- Letter to the Editor
- Open Access
MiR-194 functions as a tumor suppressor in laryngeal squamous cell carcinoma by targeting Wee1
Journal of Hematology & Oncologyvolume 10, Article number: 32 (2017)
The emerging roles of microRNAs (miRs) have been deeply investigated in cancer. However, the role of miR-194 in human laryngeal squamous cell carcinoma (LSCC) is still unclear. Here, we have demonstrated that miR-194 is significantly downregulated in LSCC tissues and cells, and overexpression of miR-194 inhibits the proliferation, migration, invasion, and drug resistance in LSCC cells. Moreover, Wee1 is identified as a novel direct target of miR-194. Ectopic expression of Wee1 at least in part overcomes the suppressive impacts of miR-194 on the malignant phenotypes of LSCC. Overall, our study provides new sights into the role of miR-194/Wee1 axis in LSCC and suggests a novel miR-194/Wee1-based clinical application for LSCC patients.
Letter to the editor
Laryngeal cancer is the fourteenth most prevalent type of malignancy worldwide in the male compared to its relative rare in the female. Laryngeal squamous cell carcinoma (LSCC) accounts for approximately 90% of all malignant tumors of the larynx . Deregulated microRNAs (miRs) are frequently demonstrated as biomarker or therapeutic target in LSCC tissues and cells, which may function as tumor suppressor or oncogene to regulate the malignancy of cancer . As one of frequently deregulated miRs in cancer, the expression and role of miR-194 in LSCC are still unknown [3, 4].
In the current study, we found that the expression level of miR-194 is significantly lower not only in two LSCC cell lines Hep-2 and KB-3-1 compared with normal human bronchial epithelial cell line 16HBE but also in clinical LSCC tissues compared with adjacent normal tissues and correlated with T stage, lymph node metastasis, and clinical stage, but not with age, tumor grades, and tumor primary locations (Fig. 1b–g, Additional file 1: Figure S1a and Table S1). The miR-194 levels could be a significant parameter to distinguish LSCC and adjacent normal tissues with an area under the ROC curve (AUC) of 0.676 (sensitivity = 63.64%, specificity = 72.72%; P = 0.004) (Fig. 1h). Kaplan–Meier analysis indicated that high miR-194 expression predicts a favorable outcome for LSCC patients (Fig. 1i). Functional assays showed that enforced expression of miR-194 inhibits the growth, migration, invasion, and drug resistance of Hep-2 and KB-3-1 cells in vitro (Additional file 1: Figure S2a–g). The data in the subcutaneous tumor model in nude mice revealed that overexpression of miR-194 significantly inhibited the growth of Hep-2 and KB-3-1 xenografts by the numbers of Ki67+ proliferating cells and CD31+ microvessels (Additional file 1: Figure S3a–d). We used three computational algorithms, including miRanda, PITA, and TargetScan in combination to identify the novel potential targets of miR-194. All three algorithms predicted Wee1 as a target gene of miR-194, which is a protein kinase to regulate cell cycle through inhibiting CDK1 by phosphorylation on its two different residues Thr14 and Tyr15 . Western blot analysis showed that ectopic expression of miR-194 in Hep-2 and KB-3-1 cells obviously downregulated the protein levels of Wee1 as well as the known targets XIAP and p27 [3, 6] (Fig. 2a). The predicted interaction between miR-194 and the target sites within the 3’-untranslated regions (3’UTR) of Wee1 was shown in Fig. 2b. We then performed luciferase assay to examine whether there is a direct interaction between miR-194 and Wee1. The wild type or mutant 3’UTR region of Wee1 were cloned into downstream of the firefly luciferase gene to generate the luciferase reporter vectors. As illustrated in Fig. 2c, ectopic expression of miR-194 significantly decreased the luciferase activity of wild type 3’UTR, but not mutant 3’UTR in Hep-2 and KB-3-1 cells, suggesting that the putative miRNA binding sites of Wee1 are responsible for this miRNA-mRNA interaction (Additional file 1: Figure S1b). Furthermore, analysis of proteins extracted from Hep-2 and KB-3-1 subcutaneous tumors in mice exhibited that the protein levels of Wee1 in miR-194-transduced tumors were remarkably lower than those in vector control (Fig. 2d). In addition, ectopic expression of Wee1 partially reverses the suppressive effects of miR-194 on LSCC cells (Additional file 1: Figure S4a–g).
In summary, our data reveal a potential suppressive role of miR-194 in LSCC by targeting Wee1 in vitro and in vivo. The clinical results indicate that miR-194 can be the potential diagnostic and prognostic biomarkers for LSCC. Our study provides new sights into the role of miR-194/Wee1 axis in LSCC and suggests a novel miR-194/Wee1-based clinical application for LSCC patients.
Laryngeal squamous cell carcinoma
Siegel RL, Miller KD, Jemal A. Cancer statistics, 2015. CA Cancer J Clin. 2015;65:5–29.
Li P, Liu H, Wang ZY, He F, Wang HF, Shi Z, Yang AK, Ye J. MicroRNAs in laryngeal cancer: implications for diagnosis, prognosis and therapy. Am J Transl Res. 2016;8:1935–44.
Wu X, Liu T, Fang O, Leach LJ, Hu X, Luo Z. miR-194 suppresses metastasis of non-small cell lung cancer through regulating expression of BMP1 and p27(kip1). Oncogene. 2014;33:1506–14.
Zhang M, Zhuang Q, Cui L. MiR-194 inhibits cell proliferation and invasion via repression of RAP2B in bladder cancer. Biomed Pharmacother. 2016;80:268–75.
Matheson CJ, Backos DS, Reigan P. Targeting WEE1 Kinase in Cancer. Trends Pharmacol Sci. 2016;37:872–81.
Zhao HJ, Ren LL, Wang ZH, Sun TT, Yu YN, Wang YC, Yan TT, Zou W, He J, Zhang Y, Hong J, Fang JY. MiR-194 deregulation contributes to colorectal carcinogenesis via targeting AKT2 pathway. Theranostics. 2014;4:1193–208.
We thank other members of Shi lab and Ye lab for their comments and critical reading of the manuscript.
This work was supported by funds from the Chinese National Natural Science Foundation Nos. 8161101021, 31271444 and No. 81201726 (Z. S.), the Guangdong Natural Science Funds for Distinguished Young Scholar No. 2014A030306001 (Z. S.) and No. 2014A030313057 (J. Y.), the Guangdong Special Support Program for Young Talent No. 2015TQ01R350 (Z. S.), the Science and Technology Program of Guangdong No. 2016A050502027 (Z. S.) and No. 2013B021800088 (J. Y.), and the Foundation for Research Cultivation and Innovation of Jinan University No. 21616119 (Z. S.).
Availability of data and materials
The datasets supporting the conclusions of this article are included within the article and its additional files.
PL, YY, HL, ZS, and JY designed the experiments, performed the experiments, analyzed the data, and wrote the paper. AKY, JMD, GMT, HFW, JGQ, WJZ, QWJ, DWZ, YC, MNW, JRH, and KW performed the experiments. All authors read and approved the final manuscript.
The authors declare that they have no competing interests.
Consent for publication
Ethics approval and consent to participate
This project was approved by the Ethics Committee of Sun Yat-sen University Cancer Center. Ethics, consent, and permissions: Informed consent was obtained from each patient enrolled in the study. All animal experimental procedures were approved by the Institutional Animal Care and Use Committee of Sun Yat-Sen University.
Supplemental data. (PDF 1209 kb)