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Abstract Background Despite advances in screening and therapy, breast cancer (BC) remains the predominant cancer in women globally. Dysregulation of microRNAs (miRNAs) is pivotal in carcinogenesis across various cancers, including BC. Evidence indicates that miR‐1307‐3p is upregulated in BC tumors, yet its target genes are not fully elucidated. This study aimed to explore how miR‐1307‐3p regulates BC proliferation, migration, invasion, and angiogenesis and to identify potential target genes. Methods Basal miR‐1307‐3p levels were quantified in BC cell lines MDA‐MB‐231 and MCF‐7, as well as MCF‐10A using quantitative real‐time reverse transcription‐PCR (RT‐qPCR). The impact of miR‐1307‐3p inhibition on BC cell proliferation, migration, invasion, and angiogenesis was assessed. Nine miRNA‐target prediction databases identified potential miR‐1307‐3p targets. Target expression was validated using RT‐qPCR, Western blot, and dual‐luciferase reporter assays. MiR‐1307‐3p was overexpressed in MDA‐MB‐231 and MCF‐7 compared to MCF‐10A. Results Inhibiting miR‐1307‐3p significantly reduced BC cell proliferation, migration, invasion, and angiogenesis. Bioinformatics analysis identified 17 potential miR‐1307‐3p targets, with protamine 2 (PRM2) overexpression confirmed via Western blot and dual‐luciferase assays. Conclusion MiR‐1307‐3p overexpression in BC promotes proliferation, migration, invasion, and angiogenesis. PRM2 emerges as a novel miR‐1307‐3p target in BC.

Abstract The development and progression of chronic lymphocytic leukemia (CLL) depend on genetic abnormalities and on the immunosuppressive microenvironment. We have explored the possibility that genetic drivers might be responsible for the immune cell dysregulation that shapes the protumor microenvironment. We performed a transcriptome analysis of coding and non-coding RNAs (ncRNAs) during leukemia progression in the Rag2−/−γc −/− MEC1-based xenotransplantation model. The DLEU2/miR-16 locus was found downmodulated in monocytes/macrophages of leukemic mice. To validate the role of this cluster in the tumor immune microenvironment, we generated a mouse model that simultaneously mimics the overexpression of hTCL1 and the germline deletion of the minimal deleted region (MDR) encoding the DLEU2/miR-15a/miR-16-1 cluster. This model provides an innovative and faster CLL system where monocyte differentiation and macrophage polarization are exacerbated, and T-cells are dysfunctional. MDR deletion inversely correlates with the levels of predicted target proteins including BCL2 and PD1/PD-L1 on murine CLL cells and immune cells. The inverse correlation of miR-15a/miR-16-1 with target proteins has been confirmed on patient-derived immune cells. Forced expression of miR-16-1 interferes with monocyte differentiation into tumor-associated macrophages, indicating that selected ncRNAs drive the protumor phenotype of non-malignant immune cells.

The exercise below aimed to familiarize students in the program with the security developments in the Black Sea Region over the previous seven years and to place them in the role of policy makers. Thus, through a series of debates and group activities, they could evaluate the main trends in the region, conceive of potential game changers and adopt policy solutions at both the national and supra-national level with the aim of realizing a vision of a secure and developed region. At the end of the exercise, students presented their own vision of the future and argued for the best policy responses which they would implement in order to achieve that vision.

Abstract Cyclin-dependent kinase 2-associated protein 1 (CDK2AP1; also known as deleted in oral cancer or DOC1) is a tumor suppressor gene known to play functional roles in both cell cycle regulation and in the epigenetic control of embryonic stem cell differentiation, the latter as a core subunit of the nucleosome remodeling and histone deacetylation (NuRD) complex. In the vast majority of oral squamous cell carcinomas (OSCC), expression of the CDK2AP1 protein is reduced or lost. Notwithstanding the latter (and the DOC1 acronym), mutations or deletions in its coding sequence are extremely rare. Accordingly, CDK2AP1 protein-deficient oral cancer cell lines express as much CDK2AP1 mRNA as proficient cell lines. Here, by combining in silico and in vitro approaches, and by taking advantage of patient-derived data and tumor material in the analysis of loss of CDK2AP1 expression, we identified a set of microRNAs, namely miR-21-5p, miR-23b-3p, miR-26b-5p, miR-93-5p, and miR-155-5p, which inhibit its translation in both cell lines and patient-derived OSCCs. Of note, no synergistic effects were observed of the different miRs on the CDK2AP1–3-UTR common target. We also developed a novel approach to the combined ISH/IF tissue microarray analysis to study the expression patterns of miRs and their target genes in the context of tumor architecture. Last, we show that CDK2AP1 loss, as the result of miRNA expression, correlates with overall survival, thus highlighting the clinical relevance of these processes for carcinomas of the oral cavity.

