SummaryThe recently discovered mitochondrial calcium uniporter (MCU) promotes Ca2+ accumulation into the mitochondrial matrix [1, 2]. We identified in silico miR-25 as a cancer-related MCU-targeting microRNA family and demonstrate that its overexpression in HeLa cells drastically reduces MCU levels and mitochondrial Ca2+ uptake, while leaving other mitochondrial parameters and cytosolic Ca2+ signals unaffected. In human colon cancers and cancer-derived cells, miR-25 is overexpressed and MCU accordingly silenced. miR-25-dependent reduction of mitochondrial Ca2+ uptake correlates with resistance to apoptotic challenges and can be reversed by anti-miR-25 overexpression. Overall, the data demonstrate that microRNA targeting of mitochondrial Ca2+ signaling favors cancer cell survival, thus providing mechanistic insight into the role of mitochondria in tumorigenesis and identifying a novel therapeutic target in neoplasia.

1MicroRNA-221 (miR-221) is one of the most frequently and consistently up-regulated microRNAs (miRNAs) in human cancer. It has been hypothesized that miR-221 may act as a tumor promoter. To demonstrate this, we developed a transgenic (TG) mouse model that exhibits an inappropriate overexpression of miR-221 in the liver. Immunoblotting and immunostaining confirmed a concomitant down-regulation of miR-221 target proteins. This TG model is characterized by the emergence of spontaneous nodular liver lesions in approximately 50% of male mice and by a strong acceleration of tumor development in 100% of mice treated with diethylnitrosamine. Similarly to human hepatocellular carcinoma, tumors are characterized by a further increase in miR-221 expression and a concomitant inhibition of its target protein-coding genes (i.e., cyclin-dependent kinase inhibitor [Cdkn]1b/p27, Cdkn1c/p57, and B-cell lymphoma 2-modifying factor). To validate the tumor-promoting effect of miR-221, we showed that in vivo delivery of anti-miR-221 oligonucleotides leads to a significant reduction of the number and size of tumor nodules. Conclusions: This study not only establishes that miR-221 can promote liver tumorigenicity, but it also establishes a valuable animal model to perform preclinical investigations for the use of anti-miRNA approaches aimed at liver cancer therapy. (HEPATOLOGY 2012;56:1025-1033

The release of circulating tumor cells (CTCs) into vasculature is an early event in the metastatic process. The analysis of CTCs in patients has recently received widespread attention because of its clinical implications, particularly for precision medicine. Accumulated evidence documents a large heterogeneity in CTCs across patients. Currently, the most accepted view is that tumor cells with an intermediate phenotype between epithelial and mesenchymal have the highest plasticity. Indeed, the existence of a meta-stable or partial epithelial–mesenchymal transition (EMT) cell state, with both epithelial and mesenchymal features, can be easily reconciled with the concept of a highly plastic stem-like state. A close connection between EMT and cancer stem cells (CSC) traits, with enhanced metastatic competence and drug resistance, has also been described. Accordingly, a subset of CTCs consisting of CSC, present a stemness profile, are able to survive chemotherapy, and generate metastases after xenotransplantation in immunodeficient mice. In the present review, we discuss the current evidence connecting CTCs, EMT, and stemness. An improved understanding of the CTC/EMT/CSC connections may uncover novel therapeutic targets, irrespective of the tumor type, since most cancers seem to harbor a pool of CSCs, and disclose important mechanisms underlying tumorigenicity.

Non-coding RNAs (ncRNAs) have been implicated in most cellular functions. The disruption of their function through somatic mutations, genomic imprinting, transcriptional and post-transcriptional regulation, plays an ever-increasing role in cancer development. ncRNAs, including notorious microRNAs, have been thus proposed to function as tumor suppressors or oncogenes, often in a context-dependent fashion. In parallel, ncRNAs with altered expression in cancer have been reported to exert a key role in determining drug sensitivity or restoring drug responsiveness in resistant cells. Acquisition of resistance to anti-cancer drugs is a major hindrance to effective chemotherapy and is one of the most important causes of relapse and mortality in cancer patients. For these reasons, non-coding RNAs have become recent focuses as prognostic agents and modifiers of chemo-sensitivity. This review starts with a brief outline of the role of most studied non-coding RNAs in cancer and then highlights the modulation of cancer drug resistance via known ncRNAs based mechanisms. We identified from literature 388 ncRNA-drugs interactions and analyzed them using an unsupervised approach. Essentially, we performed a network analysis of the non-coding RNAs with direct relations with cancer drugs. Within such a machine-learning framework we detected the most representative ncRNAs-drug associations and groups. We finally discussed the higher integration of the drug-ncRNA clusters with the goal of disentangling effectors from downstream effects and further clarify the involvement of ncRNAs in the cellular mechanisms underlying resistance to cancer treatments.

