Aldehyde dehydrogenase 1A1 confers erlotinib resistance via facilitating the reactive oxygen species-reactive carbonyl species metabolic pathway in lung adenocarcinomas
Aldehyde dehydrogenase 1A1 confers erlotinib resistance via facilitating the reactive oxygen species-reactive carbonyl species metabolic pathway in lung adenocarcinomas
复制标题
醛脱氢酶 1A1 通过促进肺腺癌中活性氧-活性羰基代谢途径赋予厄洛替尼耐药性
DOI：
10.7150/thno.35729
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发表时间：
2019-01-01
期刊：
影响因子：
12.4
通讯作者：
Zhu, Liang
中科院分区：
文献类型：
--
作者：
Lei, Hui-Min;Zhang, Ke-Ren;Zhu, Liang
背景资料：对表皮生长因子受体（EGFR）-酪氨酸激酶抑制剂（TKI）（如厄洛替尼）的获得性耐药是实现靶向治疗总体临床获益的主要挑战。最近，已发现醛脱氢酶1（ALDH 1）诱导使肺腺癌对EGFR-TKI耐药，靶向ALDH 1A 1成为克服耐药的新策略。然而，这种作用的分子机制仍然知之甚少。方法：在一组获得厄洛替尼耐药的肺腺癌细胞系和异种移植物中进行综合测定。通过体外细胞活力、凋亡、迁移和上皮-间质转化分析、离体肿瘤球形成分析以及体内肿瘤生长和播散分析来评估癌症表型。分别采用荧光氧化指示剂法和液相色谱-质谱联用技术检测活性氧和活性羰基。通过RNA干扰和药理学抑制抑制或通过基于慢病毒的克隆进行胞外过表达来抑制蛋白靶点。通过双荧光素酶报告分析测量基因启动子活性。结果：ALDH 1A 1的敲低或药理学抑制克服了体外和体内厄洛替尼耐药性。ALDH 1A 1过表达足以诱导厄洛替尼耐药。代谢组学分析表明，ALDH 1A 1成瘾的厄洛替尼耐药细胞中的ROS-RCS水平较低;与此一致，这些细胞中代谢ROS和RCS的关键酶SOD 2和GPX 4分别上调。SOD 2或GPX 4的敲低使耐药细胞对厄洛替尼重新敏感，并且该作用通过ROS-RCS清除而消除，并通过ROS-RCS诱导而模拟。ALDH 1A 1过表达的细胞，虽然抵抗厄洛替尼，但对SOD 2或GPX 4敲低更敏感。ALDH 1A 1对厄洛替尼耐药性的影响被ROS-RCS诱导消除，并被ROS-RCS清除模拟。通过检测ALDH 1A 1抑制或过表达条件下GPX 4和SOD 2的表达以及GPX 4和SOD 2启动子活性的分析，证实RCS-ROS-代谢途径受ALDH 1A 1-GPX 4-SOD 2轴控制。在ALDH 1Al诱导的抗性中的ROS-RCS代谢依赖机制在体内得到证实。对公共数据库的分析显示，在接受化疗的患者中，ALDH 1A 1、GPX 4和SOD 2高共表达的患者生存概率较低。结论：ALDH 1A 1通过促进ROS-RCS代谢途径赋予厄洛替尼抗性。ALDH 1A 1诱导的SOD 2和GPX 4上调以及ALDH 1A 1本身减轻了厄洛替尼诱导的氧化和羰基应激，并赋予TKI抗性。对厄洛替尼耐药的代谢机制的阐明为分子靶向治疗的生物学提供了新的见解，并有助于设计改进的药理学策略来克服耐药性。
Background: Acquired resistance to epidermal growth factor receptor (EGFR)-tyrosine kinase inhibitors (TKIs) such as erlotinib is a major challenge to achieve an overall clinical benefit of the targeted therapy. Recently, aldehyde dehydrogenase 1 (ALDH1) induction has been found to render lung adenocarcinomas resistant to EGFR-TKIs, and targeting ALDH1A1 becomes a novel strategy to overcome resistance. However, the molecular mechanism underlying such effect remains poorly understood. Methods: Comprehensive assays were performed in a panel of lung adenocarcinoma cell lines and xenografts that acquired resistance to erlotinib. Cancer phenotype was evaluated by cell viability, apoptosis, migration, and epithelial-mesenchymal transition analysis in vitro, tumorsphere formation analysis ex vivo, and tumor growth and dissemination analysis in vivo. Reactive oxygen species (ROS) and reactive carbonyl species (RCS) were detected based on fluorescent oxidation indicator and liquid chromatography coupled to mass spectrometry, respectively. Protein target was suppressed by RNA interference and pharmacological inhibition or ecto-overexpressed by lentivirus-based cloning. Gene promoter activity was measured by dual-luciferase reporting assay. Results: Knockdown or pharmacological inhibition of ALDH1A1 overcame erlotinib resistance in vitro and in vivo. ALDH1A1 overexpression was sufficient to induce erlotinib resistance. Metabolomic analysis demonstrated lower ROS-RCS levels in ALDH1A1-addicted, erlotinib-resistant cells; in line with this, key enzymes for metabolizing ROS and RCS, SOD2 and GPX4, respectively, were upregulated in these cells. Knockdown of SOD2 or GPX4 re-sensitized the resistant cells to erlotinib and the effect was abrogated by ROS-RCS scavenging and mimicked by ROS-RCS induction. The ALDH1A1 overexpressed cells, though resisted erlotinib, were more sensitive to SOD2 or GPX4 knockdown. The ALDH1A1 effect on erlotinib resistance was abrogated by ROS-RCS induction and mimicked by ROS-RCS scavenging. Detection of GPX4 and SOD2 expression and analysis of promoter activities of GPX4 and SOD2 under the condition of suppression or overexpression of ALDH1A1 demonstrated that the RCS-ROS-metabolic pathway was controlled by the ALDH1A1-GPX4-SOD2 axis. The ROS-RCS metabolic dependence mechanism in ALDH1Al -induced resistance was confirmed in vivo. Analysis of public databases showed that in patients undergoing chemotherapy, those with high co-expression of ALDH1A1 , GPX4, and SOD2 had a lower probability of survival. Conclusions: ALDH1A1 confers erlotinib resistance by facilitating the ROS-RCS metabolic pathway. ALDH1A1-induced upregulation of SOD2 and GPX4, as well as ALDH1A1 itself, mitigated erlotinib-induced oxidative and carbonyl stress, and imparted the TKI resistance. The elucidation of previously unrecognized metabolic mechanism underlying erlotinib resistance provides new insight into the biology of molecular targeted therapies and help to design improved pharmacological strategies to overcome the drug resistance.