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MicroRNAs (miRNAs) are a class of small noncoding RNAs that regulate gene expression by targeting mRNAs to trigger either translation repression or mRNA degradation. miR-125b is down-regulated in human breast cancer cells compared with the normal ones except highly metastatic tumor cells MDA-MB-231. However, few functional studies were designed to investigate metastatic potential of miR-125b. In this study, the effects of miR-125b on metastasis in human breast cancer cells were studied, and the targets of miR-125b were also explored. Transwell migration assay, cell wound healing assay, adhesion assay and nude mice model of metastasis were utilized to investigate the effects of miR-125b on metastasis potential in vitro and in vivo. In addition, it was implied STARD13 (DLC2) was a direct target of miR-125b by Target-Scan analysis, luciferase reporter assay and western blot. Furthermore, activation of STARD13 was identified responsible for metastasis induced by miR-125b through a siRNA targeting STARD13. qRT-PCR, immunofluorescent assay and western blot was used to observe the variation of Vimentin and α-SMA in breast cancer cells. In summary, our study provided new insights into the function of miR-125b during the metastasis of breat cancer cells and also suggested the role of miR-125b in pro-metastasis by targeting STARD13.

MicroRNAs (miRNAs or miR) have been integrated into tumorigenic programs as either oncogenes or tumor suppressor genes. The miR-124 was reported to be attenuated in several tumors, such as glioma, medulloblastoma and hepatocellular carcinoma. However, its role in cancer remains greatly elusive. In this study, we show that the miR-124 expression is significantly suppressed in human breast cancer specimens, which is reversely correlated to histological grade of the cancer. More intriguingly, ectopic expression of miR-124 in aggressive breast cancer cell lines MDA-MB-231 and BT-549 strongly inhibits cell motility and invasive capacity, as well as the epithelial–mesenchymal transition process. Also, lentivirus-delivered miR-124 endows MDA-MB-231 cells with the ability to suppress cell colony formation in vitro and pulmonary metastasis in vivo. Further studies have identified the E-cadherin transcription repressor Slug as a direct target gene of miR-124; its downregulation by miR-124 increases the expression of E-cadherin, a hallmark of epithelial cells and a repressor of cell invasion and metastasis. Moreover, knockdown of Slug notably impairs the motility of MDA-MB-231 cells, whereas re-expression of Slug abrogates the reduction of motility and invasion ability induced by miR-124 in MDA-MB-231 cells. These findings highlight an important role for miR-124 in the regulation of invasive and metastatic potential of breast cancer and suggest a potential application of miR-124 in cancer treatment.

It has become increasingly clear that microRNAs (miRNAs) play important roles in tumorigenesis and metastasis. Recently, miR-203 was reported as a suppressor microRNA often silenced in different malignancies including hepatocellular carcinoma, prostate cancer, oral cancer, and hematopoietic malignancy, but little is known about its potential role in breast carcinogenesis. In this study, we found that in breast cancer, miR-203 was upregulated in primary tumors and some nonmetastatic cell lines but was significantly downregulated in metastatic cell lines including BT549, Hs578T, and MDA-MB-231, as measured by regular and real-time PCR. Downregulation of miR-203 in metastatic breast cancer cells appeared to be caused by hypermethylation of its promoter. Functionally, ectopic expression of miR-203 in BT549 and MDA-MB-231 breast cancer cell lines caused cell cycle arrest and apoptosis and inhibited cell invasion and migration in vitro. Bioinformatic analysis predicted the snail homolog 2 (SNAI2 or SLUG), a transcription factor that promotes cell invasion and tumor metastasis, as a target of miR-203, and the prediction was validated by expression analysis and luciferase reporter assay of the 3′ untranslated region of SNAI2 that contains the miR-203 target sequences. These results suggest that in malignant breast cancer cells, miR-203 is epigenetically silenced, and the silencing promotes tumor cell growth and invasion at least in part by upregulating the SNAI2 transcription factor.

