Lady Davis Institute, McGill University, Montréal, Canada.
Department of Experimental Medicine, McGill University, Montréal, Canada.
Autophagy. 2022 Mar;18(3):540-558. doi: 10.1080/15548627.2021.1936932. Epub 2021 Jun 21.
Promoting the macroautophagy/autophagy-mediated degradation of specific proteins and organelles can potentially be utilized to induce apoptosis in cancer cells or sensitize tumor cells to therapy. To examine this concept, we enriched for autophagosomes from histone deacetylase inhibitor (HDACi)-sensitive U937 lymphoma cells and isogenic HDACi-resistant cells. Mass spectrometry on autophagosome-enriched fractions revealed that HDACi-resistant cells undergo elevated pexophagy, or autophagy of the peroxisome, an organelle that supports tumor growth. To disturb peroxisome homeostasis, we enhanced pexophagy in HDACi-resistant cells via genetic silencing of peroxisome exportomer complex components (, or ). This consequently sensitized resistant cells to HDACi-mediated apoptosis, which was rescued by inhibiting ATM/ataxia-telangiectasia mutated (ATM serine/threonine kinase), a mediator of pexophagy. We subsequently engineered melanoma cells to stably repress using CRISPR interference (CRISPRi). Melanoma cells with repressed expression showed evidence of both increased pexophagy and peroxisomal matrix protein import defects versus single guide scrambled ( controls. studies showed that melanoma xenografts recurred less compared to xenografts, following the development of resistance to mitogen-activated protein kinase (MAPK)-targeted therapy. Finally, prognostic analysis of publicly available datasets showed that low expression levels of and , were significantly associated with prolonged patient survival in lymphoma, lung cancer and melanoma cohorts. Our work highlighted that drugs designed to disrupt peroxisome homeostasis may serve as unconventional therapies to combat therapy resistance in cancer. ABCD3/PMP70: ATP binding cassette subfamily D member 3; ACOX1: acyl-CoA oxidase 1; AP: autophagosome; COX: cytochrome c oxidase; CQ: chloroquine; CRISPRi: clustered regularly interspaced short palindromic repeats interference; DLBCL: diffuse large B-cell lymphoma; GO: gene ontology; dCas9: Cas9 endonuclease dead, or dead Cas9; HDACi: histone deacetylase inhibitors; IHC: Immunohistochemistry; LAMP2: lysosomal associated membrane protein 2; LCFAs: long-chain fatty acids; LFQ-MS: label-free quantitation mass spectrometry; LPC: lysophoshatidylcholine; MAP1LC3B/LC3B: microtubule associated protein 1 light chain 3 beta; MTOR: mechanistic target of rapamycin kinase; PBD: peroxisome biogenesis disorders; PTS1: peroxisomal targeting signal 1; ROS: reactive oxygen species; sgRNA: single guide RNA; VLCFAs: very-long chain fatty acids; Vor: vorinostat; WO: wash-off.
促进特定蛋白质和细胞器的巨自噬/自噬介导的降解，可以潜在地用于诱导癌细胞凋亡或使肿瘤细胞对治疗敏感。为了检验这一概念，我们从组蛋白去乙酰化酶抑制剂 (HDACi) 敏感的 U937 淋巴瘤细胞和同源的 HDACi 耐药细胞中富集自噬体。自噬体富集部分的质谱分析显示，HDACi 耐药细胞经历了过氧化物酶体自噬的升高，或过氧化物酶体的自噬，过氧化物酶体是支持肿瘤生长的细胞器。为了扰乱过氧化物酶体的动态平衡，我们通过遗传沉默过氧化物酶体输出复合物成分 (,, 或 ) 来增强 HDACi 耐药细胞中的过氧化物酶体自噬。这继而使耐药细胞对 HDACi 介导的细胞凋亡敏感，而 ATM/ataxia-telangiectasia mutated (ATM 丝氨酸/苏氨酸激酶) 的抑制可挽救 ATM/ataxia-telangiectasia mutated (ATM 丝氨酸/苏氨酸激酶) 的抑制，ATM/ataxia-telangiectasia mutated (ATM 丝氨酸/苏氨酸激酶) 是过氧化物酶体自噬的介质。随后，我们使用 CRISPR 干扰 (CRISPRi) 使黑色素瘤细胞稳定地抑制 。与单个向导 scrambled ( 对照相比，表达受抑制的黑色素瘤细胞表现出过氧化物酶体自噬增加和过氧化物酶体基质蛋白输入缺陷的证据。研究表明，与 mitogen-activated protein kinase (MAPK)-靶向治疗耐药相比， 基因敲除的黑色素瘤异种移植物复发较少。最后，对公开可用数据集的预后分析表明，在淋巴瘤、肺癌和黑色素瘤队列中， 的低表达水平与患者生存时间延长显著相关。我们的工作强调，旨在破坏过氧化物酶体动态平衡的药物可能成为对抗癌症治疗耐药性的非常规疗法。ABCD3/PMP70：ATP 结合盒亚家族 D 成员 3；ACOX1：酰基辅酶 A 氧化酶 1；AP：自噬体；COX：细胞色素 c 氧化酶；CQ：氯喹；CRISPRi：成簇规律间隔短回文重复干扰；DLBCL：弥漫性大 B 细胞淋巴瘤；GO：基因本体论；dCas9：Cas9 内切酶失活，或失活 Cas9；HDACi：组蛋白去乙酰化酶抑制剂；IHC：免疫组织化学；LAMP2：溶酶体相关膜蛋白 2；LCFAs：长链脂肪酸；LFQ-MS：无标记定量质谱；LPC：溶血磷脂酰胆碱；MAP1LC3B/LC3B：微管相关蛋白 1 轻链 3β；MTOR：雷帕霉素靶蛋白激酶；PBD：过氧化物酶体生物发生障碍；PTS1：过氧化物酶体靶向信号 1；ROS：活性氧；sgRNA：单指导 RNA；VLCFAs：非常长链脂肪酸；Vor：伏立诺他；WO：冲洗。
