Fabre C, Carvalho G, Tasdemir E, Braun T, Adès L, Grosjean J, Boehrer S, Métivier D, Souquère S, Pierron G, Fenaux P, Kroemer G
INSERM, Unit Apoptosis, Cancer and Immunity, Villejuif, France.
Oncogene. 2007 Jun 14;26(28):4071-83. doi: 10.1038/sj.onc.1210187. Epub 2007 Jan 8.
CD34(+) bone marrow blasts from high-risk myelodysplastic syndrome (MDS) patients as well as MDS patient-derived cell lines (P39 and MOLM13) constitutively activate the nuclear factor-kappaB (NF-kappaB) pathway and undergo apoptosis when NF-kappaB is inhibited. Here, we show that the combination of conventional chemotherapeutic agents (daunorubicin, mitoxantrone, 5-azacytidine or camptothecin) with the NF-kappaB inhibitor BAY11-7082 did not yield a synergistic cytotoxicity. In contrast, BAY11-7082 (which targets the NF-kappaB-activating I-kappaB kinase (IKK) complex) or knockdown of essential components of the NF-kappaB system (such as the IKK1 and IKK2 subunits of the IKK complex and the p65 subunit of NF-kappaB), by small interfering RNAs sensitized MDS cell lines to starvation-induced apoptosis. The combination of BAY11-7082 and nutrient depletion synergistically killed the acute myeloid leukemia (AML) cell line U937 as well as primary CD34(+) bone marrow blasts from AML and high-risk MDS patients. The synergistic killing by BAY11-7082, combined with nutrient depletion, led to cell death accompanied by all hallmarks of apoptosis, including an early loss of the mitochondrial transmembrane potential, the release of cytochrome c and apoptosis-inducing factor (AIF) from mitochondria, activation of caspase-3, phosphatidylserine exposure on the plasma membrane surface and nuclear chromatin condensation. Transmission electron microscopy revealed the presence of numerous autophagic vacuoles in the cytoplasm before cells underwent nuclear apoptosis. Nonetheless, cell death was neither inhibited by the pan-caspase inhibitor z-VAD-fmk nor by knockdown of AIF or of essential components of the autophagy pathway (ATG5, ATG6/Beclin-1, ATG10, ATG12). In contrast, external supply of glucose, insulin or insulin-like growth factor-I could retard the cell death induced by BAY11-7082 combined with starvation. These results suggest that in MDS cells, NF-kappaB inhibition can precipitate a bioenergetic crisis that leads to an autophagic stress response followed by apoptotic cell death.
高危骨髓增生异常综合征（MDS）患者的CD34(+)骨髓原始细胞以及MDS患者来源的细胞系（P39和MOLM13）可组成性激活核因子-κB（NF-κB）信号通路，并且在NF-κB受到抑制时会发生凋亡。在此，我们发现传统化疗药物（柔红霉素、米托蒽醌、5-氮杂胞苷或喜树碱）与NF-κB抑制剂BAY11-7082联合使用时，并未产生协同细胞毒性。相反，BAY11-7082（靶向激活NF-κB的I-κB激酶（IKK）复合物）或通过小干扰RNA敲低NF-κB系统的关键组分（如IKK复合物的IKK1和IKK2亚基以及NF-κB的p65亚基），可使MDS细胞系对饥饿诱导的凋亡敏感。BAY11-7082与营养物质缺乏联合使用可协同杀死急性髓系白血病（AML）细胞系U937以及AML和高危MDS患者的原发性CD34(+)骨髓原始细胞。BAY11-7082与营养物质缺乏联合使用所产生的协同杀伤作用导致细胞死亡，并伴有凋亡的所有特征，包括线粒体跨膜电位的早期丧失、细胞色素c和凋亡诱导因子（AIF）从线粒体的释放、caspase-3的激活、质膜表面磷脂酰丝氨酸的暴露以及核染色质凝聚。透射电子显微镜显示，在细胞发生核凋亡之前，细胞质中存在大量自噬泡。然而，细胞死亡既未被泛caspase抑制剂z-VAD-fmk抑制，也未因敲低AIF或自噬途径的关键组分（ATG5、ATG6/Beclin-1、ATG10、ATG12）而受到抑制。相反，外源供应葡萄糖、胰岛素或胰岛素样生长因子-I可延缓BAY11-7082与饥饿联合诱导的细胞死亡。这些结果表明，在MDS细胞中，NF-κB抑制可引发生物能量危机，导致自噬应激反应，随后发生凋亡性细胞死亡。