Background

Accumulating evidence suggested that epithelial-mesenchymal transition (EMT) and cancer stem cell (CSC) characteristics, both of which contribute to tumor invasion and metastasis, are interrelated with miR-21. MiR-21 is one of the important microRNAs associated with tumor progression and metastasis, but the molecular mechanisms underlying EMT and CSC phenotype during miR-21 contributes to migration and invasion of breast cancer cells remain to be elucidated.

Methodology/Principal Findings

In this study, MDA-MB-231/anti-miR-21 cells were established by transfected hsa-miR-21 antagomir into breast cancer MDA-MB-231 cells. EMT was evaluated by the changes of mesenchymal cell markers (N-cadherin, Vimentin, and alpha-SMA), epithelial cell marker (E-cadherin), as well as capacities of cell migration and invasion; CSC phenotype was measured using the changes of CSC surface markers (ALDH1 and CD44), and the capacity of sphereforming (mammospheres). We found that antagonism of miR-21 reversed EMT and CSC phenotype, accompanied with PTEN up-regulation and AKT/ERK1/2 inactivation. Interestingly, down-regulation of PTEN by siPTEN suppressed the effects of miR-21 antagomir on EMT and CSC phenotype, confirming that PTEN is a target of miR-21 in reversing EMT and CSC phenotype. The inhibitors of PI3K-AKT and ERK1/2 pathways, LY294002 and U0126, both significantly suppressed EMT and CSC phenotype, indicating that AKT and ERK1/2 pathways are required for miR-21 mediating EMT and CSC phenotype.

Conclusions/Significance

In conclusion, our results demonstrated that antagonism of miR-21 reverses EMT and CSC phenotype through targeting PTEN, via inactivation of AKT and ERK1/2 pathways, and showed a novel mechanism of which might relieve the malignant biological behaviors of breast cancer.

MicroRNAs (miRNAs) are endogenously small non-coding RNAs which are capable of silencing gene expression at the posttranscriptional level. In this study, we report that miR-205 is significantly underexpressed in breast tumor compared to the matched normal breast tissue. Similarly, breast cancer cell lines including MCF-7 and MDA-MB-231 express a lower level miR-205 than the non-malignant MCF-10A cells. Of interest, ectopic expression of miR-205 significantly inhibits cell proliferation and anchorage independent growth as well as cell invasion. Furthermore, the animal model indicates that miR-205 suppresses lung metastasis. Finally, western blot combined with the luciferase reporter assays demonstrate that ErbB3 and vascular endothelial growth factor A (VEGF-A) are direct targets for miR-205 and this miR-205-mediated suppression is likely through the direct interaction with the putative miR-205 binding site in the 3’-untranslated region (3-UTR) of ErbB3 and VEGF-A. Together, these results suggest that miR-205 is a tumor suppressor in breast cancer.

Fenhexamid and fludioxonil are antifungal agents used in agricultural applications, which are present at measurable amounts in fruits and vegetables. Fenhexamid and fludioxonil showed endocrine disruptor activity as antiandrogens in an androgen receptor reporter assay in engineered human breast cancer cells. Little is known about how environmental chemicals regulate microRNA (miRNA) expression. This study examined the effect of fenhexamid and fludioxonil on the expression of the oncomiR miR-21 in MCF-7, T47D, and MDA-MB-231 human breast cancer cells and downstream targets of miR-21 in MCF-7 cells. Fenhexamid and fludioxonil stimulated miR-21 expression in a concentration-dependent manner and reduced the expression of miR-21 target Pdcd4 protein. Antisense to miR-21 blocked the increase in Pdcd4 protein by fenhexamid and fludioxonil. Fenhexamid and fludioxonil reduced miR-125b and miR-181a, demonstrating specificity of miRNA regulation. Induction of miR-21 was inhibited by the estrogen receptor antagonist fulvestrant, by androgen receptor antagonist bicalutamide, by actinomycin D and cycloheximide, and by inhibitors of the mitogen-activated protein kinases and phosphoinositide 3-kinase pathways. Fenhexamid activation was inhibited by the arylhydrocarbon receptor antagonist α-napthoflavone. Fenhexamid and fludioxonil did not affect dihydrotestosterone-induced miR-21 expression. Fludioxonil, but not fenhexamid, inhibited MCF-7 cell viability, and both inhibited estradiol-induced cell proliferation and reduced cell motility. Together these data indicate that fenhexamid and fludioxonil use similar and distinct mechanisms to increase miR-21 expression with downstream antiestrogenic activity.