Abstract Background Studies have shown that cancer stemness and the endoplasmic reticulum (ER) stress response are inversely regulated in colorectal cancer (CRC), but the mechanism has not been fully clarified. Long noncoding RNAs (lncRNAs) play key roles in cancer progression and metastasis. In this study we investigated lncRNA 01534 (LINC01534) as a possible modulator between cancer stemness and ER stress response. Methods In vitro experiments using CRC cell lines were performed to explore a possible role of LINC01534. The expression of LINC01534 in clinical CRC samples was assessed by quantitative reverse transcription‐polymerase chain reaction (qRT‐PCR) and in situ hybridization. Results Silencing LINC01534 led to suppression of cell proliferation, invasiveness, and cell cycle progression at the G2‐M phase, and promoted apoptosis. Moreover, we found that silencing LINC01534 suppressed cancer stemness, while it activated the ER stress response, especially through the PERK/eIF2α signaling pathway. In situ hybridization revealed LINC01534 was expressed in tumor cells and upregulated in CRC tissues compared with normal epithelium. A survival survey indicated that high LINC01534 expression was significantly associated with shorter overall survival in 187 CRC patients. Conclusion This is the first report on LINC01534 in human cancer. Our findings suggest that LINC01534 may be an important modulator of the maintenance of cancer stemness and suppression of the ER stress response, and that it could be a novel prognostic factor in CRC.

Abstract Antiangiogenic treatment targeting the vascular endothelial growth factor (VEGF) pathway is a powerful tool to combat tumor growth and progression; however, drug resistance frequently emerges. We identify CD5L (CD5 antigen-like precursor) as an important gene upregulated in response to antiangiogenic therapy leading to the emergence of adaptive resistance. By using both an RNA-aptamer and a monoclonal antibody targeting CD5L, we are able to abate the pro-angiogenic effects of CD5L overexpression in both in vitro and in vivo settings. In addition, we find that increased expression of vascular CD5L in cancer patients is associated with bevacizumab resistance and worse overall survival. These findings implicate CD5L as an important factor in adaptive resistance to antiangiogenic therapy and suggest that modalities to target CD5L have potentially important clinical utility.

Sepsis remains a leading cause of death for humans and currently has no pathogenesis-specific therapy. Hampered progress is partly due to a lack of insight into deep mechanistic processes. In the past decade, deciphering the functions of small noncoding miRNAs in sepsis pathogenesis became a dynamic research topic. To screen for new miRNA targets for sepsis therapeutics, we used samples for miRNA array analysis of PBMCs from patients with sepsis and control individuals, blood samples from 2 cohorts of patients with sepsis, and multiple animal models: mouse cecum ligation puncture–induced (CLP-induced) sepsis, mouse viral miRNA challenge, and baboon Gram+ and Gram– sepsis models. miR-93-5p met the criteria for a therapeutic target, as it was overexpressed in baboons that died early after induction of sepsis, was downregulated in patients who survived after sepsis, and correlated with negative clinical prognosticators for sepsis. Therapeutically, inhibition of miR-93-5p prolonged the overall survival of mice with CLP-induced sepsis, with a stronger effect in older mice. Mechanistically, anti–miR-93-5p therapy reduced inflammatory monocytes and increased circulating effector memory T cells, especially the CD4+ subset. AGO2 IP in miR-93–KO T cells identified important regulatory receptors, such as CD28, as direct miR-93-5p target genes. In conclusion, miR-93-5p is a potential therapeutic target in sepsis through the regulation of both innate and adaptive immunity, with possibly a greater benefit for elderly patients than for young patients.

Noncoding RNAs play regulatory roles in physiopathology, but their involvement in neurodevelopmental diseases is poorly understood. Rett syndrome is a severe, progressive neurodevelopmental disorder linked to loss-of-function mutations of the MeCP2 gene for which no cure is yet available. Analysis of the noncoding RNA profile corresponding to the brain-abundant circular RNA (circRNA) and transcribed-ultraconserved region (T-UCR) populations in a mouse model of the disease reveals widespread dysregulation and enrichment in glutamatergic excitatory signaling and microtubule cytoskeleton pathways of the corresponding host genes. Proteomic analysis of hippocampal samples from affected individuals confirms abnormal levels of several cytoskeleton-related proteins together with key alterations in neurotransmission. Importantly, the glutamate receptor GRIA3 gene displays altered biogenesis in affected individuals and in vitro human cells and is influenced by expression of two ultraconserved RNAs. We also describe post-transcriptional regulation of SIRT2 by circRNAs, which modulates acetylation and total protein levels of GluR-1. As a consequence, both regulatory mechanisms converge on the biogenesis of AMPA receptors, with an effect on neuronal differentiation. In both cases, the noncoding RNAs antagonize MeCP2-directed regulation. Our findings indicate that noncoding transcripts may contribute to key alterations in Rett syndrome and are not only useful tools for revealing dysregulated processes but also molecules of biomarker value.

