Deciphering mechanisms of drug sensitivity and resistance to Selective Inhibitor of Nuclear Export (SINE) compounds.
Deciphering mechanisms of drug sensitivity and resistance to Selective Inhibitor of Nuclear Export (SINE) compounds.
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DOI：
10.1186/s12885-015-1790-z
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发表时间：
2015-11-17
期刊：
影响因子：
3.8
通讯作者：
Landesman Y
中科院分区：
文献类型：
--
作者：
Crochiere M;Kashyap T;Kalid O;Shechter S;Klebanov B;Senapedis W;Saint-Martin JR;Landesman Y
输出蛋白1 （exouttin 1, XPO1）是一种特征明确的核输出蛋白，其表达在许多类型的癌症中上调，并具有从细胞核运输关键肿瘤抑制蛋白（tumor suppressor protein, tsp）的功能。Karyopharm Therapeutics开发了一系列小分子选择性核输出抑制剂（SINE）化合物，这些化合物在体外和体内都被证明可以阻断XPO1的功能。候选药物selinexor （KPT-330）目前正在进行ii /IIb期临床试验，用于治疗血液和实体肿瘤。本研究试图破译使细胞对以KPT-185 （KPT-330的早期类似物）为代表的SINE化合物治疗敏感或耐药的机制。使用人纤维肉瘤HT1080细胞系，随着KPT-185浓度的增加，经过10个月的持续孵育获得了对SINE的抗性。MTT法检测细胞活力。采用免疫荧光法比较tsp的核输出。荧光活化细胞分选（FACS）、定量聚合酶链反应（qPCR）和免疫印迹检测对细胞周期、基因表达和细胞死亡的影响。通过Affymetrix微阵列分析naïve和药物处理亲本细胞和耐药细胞的RNA。逐渐增加SINE浓度处理HT1080细胞，导致其对SINE细胞毒性的敏感性降低了100倍。耐药细胞表现出细胞周期延长，核中tsp积累减少，蛋白表达变化与亲本细胞相似，但蛋白表达变化的幅度在亲本细胞中更为显著。微阵列分析比较亲代和耐药细胞表明，耐药细胞中的一些关键信号通路发生了改变，包括涉及粘附、凋亡和炎症的基因的表达变化。虽然亲本细胞和耐药细胞在药物治疗后的转录变化模式相似，但亲本细胞的反应程度更强。这些结果表明，SINE抗性是由XPO1抑制下游信号通路的改变所赋予的。调节这些途径可能潜在地克服对核出口抑制剂的抗性。本文的在线版本（doi:10.1186/s12885-015-1790-z）包含补充材料，可供授权用户使用。
Exportin 1 (XPO1) is a well-characterized nuclear export protein whose expression is up-regulated in many types of cancers and functions to transport key tumor suppressor proteins (TSPs) from the nucleus. Karyopharm Therapeutics has developed a series of small-molecule Selective Inhibitor of Nuclear Export (SINE) compounds, which have been shown to block XPO1 function both in vitro and in vivo. The drug candidate, selinexor (KPT-330), is currently in Phase-II/IIb clinical trials for treatment of both hematologic and solid tumors. The present study sought to decipher the mechanisms that render cells either sensitive or resistant to treatment with SINE compounds, represented by KPT-185, an early analogue of KPT-330. Using the human fibrosarcoma HT1080 cell line, resistance to SINE was acquired over a period of 10 months of constant incubation with increasing concentration of KPT-185. Cell viability was assayed by MTT. Immunofluorescence was used to compare nuclear export of TSPs. Fluorescence activated cell sorting (FACS), quantitative polymerase chain reaction (qPCR), and immunoblots were used to measure effects on cell cycle, gene expression, and cell death. RNA from naïve and drug treated parental and resistant cells was analyzed by Affymetrix microarrays. Treatment of HT1080 cells with gradually increasing concentrations of SINE resulted in > 100 fold decrease in sensitivity to SINE cytotoxicity. Resistant cells displayed prolonged cell cycle, reduced nuclear accumulation of TSPs, and similar changes in protein expression compared to parental cells, however the magnitude of the protein expression changes were more significant in parental cells. Microarray analyses comparing parental to resistant cells indicate that a number of key signaling pathways were altered in resistant cells including expression changes in genes involved in adhesion, apoptosis, and inflammation. While the patterns of changes in transcription following drug treatment are similar in parental and resistant cells, the extent of response was more robust in the parental cells. These results suggest that SINE resistance is conferred by alterations in signaling pathways downstream of XPO1 inhibition. Modulation of these pathways could potentially overcome the resistance to nuclear export inhibitors. The online version of this article (doi:10.1186/s12885-015-1790-z) contains supplementary material, which is available to authorized users.