Okamoto S, Jiang Y, Kawamura K, Shingyoji M, Tada Y, Sekine I, Takiguchi Y, Tatsumi K, Kobayashi H, Shimada H, Hiroshima K, Tagawa M
1] Division of Pathology and Cell Therapy, Chiba Cancer Center Research Institute, Chiba, Japan [2] Department of Respirology, Graduate School of Medicine, Chiba University, Chiba, Japan.
1] Division of Pathology and Cell Therapy, Chiba Cancer Center Research Institute, Chiba, Japan [2] Department of Molecular Biology and Oncology, Graduate School of Medicine, Chiba University, Chiba, Japan.
Cell Death Dis. 2014 Nov 13;5(11):e1517. doi: 10.1038/cddis.2014.475.
Zoledronic acid (ZOL), a nitrogen-containing bisphosphonate, produced anti-tumor effects through apoptosis induction or S-phase arrest depending on human mesothelioma cells tested. An addition of isoprenoid, geranylgeraniol but not farnesol, negated these ZOL-induced effects, indicating that the ZOL-mediated effects were attributable to depletion of geranylgeranyl pyrophosphates which were substrates for prenylation processes of small guanine-nucleotide-binding regulatory proteins (small G proteins). ZOL-treated cells decreased a ratio of membrane to cytoplasmic fractions in RhoA, Cdc42 and Rab6 but less significantly Rac1 proteins, indicating that these proteins were possible targets for ZOL-induced actions. We further analyzed which small G proteins were responsible for the three ZOL-induced effects, caspase-mediated apoptosis, S-phase arrest and morphological changes, using inhibitors for respective small G proteins and siRNA for Cdc42. ZOL-induced apoptosis is due to insufficient prenylation of Rab proteins because an inhibitor of geranlygeranyl transferase II that was specific for Rab family proteins prenylation, but not others inhibitors, activated the same apoptotic pathways that ZOL did. ZOL suppressed an endogenous topoisomerase II activity, which was associated with apoptosis and S-phase arrest in respective cells because we detected the same cell cycle changes in etoposide-treated cells. Inhibitors for geranlygeranyl transferase I and for RhoA produced morphological changes and disrupted actin fiber structures, both of which were similar to those by ZOL treatments. These data demonstrated that anti-tumor effects by ZOL were attributable to inhibited functions of respective small G proteins and topoisomerase II activity, and suggested that cellular factors were involved in the differential cell cycle changes.
唑来膦酸（ZOL）是一种含氮双膦酸盐，根据所测试的人恶性间皮瘤细胞的不同，通过诱导凋亡或使细胞停滞于S期产生抗肿瘤作用。添加类异戊二烯香叶基香叶醇而非法呢醇可消除这些ZOL诱导的效应，这表明ZOL介导的效应归因于香叶基香叶基焦磷酸的消耗，而香叶基香叶基焦磷酸是小G蛋白（小GTP结合调节蛋白）异戊二烯化过程的底物。ZOL处理的细胞使RhoA、Cdc42和Rab6的膜与细胞质部分的比例降低，但Rac1蛋白降低的程度较小，这表明这些蛋白可能是ZOL诱导作用的靶点。我们进一步使用针对各个小G蛋白的抑制剂和针对Cdc42的小干扰RNA（siRNA）分析了哪些小G蛋白负责ZOL诱导的三种效应，即半胱天冬酶介导的凋亡、S期停滞和形态变化。ZOL诱导的凋亡是由于Rab蛋白的异戊二烯化不足，因为一种对Rab家族蛋白异戊二烯化具有特异性的香叶基香叶基转移酶II抑制剂，而非其他抑制剂，激活了与ZOL相同的凋亡途径。ZOL抑制内源性拓扑异构酶II活性，这与各细胞中的凋亡和S期停滞相关，因为我们在依托泊苷处理的细胞中检测到了相同的细胞周期变化。香叶基香叶基转移酶I抑制剂和RhoA抑制剂产生了形态变化并破坏了肌动蛋白纤维结构，这两者都与ZOL处理后的情况相似。这些数据表明，ZOL的抗肿瘤作用归因于各个小G蛋白功能的抑制和拓扑异构酶II活性的抑制，并提示细胞因子参与了不同的细胞周期变化。
