应激激活蛋白激酶/应激活化蛋白激酶（SAPK/JNK）的活性，另一种在许多人类癌症中上调的激酶。这些研究突出了监测间隙连接细胞间通讯以鉴定实验性抗肿瘤化合物的潜力。
