Extracellular vesicle-mediated delivery of miR-101 inhibits lung metastasis in osteosarcoma
Extracellular vesicle-mediated delivery of miR-101 inhibits lung metastasis in osteosarcoma
复制标题
细胞外囊泡介导的 miR-101 递送抑制骨肉瘤肺转移
DOI：
10.7150/thno.33482
复制
发表时间：
2020-01-01
期刊：
影响因子：
12.4
通讯作者：
Zhou, Yong
中科院分区：
文献类型：
--
作者：
Zhang, Kailiang;Dong, Chuan;Zhou, Yong
基本原理：细胞外囊泡（EV）已经成为能够稳定转移治疗性microRNA（miRNA）的细胞间通讯的新型介质，因此，EV作为癌症治疗的miRNA递送系统具有巨大的前景。此外，随着含有miRNA的EV分泌到循环中，血浆EV中含有的miRNA可能代表疾病的理想生物标志物。本研究的目的是表征一种潜在的肿瘤抑制miRNA，miR-101，并探索通过EV递送miR-101用于转移性骨肉瘤的体内治疗的潜力以及血浆EV包装的miR-101（EV-miR-101）水平预测骨肉瘤转移的潜在价值。研究方法：通过原位杂交（ISH）研究了骨肉瘤标本中miR-101表达与骨肉瘤进展的关系，并使用体内模型进一步研究了miR-101的潜在抑制作用。利用预测软件分析，通过定量逆转录聚合酶链反应（qRT-PCR）、蛋白质印迹和双荧光素酶试验探讨miR-101在骨肉瘤中的作用机制。用慢病毒颗粒转导脂肪组织来源的间充质基质细胞（AD-MSC）以获得富含miR-101的EV。采用Transwell法和骨肉瘤肺转移模型观察miR-101富集的EV对骨肉瘤侵袭和转移的影响。通过qRT-PCR检测骨肉瘤患者和健康对照者的血浆EV-miR-101水平。结果：miR-101在转移性骨肉瘤中的表达明显低于非转移性骨肉瘤。过表达miR-101的Saos-2细胞和过表达miR-101的SOSP-9607细胞注射到小鼠体内形成的转移性肺结节显著较少。随着miR-101表达的增加，B细胞淋巴瘤6（BCL 6）的mRNA和蛋白表达水平降低，发现miR-101通过直接靶向BCL 6发挥作用。AD-MSC被成功地工程化以分泌富含miR-101的EV。一旦被骨肉瘤细胞摄取，这些EV在体外显示出对细胞侵袭和迁移的抑制作用，并且全身施用这些EV有效地抑制了体内转移，没有显著的副作用。最后，EV-miR-101水平在骨肉瘤患者中低于健康对照，在有转移的骨肉瘤患者中甚至低于无转移的骨肉瘤患者。结论：miR-101可能通过下调BCL 6的表达而抑制骨肉瘤的发生。AD-MSC衍生的富含miR-101的EV代表了转移性骨肉瘤的潜在创新疗法。EV-miR-101也代表了骨肉瘤转移的有希望的循环生物标志物。
Rationale: Extracellular vesicles (EVs) have emerged as novel mediators of cell-to-cell communication that are capable of the stable transfer of therapeutic microRNAs (miRNAs), and thus, EVs hold immense promise as a miRNA delivery system for cancer therapy. Additionally, as miRNA-containing EVs are secreted into circulation, miRNAs contained within plasma EVs may represent ideal biomarkers for diseases. The objective of this study was to characterize a potential tumor suppressor miRNA, miR-101, and explore the potential of miR-101 delivery via EVs for in vivo therapy of metastatic osteosarcoma as well as the potential value of plasma EV-packaged miR-101 (EV-miR-101) level for predicting osteosarcoma metastasis. Methods: The relationship of miR-101 expression and osteosarcoma progression was investigated in osteosarcoma specimens by in situ hybridization (ISH), and the potential inhibitory effect of miR-101 was further investigated using in vivo models. Using prediction software analysis, the mechanism of action of miR-101 in osteosarcoma was explored using quantitative reverse transcription polymerase chain reaction (qRT-PCR), western blotting and dual-luciferase assay. Adipose tissue-derived mesenchymal stromal cells (AD-MSCs) were transduced with lentiviral particles to obtain miR-101-enriched EVs. A Transwell assay and lung metastasis models of osteosarcoma were used to observe the effect of miR-101-enriched EVs on osteosarcoma invasiveness and metastasis. Detection of plasma EV-miR-101 levels was carried out in osteosarcoma patients and healthy controls by qRT-PCR. Results: miR-101 expression was markedly lower in metastatic osteosarcoma specimens compared to non-metastatic specimens. Significantly fewer metastatic lung nodules were formed by Saos-2 cells overexpressing miR-101 and SOSP-9607 cells overexpressing miR-101 injected into mice. With increased miR-101 expression, B cell lymphoma 6 (BCL6) mRNA and protein expression levels were reduced, and miR-101 was found to exert its effects by directly targeting BCL6. AD-MSCs were successfully engineered to secrete miR-101-enriched EVs. Once taken up by osteosarcoma cells, these EVs showed suppressive effects on cell invasion and migration in vitro, and systemic administration of these EVs effectively suppressed metastasis in vivo with no significant side effects. Finally, the EV-miR-101 level was lower in osteosarcoma patients than in healthy controls and even lower in osteosarcoma patients with metastasis than in those without metastasis. Conclusion: Our data support the function of miR-101 as a tumor suppressor in osteosarcoma via downregulation of BCL6. AD-MSC derived miR-101-enriched EVs represent a potential innovative therapy for metastatic osteosarcoma. EV-miR-101 also represents a promising circulating biomarker of osteosarcoma metastasis.