MiR-24 induces chemotherapy resistance and hypoxic advantage in breast cancer

Breast cancer remains one of the leading causes of cancer mortality among women. It has been proved that the onset of cancer depends on a very small pool of tumor cells with a phenotype similar to that of normal adult stem cells. Cancer stem cells (CSC) possess self-renewal and multilineage differentiation potential as well as a robust ability to sustain tumorigenesis. Evidence suggests that CSCs contribute to chemotherapy resistance and to survival under hypoxic conditions. Interestingly, hypoxia in turn regulates self-renewal in CSCs and these effects may be primarily mediated by hypoxic inducible factors (HIFs). Recently, microRNAs (miRNAs) have emerged as critical players in the maintenance of pluripotency and self-renewal in normal and cancer stem cells. Here, we demonstrate that miR-24 is upregulated in breast CSCs and that its overexpression increases the number of mammospheres and the expression of stem cell markers. MiR-24 also induces apoptosis resistance through the regulation of BimL expression. Moreover, we identify a new miR-24 target, FIH1, which promotes HIFα degradation: miR-24 increases under hypoxic conditions, causing downregulation of FIH1 and upregulation of HIF1α. In conclusion, miR-24 hampers chemotherapy-induced apoptosis in breast CSCs and increases cell resistance to hypoxic conditions through an FIH1−HIFα pathway

MiR-221 promotes stemness of breast cancer cells by targeting DNMT3b

Cancer stem cells (CSCs) are a small part of the heterogeneous tumor cell population possessing self-renewal and multilineage differentiation potential as well as a great ability to sustain tumorigenesis. The molecular pathways underlying CSC phenotype are not yet well characterized. MicroRNAs (miRs) are small noncoding RNAs that play a powerful role in biological processes. Early studies have linked miRs to the control of self-renewal and differentiation in normal and cancer stem cells. We aimed to study the functional role of miRs in human breast cancer stem cells (BCSCs), also named mammospheres. We found that miR-221 was upregulated in BCSCs compared to their differentiated counterpart. Similarly, mammospheres from T47D cells had an increased level of miR-221 compared to differentiated cells. Transfection of miR-221 in T47D cells increased the number of mammospheres and the expression of stem cell markers. Among miR-221's targets, we identified DNMT3b. Furthermore, in BCSCs we found that DNMT3b repressed the expression of various stemness genes, such as Nanog and Oct 3/4, acting on the methylation of their promoters, partially reverting the effect of miR-221 on stemness. We hypothesize that miR-221 contributes to breast cancer tumorigenicity by regulating stemness, at least in part through the control of DNMT3b expression

MiR-221/222 target the DNA methyltransferase MGMT in glioma cells

Glioblastoma multiforme (GBM) is one of the most deadly types of cancer. To date, the best clinical approach for treatment is based on administration of temozolomide (TMZ) in combination with radiotherapy. Much evidence suggests that the intracellular level of the alkylating enzyme O6-methylguanine-DNA methyltransferase (MGMT) impacts response to TMZ in GBM patients. MGMT expression is regulated by the methylation of its promoter. However, evidence indicates that this is not the only regulatory mechanism present. Here, we describe a hitherto unknown microRNA-mediated mechanism of MGMT expression regulation. We show that miR-221 and miR-222 are upregulated in GMB patients and that these paralogues target MGMT mRNA, inducing greater TMZ-mediated cell death. However, miR-221/miR-222 also increase DNA damage and, thus, chromosomal rearrangements. Indeed, miR-221 overexpression in glioma cells led to an increase in markers of DNA damage, an effect rescued by re-expression of MGMT. Thus, chronic miR-221/222-mediated MGMT downregulation may render cells unable to repair genetic damage. This, associated also to miR-221/222 oncogenic potential, may poor GBM prognosis

RYK promotes the stemness of glioblastoma cells via the WNT/ \u3b2-catenin pathway