Poly(ADP-ribose) polymerase inhibitors (PARP inhibitors) have had an increasing role in the treatment of ovarian and breast cancers. PARP inhibitors are selectively active in cells with homologous recombination DNA repair deficiency caused by mutations in BRCA1/2 and other DNA repair pathway genes. Cancers with homologous recombination DNA repair proficiency respond poorly to PARP inhibitors. Cancers that initially respond to PARP inhibitors eventually develop drug resistance. We have identified salt-inducible kinase 2 (SIK2) inhibitors, ARN3236 and ARN3261, which decreased DNA double-strand break (DSB) repair functions and produced synthetic lethality with multiple PARP inhibitors in both homologous recombination DNA repair deficiency and proficiency cancer cells. SIK2 is required for centrosome splitting and PI3K activation and regulates cancer cell proliferation, metastasis, and sensitivity to chemotherapy. Here, we showed that SIK2 inhibitors sensitized ovarian and triple-negative breast cancer (TNBC) cells and xenografts to PARP inhibitors. SIK2 inhibitors decreased PARP enzyme activity and phosphorylation of class-IIa histone deacetylases (HDAC4/5/7). Furthermore, SIK2 inhibitors abolished class-IIa HDAC4/5/7–associated transcriptional activity of myocyte enhancer factor-2D (MEF2D), decreasing MEF2D binding to regulatory regions with high chromatin accessibility in FANCD2, EXO1, and XRCC4 genes, resulting in repression of their functions in the DNA DSB repair pathway. The combination of PARP inhibitors and SIK2 inhibitors provides a therapeutic strategy to enhance PARP inhibitor sensitivity for ovarian cancer and TNBC.

Deregulation of noncoding RNAs, including microRNAs (miRs), is implicated in the pathogenesis of many human cancers, including breast cancer. Through extensive analysis of The Cancer Genome Atlas, we found that expression of miR-22-3p is markedly lower in triple-negative breast cancer (TNBC) than in normal breast tissue. The restoration of miR-22-3p expression led to significant inhibition of TNBC cell proliferation, colony formation, migration, and invasion. We demonstrated that miR-22-3p reduces eukaryotic elongation factor 2 kinase (eEF2K) expression by directly binding to the 3′ untranslated region of eEF2K mRNA. Inhibition of EF2K expression recapitulated the effects of miR-22-3p on TNBC cell proliferation, motility, invasion, and suppression of phosphatidylinositol 3-kinase/Akt and Src signaling. Systemic administration of miR-22-3p in single-lipid nanoparticles significantly suppressed tumor growth in orthotopic MDA-MB-231 and MDA-MB-436 TNBC models. Evaluation of the tumor response, following miR-22-3p therapy in these models using a novel mathematical model factoring in various in vivo parameters, demonstrated that the therapy is highly effective against TNBC. These findings suggest that miR-22-3p functions as a tumor suppressor by targeting clinically significant oncogenic pathways and that miR-22-3p loss contributes to TNBC growth and progression. The restoration of miR-22-3p expression is a potential novel noncoding RNA-based therapy for TNBC.

Noncoding transcripts contribute to the adaptation of cellular processes to oxygen levels. Here the authors characterize a hypoxia responsive lncRNA lincNORS and show that it has a role in cellular sterol homeostasis.

The mechanisms underlying resistance to immunotherapy are still poorly understood. Here, the authors show that BMP7, a molecule part of the TGF-beta superfamily, suppresses proinflammatory antitumor responses and may represent a target for overcoming resistance to PD1 inhibitors.

Abstract Background Non-coding RNAs and especially microRNAs have been discovered to act as master regulators of cancer initiation and progression. The aim of our study was to discover and characterize the function of yet functionally uncharacterized microRNAs in human breast carcinogenesis. Methods In an unbiased approach, we utilized an established model system for breast cancer (BC) stem cell formation (“mammosphere assay”) to identify whole miRNome alterations in breast carcinogenesis. Clinical samples of BC patients were used to evaluate the human relevance of the newly identified miRNA candidates. One promising candidate, miR-1287-5p, was further explored on its impact on several hallmarks of cancer. The molecular mode of action was characterized by whole transcriptome analysis, in silico prediction tools, miRNA-interaction assays, pheno-copy assays, and drug sensitivity assays. Results Among several other microRNAs, miR-1287-5p was significantly downregulated in mammospheres and human BC tissue compared to normal breast tissue (p

Background: Inflammatory mediator prostaglandin E2–prostaglandin E2 receptor EP3 (PTGER3) signaling is critical for tumor-associated angiogenesis, tumor growth, and chemoresistance. However, the mechanism underlying these effects in ovarian cancer is not known. Methods: An association between higher tumoral expression of PTGER3 and shorter patient survival in the ovarian cancer dataset of The Cancer Genome Atlas prompted investigation of the antitumor effects of PTGER3 downmodulation. PTGER3 mRNA and protein levels were higher in cisplatin-resistant ovarian cancer cells than in their cisplatin-sensitive counterparts. Findings: Silencing of PTGER3 via siRNA in cancer cells was associated with decreased cell growth and less invasiveness, as well as cell-cycle arrest and increased apoptosis, mediated through the Ras-MAPK/Erk-ETS1-ELK1/CFTR1 axis. Furthermore, sustained PTGER3 silencing with multistage vector and liposomal 2’-F-phosphorodithioate-siRNA–mediated silencing of PTGER3 combined with cisplatin resulted in robust antitumor effects in cisplatin-resistant ovarian cancer models. Interpretation: These findings identify PTGER3 as a potential therapeutic target in chemoresistant ovarian cancers expressing high levels of this oncogenic protein. Fund: National Institutes of Health/National Cancer Institute, USA. Keywords: PTGER3, Ovarian cancer, RNA interference, Chemically modified siRNA, Cisplatin resistance, ETS1, ELK1, CFTR

