MiR-20a Was Identified As A Key Molecule Triggering Metastasis Of Gallbladder Carcinoma

Part I: miR-20a was Identified as A Key Molecule Triggering Metastasisof Gallbladder Carcinoma by High Content ScreeningBackground&Aims: Gallbladder carcinoma(GBC) is one of the most aggressive and lethaltumors, with extremely high metastatic activity and poor prognosis clinically. Thedysfunction of miRNAs has been demonstrated participating in the regulation of varioustumor development. However, whether miRNAs are involved in metastasis and progressionof GBC remains obscure. We plan to thoroughly explore critical miRNAs involved inmetastasis and progression of GBC.Methods: A new designed high content miRNAs screening technology was applied for thefirst time to filter out pro-metastatic miRNAs in GBC. The biological functions of miRNAswere intensively studied by transwell, immunoblot, immunohistochemical and in situhybridization assays. Tumorigenicity and liver metastasis were further examined in nudemice.Results: Of880miRNAs,17were selected as the prominent metastatic inducers for GBCs.Among them, the up-regulation of miR-20a in tumor tissues showed tight association with thelocal invasion, distant metastasis and poor prognosis of GBCs clinically. The ectopicexpression of miR-20a could induce Epithelial-mesenchymal transition(EMT) and enhancethe metastasis of GBC-SD cells in vitro and in vivo, through directly targeting3’UTR ofSmad7and promoting nucleus translocation of β-catenin subsequently. Conversely, theblockage of miR-20a effectively restored the expression of Smad7and attenuatedTGF-β1-induced cell metastasis. The significant correlation between abnormal miR-20aexpression and the absence of Smad7in GBC patients suggested that the combination of thesetwo parameters has more powerful predictive value for the progression of GBCs.Conclusions: TGF-β1-mediated activation of miR-20a/Smad7/β-catenin axis plays a pivotalrole in the pathogenesis and worse prognosis of GBCs, which may serve as a potentialtherapeutic target in future. Part II: Inhibition of Autophagy may Suppress the Development ofHepatoblastomaBackground&Aims: Hepatoblastoma (HB) is a rare cancer but represents the mostcommon liver malignancy in children under3years of age. Nevertheless, a clearunderstanding of the pathogenesis is lacking. Although the treatment of HB has beendramatically improved by combining chemotherapy regimens with surgery, its fatal outcomeof fast development and recurrence makes new treatment strategies for HB, based on animproved understanding of the pathogenesis, essential. Autophagy is believed to be importantin the progression of cancers. However, the role of autophagy in HB remains to be elucidated.Methods: Transmission electron microscopy (TEM) was used to examine the accumulationof autophagosomes in HB tissues. The activity of autophagy was then examined by real-timePCR, western blot, immunohistochemistry, GFP-LC3assay. Next, the role of autophagy inHB was elucidated under nutrition deprivation and chemical-therapy.3-MA, wortmannin,rapamycin and siRNA transfection were used to inhibit the activity of autophagy. Thenxenograft tumor model in nude mice was employed to investigate the role of autophagy invivo.Results: Here, we show that autophagy is activated in HB tissues and cells under theconditions of starvation or chemotherapy, coupled with the over-expression ofautophagic-related genes BECN1and ATG5. Suppression of autophagy with pharmacologicalagents and small interfering RNAs significantly increased cell apoptosis and retardedproliferation in response to nutrition deprivation and treatment with chemotherapeutics.Conclusions: Our data demonstrate that the BECN1and ATG5-dependent phosphoinositide3-kinase (PI3K) signaling pathway is essential for the survival of HB cells and their toleranceto chemotherapy and starvation-induced death, and suggests that modifying such autophagicgenes may suppress the development of HB, thus offering a therapeutic potential for patientswith HB.