Background

MicroRNAs are non-coding RNA molecules that posttranscriptionally regulate expression of target genes and have been implicated in the progress of cancer proliferation, differentiation and apoptosis. The aim of this study was to determine whether microRNA-21 (miR-21), a specific microRNA implicated in multiple aspects of carcinogenesis, impacts breast cancer invasion by regulating the tissue inhibitor of metalloproteinase 3 (TIMP3) gene.

Methods

miR-21 expression was investigated in 32 matched breast cancer and normal breast tissues, and in four human breast cancer cell lines, by Taqman quantitative real-time PCR. Cell invasive ability was determined by matrigel invasion assay in vitro, in cells transfected with miR-21 or anti-miR-21 oligonucleotides. In addition, the regulation of tissue inhibitor of metalloproteinase 3 (TIMP3) by miR-21 was evaluated by western blotting and luciferase assays.

Results

Of the 32 paired samples analyzed, 25 breast cancer tissues displayed overexpression of miR-21 in comparison with matched normal breast epithelium. Additionally, incidence of lymph node metastasis closely correlated with miR-21 expression, suggesting a role for miR-21 in metastasis. Similarly, each of the four breast cancer cell lines analyzed overexpressed miR-21, to varied levels. Further, cells transfected with miR-21 showed significantly increased matrigel invasion compared with control cells, whereas transfection with anti-miR-21 significantly decreased cell invasion. Evaluation of TIMP3 protein levels, a peptidase involved in extarcellular matrix degredation, inversely correlated with miR-21 expression.

Conclusion

As knockdown of miR-21 increased TIMP3 protein expression and luciferase reporter activity, our data suggests that miR-21 could promote invasion in breast cancer cells via its regulation of TIMP3.

Background

Breast cancer and its metastatic progression is mainly directed by epithelial to mesenchymal transition (EMT), a phenomenon supported by specific transcription factors and miRNAs.

Methods

In order to investigate a possible correlation between Slug transcription factor and miR-221, we performed Slug gene silencing in MDA-MB-231 breast cancer cells and evaluated the expression of genes involved in supporting the breast cancer phenotype, using qRT-PCR and Western blot analysis. Chromatin immunoprecipitation and wound healing assays were employed to determine a functional link between these two molecules.

Results

We showed that Slug silencing significantly decreased the level of miR-221 and vimentin, reactivated Estrogen Receptor α and increased E-cadherin and TRPS1 expression. We demonstrated that miR-221 is a Slug target gene, and identified a specific region of miR-221 promoter that is transcriptionally active and binds the transcription factor Slug “in vivo”. In addition, we showed that in Slug-silenced cells, wich retained residual miR-221 (about 38%), cell migration was strongly inhibited. Cell migration was inhibited, but to a less degree, following complete knockdown of miR-221 expression by transfection with antagomiR-221.

Conclusions

We report for the first time evidence of a correlation between Slug transcription factor and miR-221 in breast cancer cells. These studies suggest that miR-221 expression is, in part, dependent on Slug in breast cancer cells, and that Slug plays a more important role than miR-221 in cell migration and invasion.

