Division of Hematology/Oncology, Department of Medicine, Virginia Commonwealth University, Richmond, VA 23298, USA.
Clin Cancer Res. 2013 Jan 15;19(2):404-14. doi: 10.1158/1078-0432.CCR-12-2799. Epub 2012 Nov 30.
To determine whether Polo-like kinase 1 (PLK1) inhibitors (e.g., BI2536) and histone deacetylase (HDAC) inhibitors (e.g., vorinostat) interact synergistically in the BCR/ABL(+) leukemia cells sensitive or resistant to imatinib mesylate (IM) in vitro and in vivo.
K562 and LAMA84 cells sensitive or resistant to imatinib mesylate and primary CML cells were exposed to BI2536 and vorinostat. Effects on cell viability and signaling pathways were determined using flow cytometry, Western blotting, and gene transfection. K562 and BV173/E255K animal models were used to test in vivo efficacy.
Cotreatment with BI2536 and vorinostat synergistically induced cell death in parental or imatinib mesylate-resistant BCR/ABL(+) cells and primary CD34(+) bone marrow cells but was minimally toxic to normal cells. BI2536/vorinostat cotreatment triggered pronounced mitochondrial dysfunction, inhibition of p-BCR/ABL, caspase activation, PARP cleavage, reactive oxygen species (ROS) generation, and DNA damage (manifest by increased expression of γH2A.X, p-ATM, p-ATR), events attenuated by the antioxidant TBAP. PLK1 short hairpin RNA (shRNA) knockdown significantly increased HDACI lethality, whereas HDAC1-3 shRNA knockdown reciprocally increased BI2536-induced apoptosis. Genetic interruption of the DNA damage linker H1.2 partially but significantly reduced PLK1/HDAC inhibitor-mediated cell death, suggesting a functional role for DNA damage in lethality. Finally, BI2536/vorinostat cotreatment dramatically reduced tumor growth in both subcutaneous and systemic BCR/ABL(+) leukemia xenograft models and significantly enhanced animal survival.
These findings suggest that concomitant PLK1 and HDAC inhibition is active against imatinib mesylate-sensitive or refractory CML and ALL cells both in vitro and in vivo and that this strategy warrants further evaluation in the setting of BCR/ABL(+) leukemias.
确定 Polo 样激酶 1（PLK1）抑制剂（如 BI2536）和组蛋白去乙酰化酶（HDAC）抑制剂（如伏立诺他）在体外和体内对伊马替尼敏感或耐药的 BCR/ABL（+）白血病细胞是否具有协同作用。
用 BI2536 和伏立诺他处理对伊马替尼敏感或耐药的 K562 和 LAMA84 细胞以及原代 CML 细胞。采用流式细胞术、Western blot 和基因转染检测细胞活力和信号通路的变化。用 K562 和 BV173/E255K 动物模型检测体内疗效。
BI2536 和伏立诺他联合用药可协同诱导亲本或伊马替尼耐药的 BCR/ABL（+）细胞及原代 CD34（+）骨髓细胞死亡，而对正常细胞的毒性较小。BI2536/伏立诺他联合用药可触发明显的线粒体功能障碍、p-BCR/ABL 抑制、半胱天冬酶激活、PARP 切割、活性氧（ROS）生成和 DNA 损伤（表现为 γH2A.X、p-ATM、p-ATR 表达增加），抗氧化剂 TBAP 可减弱这些作用。PLK1 短发夹 RNA（shRNA）敲低显著增加了 HDACI 的致死率，而 HDAC1-3 shRNA 敲低则相反，增加了 BI2536 诱导的凋亡。DNA 损伤连接蛋白 H1.2 的遗传中断部分但显著降低了 PLK1/HDAC 抑制剂介导的细胞死亡，表明 DNA 损伤在致死性中起作用。最后，BI2536/伏立诺他联合用药显著抑制了皮下和全身 BCR/ABL（+）白血病异种移植模型中的肿瘤生长，并显著提高了动物存活率。
这些发现表明，PLK1 和 HDAC 抑制的联合应用对体外和体内的伊马替尼敏感或耐药的 CML 和 ALL 细胞均具有活性，这种策略在 BCR/ABL（+）白血病中值得进一步评估。