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-gp

Summary: Tumor and stromal interactions consist of reciprocal signaling through cytokines, growth factors, direct cell-cell interactions, and extracellular vesicles (EVs). Small EVs (≤200 nm) have been considered critical messengers of cellular communication during tumor development. Here, we demonstrate that gain-of-function (GOF) p53 protein can be packaged into small EVs and transferred to fibroblasts. GOF p53 protein is selectively bound by heat shock protein 90 (HSP90), a chaperone protein, and packaged into small EVs. Inhibition of HSP90 activity blocks packaging of GOF, but not wild-type, p53 in small EVs. GOF p53-containing small EVs result in their conversion to cancer-associated fibroblasts. In vivo studies reveal that GOF p53-containing small EVs can enhance tumor growth and promote fibroblast transformation into a cancer-associated phenotype. These findings provide a better understanding of the complex interactions between cancer and stromal cells and may have therapeutic implications.

Digital communication channels can be subject to exploitation, resulting in harming individuals or societies by promoting fake news, disinformation, radicalization, or social polarization. It is important that communicators are well prepared in identifying, preventing, and responding to such actions, in understanding the vulnerabilities of their audience, while respecting the rights and liberties of the population. Thus, ethical and strategic communication should be of top priority for institutional communicators, academia, journalists, or stakeholders when addressing security or sensitive issues. The article presents the results of an empirical sociological study, part of an extensive exploratory research within the CRESCEnt project, aiming at identifying elements of enhancing critical thinking, responsible communication and accountable behaviour. Three European states were targeted by the study: Romania, Spain and Greece, while 28 practitioners in communication, intelligence and security and law enforcement contributed to the research with significant input on topics related to ethical, successful, and strategic communication characteristics. Ethical communication implies acting by ethical principles, like being truthful, accurate, and well-intentioned. Responsibility is the central element of ethical engagement when informing the public. Effective messages are characterized by clearness, validity of information, novelty and presented in real time. Different types of authorities come into action for the communicators, such as the journalism code of practice, European values, or possible negative social impact.

Tacrolimus has a narrow therapeutic window; a whole-blood trough target concentration of between 5 and 8 ng/mL is considered a safe level for stable kidney transplant recipients. Tacrolimus serum levels must be closely monitored to obtain a balance between maximizing efficacy and minimizing dose-related toxic effects. Currently, there is no specific tacrolimus toxicity biomarker except a graft biopsy. Our study aimed to identify specific serum metabolites correlated with tacrolinemia levels using serum high-precision liquid chromatography–mass spectrometry and standard laboratory evaluation. Three machine learning algorithms were used (Naïve Bayes, logistic regression, and Random Forest) in 19 patients with high tacrolinemia (8 ng/mL) and 23 patients with low tacrolinemia (5 ng/mL). Using a selected panel of five lipid metabolites (phosphatidylserine, phosphatidylglycerol, phosphatidylethanolamine, arachidyl palmitoleate, and ceramide), Mg2+, and uric acid, all three machine learning algorithms yielded excellent classification accuracies between the two groups. The highest classification accuracy was obtained by Naïve Bayes, with an area under the curve of 0.799 and a classification accuracy of 0.756. Our results show that using our identified five lipid metabolites combined with Mg2+ and uric acid serum levels may provide a novel tool for diagnosing tacrolimus toxicity in kidney transplant recipients. Further validation with targeted MS and biopsy-proven TAC toxicity is needed.

As one of the most recently (re)discovered types of non-coding RNAs (ncRNA), circular RNAs (circRNAs) differentiate from other ncRNAs by a specific biogenesis, high stability, and distinct functions. The biogenesis of circRNAs can be categorized into three mechanisms that permit the back-splicing reaction: exon-skipping, pairing of neighboring introns, and dimerization of RNA-binding proteins. Regarding their stability, circRNAs have no free ends, specific to linear RNA molecules, prompting a longer half-life and resistance to exonuclease-mediated activity by RNase R, bypassing the common RNA turnover process. Regarding their functions, circular transcripts can be categorized into four broad roles: miRNA sponging, protein binding, regulation of transcription, and coding for proteins and peptides. Female reproductive system (including mainly ovarian, corpus, and cervix uteri cancers) and breast cancers are the primary causes of death in women worldwide, accounting for over 1,212,772 deaths in 2018. We consider that a better understanding of the molecular pathophysiology through the study of coding and non-coding RNA regulators could improve the diagnosis and therapeutics of these cancers. Developments in the field of circRNA in regard to breast or gynecological cancers are recent, with most circRNA-related discoveries having been made in the last 2 years. Therefore, in this review we summarize the newly detected roles of circRNAs in female reproductive system (cervical cancer, ovarian cancer, and endometrial cancer) and breast cancers. We argue that circRNAs can become essential elements of the diagnostic and therapeutic tools for female reproductive system cancers in the future.