To study the roles of microRNA-223 (miR-223) in regulation of cell growth, we established a miR-223 over-expression model in HeLa cells infected with miR-223 by Lentivirus pLL3.7 system. We observed in this model that miR-223 significantly suppressed the proliferation, growth rate, colony formation of HeLa cells in vitro, and in vivo tumorigenicity or tumor formation in nude mice. To investigate the mechanisms involved, we scanned and examined the potential and putative target molecules of miR-223 by informatics, quantitative PCR and Western blot, and found that insulin-like growth factor-1 receptor (IGF-1R) was the functional target of miR-223 inhibition of cell proliferation. Targeting IGF-1R by miR-223 was not only seen in HeLa cells, but also in leukemia and hepatoma cells. The downstream pathway, Akt/mTOR/p70S6K, to which the signal was mediated by IGF-1R, was inhibited as well. The relative luciferase activity of the reporter containing wild-type 3′UTR(3′untranslated region) of IGF-1R was significantly suppressed, but the mutant not. Silence of IGF-1R expression by vector-based short hairpin RNA resulted in the similar inhibition with miR-223. Contrarily, rescued IGF-1R expression in the cells that over-expressed miR-223, reversed the inhibition caused by miR-223 via introducing IGF-1R cDNA that didn't contain the 3′UTR. Meanwhile, we also noted that miR-223 targeted Rasa1, but the downstream molecules mediated by Rasa1 was neither targeted nor regulated. Therefore we believed that IGF-1R was the functional target for miR-223 suppression of cell proliferation and its downstream PI3K/Akt/mTOR/p70S6K pathway suppressed by miR-223 was by targeting IGF-1R.

Developing vehicles for the delivery of therapeutic molecules, like siRNA, is an area of active research. Nanoparticles composed of bovine serum albumin, stabilized via the adsorption of poly-L-lysine (PLL), have been shown to be potentially inert drug-delivery vehicles. With the primary goal of reducing nonspecific protein adsorption, the effect of using comb-type structures of poly(ethylene glycol) (1 kDa, PEG) units conjugated to PLL (4.2 and 24 kDa) on BSA-NP properties, apparent siRNA release rate, cell viability, and cell uptake were evaluated. PEGylated PLL coatings resulted in NPs with ζ-potentials close to neutral. Incubation with platelet-poor plasma showed the composition of the adsorbed proteome was similar for all systems. siRNA was effectively encapsulated and released in a sustained manner from all NPs. With 4.2 kDa PLL, cellular uptake was not affected by the presence of PEG, but PEG coating inhibited uptake with 24 kDa PLL NPs. Moreover, 24 kDa PLL systems were cytotoxic and this cytotoxicity was diminished upon PEG incorporation. The overall results identified a BSA-NP coating structure that provided effective siRNA encapsulation while reducing ζ-potential, protein adsorption, and cytotoxicity, necessary attributes for in vivo application of drug-delivery vehicles.

MicroRNAs (miRNAs) are important gene regulators that could play a profound role in tumorigenesis. Our previous studies indicate that miR-145 is a tumor suppressor capable of inhibiting tumor cell growth both in vitro and in vivo. In this study, we show that miR-145 exerts its function in a cell-specific manner. Although miR-145 inhibits cell growth in MCF-7 and HCT-116 cells, it has no significant effect on cell growth in metastatic breast cancer cell lines. However, miR-145 significantly suppresses cell invasion in these cells; in contrast, the antisense oligo against miR-145 increases cell invasion. miR-145 is also able to suppress lung metastasis in an experimental metastasis animal model. This miR-145-mediated suppression of cell invasion is in part due to the silencing of the metastasis gene mucin 1 (MUC1). Using luciferase reporters carrying the 3'-untranslated region of MUC1 combined with western blot and immunofluorescence staining, we identify MUC1 as a direct target of miR-145. Moreover, ectopic expression of MUC1 enhances cell invasion, which can be blocked by miR-145. Of interest, suppression of MUC1 by miR-145 causes a reduction of β-catenin as well as the oncogenic cadherin 11. Finally, suppression of MUC1 by RNAi mimics the miR-145 action in suppression of invasion, which is associated with downregulation of β-catenin and cadherin 11. Taken together, these results suggest that as a tumor suppressor, miR-145 inhibits not only tumor growth, but also cell invasion and metastasis.

