Objective: To investigate the influence of novel CRM1 inhibitor KPT-330 on the autophagy of mantle cell lymphoma (MCL) cells, and effect of KPT-330 on the prolifiration of MCL cells in the presence or absence of autophagy inhibitor.
Methods: CCK-8 assay was used to detect the effect of KPT-330 on MCL cell lines Z-138， Jeko-1， Granta-519， Rec-1. The effect of KPT-330 on autophagy features were determined by detecting acidic vesicular organelles (AVO) by MDC staining under fluorescence microscope and detecting protein expression of LC3B-II assessed by Western blot. Further combined application of lysosomal inhibitor Chloroquine (CQ) was used to observe its effect on the increase of LC3B-Ⅱ caused by KPT-330. CalcuSyn 2.0 software was used to detected the Combination index (CI) of KPT-330 combined with CQ.
Results: The proliferation of MCL cell lines （Z-138, Jeko-1, Grant-519, Rec-1） could be inhibited by KPT-330 in a dose-dependent manner (r =0.930, 0.946, 0.691, 0.968 respectively). The number of acidic vesicular organelles (AVO) and the expression of LC3B-II were increased in KPT-330 treated Jeko-1 and Granta-519 cells in a dose-dependent manner (r Jeko-1=0.993, r Granta-519=0.971). LC3B-II protein amounts still increased upon KPT-330 treatment with the existence of lysosomal inhibitor CQ in Jeko-1 and Granta-519 cells, which was higher than CQ or KPT-330 single drug group. The combination of KPT-330 and CQ produced the synergistic effects on cells proliferation inhibition with CalcuSyn 2.0 analysis.
Conclusion: KPT-330 can inhibit MCL cell proliferation and induce autophagy. KPT-330 combined with autophagy inhibitor CQ could produce synergistic anti MCL effects, providing experimental basis for clinical combination therapy.
题目: 抑制细胞自噬提高KPT-330抗套细胞淋巴瘤效应.
目的: 探讨新型CRM1抑制剂（KPT-330）对套细胞淋巴瘤（MCL）细胞自噬的影响，以及有无自噬溶酶体抑制剂，KPT-330对MCL细胞增殖的影响。.
方法: 应用CCK-8法检测KPT-330体外对MCL细胞（Z-138、Jeko-1、Granta-519、Rec-1）增殖的影响；通过MDC染色分析自噬体的形成和Western blot检测LC3B的切割以探讨KPT-330对MCL细胞自噬的影响； 进一步联合应用自噬溶酶体抑制剂氯喹（CQ），观察其对KPT-330导致LC3B-Ⅱ增加的影响。通过CalcuSyn联合用药软件分析CQ与KPT-330的联合用药指数（Combination index，CI）。.
结果: 新型CRM1抑制剂KPT-330对MCL细胞株（Z-138、Jeko-1、Granta-519、Rec-1）的增殖有明显的抑制作用，且呈剂量依赖性(r 分别为：0.930，0.946，0.691，0.968)。KPT-330作用于Jeko-1细胞和Granta-519细胞后，导致自噬泡形成增加，LC3B-Ⅱ表达增加，并具有浓度依赖性(r =0.993, r =0.971)，表明，KPT-330导致MCL细胞自噬体形成增加。CQ联合KPT-330导致Jeko-1、Granta-519细胞LC3B-Ⅱ表达高于单药CQ或单药KPT-330（P < 0.05）。CalcuSyn 2.0软件分析结果提示KPT-330和CQ联合应用可以产生协同抗MCL效应。.
结论: 新型CRM1小分子靶向抑制剂KPT-330具有体外抑制MCL细胞株Z-138、Jeko-1、Granta-519、Rec-1增殖，诱导Jeko-1、Granta-519细胞自噬的作用；KPT-330联合CQ可产生协同抗MCL作用，为临床联合用药提供了实验依据。.
Keywords: novel CRM1 inhibitor; Mantle Cell Lymphoma; autophagy; chloroquine.