Background: TGF-β promotes tumor invasion and metastasis through inducing epithelial-mesenchymal transition (EMT) in non-small cell lung cancer (NSCLC). Circular RNAs (circRNAs) are recognized as functional non-coding RNAs involved in human cancers. However, whether and how circRNAs contribute to TGF-β-induced EMT and metastasis in NSCLC remain vague. Here, we investigated the regulation and function of Circular RNA hsa_circ_0008305 (circPTK2) in TGF-β-induced EMT and tumor metastasis, as well as a link between circPTK2 and transcriptional intermediary factor 1 γ (TIF1γ) in NSCLC. Methods: Circular RNAs were determined by human circRNA Array analysis, real-time quantitative reverse transcriptase PCR and northern blot. Luciferase reporter, RNA-binding protein immunoprecipitation (RIP), RNA pull-down and fluorescence in situ hybridization (FISH) assays were employed to test the interaction between circPTK2 and miR-429/miR-200b-3p. Ectopic overexpression and siRNA-mediated knockdown of circPTK2, TGF-β-induced EMT, Transwell migration and invasion in vitro, and in vivo experiment of metastasis were used to evaluate the function of circPTK2. Transcription and prognosis analyses were done in public databases. Results: CircPTK2 and TIF1γ were significantly down-regulated in NSCLC cells undergoing EMT induced by TGF-β. CircPTK2 overexpression augmented TIF1γ expression, inhibited TGF-β-induced EMT and NSCLC cell invasion, whereas circPTK2 knockdown had the opposite effects. CircPTK2 functions as a sponge of miR-429/miR-200b-3p, and miR-429/miR-200b-3p promote TGF-β-induced EMT and NSCLC cell invasion by targeting TIF1γ. CircPTK2 overexpression inhibited the invasion-promoting phenotype of endogenous miR-429/miR-200b-3p in NSCLC cells in response to TGF-β. CircPTK2 overexpression significantly decreased the expression of Snail, an important downstream transcriptional activator of TGF-β/Smad signaling. In an in vivo experiment of metastasis, circPTK2 overexpression suppressed NSCLC cell metastasis. Moreover, circPTK2 expression was dramatically down-regulated and positively correlated with TIF1γ expression in human NSCLC tissues. Especially, circPTK2 was significantly lower in metastatic NSCLC tissues than non-metastatic counterparts. Conclusion: Our findings show that circPTK2 (hsa_circ_0008305) inhibits TGF-β-induced EMT and metastasis by controlling TIF1γ in NSCLC, revealing a novel mechanism by which circRNA regulates TGF-β-induced EMT and tumor metastasis, and suggesting that circPTK2 overexpression could provide a therapeutic strategy for advanced NSCLC.

CD73 (ecto-5′-nucleotidase) is implicated in the development of many types of cancer. CD73 inhibitors are currently being tested in clinical trials for the treatment of cancer. Understanding the molecular and cellular actions of CD73 inhibitors is the key to improving this line of therapy. Quantitative real-time PCR (qRT-PCR) was used to detect the expression of CD73 and miR-30a-5p; Western blot and immunohistochemical assays were used to investigate the levels of CD73 and other proteins. Flow cytometry was used to determine cell cycle stage and apoptosis. CCK-8 and clonogenic assays were used to investigate cell proliferation. Wound healing, migration and invasion assays were used to investigate the motility of cells. A lung carcinoma xenograft mouse model was used to investigate the in vivo effects of CD73 and miR-30a-5p. In the present study, we found that CD73 is overexpressed and miR-30a-5p is underexpressed in non-small cell lung cancer tissues compared with adjacent noncancerous. Further, we showed that CD73 is a direct target of miR-30a-5p by luciferase reporter assays, qRT-PCR and western blot analysis. We also found that overexpression of miR-30a-5p in these non-small cell lung cancer cell lines inhibited cell proliferation in vitro and in vivo. Moreover, the epithelial-to-mesenchymal phenotype was suppressed and cell migration and invasion were inhibited; these effects were brought about via the EGF signaling pathway. Our findings reveal a new post-transcriptional mechanism of CD73 regulation via miR-30a-5p and EGFR-related drug resistance in non-small cell lung cancer.

The molecular underpinnings of lung adenocarcinoma (LUAD) metastasis remain poorly defined. Here, using human LUAD cell lines, we find that transcriptional intermediary factor 1 γ (TIF1γ) binds to TATA box binding protein (TBP) in competition with TBP-associated factor 15 (TAF15) and impedes TAF15/TBP-mediated interleukin 6 (IL-6) transactivation. TIF1γ modifies TAF15 through multi-mono-ubiquitylation and drives nuclear export of TAF15. Functionally, TAF15 accelerates epithelial-mesenchymal transition (EMT) and metastasis of LUAD cells, acting in just the opposite way as TIF1γ. Low TIF1γ or high TAF15 expression levels are shown in metastatic LUAD specimens and correlate with poor survival of individuals with LUAD. Our findings suggest that the TAF15/TBP complex is required for IL-6 activation-induced EMT and invasion, which are inhibited by TIF1γ. This study highlights the crucial interaction between TIF1γ and the TAF15/TBP complex for regulating EMT and metastasis in LUAD.

Increased evidence reveals that miR-940 inhibits the migration and invasion of cancer cells. Considering transforming growth factor β (TGF-β) signaling is crucial to cellular epithelial-mesenchymal transition (EMT) process and metastasis of cancer, it is in urgent to explore whether and how miR-940 plays an essential role in regulating TGF-β-induced EMT in lung cancer progression. In the present study, we observed a reciprocal expression with down-regulated miR-940 and up-regulated Snail mRNA in non-small-cell lung cancer (NSCLC) tissues. we further found that the expression of miR-940 was decreased in NSCLC tissues with lymph node metastasis, advanced TNM stages and poor cell differentiation, in which, on the contrary, the expression of Snail was increased. Overexpression of miR-940 significantly inhibited Snail mRNA and protein expression in A549 and H226 cells. Mechanistically, Snail mRNA was identified as target of miR-940. In addition, miR-940 repressed TGF-β-induced EMT and further hampered the cell migration and invasion. Finally, siRNA-mediated knockdown of Snail copied the phenotype of miR-940 overexpression in A549 and H226 cells. Taken together, our study reveals that miR-940 can suppress TGF-β-induced EMT and cell invasion by targeting Snail 3'-UTR mRNA in NSCLC.