[MELK抑制剂OTSSP167对弥漫性大B细胞淋巴瘤的作用]

[Effect of MELK Inhibitor OTSSP167 on Diffuse Large B-Cell Lymphoma].

作者信息

Zhou Jun-Yi, Huang Hao, Zhuang Yan, Zhong Xiao-Min

机构信息

The Huaian Clinical College of Xuzhou Medical University, Huai'an 223300, Jiangsu Province, China.

The Huaian Clinical College of Xuzhou Medical University, Huai'an 223300, Jiangsu Provinc China.E-mail：

出版信息

Zhongguo Shi Yan Xue Ye Xue Za Zhi. 2023 Jun;31(3):739-745. doi: 10.19746/j.cnki.issn.1009-2137.2023.03.018.

DOI:10.19746/j.cnki.issn.1009-2137.2023.03.018

PMID:37356934

Abstract

OBJECTIVE

To investigate the effect of MELK inhibitor OTSSP167 against diffuse large B-cell lymphoma (DLBCL).

METHODS

The effect of OTSSP167 on activity, proliferation, and apoptosis of DLBCL cell line (SUDHL2 and HBL1) was detected by CCK-8 assay, 5-ethynyl-2'-deoxyuridine (EdU) staining, and Annexin V-FITC/PI double staining, respectively. DLBCL cells were inoculated into nude mice, after 4 weeks of OTSSP167 treatment, the effect of OTSSP167 on DLBCL growth was detected. Caspase-Glo 3/7 enzyme activity assay kit was used to detect the effect of OTSSP167 on Caspase 3/7 enzyme activity of DLBCL cells. The expression levels of apoptosis and cycle-related proteins were detected by Western blot.

RESULTS

OTSSP167 significantly inhibited the activity of SUDHL2 and HBL1 cells in a dose-dependent manner ( =-0.61, =-0.52). EdU staining showed that OTSSP167 could significantly inhibit the proliferation of SUDHL2 and HBL1 cells. Annexin V-FITC/PI result showed that OTSSP167 could significantly promote the apoptosis of SUDHL2 and HBL1 cells ( <0.001). The result of experiment showed that OTSSP167 could inhibit the growth of SUDHL2 cells in nude mice. The result of TUNEL staining of tumor further confirmed that OTSSP167 could promote the apoptosis of SUDHL2 cells. Caspase 3/7 enzyme activity test demonstrated that OTSSP167 could significantly increase caspase activity in SUDHL2 and HBL1 cells ( =0.98, =0.87). Western blot showed that OTSSP167 could dose-dependently inhibit the expression of PARP, Bcl-xL, and Bcl-2 in apoptosis signaling pathway ( =-0.93, =-0.66, =-0.87), while p53 protein was significantly up-regulated ( =0.82). The expression of cell cycle-related proteins cdc2, Cyclin E1, Cyclin A2, and Cyclin B1 also showed a dose-dependent down-regulation ( =-0.89, =-0.83, =-0.61, =-0.93).

CONCLUSION

The MELK inhibitor OTSSP167 can inhibit the proliferation and promote the apoptosis of DLBCL cells by inhibiting the expression of cycle-related proteins and anti-apoptosis-related proteins.

摘要

目的

研究MELK抑制剂OTSSP167对弥漫性大B细胞淋巴瘤（DLBCL）的作用。

方法

分别采用CCK-8法、5-乙炔基-2'-脱氧尿苷（EdU）染色法和膜联蛋白V-异硫氰酸荧光素/碘化丙啶（Annexin V-FITC/PI）双染法检测OTSSP167对DLBCUDHL2和HBL1细胞活性、增殖及凋亡的影响。将DLBCL细胞接种于裸鼠体内，经OTSSP167处理4周后，检测OTSSP167对DLBCL生长的影响。使用Caspase-Glo 3/7酶活性检测试剂盒检测OTSSP167对DLBCL细胞Caspase 3/7酶活性的影响。通过蛋白质免疫印迹法检测凋亡和细胞周期相关蛋白的表达水平。

结果

OTSSP167能以剂量依赖性方式显著抑制SUDHL2和HBL1细胞的活性（r=-0.61，r=-0.52）。EdU染色显示，OTSSP167可显著抑制SUDHL2和HBL1细胞的增殖。Annexin V-FITC/PI结果显示，OTSSP167可显著促进SUDHL2和HBL1细胞的凋亡（P<0.001）。体内实验结果显示，OTSSP167可抑制裸鼠体内SUDHL2细胞的生长。肿瘤组织的TUNEL染色结果进一步证实，OTSSP167可促进SUDHL2细胞的凋亡。Caspase 3/7酶活性检测表明，OTSSP167可显著增加SUDHL2和HBL1细胞中的半胱天冬酶活性（r=0.98，r=0.87）。蛋白质免疫印迹法显示OTSSP167可剂量依赖性抑制凋亡信号通路中PARP、Bcl-xL和Bcl-2的表达（r=-0.93，r=-0.66，r=-0.87），而p53蛋白显著上调（r=0.82）。细胞周期相关蛋白cdc2、细胞周期蛋白E1、细胞周期蛋白A2和细胞周期蛋白B1的表达也呈剂量依赖性下调（r=-0.89，r=-0.83，r=-0.61，r=-0.93）。