Tipanee Jaitip, Di Matteo Mario, Tulalamba Warut, Samara-Kuko Ermira, Keirsse Jiri, Van Ginderachter Jo A, Chuah Marinee Khim, VandenDriessche Thierry
Department of Gene Therapy & Regenerative Medicine, Vrije Universiteit Brussel, 1090 Brussels, Belgium.
Department of Gene Therapy & Regenerative Medicine, Vrije Universiteit Brussel, 1090 Brussels, Belgium; Center for Molecular & Vascular Biology, Department of Cardiovascular Sciences, University of Leuven, 3000 Leuven, Belgium.
Mol Ther Nucleic Acids. 2020 Mar 6;19:1309-1329. doi: 10.1016/j.omtn.2020.01.015. Epub 2020 Jan 22.
We established a semi-high-throughput in vivo screening platform using hyperactive piggyBac (hyPB) transposons (designated as PB-miR) to identify microRNAs (miRs) that inhibit hepatocellular carcinoma (HCC) development in vivo, following miR overexpression in hepatocytes. PB-miRs encoding six different miRs from the miR-17-92 cluster and nine miRs from outside this cluster were transfected into mouse livers that were chemically induced to develop HCC. In this slow-onset HCC model, miR-20a significantly inhibited HCC. Next, we developed a more aggressive HCC model by overexpression of oncogenic Harvey rat sarcoma viral oncogene homolog (HRAS) and c-MYC oncogenes that accelerated HCC development after only 6 weeks. The tumor suppressor effect of miR-20a could be demonstrated even in this rapid-onset HRAS/c-MYC HCC model, consistent with significantly prolonged survival and decreased HCC tumor burden. Comprehensive RNA expression profiling of 95 selected genes typically associated with HCC development revealed differentially expressed genes and functional pathways that were associated with miR-20a-mediated HCC suppression. To our knowledge, this is the first study establishing a direct causal relationship between miR-20a overexpression and liver cancer inhibition in vivo. Moreover, these results demonstrate that hepatocyte-specific hyPB transposons are an efficient platform to screen and identify miRs that affect overall survival and HCC tumor regression.
我们利用超活性猪尾巴（hyPB）转座子（命名为PB-miR）建立了一个半高通量体内筛选平台，以在肝细胞中过表达miR后，在体内鉴定抑制肝细胞癌（HCC）发展的微小RNA（miR）。将编码来自miR-17-92簇的六种不同miR和来自该簇外的九种miR的PB-miR转染到化学诱导发生HCC的小鼠肝脏中。在这个缓慢发生的HCC模型中，miR-20a显著抑制了HCC。接下来，我们通过过表达致癌性哈维大鼠肉瘤病毒癌基因同源物（HRAS）和c-MYC癌基因，开发了一种更具侵袭性的HCC模型，该模型在仅6周后就加速了HCC的发展。即使在这种快速发生的HRAS/c-MYC HCC模型中，也能证明miR-20a的肿瘤抑制作用，这与显著延长的生存期和降低的HCC肿瘤负担一致。对95个通常与HCC发展相关的选定基因进行的综合RNA表达谱分析揭示了与miR-20a介导的HCC抑制相关的差异表达基因和功能途径。据我们所知，这是第一项在体内建立miR-20a过表达与肝癌抑制之间直接因果关系的研究。此外，这些结果表明，肝细胞特异性hyPB转座子是筛选和鉴定影响总体生存和HCC肿瘤消退的miR的有效平台。