Ultraconserved regions (UCRs) are 481 genome segments, with length longer than 200 bp, that are 100% conserved among humans, mice, and rats. The majority of UCRs are transcriptionally active (T-UCRs) as many of them produce non-coding RNAs. In a previous study, we evaluated the expression level of T-UCRs in breast cancer (BC) patients and found that 63% of transcripts correlated with some clinical and/or molecular parameter of BC. In this study, we delved into the expression levels of 12 T-UCRs and correlated them with clinicopathological parameters, immunohistochemical markers, and overall survival in two breast cancer cohorts: TCGA and Brazilian patients. We found that uc.268 is more expressed in TCGA patients under 40 years of age, associated with progesterone receptor (PR) and estrogen receptor (ER), and its high expression is found in luminal A. Lower uc.84 and uc.376 were respectively observed in metastatic and stage IV tumors associated with good prognostic in luminal B. Moreover, uc.84 was only related to the HER2+, while uc.376 was related to ER+ and PR+, and HER2+. A panel composed of uc.147, uc.271, and uc.427 distinguished luminal A from triple negative patients with an AUC of 0.9531 (sensitivity 92.19% and specificity 86.76%). These results highlight the potential role of T-UCRs in BC and provide insights into the potential application of T-UCRs as biomarkers.

In ovarian cancer, metastatic phenotype may impact surgical outcomes. Here, the authors show miR-409-3p regulates FABP4 which can increase metastatic potential of ovarian cancer, and treatment with DOPC nanoliposomes containing either miR-409--3p mimic or FABP4 siRNA inhibits tumor progression in mice.

Abstract Background The disruption of normal gene regulation due to microRNA dysfunction is a common event in cancer pathogenesis. MicroRNA-27b is an example of an oncogenic miRNA, and it is frequently upregulated in breast cancer. MicroRNAs have been found to deregulate tumor metabolism, which typically manifests as heightened cellular glucose uptake in consort with increased flux through glycolysis, followed by the preferential conversion of glycolytic pyruvate into lactate (a phenomenon known as the Warburg Effect). Pyruvate Dehydrogenase, an enzyme complex linking glycolysis with downstream oxidative metabolism, represents a key location where regulation of metabolism occurs; PDHX is a key structural component of this complex and is essential for its function. Methods We sought to characterize the role of miR-27b in breast cancer by identifying novel transcripts under its control. We began by utilizing luciferase, RNA, and protein assays to establish PDHX as a novel target of miR-27b. We then tested whether miR-27b could alter metabolism using several metabolite assay kits and performed a seahorse analysis. We also examined how the altered metabolism might affect cell proliferation. Lastly, we confirmed the relevance of our findings in human breast tumor samples. Results Our data indicate that Pyruvate Dehydrogenase Protein X is a credible target of miR-27b in breast cancer. Mechanistically, by suppressing PDHX, miR-27b altered levels of pyruvate, lactate and citrate, as well as reducing mitochondrial oxidation and promoting extracellular acidification. These changes corresponded with an increased capacity for cell proliferation. In human breast tumor samples, PDHX expression was deficient, and low levels of PDHX were associated with reduced patient survival. Conclusions MicroRNA-27b targets PDHX, resulting in an altered metabolic configuration that is better suited to fuel biosynthetic processes and cell proliferation, thereby promoting breast cancer progression.

In melanoma, reduced ADAR1 impairs A-to-I microRNA editing. Here, the authors show that miR-378a-3p undergoes this editing in non-metastatic cells and the edited form of miR-378a-3p binds to the PARVA oncogene, inhibiting its expression and preventing melanoma progression and metastasis.

Despite substantial improvements in the treatment strategies, ovarian cancer is still the most lethal gynecological malignancy. Identification of drug treatable therapeutic targets and their safe and effective targeting is critical to improve patient survival in ovarian cancer. AXL receptor tyrosine kinase (RTK) has been proposed to be an important therapeutic target for metastatic and advanced-stage human ovarian cancer. We found that AXL-RTK expression is associated with significantly shorter patient survival based on the The Cancer Genome Atlas patient database. To target AXL-RTK, we developed a chemically modified serum nuclease-stable AXL aptamer (AXL-APTAMER), and we evaluated its in vitro and in vivo antitumor activity using in vitro assays as well as two intraperitoneal animal models. AXL-aptamer treatment inhibited the phosphorylation and the activity of AXL, impaired the migration and invasion ability of ovarian cancer cells, and led to the inhibition of tumor growth and number of intraperitoneal metastatic nodules, which was associated with the inhibition of AXL activity and angiogenesis in tumors. When combined with paclitaxel, in vivo systemic (intravenous [i.v.]) administration of AXL-aptamer treatment markedly enhanced the antitumor efficacy of paclitaxel in mice. Taken together, our data indicate that AXL-aptamers successfully target in vivo AXL-RTK and inhibit its AXL activity and tumor growth and progression, representing a promising strategy for the treatment of ovarian cancer.

T-UCRs encode long non-coding RNAs implicated in human carcinogenesis, but the underlying mechanisms are poorly understood. Here, the authors identify uc.339 as an oncogene in lung cancer that is upregulated through the loss of TP53 and promotes Cyclin E activation by entrapping regulatory miRNAs.

TRPA1 has been reported to contribute lung cancer adenocarcinoma (LUAD), but the mechanisms are unclear. Here the authors propose that TRPA1/FGFR2 interaction is functional in LUAD and show that astrocytes oppose brain metastasis by mediating the downregulation of TRPA1 through exosome-delivered miRNA-142-3p.

