MicroRNA 603 acts as a tumor suppressor and inhibits triple-negative breast cancer tumorigenesis by targeting elongation factor 2 kinase

Triple negative breast cancer (TNBC) is an aggressive type of breast cancer characterized by the absence of defined molecular targets, including estrogen receptor (ER), progesterone receptor (PR), human epidermal growth factor receptor 2 (HER2) and is associated with high rates of relapse and distant metastasis despite surgery and adjuvant chemotherapy. The lack of effective targeted therapies for TNBC represents an unmet therapeutic challenge. Eukaryotic elongation factor 2 kinase (eEF2K) is an atypical calcium/calmodulin-dependent serine/threonine kinase that promotes TNBC tumorigenesis, progression, and drug resistance, representing a potential novel molecular target. However, the mechanisms regulating eEF2K expression are unknown. Here, we report that eEF2K protein expression is highly up-regulated in TNBC cells and patient tumors and it is associated with poor patient survival and clinical outcome. We found that loss/reduced expression of miR-603 leads to eEF2K overexpression in TNBC cell lines. Its expression results in inhibition of eEF2K by directly targeting the 3-UTR and the inhibition of tumor cell growth, migration and invasion in TNBC. In vivo therapeutic gene delivery of miR-603 into TNBC xenograft mouse models by systemic administration of miR-603-nanoparticles led to a significant inhibition of eEF2K expression and tumor growth, which was associated with decreased activity of the downstream targets of eEF2K, including Src, Akt, cyclin D1 and c-myc. Our findings suggest that miR-603 functions as a tumor suppressor and loss of miR-603 expression leads to increase in eEF2K expression and contributes to the growth, invasion, and progression of TNBC. Taken together, our data suggest that miR-603-based gene therapy is a potential strategy against TNBC

Exosomal miR-940 maintains SRC-mediated oncogenic activity in cancer cells: a possible role for exosomal disposal of tumor suppressor miRNAs

Exosomes have emerged as important mediators of diverse biological functions including tumor suppression, tumor progression, invasion, immune escape and cell-to-cell communication, through the release of molecules such as mRNAs, miRNAs, and proteins. Here, we identified differentially expressed exosomal miRNAs between normal epithelial ovarian cell line and both resistant and sensitive ovarian cancer (OC) cell lines. We found miR-940 as abundant in exosomes from SKOV3-IP1, HeyA8, and HeyA8-MDR cells. The high expression of miR-940 is associated with better survival in patients with ovarian serous cystadenocarcinoma. Ectopic expression of miR-940 inhibited proliferation, colony formation, invasion, and migration and triggered G0/G1 cell cycle arrest and apoptosis in OC cells. Overexpression of miR-940 also inhibited tumor cell growth in vivo. We showed that proto-oncogene tyrosine-protein kinase (SRC) is directly targeted by miR-940 and that miR-940 inhibited SRC expression at mRNA and protein levels. Following this inhibition, the expression of proteins downstream of SRC, such as FAK, paxillin and Akt was also reduced. Collectively, our results suggest that OC cells secrete the tumor-suppressive miR-940 into the extracellular environment via exosomes, to maintain their invasiveness and tumorigenic phenotype

Effect of black cumin oil (Nigella sativa L.) on fresh fish (Barbus grypus) fillets during storage at 2 ± 1 °C

Abstract It is important that the methods and materials used for to lengthen the shelf-life of food are simple, inexpensive and safe. For these reasons, herbal additives like cumin oil (Nigella sativa L.) have been preferred recently. Thus, the present study focuses on the influence of black cumin oil on chemical, microbiological and sensory quality of fish (Barbus grypus) fillets during storage at 2 ± 1 °C. Acceptability scores for sensory quality of all described treatment groups decreased with storage time. Defined limits for mesophilic bacteria and Total volatile basic nitrogen (TVB-N) were reached after 21st days for the control group, after 24th days for with 0.2% black cumin oil treated group and after 28th days for with 0.4% and 0.6% black cumin oil treated groups. No difference was found among groups during storage in terms of defined thiobarbituric acid (TBA) values. Consequently, it was found that black cumin oil treated groups had longer shelf-life and higher sensory quality than the untreated control group