Small interfering RNA (siRNA) has been widely proposed to treat various diseases by silencing genes, but its delivery remains a challenge. A well-controlled assembly approach was applied to prepare a protease assisted nanodelivery system. Onto a gold nanoparticles (AuNPs), protease degradable poly-L-lysine (PLL) and siRNA were fabricated by alternating the charged polyelectrolytes. In this study, up to 4 layers of PLL and 3 layers of siRNA (sR3P) were coated. Due to slow degradation of PLL, the incorporated siRNA was released gradually and showed extended gene silencing effect. Importantly, the inhibition effect in cells was found to correlate with the number of siRNA layers.

Background

MicroRNAs (miRNAs) are small, non-coding RNAs (20–24 nucleotides) that post-transcriptionally modulate gene expression by negatively regulating the stability or translational efficiency of their target mRNAs. The aim of this study was to investigate the expression pattern of microRNA-26b (miR-26b) in human breast cancer, and its potential role in disease pathogenesis.

Methods

Quantitative reverse transcription-polymerase chain reaction (qRT-PCR) was performed to determine the expression level of miR-26b in 38 breast cancer specimens and adjacent normal breast tissues. MTT assays were conducted to explore the impact of miR-26b overexpression on the proliferation of human MDA-MB-231 breast cancer cells. Luciferase reporter assays were employed to validate regulation of a putative target of miR-26b. The effect of modulating miR-26b on endogenous levels of this target were subsequently confirmed via qRT-PCR and Western blot.

Results

MiR-26b expression was relatively decreased in breast cancer specimens compared with adjacent normal tissues (P<0.01). Overexpression of miR-26b suppressed MDA-MB-231 cell growth. Luciferase assays using a reporter carrying a putative miR-26b target site in the 3' untranslated region of PTGS2 revealed that miR-26b directly targets PTGS2. Overexpression of miR-26b led to downregulation of PTGS2 at the mRNA and protein level, as assessed by qRT-PCR and Western blot. Targeted knockdown of PTGS2 by siRNA significantly inhibited the proliferation of MDA-MB-231 breast cancer cells.

Conclusions

MiR-26b may act as a tumor suppressor in breast cancer. The overexpression of miR-26b inhibits cellular growth by targeting PTGS2, suggesting its use as a potential therapeutic target for breast cancer.

MicroRNAs (miRNAs) are increasingly implicated in regulating metastasis. Despite progress in silencing miRNAs in normal tissues of rodents and non-human primates, the development of effective approaches for sequence-specific inhibition of miRNAs in fast-growing tumors remains a significant scientific and clinical challenge. Here we show that systemic treatment of tumor-bearing mice with miR-10b antagomirs – a class of chemically modified anti-miRNA oligonucleotides – suppresses breast cancer metastasis. Silencing of miR-10b both in vitro and in vivo with antagomirs significantly decreases miR-10b levels and increases levels of a functionally important miR-10b target, Hoxd10. Administration of miR-10b antagomirs to mice bearing highly metastatic cells does not reduce primary mammary tumor growth but instead markedly suppresses formation of lung metastases. This metastasis-suppressing effect is sequence-specific. The miR-10b antagomir, which is well tolerated by normal animals, appears to be a promising candidate and a starting point for the development of new anti-metastasis agents.

The aryl hydrocarbon receptor (AHR) was initially identified as a receptor that bound 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) and related environmental toxicants; however, there is increasing evidence that the AHR is an important new drug target for treating multiple diseases including breast cancer. Treatment of estrogen receptor (ER)-negative MDA-MB-231 and BT474 breast cancer cells with TCDD or the selective AHR modulator 6-methyl-1,3,-trichlorodibenzofuran (MCDF) inhibited breast cancer cell invasion in a Boyden chamber assay. These results were similar to those previously reported for the antimetastic microRNA-335 (miR-335). Both TCDD and MCDF induced miR-335 in MDA-MB-231 and BT474 cells and this was accompanied by downregulation of SOX4, a miR-335-regulated (inhibited) gene. The effects of TCDD and MCDF on miR-335 and SOX4 expression and interactions of miR-335 with the 3'-UTR target sequence in the SOX4 gene were all inhibited in cells transfected with an oligonucleotide (iAHR) that knocks down the AHR, thus confirming AHR-miR-335 interactions. MCDF (40 mg/kg/day) also inhibited lung metastasis of MDA-MB-231 cells in a tail vein injection model, demonstrating that the AHR is a potential new target for treating patients with ER-negative breast cancer, a disease where treatment options and their effectiveness are limited.

