Combined treatment with bortezomib plus bafilomycin A1 enhances the cytocidal effect and induces endoplasmic reticulum stress in U266 myeloma cells: Crosstalk among proteasome, autophagy-lysosome and ER stress
Combined treatment with bortezomib plus bafilomycin A1 enhances the cytocidal effect and induces endoplasmic reticulum stress in U266 myeloma cells: Crosstalk among proteasome, autophagy-lysosome and ER stress
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DOI：
10.3892/ijo.2010.882
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发表时间：
2011-03-01
影响因子：
5.2
通讯作者：
Tomoda, Akio
中科院分区：
文献类型：
--
作者：
Kawaguchi, Tomohiro;Miyazawa, Keisuke;Tomoda, Akio
Bortezomib (BZ) 是一线 26S 蛋白酶体抑制剂，在多发性骨髓瘤 (MM) 细胞系中通过 caspase-3 激活诱导有效的杀细胞作用。由于 I kappa B α 是蛋白酶体的底物，因此在 MM 中使用 BZ 的最初理由是抑制 NE-kappa B。然而，BZ 在 U266 细胞中反而激活了 NF-kappa B 活性。 BZ 在测试的各种细胞系中诱导自噬以及内质网 (ER) 应激。在 U266 细胞中用 3-甲基腺嘌呤或针对 LC3B 的 siRNA 处理来抑制初始自噬体形成，以及在鼠胚胎成纤维细胞系中敲低 atg5 基因，均可导致 BZ 诱导的细胞死亡减弱。相比之下，与单独使用 BZ 或 BAF 相比，BZ 和巴弗洛霉素 A(1) (BAF)（一种液泡 ATP 酶特异性抑制剂，在后期用作自噬抑制剂）联合治疗可产生协同细胞毒性。 BAF 处理也诱导 ER 应激，但 ER 应激相关基因的诱导动力学 [例如CHOP (GADD153) 和 GRP78] BZ 和 BAF 处理之间完全不同：BZ 在 8 小时内诱导这些 ER 应激标记物，而 BAF 处理在 U266 细胞中需要超过 48 小时。为了同步 ER 应激，我们用 BAF 预处理 U266 细胞 48 小时，然后用 BZ 预处理 48 小时。与同时组合 BAF 和 BZ 相比，BAF 和 BZ 连续治疗诱导了进一步增强的细胞毒性。这些数据表明泛素-蛋白酶体系统、自噬-溶酶体系统和内质网应激之间存在串扰。控制这些相互作用和动力学似乎对于优化临床癌症治疗（包括 MM 治疗）具有重要意义。
Bortezomib (BZ), a first line 26S proteasome inhibitor, induces a potent cytocidal effect with caspase-3 activation in multiple myeloma (MM) cell lines. Since I kappa B alpha is a substrate of the proteasome, the initial rationale for using BZ in MM has been to inhibit NE-kappa B. However, BZ rather activated NF-kappa B activity in U266 cells. BZ induces autophagy as well as endoplasmic reticulum (ER) stress in various cell lines tested. Inhibition of initial autophagosome formation by treatment with either 3-methyladenine or siRNA for LC3B in U266 cells and knockdown of the atg5 gene in a murine embryonic fibroblastic cell line all resulted in attenuation of BZ-induced cell death. In contrast, combined treatment with BZ and bafilomycin A(1) (BAF), which is a specific inhibitor of vacuolar-ATPase and is used as an autophagy inhibitor at the late stage, resulted in synergistic cytotoxicity, compared with that by either BZ or BAF alone. BAF treatment also induced ER stress, but the kinetics of inductions of ER stress-related genes [e.g. CHOP (GADD153) and GRP78] completely differed between BZ- and BAF-treatments: BZ induced these ER stress markers within 8 h, whereas treatment with BAF required more than 48 h in U266 cells. In order to synchronize ER stress, we pre-treated U266 cells with BAF for 48 h, followed with BZ for 48 h. The sequential treatment with BAF and BZ induced a further enhanced cytotoxicity, compared with the simultaneous combination of BAF and BZ. These data suggest crosstalk among the ubiquitin-proteasome system, the autophagy-lysosome system, and ER stress. Controlling these interactions and kinetics appears to have important implications for optimizing clinical cancer treatment including MM-therapy.