Smac mimetic sensitizes glioblastoma cells to Temozolomide-induced apoptosis in a RIP1- and NF-κB-dependent manner
Smac mimetic sensitizes glioblastoma cells to Temozolomide-induced apoptosis in a RIP1- and NF-κB-dependent manner
复制标题
Smac 模拟物以 RIP1 和 NF-κB 依赖性方式使胶质母细胞瘤细胞对替莫唑胺诱导的细胞凋亡敏感
作者：
Wagner L;Marschall V;Karl S;Cristofanon S;Zobel K;Deshayes K;Vucic D;Debatin KM;Fulda S
凋亡抑制剂 (IAP) 蛋白在许多癌症中高水平表达，因此代表了治疗干预的有吸引力的靶标。在这里，我们首次报告第二种线粒体衍生的半胱天冬酶激活剂（Smac）模拟物 BV6 使胶质母细胞瘤细胞对替莫唑胺（TMZ）敏感，替莫唑胺是治疗胶质母细胞瘤的一线化疗药物。 BV6 和 TMZ 协同降低细胞活力并引发胶质母细胞瘤细胞凋亡（组合指数< 0.4-0.8），伴随着线粒体膜电位损失增加、细胞色素 C 释放、Caspase 激活和 Caspase 依赖性细胞凋亡。分子机制分析表明，BV6 导致 cIAP1 快速降解，从而稳定 NF-κB 诱导激酶和 NF-κB 激活。 BV6 刺激的 NF-κB 激活对于 TMZ 致敏至关重要，因为突变 IκBα 超级阻遏物的过度表达对 NF-κB 的抑制可显着减少线粒体膜电位的损失、细胞色素 C 的释放、Caspase 激活和细胞凋亡。值得注意的是，BV6 介导的 TMZ 致敏与肿瘤坏死因子 α (TNFα) 产生的增加无关。此外，TNFα、CD95 或 TRAIL 阻断抗体或 TNFR1 敲低对联合治疗诱导的细胞凋亡没有影响或影响很小。有趣的是，BV6 和 TMZ 协同触发 RIP1（受体激活蛋白 1）/caspase-8/FADD 复合物的形成。通过小干扰 RNA 敲低 RIP1 显着降低 BV6 和 TMZ 诱导的 caspase-8 激活和细胞凋亡，表明 RIP1 对于细胞凋亡诱导是必需的。通过证明 BV6 以 NF-κB 和 RIP1 依赖性方式为胶质母细胞瘤细胞启动 TMZ，这些发现为 Smac 模拟物与 TMZ 联合的进一步（预）临床开发奠定了基础。
Inhibitor of apoptosis (IAP) proteins are expressed at high levels in many cancers and therefore represent attractive targets for therapeutic intervention. Here, we report for the first time that the second mitochondria-derived activator of caspases (Smac) mimetic BV6 sensitizes glioblastoma cells toward Temozolomide (TMZ), the first-line chemotherapeutic agent in the treatment of glioblastoma. BV6 and TMZ synergistically reduce cell viability and trigger apoptosis in glioblastoma cells (combination index< 0.4–0.8), which is accompanied by increased loss of mitochondrial-membrane potential, cytochrome c release, caspase activation and caspase-dependent apoptosis. Analysis of the molecular mechanisms reveals that BV6 causes rapid degradation of cIAP1, leading to stabilization of NF-κB-inducing kinase and NF-κB activation. BV6-stimulated NF-κB activation is critically required for sensitization toward TMZ, as inhibition of NF-κB by overexpression of the mutant IκBα super-repressor profoundly reduces loss of mitochondrial membrane potential, cytochrome c release, caspase activation and apoptosis. Of note, BV6-mediated sensitization to TMZ is not associated with increased tumor necrosis factor alpha (TNFα) production. Also, TNFα, CD95 or TRAIL-blocking antibodies or knockdown of TNFR1 have no or little effect on combination treatment-induced apoptosis. Interestingly, BV6 and TMZ cooperate to trigger the formation of a RIP1 (receptor activating protein 1)/caspase-8/FADD complex. Knockdown of RIP1 by small interfering RNA significantly reduces BV6-and TMZ-induced caspase-8 activation and apoptosis, showing that RIP1 is necessary for apoptosis induction. By demonstrating that BV6 primes glioblastoma cells for TMZ in a NF-κB-and RIP1-dependent manner, these findings build the rationale for further (pre) clinical development of Smac mimetics in combination with TMZ.