Effect of black cumin oil (Nigella sativa L.) on fresh fish (Barbus grypus) fillets during storage at 2 ± 1 °C

Abstract It is important that the methods and materials used for to lengthen the shelf-life of food are simple, inexpensive and safe. For these reasons, herbal additives like cumin oil (Nigella sativa L.) have been preferred recently. Thus, the present study focuses on the influence of black cumin oil on chemical, microbiological and sensory quality of fish (Barbus grypus) fillets during storage at 2 ± 1 °C. Acceptability scores for sensory quality of all described treatment groups decreased with storage time. Defined limits for mesophilic bacteria and Total volatile basic nitrogen (TVB-N) were reached after 21st days for the control group, after 24th days for with 0.2% black cumin oil treated group and after 28th days for with 0.4% and 0.6% black cumin oil treated groups. No difference was found among groups during storage in terms of defined thiobarbituric acid (TBA) values. Consequently, it was found that black cumin oil treated groups had longer shelf-life and higher sensory quality than the untreated control group

Investigating the effect of decontaminants on microbiological and chemical properties of rainbow trouts

This study was designed to determine the effect of decontamination on the shelf life of whole rainbow trouts. For this purpose 0.5% cetylpyridinium chloride (CPC), 10% trisodium phosphate (TSP), 2.5% acetic acid (AA), 2.5% lactic acid (LA), 1200 ppm acidified sodium chloride (ASC) and control (tap water) were used as decontaminants. After the decontamination process, the samples were stored in cold storage and subjected to microbiological and chemical analyzes on days 0, 3, 6, 9, 12 and 15. Mesophilic bacteria, psychrophilic bacteria, lactic acid bacteria, Pseudomonas spp., Enterobacteriaceae and coliform bacteria were enumerated for the evaluation of microbiological quality, whereas pH, total volatile alkaline nitrogen (TVB-N), thiobarbituric acid (TBA) were determined for the evaluation of the chemical quality of fish samples. The study was repeated 3 times and 6 fish were used in each group corresponding to 108 fish in total. Microbiological samples were evaluated with a modification in USDA/FSIS chicken carcass method. The data of microbiological analysis showed that decontamination provided a significant improvement on the microbiological quality and the decontaminants used in this study extended the microbiological shelf life of rainbow trout. However, acidic decontaminants and TSP caused some changes in the physical properties of rainbow trouts. On the other hand, the use of CPC extended the shelf life of rainbow trouts without adversely affecting the texture. The microbiological sampling protocol used in this study was proved to be easier to apply and gave coherent results

MicroRNA 603 acts as a tumor suppressor and inhibits triplenegative breast cancer tumorigenesis by targeting elongation factor 2 kinase

Ivan, Cristina/0000-0002-4848-0168; Calin, George/0000-0001-6704-5615; Calin, George/0000-0002-7427-0578WOS: 000394187400082PubMed: 28036267Triple negative breast cancer (TNBC) is an aggressive type of breast cancer characterized by the absence of defined molecular targets, including estrogen receptor (ER), progesterone receptor (PR), human epidermal growth factor receptor 2 (HER2) and is associated with high rates of relapse and distant metastasis despite surgery and adjuvant chemotherapy. The lack of effective targeted therapies for TNBC represents an unmet therapeutic challenge. Eukaryotic elongation factor 2 kinase (eEF2K) is an atypical calcium/calmodulin-dependent serine/threonine kinase that promotes TNBC tumorigenesis, progression, and drug resistance, representing a potential novel molecular target. However, the mechanisms regulating eEF2K expression are unknown. Here, we report that eEF2K protein expression is highly up-regulated in TNBC cells and patient tumors and it is associated with poor patient survival and clinical outcome. We found that loss/reduced expression of miR-603 leads to eEF2K overexpression in TNBC cell lines. Its expression results in inhibition of eEF2K by directly targeting the 3-UTR and the inhibition of tumor cell growth, migration and invasion in TNBC. In vivo therapeutic gene delivery of miR-603 into TNBC xenograft mouse models by systemic administration of miR-603-nanoparticles led to a significant inhibition of eEF2K expression and tumor growth, which was associated with decreased activity of the downstream targets of eEF2K, including Src, Akt, cyclin D1 and c-myc. Our findings suggest that miR-603 functions as a tumor suppressor and loss of miR-603 expression leads to increase in eEF2K expression and contributes to the growth, invasion, and progression of TNBC. Taken together, our data suggest that miR-603-based gene therapy is a potential strategy against TNBC.non-Coding RNA center; U54 NIH/NCIThis study was supported by the funding from non-Coding RNA center and U54 NIH/NCI and used the Functional Proteomics RPPA Core Facility"

