Warburg Effect-Promoted Exosomal Circ_0072083 Releasing Up-Regulates Nango Expression Through Multiple Pathways and Enhances Temozolomide Resistance in Glioma

Abstract

BackgroundTemozolomide (TMZ) resistance limits its application in glioma. Exosome can carry circular RNAs (circRNAs) to regulate drug resistance via sponging microRNAs (miRNAs). miRNAs can control mRNA expression by regulate the interaction with 3’UTR and methylation. Nanog homeobox (NANOG) is an important biomarker for TMZ resistance. Hitherto, it is unknown about the role of exosomal hsa_circ_0072083 (circ_0072083) in TMZ resistance in glioma, and whether it is associated with NANOG via regulating miRNA sponge and methylation.MethodsTMZ-resistant (n=36) and sensitive (n=33) patients were recruited. The sensitive cells and constructed resistant cells were cultured and exposed to TMZ. circ_0072083, miR-1252-5p, AlkB homolog H5 (ALKBH5) and NANOG levels were examined via quantitative reverse transcription polymerase chain reaction and western blot. The half maximal inhibitory concentration (IC50) of TMZ, cell proliferation, apoptosis, migration and invasion were analyzed via Cell Counting Kit-8, colony formation, flow cytometry, wound healing and transwell assays. The in vivo function was assessed using xenograft model. The N6-methyladenosine (m6A) level was analyzed via methylated RNA immunoprecipitation (MeRIP). Target relationship was investigated via dual-luciferase reporter assay and RNA immunoprecipitation. Warburg effect was investigated via lactate production, glucose uptake and key enzymes expression. Exosome was isolated and confirmed via transmission electron microscopy and specific protein expression.Resultscirc_0072083 expression was increased in TMZ-resistant glioma tissues and cells. circ_0072083 knockdown restrained the resistance of resistant cells via decreasing IC50 of TMZ, proliferation, migration, invasion and xenograft tumor growth and increasing apoptosis. circ_0072083 silence reduced NANOG expression via blocking ALKBH5-mediated demethylation. circ_0072083 could regulate NANOG and ALKBH5 via targeting miR-1252-5p to control TMZ resistance. Warburg effect promoted the release of exosomal circ_0072083 in resistant cells. Exosomal circ_0072083 from resistant cells increased the resistance of sensitive cells to TMZ in vitro and xenograft model. Exosomal circ_0072083 level was enhanced in resistant patients, and it had a diagnostic value and indicated a lower overall survival in glioma. ConclusionExosomal circ_0072083 promoted TMZ resistance via increasing NANOG via regulating miR-1252-5p-mediated degradation and demethylation in glioma.

Circular RNAs in eukaryotic cells: origin, characteristics, mechanisms of molecular functioning in human malignant diseases

“…Показано, что РСК обладают многими общими характеристиками с регенеративными стволовыми клетками, такими как самообновление, мультипотентность и обратимость их состояния покоя [17,309]. После химиотерапии остаются остаточные опухолевые фрагменты, обогащенные РСК, которые могут способствовать рецидиву опухоли, повторному росту опухолевых метастазов и повышению устойчивости к лекарственным препаратам [25,71].…”

Roles of RNA alternative splicing during the occurrence and development of lung cancer

“…表型，帮助肿瘤细胞在不利的环境下生长 [73] 。Warburg 效应在肿瘤细胞转移、增 殖、免疫逃逸、耐药等方面发挥着重要作用 [74][75][76][77] ，同时该效应也被诸多因素所影 响，其中包括可变剪接。 微阵列研究表明，糖酵解基因是癌症中上调率最高的基因之一，其中包括糖 酵解最后一步的限速酶-丙酮酸激酶(PK) [78,79] 。哺乳动物中存在四种丙酮酸 激酶亚型，L 型和 R 型在肝脏和红细胞中表达，M1 亚型在成人大多数组织中表 达，而 M2 亚型是 M1 亚型的剪接异构体，在胚胎发育过程中表达 [80] 。在癌前病 变的组织中，主要表达 PKM1，而 PKM2 主要在肿瘤组织中表达，且在肺癌细胞 系 A549 中，PKM2 高表达，PKM1 则几乎不表达 [81]…”