磷酸肌醇-3'-激酶δ抑制剂CAL-101联合硼替佐米对套细胞淋巴瘤细胞的作用及其相关机制探讨

[Effects of the phosphoinostitide-3'-kinase delta inhibitor, CAL-101, in combination with Bortezomib on mantle lymophma cells and exploration of its related mechanism].

作者信息

Qu Fulian, Xia Bing, Li Xiaowu, Guo Shanqi, Zhang Le, Tian Chen, Yu Yong, Zhang Yizhuo

机构信息

Department of Hematology, Tianjin Cancer Institute and Hospital, Tianjin 300060, China.

Department of Hematology, Tianjin Cancer Institute and Hospital, Tianjin 300060, China; Email:

出版信息

Zhonghua Zhong Liu Za Zhi. 2015 Jun;37(6):412-7.

PMID:26463142

Abstract

OBJECTIVE

To investigate the effect of CAL-101, a selective inhibitor of PI3Kδ, in combination with bortezomib on the proliferation and apoptosis in human mantle cell lymphoma cell lines Z138, HBL-2 and Jeko-1 in vitro, to explore its mechanisms and provide the foundation for effective treatment strategies against mantle cell lymphoma.

METHODS

MTT assay was applied to detect the inhibitory effects of CAL-101 and bortezomib either alone or combined on Z138, HBL-2 and Jeko-1 cells. Calcusyn software was used to analyze the synergistic cytotoxicity. Western blot was used to detect the expression of PI3K-p110σ and p-Akt, Akt, p-ERK and ERK proteins after the cells were exposed to different concentrations of CAL-101. Flow cytometry was employed to assess the apoptosis rate. NF-κB kit was used to determine the changes of location of NF-κB P65, and Western blot was applied to detect the level of caswpase-3 and the phosphorylation of Akt in different groups.

RESULTS

CAL-101 and BTZ inhibited the proliferation of Z138, HBL-2 and Jeko-1 cells in a dose- and time-dependent manner. CAL-101/BTZ combination induced significantly synergistic cytotoxicity in the MCL cells. The results of Western blot assay showed that CAL-101 significantly blocked the phosphorylation of Akt and ERK in the MCL cell lines. In addition, CAL-101 combined with BTZ induced pronounced apoptosis (P < 0.01). For example, after the Z138 cells exposed to the drugs for 48 h, the apoptosis rates of the control, CAL-101, BTZ and CAL-101 + BTZ groups were: (2.6 ± 1.8)%, (40.0 ± 3.0)%, (34.0 ± 1.0)%, and (67.4 ± 1.0)%, respectively; and when drug treatment was given to HBL-2 cells over 96 h, the apoptosis rates of these four cell groups were (7.4 ± 0.6)%, (30.7 ± 5.7)%, (12.0 ± 1.0)%, and (85.0 ± 4.0)%, respectively. The combination therapy contributed to the enhanced inactivity of nuclear factor-κB (NF-κB) and Akt inactivation in the MCL cell lines (P < 0.05), however, the casepase-3 activity was up-regulated.

CONCLUSIONS

The combination of CAL-101 and bortezomib is muchmore effective in inhibiting proliferation and promoting apoptosis of mantle cell lymphoma cell lines (Z138, HBL-2 and Jeko-1), which may be mediated through inhibiting PI3K/Akt signaling pathway and the transcription of NF-κB.

摘要

目的

研究PI3Kδ选择性抑制剂CAL-101与硼替佐米联合应用对人套细胞淋巴瘤细胞系Z138、HBL-2和Jeko-1体外增殖和凋亡的影响，探讨其作用机制，为套细胞淋巴瘤的有效治疗策略提供依据。

方法

采用MTT法检测CAL-101和硼替佐米单独及联合应用对Z138、HBL-2和Jeko-1细胞的抑制作用。用Calcusyn软件分析协同细胞毒性。细胞暴露于不同浓度的CAL-101后，采用蛋白质免疫印迹法检测PI3K-p110σ、p-Akt、Akt、p-ERK和ERK蛋白的表达。采用流式细胞术评估凋亡率。用NF-κB试剂盒检测NF-κB P65的定位变化，采用蛋白质免疫印迹法检测不同组中caspase-3水平和Akt的磷酸化水平。

结果

CAL-101和硼替佐米以剂量和时间依赖性方式抑制Z138、HBL-2和Jeko-1细胞的增殖。CAL-101/硼替佐米联合应用在套细胞淋巴瘤细胞中诱导出显著的协同细胞毒性。蛋白质免疫印迹分析结果显示，CAL-101显著阻断套细胞淋巴瘤细胞系中Akt和ERK的磷酸化。此外，CAL-101与硼替佐米联合应用诱导明显的凋亡（P<0.01）。例如，Z138细胞经药物处理48小时后，对照组、CAL-101组、硼替佐米组和CAL-101+硼替佐米组的凋亡率分别为：(2.6±1.8)%、(40.0±3.0)%、(34.0±1.0)%和(67.4±1.0)%；当对HBL-2细胞进行96小时的药物处理时，这四个细胞组的凋亡率分别为(7.4±0.6)%、(30.7±5.7)%、(12.0±1.0)%和(85.0±4.0)%。联合治疗导致套细胞淋巴瘤细胞系中核因子-κB（NF-κB）活性增强和Akt失活（P<0.05），然而，caspase-3活性上调。