Glioblastoma multiforme (GBM) is characterized by a strong self-renewal potential and a poor differentiation state. Since receptor-like tyrosine kinase (RYK) activates the WNT/\u3b2-catenin pathway essential for cancer stem cell maintenance, we evaluated its contribution in conferring stemness to GBM cells. Here, we report that Ryk (related-to-receptor tyrosine kinase), an atypical tyrosine kinase receptor, is upregulated in samples from GBM patients as well as in GSCs. Ryk overexpression confers stemness properties to GBM cells through the modulation of the canonical Wnt signaling and by promoting the activation of pluripotency-related transcription factor circuitry and neurosphere formation ability. In contrast, siRNA-mediated knockdown of Ryk expression suppresses this stem-like phenotype. Rescue experiments reveal that stemness-promoting activity of Ryk is attributable, at least in part, to \u3b2-catenin stabilization. Furthermore, Ryk overexpression improves cell motility and anchorage independent cell growth. Taken together, our findings demonstrate that Ryk promotes stem cell-like and tumorigenic features to glioma cells its essential for the maintenance of GSCs and could be a target of novel therapies

miR-340 predicts glioblastoma survival and modulates key cancer hallmarks through down-regulation of NRAS

Glioblastoma is the most common primary brain tumor in adults; with a survival rate of 12 months from diagnosis. However, a small subgroup of patients, termed long-term survivors (LTS), has a survival rate longer then 12–14 months. There is thus increasing interest in the identification of molecular signatures predicting glioblastoma prognosis and in how to improve the therapeutic approach. Here, we report miR-340 as prognostic tumor-suppressor microRNA for glioblastoma. We analyzed microRNA expression in > 500 glioblastoma patients and found that although miR-340 is strongly down-regulated in glioblastoma overall, it is up-regulated in LTS patients compared to short-term survivors (STS). Indeed, miR-340 expression predicted better prognosis in glioblastoma patients. Coherently, overexpression of miR-340 in glioblastoma cells was found to produce a tumor-suppressive activity. We identified NRAS mRNA as a critical, direct target of miR-340: in fact, miR-340 negatively influenced multiple aspects of glioblastoma tumorigenesis by down-regulating NRAS and downstream AKT and ERK pathways. Thus, we demonstrate that expression of miR-340 in glioblastoma is responsible for a strong tumor-suppressive effect in LTS patients by down-regulating NRAS. miR-340 may thus represent a novel marker for glioblastoma diagnosis and prognosis, and may be developed into a tool to improve treatment of glioblastoma

MiR-221 promotes stemness of breast cancer cells by targeting DNMT3b

Cancer stem cells (CSCs) are a small part of the heterogeneous tumor cell population possessing self-renewal and multilineage differentiation potential as well as a great ability to sustain tumorigenesis. The molecular pathways underlying CSC phenotype are not yet well characterized. MicroRNAs (miRs) are small noncoding RNAs that play a powerful role in biological processes. Early studies have linked miRs to the control of self-renewal and differentiation in normal and cancer stem cells. We aimed to study the functional role of miRs in human breast cancer stem cells (BCSCs), also named mammospheres. We found that miR-221 was upregulated in BCSCs compared to their differentiated counterpart. Similarly, mammospheres from T47D cells had an increased level of miR-221 compared to differentiated cells. Transfection of miR-221 in T47D cells increased the number of mammospheres and the expression of stem cell markers. Among miR-221's targets, we identified DNMT3b. Furthermore, in BCSCs we found that DNMT3b repressed the expression of various stemness genes, such as Nanog and Oct 3/4, acting on the methylation of their promoters, partially reverting the effect of miR-221 on stemness. We hypothesize that miR-221 contributes to breast cancer tumorigenicity by regulating stemness, at least in part through the control of DNMT3b expression