BackgroundThe microRNA 125b is a double-faced gene expression regulator described both as a tumor suppressor gene (in solid tumors) and an oncogene (in hematologic malignancies). In human breast cancer, it is one of the most down-regulated miRNAs and is able to modulate ERBB2/3 expression. Here, we investigated its targets in breast cancer cell lines after miRNA-mimic transfection. We examined the interactions of the validated targets with ERBB2 oncogene and the correlation of miR-125b expression with clinical variables.MethodsMiR-125b possible targets were identified after transfecting a miRNA-mimic in MCF7 cell line and analyzing gene expression modifications with Agilent microarrays and Sylamer bioinformatic tool. Erythropoietin (EPO) and its receptor (EPOR) were validated as targets of miR-125b by luciferase assay and their expression was assessed by RT-qPCR in 42 breast cancers and 13 normal samples. The molecular talk between EPOR and ERBB2 transcripts, through miR-125b, was explored transfecting MDA-MD-453 and MDA-MB-157 with ERBB2 RNA and using RT-qPCR.ResultsWe identified a panel of genes down-regulated after miR-125b transfection and putative targets of miR-125b. Among them, we validated erythropoietin (EPO) and its receptor (EPOR) - frequently overexpressed in breast cancer - as true targets of miR-125b. Moreover, we explored possible correlations with clinical variables and we found a down-regulation of miR-125b in metastatic breast cancers and a significant positive correlation between EPOR and ERBB2/HER2 levels, that are both targets of miR-125b and function as competing endogenous RNAs (ceRNAs).ConclusionsTaken together our results show a mechanism for EPO/EPOR and ERBB2 co-regulation in breast cancer and confirm the importance of miR-125b in controlling clinically-relevant cancer features.

In the last decade, it has been demonstrated that long non-coding RNAs (lncRNAs) are involved in cancer development. The great majority of studies on lncRNAs report alterations, principally on their expression profiles, in several tumor types with respect to the normal tissues of origin. Conversely, since lncRNAs constitute a relatively novel class of RNAs compared to protein-coding transcripts (mRNAs), the landscape of their mutations and variations has not yet been extensively studied. However, in recent years an ever-increasing number of articles have described mutations of lncRNAs. Single-nucleotide polymorphisms (SNPs) that occur within the lncRNA transcripts can affect the structure and function of these RNA molecules, while the presence of a SNP in the promoter region of a lncRNA could alter its expression level. Also, somatic mutations that occur within lncRNAs have been shown to exert important effects in cancer and preliminary data are promising. Overall, the evidence suggests that SNPs and somatic mutation on lncRNAs may play a role in the pathogenesis of cancer, and indicates strong potential for further development of lncRNAs as biomarkers.

The recent advantage obtained by next generation sequencing allows a depth investigation of a new “old” kind of noncoding transcript, the circular RNAs. Circular RNAs are nontranslated RNAs, typically nonpolyadenylated, with a resistance to exonucleases that gives them the ability to be more stable than the common linear RNA isoforms. We used a bioinformatic detection tool (CIRCexplorer) to research predictive circRNAs from the next generation sequenced data of five samples of ductal in situ carcinoma (DCIS) and matched adjacent invasive ductal carcinoma (IDC). Furthermore, we also investigated the circular RNAs expressed in MCF7, an invasive breast ductal carcinoma cell line. We described the genomic context of the predicted circular RNAs and we address the hypothetical possible functional roles. This study showed a perspective of a panel of predictive circRNAs identified and the function that circRNAs could exert.

Background: Breast cancer (BC) represents the most common cancer in women worldwide. Due to its heterogeneous nature, breast cancer management might benefit from differential treatments toward personalized medicine. Additionally, drug resistance is a common phenomenon. We systematically investigated the effect of 14 different drugs administered on BC cell lines in combination with microRNAs (miRNA, miR).Methods: Thirty-eight miRNAs, all associated with BC by clinical and molecular parameters including progression, prognosis and subtypes, were tested for their effects on the viability of 12 different BC cell lines. Four miRNAs with the strongest impact on viability were further assayed in combination with 14 BC drugs. Mann–Whitney U-test with Bonferroni correction was used for statistical analysis.Results: In a miRNA only pre-screen we observed effects on BC cell lines' viability for 34 out of 38 candidate miRNAs. We then identified 14 miRNA/drug combinations for which the combination IC50 was lower than that of both miRNA and drug as single agents. miR-181a, paired with GSK1070916, Doxorubicin, XL765 and AMG511, was the only miRNA active on the triple negative (TNBC) MDA-MB-468 cell line. miR-126 was the only miRNA (in combination with CDK4/6 or PIK3CA inhibitors) with significant effects on cell lines from different subtypes: MCF7 (Luminal) and MDA-MB-453 (HER2+). Because of its activity on different BC subtypes, we investigated the genome wide effects of miR-126 using transcriptomics and confirmed that expression of miR-126 in BC cell lines affected cell cycle and mitosis.Conclusion: Our results show that a combination treatment with miRNAs, in particular miR-181a, miR-326, miR-9 and miR-126, enhance the activity of specific BC drugs in vitro, even on the most aggressive BC subtypes, HER2+ and TNBC. Finally, as expected from its drug interactions, based on a whole transcriptome study we could confirm a role for miR-126 in cell cycle regulation.