Ubiquitous release of exosomal tumor suppressor miR-6126 from ovarian cancer cells

Abstract Cancer cells actively promote their tumorigenic behavior by reprogramming gene expression. Loading intraluminal vesicles with specific miRNAs and releasing them into the tumor microenvironment as exosomes is one mechanism of reprogramming whose regulation remains to be elucidated. Here, we report that miR-6126 is ubiquitously released in high abundance from both chemosensitive and chemoresistant ovarian cancer cells via exosomes. Overexpression of miR-6126 was confirmed in healthy ovarian tissue compared with ovarian cancer patient samples and correlated with better overall survival in patients with high-grade serous ovarian cancer. miR-6126 acted as a tumor suppressor by directly targeting integrin-β1, a key regulator of cancer cell metastasis. miR-6126 mimic treatment of cancer cells resulted in increased miR-6126 and decreased integrin-β1 mRNA levels in the exosome. Functional analysis showed that treatment of endothelial cells with miR-6126 mimic significantly reduced tube formation as well as invasion and migration capacities of ovarian cancer cells in vitro. Administration of miR-6126 mimic in an orthotopic mouse model of ovarian cancer elicited a relative reduction in tumor growth, proliferating cells, and microvessel density. miR-6126 inhibition promoted oncogenic behavior by leading ovarian cancer cells to release more exosomes. Our findings provide new insights into the role of exosomal miRNA-mediated tumor progression and suggest a new therapeutic approach to disrupt oncogenic phenotypes in tumors. Cancer Res; 76(24); 7194–207. ©2016 AACR.</jats:p

Exosomal miRNA confers chemo resistance via targeting Cav1/p-gp/M2-type macrophage axis in ovarian cancerResearch in context

Background: Circulating miRNAs are known to play important roles in intercellular communication. However, the effects of exosomal miRNAs on cells are not fully understood. Methods: To investigate the role of exosomal miR-1246 in ovarian cancer (OC) microenvironment, we performed RPPA as well as many other in vitro functional assays in ovarian cancer cells (sensitive; HeyA8, Skov3ip1, A2780 and chemoresistant; HeyA8-MDR, Skov3-TR, A2780-CP20). Therapeutic effect of miR-1246 inhibitor treatment was tested in OC animal model. We showed the effect of OC exosomal miR-1246 uptake on macrophages by co-culture experiments. Findings: Substantial expression of oncogenic miR-1246 OC exosomes was found. We showed that Cav1 gene, which is the direct target of miR-1246, is involved in the process of exosomal transfer. A significantly worse overall prognosis were found for OC patients with high miR-1246 and low Cav1 expression based on TCGA data. miR-1246 expression were significantly higher in paclitaxel-resistant OC exosomes than in their sensitive counterparts. Overexpression of Cav1 and anti-miR-1246 treatment significantly sensitized OC cells to paclitaxel. We showed that Cav1 and multi drug resistance (MDR) gene is involved in the process of exosomal transfer. Our proteomic approach also revealed that miR-1246 inhibits Cav1 and acts through PDGFβ receptor at the recipient cells to inhibit cell proliferation. miR-1246 inhibitor treatment in combination with chemotherapy led to reduced tumor burden in vivo. Finally, we demonstrated that when OC cells are co-cultured with macrophages, they are capable of transferring their oncogenic miR-1246 to M2-type macrophages, but not M0-type macrophages. Interpretation: Our results suggest that cancer exosomes may contribute to oncogenesis by manipulating neighboring infiltrating immune cells. This study provide a new mechanistic therapeutic approach to overcome chemoresistance and tumor progression through exosomal miR-1246 in OC patients. Keywords: Exosome, oncomiR, miR-1246, Ovarian cancer, Cav1, P-g