miR-221 acts as stemness promoter in breast cancer cells by targeting DNMT3b

Current hypotheses suggest that tumor originates from cells that carry out a process of “malignant reprogramming” driven by genetic and epigenetic alterations. Multiples studies reported the existence of stem-cell-like cells that acquire the ability to self-renew and are able to generate the bulk of more differentiated cells that form the tumor. This population of cancer cells, called cancer stem cells (CSC), is responsible for sustaining the tumor growth and is able to disseminate and migrate giving metastases to distant organs. Furthermore, CSCs have shown to be more resistant to anti-tumor treatments than the differentiated cells, suggesting that surviving CSCs could be responsible for tumor relapse after therapy. Nevertheless, the cancer stem-like properties are not well characterized yet. MicroRNAs (miRs) are small, noncoding RNAs (20-25 nucleotidies) that play a crucial role in biological processes including development, proliferation, and apoptosis. Previous investigations have linked miRs to the control of self-renewal and differentiation of normal stem cells. The aim of this study was to test the functional role of miRs in human Breast Cancer Stem Cells (BCSCs) also named mammospheres. We analyzed, by miR-Array, the miRs differentially expressed in BCSCs and their differentiated counterpart. Among several miRs, we focused our attention on miR-221 that was found increased in mammospheres. In order to validate data achieved in primary cultures, we obtained mammospheres from T47D, an immortalized breast cancer cell line. Interestingly, like primary cultures mammospheres, also T47D mammospheres exhibited increased levels of miR-221 compared to T47D differentiated cells. Moreover, the overexpression of miR-221 by a miR mimic in T47D differentiated cells was able to increase the number of mammospheres and the expression of stem cell protein markers. Among miR-221 targets, we demonstrated, by luciferase-assay, that miR-221 targets the 3' untranslated region of DNMT3b, a DNA Methyl Transferase. Furthermore, our data showed that DNMT3b was able to repress the expression of some stemness genes, such as Nanog and Oct3/4, and mammospheres formation partially reverting miR-221 mediated effects on stemness properties. In conclusion, we hypothesize that miR-221 may contribute to breast cancer tumorigenicity regulating the stemness properties through DNMT3b expression

Modulating the Crosstalk between the Tumor and the Microenvironment Using SiRNA: A Flexible Strategy for Breast Cancer Treatment

Tumorigenesis is a complex and multistep process in which sequential mutations in oncogenes and tumor-suppressor genes result in enhanced proliferation and apoptosis escape. Over the past decades, several studies have provided evidence that tumors are more than merely a mass of malignant cancer cells, with the tumor microenvironment (TME) also contributing to cancer progression. For this reason, the focus of cancer research in recent years has shifted from the malignant cancer cell itself to the TME and its interactions. Since the TME actively participates in tumor progression, therapeutic strategies targeting it have created great interest. In this context, much attention has been paid to the potential application of small interfering RNA (siRNA), a class of non-coding RNA that has the ability to downregulate the expression of target genes in a sequence-specific way. This is paving the way for a novel therapeutic approach for the treatment of several diseases, including cancer. In this review, we describe recent efforts in developing siRNA therapeutics for the treatment of breast cancer, with particular emphasis on TME regulation. We focus on studies that adapt siRNA design to reprogram/re-educate the TME and eradicate the interplay between cancer cells and TME

The Role of Exo-miRNAs in Cancer: A Focus on Therapeutic and Diagnostic Applications

Exosomes are extracellular vesicles released into biological fluids where they act as carriers of various molecules, including proteins, lipids, and RNAs, between cells, modulating or perturbing specific physiological processes. Recently, it has been suggested that tumoral cells release excessive amounts of exosomes that, through their cargo, promote tumor progression, stimulating growth, angiogenesis, metastasis, insensitivity to chemotherapy, and immune evasion. Increasing evidence highlights exosomal microRNAs (exo-miRNAs) as important players in tumorigenesis. MicroRNA (miRNA) are a class of small non-coding RNA able to regulate gene expression, targeting multiple mRNAs and inducing translational repression and/or mRNA degradation. Exo-miRNAs are highly stable and easily detectable in biological fluids, and for these reasons, miRNAs are potential cancer biomarkers useful diagnostically and prognostically. Furthermore, since exosomes are natural delivery systems between cells, they can be appropriately modified to carry therapeutic miRNAs to specific recipient cells. Here we summarize the main functions of exo-miRNAs and their possible role for diagnostic and therapeutic applications