Cancer cells often acquire a constitutively active nuclear factor-κB (NF-κB) program to promote survival, proliferation and metastatic potential by mechanisms that remain largely unknown. Extending observations from an immunologic setting, we demonstrate that microRNA-146a and microRNA-146b (miR-146a/b) when expressed in the highly metastatic human breast cancer cell line MDA-MB-231 function to negatively regulate NF-κB activity. Lentiviral-mediated expression of miR-146a/b significantly downregulated interleukin (IL)-1 receptor-associated kinase and TNF receptor-associated factor 6, two key adaptor/scaffold proteins in the IL-1 and Toll-like receptor signaling pathway, known to positively regulate NF-κB activity. Impaired NF-κB activity was evident from reduced phosphorylation of the NF-κB inhibitor IκBα, reduced NF-κB DNA-binding activity and suppressed expression of the NF-κB target genes IL-8, IL-6 and matrix metalloproteinase-9. Functionally, miR-146a/b-expressing MDA-MB-231 cells showed markedly impaired invasion and migration capacity relative to control cells. These findings implicate miR-146a/b as a negative regulator of constitutive NF-κB activity in a breast cancer setting and suggest that modulating miR-146a/b levels has therapeutic potential to suppress breast cancer metastases.

Background

MicroRNAs (miRNAs) play an important role in the regulation of cell growth, differentiation, apoptosis, and carcinogenesis. Detection of their expression may lead to identifying novel markers for breast cancer.

Methods

We profiled miRNA expression in three breast cancer cell lines (MCF-7, MDA-MB-231, and MDA-MB-468) and then focused on one miRNA, miR-339-5p, for its role in regulation of tumor cell growth, migration, and invasion and target gene expression. We then analyzed miR-339-5p expression in benign and cancerous breast tissue specimens.

Results

A number of miRNAs were differentially expressed in these cancer cell lines. Real-time PCR indicated that miR-339-5p expression was downregulated in the aggressive cell lines MDA-MB-468 and MDA-MB-231 and in breast cancer tissues compared with benign tissues. Transfection of miR-339-5p oligonucleotides reduced cancer cell growth only slightly but significantly decreased tumor cell migration and invasion capacity compared with controls. Real-time PCR analysis showed that BCL-6, a potential target gene of miR-339-5p, was downregulated in MDA-MB-231 cells by miR-339-5p transfection. Furthermore, the reduced miR-339-5p expression was associated with an increase in metastasis to lymph nodes and with high clinical stages. Kaplan-Meier analyses found that the patients with miR-339-5p expression had better overall and relapse-free survivals compared with those without miR-339-5p expression. Cox proportional hazards analyses showed that miR-339-5p expression was an independent prognostic factor for breast cancer patients.

Conclusions

MiR-339-5p may play an important role in breast cancer progression, suggesting that miR-339-5p should be further evaluated as a biomarker for predicting the survival of breast cancer patients.

A search for general regulators of cancer metastasis has yielded a set of microRNAs for which expression is specifically lost as human breast cancer cells develop metastatic potential. Here we show that restoring the expression of these microRNAs in malignant cells suppresses lung and bone metastasis by human cancer cells in vivo. Of these microRNAs, miR-126 restoration reduces overall tumour growth and proliferation, whereas miR-335 inhibits metastatic cell invasion. miR-335 regulates a set of genes whose collective expression in a large cohort of human tumours is associated with risk of distal metastasis. miR-335 suppresses metastasis and migration through targeting of the progenitor cell transcription factor SOX4 and extracellular matrix component tenascin C. Expression of miR-126 and miR-335 is lost in the majority of primary breast tumours from patients who relapse, and the loss of expression of either microRNA is associated with poor distal metastasis-free survival. miR-335 and miR-126 are thus identified as metastasis suppressor microRNAs in human breast cancer.