Platelets have been associated with increased tumor growth and metastasis but the mechanistic details of this interaction are still unclear. Here the authors show that platelets improve anoikis resistance of cancer cells and increase metastasis by activating Yap through a RhoA/MYPT-PP1 pathway.

Prevalence of Kaposi sarcoma-associated herpesvirus (KSHV/HHV-8) varies greatly in different populations. We hypothesized that the actual prevalence of KSHV/HHV8 infection in humans is underestimated by the currently available serological tests. We analyzed four independent patient cohorts with post-surgical or post-chemotherapy sepsis, chronic lymphocytic leukemia and post-surgical patients with abdominal surgical interventions. Levels of specific KSHV-encoded miRNAs were measured by reverse transcription-quantitative polymerase chain reaction (RT-qPCR), and KSHV/HHV-8 IgG were measured by immunoassay. We also measured specific miRNAs from Epstein Barr Virus (EBV), a virus closely related to KSHV/HHV-8, and determined the EBV serological status by ELISA for Epstein-Barr nuclear antigen 1 (EBNA-1) IgG. Finally, we identified the viral miRNAs by in situ hybridization (ISH) in bone marrow cells. In training/validation settings using independent multi-institutional cohorts of 300 plasma samples, we identified in 78.50% of the samples detectable expression of at least one of the three tested KSHV-miRNAs by RT-qPCR, while only 27.57% of samples were found to be seropositive for KSHV/HHV-8 IgG (P

Abstract Background Non-coding RNAs have been drawing increasing attention in recent years as functional data suggest that they play important roles in key cellular processes. N-BLR is a primate-specific long non-coding RNA that modulates the epithelial-to-mesenchymal transition, facilitates cell migration, and increases colorectal cancer invasion. Results We performed multivariate analyses of data from two independent cohorts of colorectal cancer patients and show that the abundance of N-BLR is associated with tumor stage, invasion potential, and overall patient survival. Through in vitro and in vivo experiments we found that N-BLR facilitates migration primarily via crosstalk with E-cadherin and ZEB1. We showed that this crosstalk is mediated by a pyknon, a short ~20 nucleotide-long DNA motif contained in the N-BLR transcript and is targeted by members of the miR-200 family. In light of these findings, we used a microarray to investigate the expression patterns of other pyknon-containing genomic loci. We found multiple such loci that are differentially transcribed between healthy and diseased tissues in colorectal cancer and chronic lymphocytic leukemia. Moreover, we identified several new loci whose expression correlates with the colorectal cancer patients’ overall survival. Conclusions The primate-specific N-BLR is a novel molecular contributor to the complex mechanisms that underlie metastasis in colorectal cancer and a potential novel biomarker for this disease. The presence of a functional pyknon within N-BLR and the related finding that many more pyknon-containing genomic loci in the human genome exhibit tissue-specific and disease-specific expression suggests the possibility of an alternative class of biomarkers and therapeutic targets that are primate-specific.

An amendment to this paper has been published and can be accessed via a link at the top of the paper.

The clinical significance of long noncoding RNAs (lncRNAs) in colorectal cancer (CRC) remains largely unexplored. Here, we analyzed a large panel of lncRNA candidates with The Cancer Genome Atlas (TCGA) CRC dataset, and identified H19 as the most significant lncRNA associated with CRC patient survival. We further validated such association in two independent CRC cohorts. H19 silencing blocked G1-S transition, reduced cell proliferation, and inhibited cell migration. We profiled gene expression changes to gain mechanism insight of H19 function. Transcriptome data analysis revealed not only previously identified mechanisms such as Let-7 regulation by H19, but also RB1-E2F1 function and β-catenin activity as essential upstream regulators mediating H19 function. Our experimental data showed that H19 affects phosphorylation of RB1 protein by regulating gene expression of CDK4 and CCND1. We further demonstrated that reduced CDK8 expression underlies changes of β-catenin activity, and identified that H19 interacts with macroH2A, an essential regulator of CDK8 gene transcription. However, the relevance of H19-macroH2A interaction in CDK8 regulation remains to be experimentally determined. We further explored the clinical relevance of above mechanisms in clinical samples, and showed that combined analysis of H19 with its targets improved prognostic value of H19 in CRC.

Resistance to chemotherapy is a hallmark of pancreatic ductal adenocarcinoma (PDA) and has been partly attributed to the dense desmoplastic stroma, which forms a protective niche for cancer cells. Tissue transglutaminase (TG2), a Ca2+-dependent enzyme, is secreted by PDA cells and cross-links proteins in the tumor microenvironment (TME) through acyl-transfer between glutamine and lysine residues, promoting PDA growth. The objective of the current study was to determine whether secreted TG2 by PDA cells alters the response of pancreatic tumors to gemcitabine. Orthotopic pancreatic xenografts and co-culture of PDA and stromal cells were employed to determine the mechanisms by which TG2 alters tumor-stroma interactions and response to gemcitabine. Analysis of the pancreatic The Cancer Genome Atlas (TCGA) database demonstrated that increased TG2 expression levels correlate with worse overall survival (hazard ratio = 1.37). Stable TG2 knockdown in PDA cells led to decreased size of pancreatic xenografts and increased sensitivity to gemcitabine in vivo. However, TG2 downregulation did not increase cytotoxicity of gemcitabine in vitro. Additionally, multivessel density and gemcitabine uptake in pancreatic tumor tissue, as measured by mass spectrometry (MS-HPLC), were not significantly different in tumors expressing TG2 versus tumors in which TG2 was knocked down. Fibroblasts, stimulated by TG2 secreted by PDA cells, secrete laminin A1, which protects cancer cells from gemcitabine-induced cytotoxicity. In all, our results demonstrate that TG2 secreted in the pancreatic TME orchestrates the cross talk between cancer cells and stroma, impacting tumor growth and response to chemotherapy. Our study supports TG2 inhibition to increase the antitumor effects of gemcitabine in PDA.

