NEK4 kinase regulates EMT to promote lung cancer metastasis.

Epithelial-to-mesenchymal transition (EMT) is a dynamic transitional state from the epithelial to mesenchymal phenotypes. Numerous studies have suggested that EMT and its intermediate states play important roles in tumor invasion and metastasis. To identify novel regulatory molecules of EMT, we screened a siRNA library targeting human 720 kinases in A549 lung adenocarcinoma cells harboring E-cadherin promoter-luciferase reporter vectors. NIMA-related kinase-4 (NEK4) was identified and characterized as a positive regulator of EMT in the screening. Suppression of NEK4 resulted in the inhibition of cell migration and invasion, accompanying with an increased expression of cell adhesion-related proteins such as E-cadherin and ZO1. Furthermore, NEK4 knockdown caused the decreased expression of the transcriptional factor Zeb1 and Smads proteins, which are known to play key roles in EMT regulation. Consistently, overexpression of NEK4 resulted in the decreased expression of E-cadherin and increased expression of Smad3. Using a mouse model with tail vein injection of NEK4 knockdown stable cell line, we found a lower rate of tumor formation and metastasis of the NEK4-knockdown cells in vivo. Thus, this study demonstrates NEK4 as a novel kinase involved in regulation of EMT and suggests that NEK4 may be further explored as a potential therapeutic target for lung cancer metastasis.

Heterozygous p53-R280T Mutation Enhances the Oncogenicity of NPC Cells Through Activating PI3K-Akt Signaling Pathway.

A heterozygous point mutation of p53 gene at codon 280 from AGA to ACA (R280T) frequently occurs in nasopharyngeal carcinoma (NPC) cell lines, and about 10% NPC tissues. However, the role of this mutation in the pathogenesis of NPC remains unclear. In this study, we generated p53 knockout (KO) NPC cell lines from CNE2 cells carrying heterozygous p53 R280T (p53-R280T) mutation and C666-1 cells carrying wild-type p53 by CRISPR-Cas9 gene editing system, and found that KO of heterozygous p53-R280T significantly decreased NPC cell proliferation and increased NPC cell apoptosis, whereas KO of wild-type p53 had opposite effects on NPC cell proliferation and apoptosis. Moreover, KO of heterozygous p53-R280T inhibited the anchorage-independent growth and in vivo tumorigenicity of NPC cells. mRNA sequencing of heterozygous p53-R280T KO and control CNE2 cells revealed that heterozygous p53-R280T mutation activated PI3K-Akt signaling pathway. Moreover, blocking of PI3K-Akt signaling pathway abolished heterozygous p53-R280T mutation-promoting NPC cell proliferation and survival. Our data indicate that p53 with heterozygous R280T mutation functions as an oncogene, and promotes the oncogenicity of NPC cells by activating PI3K-Akt signaling pathway.

m6A Reader YTHDC2 Promotes Radiotherapy Resistance of Nasopharyngeal Carcinoma via Activating IGF1R/AKT/S6 Signaling Axis.

N6-methyladenosine (m6A) modification has been reported as a critical regulator of gene transcript expression. Although m6A modification plays important roles in tumor development, its role in therapeutic resistance remains unknown. In this study, we aimed to examine the expression level of m6A-modification related proteins and elucidate the effect of m6A-related proteins on radiation response in nasopharyngeal carcinoma (NPC). Among the genes that participated in m6A modification, YTHDC2, a m6A reader, was found to be consistently highly expressed in radioresistant NPC cells. Knocking down of YTHDC2 expression in radioresistant NPC cells improved the therapeutic effect of radiotherapy in vitro and in vivo, whereas overexpression of YTHDC2 in radiosensitive NPC cells exerted an opposite effect. Bioinformatics and mechanistic studies revealed that YTHDC2 could physically bound to insulin-like growth factor 1 receptor (IGF1R) messenger RNA and promoted translation initiation of IGF1R mRNA, which in turn activated the IGF1R-AKT/S6 signaling pathway. Thus, the present study suggests that YTHDC2 promotes radiotherapy resistance of NPC cells by activating the IGF1R/ATK/S6 signaling axis and may serve as a potential therapeutic target in radiosensitization of NPC cells.

Downregulation of Fat Mass and Obesity Associated (FTO) Promotes the Progression of Intrahepatic Cholangiocarcinoma.

Intrahepatic cholangiocarcinoma (ICC) ranks as the second most malignant type of primary liver cancer with a high degree of incidence and a very poor prognosis. Fat mass and obesity-associated protein (FTO) functions as an eraser of the RNA m6A modification, but its roles in ICC tumorigenesis and development remain unknown. We showed here that the protein level of FTO was downregulated in clinical ICC samples and cell lines and that FTO expression was inversely correlated with the expression of CA19-9 and micro-vessel density (MVD). A Kaplan-Meier survival analysis showed that a low expression of FTO predicted poor prognosis in ICC. in vitro, decreased endogenous expression of FTO obviously reduced apoptosis of ICC cells. Moreover, FTO suppressed the anchorage-independent growth and mobility of ICC cells. Through mining the database, FTO was found to regulate the integrin signaling pathway, inflammation signaling pathway, epidermal growth factor receptor (EGFR) signaling pathway, angiogenesis, and the pyrimidine metabolism pathway. RNA decay assay showed that oncogene TEAD2 mRNA stability was impaired by FTO. In addition, the overexpression of FTO suppressed tumor growth in vivo. In conclusion, our study demonstrated the critical roles of FTO in ICC.