The expression of small, non-coding RNA, or microRNAs (miR), is frequently deregulated in human cancer, but how these pathways affect disease progression is still largely elusive. Here, we report on a microRNA, miR-296, which is progressively lost during tumor progression, and correlates with metastatic disease in colorectal, breast, lung, gastric, parathyroid, liver and bile ducts cancers. Functionally, miR-296 controls a global cell motility gene signature in epithelial cells by transcriptionally repressing the cell polarity-cell plasticity module, Scrib. In turn, loss of miR-296 causes aberrantly increased and mislocalized Scrib in human tumors, resulting in exaggerated random cell migration, and tumor cell invasiveness. Re-expression of miR-296 in MDA-MB231 cells inhibits tumor growth, in vivo. Finally, miR-296 or Scrib levels predict tumor relapse in hepatocellular carcinoma patients.

These data identify miR-296 as a global repressor of tumorigenicity, and uncover a previously unexplored exploitation of Scrib in tumor progression in humans.

MicroRNAs (miRNAs) are a major class of small endogenous RNA molecules that post-transcriptionally inhibit gene expression. Many miRNAs have been implicated in several human cancers, including breast cancer. Here we describe the association between altered miRNA signatures and breast cancer tumorigenesis and metastasis. The loss of several tumor suppressor miRNAs (miR-206, miR-17-5p, miR-125a, miR-125b, miR-200, let-7, miR-34 and miR-31) and the overexpression of certain oncogenic miRNAs (miR-21, miR-155, miR-10b, miR-373 and miR-520c) have been observed in many breast cancers. The gene networks orchestrated by these miRNAs are still largely unknown, although key targets have been identified that may contribute to the disease phenotype. Here we report how the observed perturbations in miRNA expression profiles may lead to disruption of key pathways involved in breast cancer.

Poly-L-lysine (PLL), a homopolymer of amino acid L-lysine (LL), has been frequently used for drug delivery. Here, we report that PLL is an effective agent to inhibit propagation of prions that cause fatal and incurable neurologic disorders in humans and animals, termed prion diseases. In our recent investigation on prion propagation facilitated by conversion of the cellular prion protein (PrP) to the misfolded, disease-associated PrP (PrPSc), we demonstrated that plasminogen stimulates PrP conversion as a cellular cofactor. In the current study, we targeted plasminogen using PLL and assessed its anti-prion efficacy. The results showed that PLL strongly inhibited PrPSc propagation in the cell-free, cell culture, and mouse models of prion disease. These results confirm the role of plasminogen in PrPSc propagation, validates plasminogen as a therapeutic target to combat prion disease, and suggests PLL as a potential anti-prion agent. Therefore, our study represents a proof-of-concept that targeting plasminogen, a cofactor for PrP conversion, using PLL results in suppression of prion propagation, which represents a successful translation of our understanding on details of prion propagation into a potential therapeutic strategy for prion diseases.

MicroRNAs (miRNAs) are a class of small non coding regulatory RNAs that have key functions in multiple cell processes. Deregulation of these tiny miRNAs is involved in various human diseases. MiR-155 is one of the multifunctional miRNA that its over-expression has been found to be associated with different kinds of cancer such as leukemia, breast and colon cancers. It is thought that deregulation and over-expression of this microRNA may be associated with PC12 cell proliferation. So, the aim of this study was to investigate the role of miR-155 expression on PC12 cell growth. For this reason, PC12 cells were cultured and transfected by 3 different concentration (25, 50 and 75 nmol) of either LNA anti-miR-155 or scramble antisense in 24-well plate. Then, total RNA was extracted from transfected cells. miRNA cDNAs were synthesized from isolated total RNA. In the second step, miR-155 expression level was analyzed using the quantitative real-time polymerase chain reaction (QRT-PCR). MTT test was performed to evaluate cell viability. In the next step, apoptosis assay was assessed to investigate anti miR-155 effect on PC12 cells death. Obtained results were analyzed with t-test. MTT test revealed that cell viability of transfected cells with 75 nM of anti-miR- 155 to be reduced by half of the control and scramble groups (0.5 vs. 0.97 and 0.94). Our data suggest that miR-155 over-expression is associated with PC12 cell growth. So, miR-155 down regulation by anti-miR-155 could open up new ways to restrain brain tumor growth, as anti-miR-155 causes PC12 cells to repress.

