Lyndon Lee

Introduction: Epithelial-to-mesenchymal transition (EMT) is implicated in the acquisition of an invasive and treatment resistant phenotype in cancer, and is characterized by the loss epithelial characteristics.^1-3 Mesenchymal characteristics include the gain of a leading/trailing edge, upregulation of mesenchymal markers and the formation of migratory protrusions.^1-3 Transforming growth factor β1 (TGF-β1) promotes EMT, whereas multiple miRNA families including let-7, miR-200, and miR-138 suppress EMT and other stages of carcinogenesis.^4-9 Multiple studies discuss a possible mechanism by which miRNA suppression of EMT is inhibited, in which long non-coding RNA H19 (LncRNA H19) acts as a molecular sponge that binds and suppresses let-7, mir-200, and miR-138.^10,11  Methods: Through bioinformatics analysis, Kallen identified four potential binding sites on H19 to let-7.¹⁰ A dual-reporter luciferase plasmid was generated with partial or complete H19 sequences embedded in the 3’-UTR of the Renilla luciferase (Rluc) reporter and an unmodified Firefly luciferase reporter.¹⁰ Co-transfection of the plasmid with let-7 sequences was performed to determine whether Rluc harboring H19 sequences would exhibit diminished activity.¹⁰ qRT-PCR was performed to assess HMGA2 mRNA, a let-7 target, after H19 overexpression.¹⁰ Liang assessed H19 expression after TGF-β1 stimulation.¹¹ Further experiments involved overexpression and knockdown of H19 to assess invasiveness, differential gene expression and morphological changes.¹¹ Vimentin, ZEB1, and ZEB2 mRNA and protein levels were assessed after miR-138 and miR-200 treatment, and again after ectopic H19 expression in miRNA-treated cells.¹¹ H19 RNA levels were assessed after miR-138 and miR-200 treatment as well.¹¹ Sorin sought a therapeutic application for H19 overexpression and generated a plasmid containing the diphtheria toxin A gene under regulatory control of the H19 promoter region.^12,13  Results and Discussion: Kallen found that Let-7 suppressed Rluc activity, and mutation of the predicted binding sites on H19 to let-7 resulted in insignificant changes in reporter activity. Overexpression of H19 resulted in derepression of let-7 target HMGA2. TGF-β1 stimulation resulted in elevated H19 expression, and similar interactions were reported by Liang between H19 and miR-200 and miR-138. H19 overexpression resulted in EMT-like changes and increased invasiveness. H19 levels were unaffected by miR-138 and miR-200 expression. H19 was also shown to derepress vimentin, ZEB1, and ZEB2. Sorin. reported that the plasmid could selectively target tumor cells and resulted in reduced tumor size. Therefore, current evidence suggests that H19 functions as a decoy that binds and antagonizes let-7, miR-138, and miR-200 and drives EMT, and that it is a potential therapeutic target.

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2. Nitta T, Mitsuhashi T, Hatanaka Y, et al. Prognostic significance of epithelial–mesenchymal transition-related markers in extrahepatic cholangiocarcinoma: comprehensive immunohistochemical study using a tissue microarray. British Journal of Cancer. 2014;111(7):1363-1372. doi:10.1038/bjc.2014.415.
3. Sowa T, Menju T, Sonobe M, et al. Association between epithelial‐mesenchymal transition and cancer stemness and their effect on the prognosis of lung adenocarcinoma. Cancer Medicine. 2015;4(12):1853-1862. doi:10.1002/cam4.556.
4. Farazi TA, Hoell JI, Morozov P, Tuschl T. microRNAs in Human Cancer. Advances in experimental medicine and biology. 2013;774:1-20. doi:10.1007/978-94-007-5590-1_1.
5. Perdigão-Henriques R, Petrocca F, Altschuler G, et al. miR-200 promotes the mesenchymal to epithelial transition by suppressing multiple members of the Zeb2 and Snail1 transcriptional repressor complexes. Oncogene. 2016:35(2):158-172.d oi: 10.1038/onc.2015.69.
6. Peter ME. Let-7 and miR-200 microRNAs: Guardians against pluripotency and cancer progression. Cell cycle (Georgetown, Tex). 2009;8(6):843-852.
7. Gregory PA, Bracken CP, Smith E, et al. An autocrine TGF-β/ZEB/miR-200 signaling network regulates establishment and maintenance of epithelial-mesenchymal transition. Bronner-Fraser M, ed. Molecular Biology of the Cell. 2011;22(10):1686-1698. doi:10.1091/mbc.E11-02-0103.
8. Li J, Wang Q, Wen R, et al. MiR‐138 inhibits cell proliferation and reverses epithelial‐mesenchymal transition in non‐small cell lung cancer cells by targeting GIT1 and SEMA4C. Journal of Cellular and Molecular Medicine. 2015;19(12):2793-2805. doi:10.1111/jcmm.12666.
9. Liu X, Wang C, Chen Z, et al. MicroRNA-138 suppresses epithelial–mesenchymal transition in squamous cell carcinoma cell lines. Biochemical Journal. 2011;440(1):23-31. doi:10.1042/BJ20111006.
10. Kallen AN, Zhou X-B, Xu J, et al. The Imprinted H19 LncRNA Antagonizes Let-7 MicroRNAs. Molecular cell. 2013;52(1):10.1016/j.molcel.2013.08.027. doi:10.1016/j.molcel.2013.08.027.
11. Liang W-C, Fu W-M, Wong C-W, et al. The lncRNA H19 promotes epithelial to mesenchymal transition by functioning as miRNA sponges in colorectal cancer. Oncotarget. 2015;6(26):22513-22525.
12. Sorin V, Ohana P, Mizrahi A, et al. Regional therapy with DTA-H19 vector suppresses growth of colon adenocarcinoma metastases in the rat liver. International Journal of Oncology. 2011;39(6):1407-1412. doi:10.3892/ijo.2011.1171.
13. Sorin V, Ohana P, Gallula J, et al. H19-Promoter-Targeted Therapy Combined with Gemcitabine in the Treatment of Pancreatic Cancer. ISRN Oncology. 2012;2012:351750. doi:10.5402/2012/351750.