Downregulation of ULK1 by microRNA-372 inhibits the survival of human pancreatic adenocarcinoma cells.

Dysregulation of microRNA (miRNA) expression in various cancers and their role in cancer progression is well documented. The purpose of this study was to investigate the biological role of miR-372 in human pancreatic adenocarcinoma (HPAC). We collected 20 pairs of HPAC tissues and adjacent non-cancerous tissues to detect miR-372 expression levels. We transfected BXPC-3 and PANC-1 cells with miR-372 inhibitor/mimics to study their effect on cell proliferation, apoptosis, invasion, migration and autophagy. In addition, miR-372 mimics and a tumor protein UNC51-like kinase 1 (ULK1) siRNA were co-transfected into BXPC-3 and PANC-1 cells to explore the mechanism of miR-372 and ULK1 on HPAC tumorigenesis. We found that the expression of miR-372 was markedly downregulated in HPAC cells compared to adjacent normal tissues. Furthermore, functional assays showed that miR-372 inhibited cell proliferation, invasion, migration and autophagy in BXPC-3 and PANC-1 cells. An inverse correlation between miR-372 expression and ULK1 expression was observed in HPAC tissues. Downregulation of ULK1 inhibited the overexpression effects of miR-372, and upregulation of ULK1 reversed the effects of overexpressed miR-372. Finally, we found that silencing ULK1 or inhibiting autophagy partly rescued the effects of miR-372 knockdown in HPAC cells, which may explain the influence of miR-372/ULK1 in HPAC development. Taken together, these results revealed a significant role of the miR-372/ULK1 axis in suppressing HPAC cell proliferation, migration, invasion and autophagy.

Chen H ，Zhang Z ，Lu Y ，Song K ，Liu X ，Xia F ，Sun W ... -  《-》

MicroRNA-217 inhibits cell proliferation, invasion and migration by targeting Tpd52l2 in human pancreatic adenocarcinoma.

Chen Q ，Wang P ，Fu Y ，Liu X ，Xu W ，Wei J ，Gao W ，Jiang K ，Wu J ，Miao Y ... -  《-》

miR-373 inhibits autophagy and further promotes apoptosis of cholangiocarcinoma cells by targeting ULK1.

Intrahepatic cholangiocarcinoma is a malignant tumor originating from intrahepatic bile ducts. Surgical therapy, radiotherapy, and chemotherapy are taken to treat this disease, but it is prone to recurrence and metastasis, with poor prognosis. Therefore, it is of great significance to explore new targets and molecular mechanisms for the development of cholangiocarcinoma cells. Clinical cholangiocarcinoma tissues from patients and four human cholangiocarcinoma cell lines were analyzed for microRNA-373 (miR-373) expression. For investigating whether miR-373 directly modulated unc-51 like autophagy activating kinase 1 (ULK1), dual-luciferase reporter assay was performed. In addition, CCK-8 assay, flow cytometry, western blot, and immunofluorescence were applied to evaluate the proliferation, apoptosis, and autophagy of cholangiocytic hepatocellular carcinoma cells. miR-373 downregulation was observed in clinical tissues and cell lines of cholangiocarcinoma. Overexpression of miR-373 reduced proliferation, enhanced apoptosis, and raised expression levels of pro-apoptosis proteins including BCL2 associated X (Bax), Caspase-3, and Caspase-9. Moreover, overexpression of miR-373 downregulated expression levels of microtubule-associated protein 1A/1B-light chain 3 (LC3)-II, Beclin-1, and promoted P62 expression on mRNA and protein levels. After miR-373 knockdown, all indexes of apoptosis and autophagy mentioned above were reversed. Luciferase activity was decreased after cotransfection of miR-373 mimic and wild-type ULK1 vector. Also, miR-373 overexpression inhibited ULK1 expression. Importantly, overexpression of miR-373 weakened expressions of ULK1, LC3, Beclin-1, and Bcl-2, and enhanced expressions of P62, Bax, Caspase-3, and Caspase-9. miR-373 mimic treatment and subsequent ULK1 overexpression, induced reverse regulation in expressions of these proteins, compared with overexpression of miR-373 only. miR-373 targeted ULK1 to initiate inhibition of autophagy and subsequent promotion of apoptosis in cholangiocarcinoma cells.

Lv P ，Luo YF ，Zhou WY ，Liu B ，Zhou Z ，Shi YZ ，Huang R ，Peng C ，He ZL ，Wang J ，Zhang HH ，Nie SD ... -  《-》

MicroRNA-106a targets autophagy and enhances sensitivity of lung cancer cells to Src inhibitors.

Src tyrosine kinase inhibitors (TKIs) significantly inhibit cell migration and invasion in lung cancer cell lines with minor cytotoxic effects. In clinical trials, however, they show modest activity in combination with chemotherapeutic agents. Possible resistance mechanisms include the induction of cytoprotective autophagy upon Src inhibition. Autophagy is a cellular recycling process that allows cell survival in response to a variety of stress stimuli including responses to various treatments. We screened autophagic activity in A549, H460, and H1299 NSCLC cell lines treated with two different Src-TKIs (saracatinib, dasatinib) or shRNA targeting SRC. The autophagy response was determined by LC3B-I to -II conversion, increased ULK1 epxression and increased GFP-LC3B dot formation. Autophagy was inhibited by pharmacological (bafilomycin A, chloroquine) or genetic (ULK1 shRNA) means. Expression of miR-106a and miR-20b was analyzed by qPCR, and we used different lentivral vectors for ectopic expression of either miR-106a mimetics, anti-sense miR-106a or different miR-106a-363 cluster constructs. In the current study we found that Src-TKIs induce autophagy in lung adenocarcinoma cell lines and that a combination of autophagy and Src tyrosine kinase inhibition results in cell death. Moreover, Src-TKI induced autophagy depends on the induction of the key autophagy kinase ULK1. This ULK1 upregulation is caused by downregulation of the ULK1-targeting microRNA-106a. An inverse correlation of miR-106a and ULK1 expression was seen in lung adenocarcinoma. Accordingly, ectopic expression of miR-106a in combination with Src-TKI treatment resulted in significant cell death as compared to control transduced cells. Autophagy protects lung adenocarcinoma cells from Src-TKIs via a newly identified miR-106a-ULK1 signaling pathway. The combined inhibition of Src and ULK1/autophagy might represent a promising treatment option for future clinical trials. Lastly, our data might challenge the term "oncogenic" miR-106a as it can promote sensitivity to Src-TKIs thereby underlining the context-dependent function of miRNAs.

Rothschild SI ，Gautschi O ，Batliner J ，Gugger M ，Fey MF ，Tschan MP ... -  《-》

LncRNA PVT1 triggers Cyto-protective autophagy and promotes pancreatic ductal adenocarcinoma development via the miR-20a-5p/ULK1 Axis.

Huang F ，Chen W ，Peng J ，Li Y ，Zhuang Y ，Zhu Z ，Shao C ，Yang W ，Yao H ，Zhang S ... -  《Molecular Cancer》