Late-stage breast cancer metastasis is driven by dysregulated TGF-β signaling, but the underlying molecular mechanisms have not been fully elucidated. We attempted to recapitulate tumor and metastatic microenvironments via the use of biomechanically compliant or rigid 3D organotypic cultures and combined them with global microRNA (miR) profiling analyses to identify miRs that were upregulated in metastatic breast cancer cells by TGF-β. Here we establish miR-181a as a TGF-β–regulated “metastamir” that enhanced the metastatic potential of breast cancers by promoting epithelial-mesenchymal transition, migratory, and invasive phenotypes. Mechanistically, inactivation of miR-181a elevated the expression of the proapoptotic molecule Bim, which sensitized metastatic cells to anoikis. Along these lines, miR-181a expression was essential in driving pulmonary micrometastatic outgrowth and enhancing the lethality of late-stage mammary tumors in mice. Finally, miR-181a expression was dramatically and selectively upregulated in metastatic breast tumors, particularly triple-negative breast cancers, and was highly predictive for decreased overall survival in human breast cancer patients. Collectively, our findings strongly implicate miR-181a as a predictive biomarker for breast cancer metastasis and patient survival, and consequently, as a potential therapeutic target in metastatic breast cancer.

MicroRNAs (miRNAs) are small endogenous non-coding RNAs that regulate post-transcriptional gene expression and are important in many biological processes. Disease-associated miRNAs have been shown to become potential targets for therapeutic intervention. Functions of miRNAs can be inhibited by using antisense oligonucleotides, called anti-miRs, complimentary to the miRNA sequences. Here, we show that systemic delivery of a chemically stabilized anti-miR-122 complexed with interfering nanoparticles (iNOPs) effectively silences the liver-expressed miR-122 in mice. Intravenous administration of 2 mg kg−1 chemically modified anti-miR-122 complexed with iNOP-7 resulted in 83.2 ± 3.2% specific silencing of miR-122, which was accompanied by regulating gene expression in liver and lowering of plasma cholesterol. The specific silencing of miR-122 was long lasting and did not induce an immune response. Our results demonstrate that iNOPs can successfully deliver anti-miR to specifically target and silence miRNA in clinically acceptable and therapeutically affordable doses.

SUMMARY

MicroRNAs are well-suited to regulate tumor metastasis due to their capacity to coordinately repress numerous target genes, thereby potentially enabling their intervention at multiple steps of the invasion-metastasis cascade. We identify a microRNA exemplifying these attributes, miR-31, whose expression correlates inversely with metastasis in human breast cancer patients. Overexpression of miR-31 in otherwise-aggressive breast tumor cells suppresses metastasis. We deploy a stable microRNA sponge strategy to stably inhibit miR-31 in vivo; this allows otherwise-non-aggressive breast cancer cells to metastasize. These phenotypes do not involve confounding influences on primary tumor development and are specifically attributable to miR-31-mediated inhibition of several steps of metastasis, including local invasion, extravasation or initial survival at a distant site, and metastatic colonization. Such pleiotropy is achieved via coordinate repression of a cohort of metastasis-promoting genes, including RhoA. Indeed, RhoA re-expression partially reverses miR-31-imposed metastasis-suppression. These findings indicate that miR-31 uses multiple mechanisms to oppose metastasis.