Thor Theresa, Künkele Annette, Pajtler Kristian W, Wefers Annika K, Stephan Harald, Mestdagh Pieter, Heukamp Lukas, Hartmann Wolfgang, Vandesompele Jo, Sadowski Natalie, Becker Lore, Garrett Lillian, Hölter Sabine M, Horsch Marion, Calzada-Wack Julia, Klein-Rodewald Tanja, Racz Ildiko, Zimmer Andreas, Beckers Johannes, Neff Frauke, Klopstock Thomas, De Antonellis Pasqualino, Zollo Massimo, Wurst Wolfgang, Fuchs Helmut, Gailus-Durner Valérie, Schüller Ulrich, de Angelis Martin Hrabě, Eggert Angelika, Schramm Alexander, Schulte Johannes H
Department of Pediatric Oncology and Hematology, University Children's Hospital Essen, Hufelandstr. 55 45147, Essen, Germany; German Cancer Consortium (DKTK), Heidelberg, Germany; German Cancer Research Center (DKFZ), D-69120, Heidelberg, Germany.
Int J Cancer. 2015 May 15;136(10):2293-303. doi: 10.1002/ijc.29294. Epub 2014 Nov 25.
Previous studies have evaluated the role of miRNAs in cancer initiation and progression. MiR-34a was found to be downregulated in several tumors, including medulloblastomas. Here we employed targeted transgenesis to analyze the function of miR-34a in vivo. We generated mice with a constitutive deletion of the miR-34a gene. These mice were devoid of mir-34a expression in all analyzed tissues, but were viable and fertile. A comprehensive standardized phenotypic analysis including more than 300 single parameters revealed no apparent phenotype. Analysis of miR-34a expression in human medulloblastomas and medulloblastoma cell lines revealed significantly lower levels than in normal human cerebellum. Re-expression of miR-34a in human medulloblastoma cells reduced cell viability and proliferation, induced apoptosis and downregulated the miR-34a target genes, MYCN and SIRT1. Activation of the Shh pathway by targeting SmoA1 transgene overexpression causes medulloblastoma in mice, which is dependent on the presence and upregulation of Mycn. Analysis of miR-34a in medulloblastomas derived from ND2:SmoA1(tg) mice revealed significant suppression of miR-34a compared to normal cerebellum. Tumor incidence was significantly increased and tumor formation was significantly accelerated in mice transgenic for SmoA1 and lacking miR-34a. Interestingly, Mycn and Sirt1 were strongly expressed in medulloblastomas derived from these mice. We here demonstrate that miR-34a is dispensable for normal development, but that its loss accelerates medulloblastomagenesis. Strategies aiming to re-express miR-34a in tumors could, therefore, represent an efficient therapeutic option.
以往的研究评估了微小RNA（miRNA）在癌症发生和发展中的作用。研究发现，miR-34a在包括髓母细胞瘤在内的多种肿瘤中表达下调。在此，我们采用靶向转基因技术在体内分析miR-34a的功能。我们构建了miR-34a基因组成型缺失的小鼠。这些小鼠在所有分析的组织中均无miR-34a表达，但可存活且能生育。一项包括300多个单一参数的全面标准化表型分析未发现明显的表型。对人类髓母细胞瘤和髓母细胞瘤细胞系中miR-34a表达的分析显示，其水平明显低于正常人类小脑。在人类髓母细胞瘤细胞中重新表达miR-34a可降低细胞活力和增殖，诱导细胞凋亡，并下调miR-34a的靶基因MYCN和SIRT1。通过靶向SmoA1转基因过表达激活Shh信号通路可导致小鼠发生髓母细胞瘤，这依赖于Mycn的存在和上调。对源自ND2:SmoA1（tg）小鼠的髓母细胞瘤中miR-34a的分析显示，与正常小脑相比，miR-34a受到显著抑制。在转基因表达SmoA1且缺乏miR-34a的小鼠中，肿瘤发生率显著增加，肿瘤形成明显加速。有趣的是，Mycn和Sirt1在源自这些小鼠的髓母细胞瘤中强烈表达。我们在此证明，miR-34a对正常发育并非必需，但其缺失会加速髓母细胞瘤的发生。因此，旨在使肿瘤中重新表达miR-34a的策略可能代表一种有效的治疗选择。