The formation of blood vessels in tumours, angiogenesis, is a promising target for therapy. Here, the authors show that microRNA192 has anti-angiogenic functions and negatively regulates EGR1 and HOXB9, and that delivery of this microRNA to tumours in vivocan reduce angiogenesis and tumour growth.

Abstract Large‐scale molecular annotation of epithelial ovarian cancer (EOC) indicates remarkable heterogeneity in the etiology of that disease. This diversity presents a significant obstacle against intervention target discovery. However, inactivation of miRNA biogenesis is commonly associated with advanced disease. Thus, restoration of miRNA activity may represent a common vulnerability among diverse EOC oncogenotypes. To test this, we employed genome‐scale, gain‐of‐function, miRNA mimic toxicity screens in a large, diverse spectrum of EOC cell lines. We found that all cell lines responded to at least some miRNA mimics, but that the nature of the miRNA mimics provoking a response was highly selective within the panel. These selective toxicity profiles were leveraged to define modes of action and molecular response indicators for miRNA mimics with tumor‐suppressive characteristics in vivo. A mechanistic principle emerging from this analysis was sensitivity of EOC to miRNA‐mediated release of cell fate specification programs, loss of which may be a prerequisite for development of this disease.

Long noncoding RNAs (lncRNAs) significantly influence the development and regulation of genome expression in cells. Here, we demonstrate the role of lncRNA ceruloplasmin (NRCP) in cancer metabolism and elucidate functional effects leading to increased tumor progression. NRCP was highly upregulated in ovarian tumors, and knockdown of NRCP resulted in significantly increased apoptosis, decreased cell proliferation, and decreased glycolysis compared with control cancer cells. In an orthotopic mouse model of ovarian cancer, siNRCP delivered via a liposomal carrier significantly reduced tumor growth compared with control treatment. We identified NRCP as an intermediate binding partner between STAT1 and RNA polymerase II, leading to increased expression of downstream target genes such as glucose-6-phosphate isomerase. Collectively, we report a previously unrecognized role of the lncRNA NRCP in modulating cancer metabolism. As demonstrated, DOPC nanoparticle-incorporated siRNA-mediated silencing of this lncRNA in vivo provides therapeutic avenue toward modulating lncRNAs in cancer.

An amendment to this paper has been published and can be accessed via a link at the top of the paper.

An amendment to this paper has been published and can be accessed via a link at the top of the paper.

Non-coding RNAs (ncRNAs) are essential players in many cellular processes, from normal development to oncogenic transformation. Initially, ncRNAs were defined as transcripts that lacked an open reading frame (ORF). However, multiple lines of evidence suggest that certain ncRNAs encode small peptides of less than 100 amino acids. The sequences encoding these peptides are known as small open reading frames (smORFs), many initiating with the traditional AUG start codon but terminating with atypical stop codons, suggesting a different biogenesis. The ncRNA-encoded peptides (ncPEPs) are gradually becoming appreciated as a new class of functional molecules that contribute to diverse cellular processes, and are deregulated in different diseases contributing to pathogenesis. As multiple publications have identified unique ncPEPs, we appreciated the need for assembling a new web resource that could gather information about these functional ncPEPs. We developed FuncPEP, a new database of functional ncRNA encoded peptides, containing all experimentally validated and functionally characterized ncPEPs. Currently, FuncPEP includes a comprehensive annotation of 112 functional ncPEPs and specific details regarding the ncRNA transcripts that encode these peptides. We believe that FuncPEP will serve as a platform for further deciphering the biologic significance and medical use of ncPEPs. The link for FuncPEP database can be found at the end of the Introduction Section.

Inflammatory breast cancer (IBC) is a rare and aggressive presentation of invasive breast cancer with a 62% to 68% 5-year survival rate. It is the most lethal form of breast cancer, and early recognition and treatment is important for patient survival. Like non-inflammatory breast cancer, IBC comprises multiple subtypes, with the triple-negative subtype being overrepresented. Although the current multimodality treatment regime of anthracycline- and taxane-based neoadjuvant therapy, surgery, and radiotherapy has improved the outcome of patients with triple-negative IBC, overall survival continues to be worse than in patients with non-inflammatory locally advanced breast cancer. Translation of new therapies into the clinics to successfully treat IBC has been poor, in part because of the lack of in vitro preclinical models that can accurately predict the response of the original tumor to therapy. We report the generation of a preclinical IBC patient-derived xenograft (PDX)-derived ex vivo (PDXEx) model and show that it closely replicates the tissue architecture of the original PDX tumor harvested from mice. The gene expression profile of our IBC PDXEx model had a high degree of correlation to that of the original tumor. This suggests that the process of generating the PDXEx model did not significantly alter the molecular signature of the original tumor. We demonstrate a high degree of similarity in drug response profile between a PDX mouse model and our PDXEx model generated from the same original PDX tumor tissue and treated with the same panel of drugs, indicating that our PDXEx model had high predictive value in identifying effective tumor-specific therapies. Finally, we used our PDXEx model as a platform for a robotic-based high-throughput drug screen of a 386-drug anti-cancer compound library. The top candidates identified from this drug screen all demonstrated greater therapeutic efficacy than the standard-of-care drugs used in the clinic to treat triple-negative IBC, doxorubicin and paclitaxel. Our PDXEx model is simple, and we are confident that it can be incorporated into a PDX mouse system for use as a first-pass screening platform. This will permit the identification of effective tumor-specific therapies with high predictive value in a resource-, time-, and cost-efficient manner.

Although numerous studies highlighted the role of Epstein–Barr Virus (EBV) in B-cell transformation, the involvement of EBV proteins or genome in the development of the most frequent adult leukemia, chronic lymphocytic leukemia (CLL), has not yet been defined. We hypothesized that EBV microRNAs contribute to progression of CLL and demonstrated the presence of EBV miRNAs in B-cells, in paraffin-embedded bone marrow biopsies and in the plasma of patients with CLL by using three different methods (small RNA-sequencing, quantitative reverse transcription PCR [q-RT-PCR] and miRNAs in situ hybridization [miRNA-ISH]). We found that EBV miRNA BHRF1-1 expression levels were significantly higher in the plasma of patients with CLL compared with healthy individuals (p

Cells adjust to hypoxic stress within the tumor microenvironment by downregulating energy-consuming processes including translation. To delineate mechanisms of cellular adaptation to hypoxia, we performed RNA-Seq of normoxic and hypoxic head and neck cancer cells. These data revealed a significant down regulation of genes known to regulate RNA processing and splicing. Exon-level analyses classified > 1,000 mRNAs as alternatively spliced under hypoxia and uncovered a unique retained intron (RI) in the master regulator of translation initiation, EIF2B5. Notably, this intron was expressed in solid tumors in a stage-dependent manner. We investigated the biological consequence of this RI and demonstrate that its inclusion creates a premature termination codon (PTC), that leads to a 65kDa truncated protein isoform that opposes full-length eIF2Bε to inhibit global translation. Furthermore, expression of 65kDa eIF2Bε led to increased survival of head and neck cancer cells under hypoxia, providing evidence that this isoform enables cells to adapt to conditions of low oxygen. Additional work to uncover -cis and -trans regulators of EIF2B5 splicing identified several factors that influence intron retention in EIF2B5: a weak splicing potential at the RI, hypoxia-induced expression and binding of the splicing factor SRSF3, and increased binding of total and phospho-Ser2 RNA polymerase II specifically at the intron retained under hypoxia. Altogether, these data reveal differential splicing as a previously uncharacterized mode of translational control under hypoxia and are supported by a model in which hypoxia-induced changes to cotranscriptional processing lead to selective retention of a PTC-containing intron in EIF2B5.

Posttranscriptional maturation is a critical step in microRNA (miRNA) biogenesis that determines mature miRNA levels. In addition to core components (Drosha and DGCR8 [DiGeorge syndrome critical region gene 8]) in the microprocessor, regulatory RNA-binding proteins may confer the specificity for recruiting and processing of individual primary miRNAs (pri-miRNAs). Here, we identify DDX1 as a regulatory protein that promotes the expression of a subset of miRNAs, including five members in the microRNA-200 (miR-200) family and four miRNAs in an eight-miRNA signature of a mesenchymal ovarian cancer subtype. A majority of DDX1-dependent miRNAs are induced after DNA damage. This induction is facilitated by the ataxia telangiectasia mutated (ATM)-mediated phosphorylation of DDX1. Inhibiting DDX1 promotes ovarian tumor growth and metastasis in a syngeneic mouse model. Analysis of The Cancer Genome Atlas (TCGA) reveals that low DDX1 levels are associated with poor clinical outcome in patients with serous ovarian cancer. These findings suggest that DDX1 is a key modulator in miRNA maturation and ovarian tumor suppression.

We describe a role for the complement system in enhancing cancer growth. Cancer cells secrete complement proteins that stimulate tumor growth upon activation. Complement promotes tumor growth via a direct autocrine effect that is partially independent of tumor-infiltrating cytotoxic T cells. Activated C5aR and C3aR signal through the PI3K/AKT pathway in cancer cells, and silencing the PI3K or AKT gene in cancer cells eliminates the progrowth effects of C5aR and C3aR stimulation. In patients with ovarian or lung cancer, higher tumoral C3 or C5aR mRNA levels were associated with decreased overall survival. These data identify a role for tumor-derived complement proteins in promoting tumor growth, and they therefore have substantial clinical and therapeutic implications.