The endocrine disruptor cadmium modulates the androgen-estrogen receptors ratio and induces inflammatory cytokines in luminal (A) cell models of breast cancer.

PURPOSE: Breast cancer (BC) is the most common malignancy that affects women, and it is, to date, their leading cause of death. Luminal A molecular subtype accounts for 40% of BC and is characterized by hormone receptors positive/human epidermal growth factor 2 expression and current treatment consists of surgery plus aromatase inhibitor therapy. Interestingly, several studies demonstrated that the heavy metal cadmium (Cd), classified as a group 1 human carcinogen and widely spread in the environment, exerts estrogen-like activities in several tissues and suggested an intriguing relationship between increased Cd exposure and BC incidence. Thus, aim of this study was to evaluate effects of Cd on Luminal A BC estrogen receptor (ER) positive/progesterone receptor positive cell models in vitro to characterize the mechanism(s) involved in breast cell homeostasis disruption. METHODS: T47D and MCF7 were exposed to Cd (0.5-1 µM) for 6-24 h to evaluate potential alterations in: cells viability, steroid receptors and intracellular signaling by western blot. Moreover, we evaluated the expression of inflammatory cytokines interleukin by RT-PCR. RESULTS: Our results showed a significant induction of androgen receptor (AR) and an increased AR/ER ratio. Further, Cd exposure increased pro-inflammatory cytokines interleukin (IL)6, IL8 and tumor necrosis factor α levels. Finally, as previously demonstrated by our group, Cd alters pathways such as mitogen-activated protein kinase family and protein kinase B. CONCLUSION: In conclusion, our study demonstrates that Cd modifies the expression and pattern of ERs and AR in BC cell lines, suggesting an alteration of BC cells homeostasis, likely predisposing to a carcinogenetic microenvironment.

Network analysis identifies circulating miR-155 as predictive biomarker of type 2 diabetes mellitus development in obese patients: a pilot study.

Obesity is the main risk factor for many non-communicable diseases. In clinical practice, unspecific markers are used for the determination of metabolic alterations and inflammation, without allowing the characterization of subjects at higher risk of complications. Circulating microRNAs represent an attractive approach for early screening to identify subjects affected by obesity more at risk of developing connected pathologies. The aim of this study was the identification of circulating free and extracellular vesicles (EVs)-embedded microRNAs able to identify obese patients at higher risk of type 2 diabetes (DM2). The expression data of circulating microRNAs derived from obese patients (OB), with DM2 (OBDM) and healthy donors were combined with clinical data, through network-based methodology implemented by weighted gene co-expression network analysis. The six circulating microRNAs overexpressed in OBDM patients were evaluated in a second group of patients, confirming the overexpression of miR-155-5p in OBDM patients. Interestingly, the combination of miR-155-5p with serum levels of IL-8, Leptin and RAGE was useful to identify OB patients most at risk of developing DM2. These results suggest that miR-155-5p is a potential circulating biomarker for DM2 and that the combination of this microRNA with other inflammatory markers in OB patients can predict the risk of developing DM2.

HHV-6A Infection of Papillary Thyroid Cancer Cells Induces Several Effects Related to Cancer Progression.

Recent studies have shown that thyrocytes are permissive to HHV-6A infection and that the virus may contribute to the pathogenesis of autoimmune thyroiditis. Thyroid autoimmune diseases increase the risk of papillary cancer, which is not surprising considering that chronic inflammation activates pathways that are also pro-oncogenic. Moreover, in this condition, cell proliferation is stimulated as an attempt to repair tissue damage caused by the inflammatory process. Interestingly, it has been reported that the well-differentiated papillary thyroid carcinoma (PTC), the less aggressive form of thyroid tumor, may progress to the more aggressive follicular thyroid carcinoma (FTC) and eventually to the anaplastic thyroid carcinoma (ATC), and that to such progression contributes the presence of an inflammatory/immune suppressive tumor microenvironment. In this study, we investigated whether papillary tumor cells (BCPAP) could be infected by human herpes virus-6A (HHV-6A), and if viral infection could induce effects related to cancer progression. We found that the virus dysregulated the expression of several microRNAs, such as miR-155, miR-9, and the miR-221/222 cluster, which are involved in different steps of carcinogenesis, and increased the secretion of pro-inflammatory cytokines, particularly IL-6, which may also sustain thyroid tumor cell growth and promote cancer progression. Genomic instability and the expression of PTEN, reported to act as an oncogene in mutp53-carrying cells such as BCPAP, also increased following HHV-6A-infection. These findings suggest that a ubiquitous herpesvirus such as HHV-6A, which displays a marked tropism for thyrocytes, could be involved in the progression of PTC towards more aggressive forms of thyroid tumor.

The inhibition of IRE1alpha/XBP1 axis prevents EBV-driven lymphomagenesis in NSG mice.

IMPORTANCE: The novelty of this study lies in the fact that it shows that IRE1 alpha endoribonuclease inhibition by 4µ8C was able to counteract Epstein-Barr virus-driven lymphomagenesis in NOD SCID gamma mice and prevent B-cell immortalization in vitro, unveiling that this drug may be a promising therapeutic approach to reduce the risk of post-transplant lymphoproliferative disorders (PTLD) onset in immune-deficient patients. This hypothesis is further supported by the fact that 4µ8C impaired the survival of PTLD-like cells derived from mice, meaning that it could be helpful also in the case in which there is the possibility that these malignancies have begun to arise.

Circulating cell-free DNA (cfDNA) in patients with medullary thyroid carcinoma is characterized by specific fragmentation and methylation changes with diagnostic value.

Medullary Thyroid Carcinoma (MTC) is a rare neuroendocrine tumour whose diagnosis includes evaluating calcitonin serum levels, which can present fluctuations unrelated to MTC. Here, we investigated circulating DNA fragmentation and methylation changes as potential biomarkers using ddPCR on cell-free DNA (cfDNA) isolated from the plasma of MTC patients. For cfDNA fragmentation analysis, we investigated the fragment size distribution of a gene family and calculated short fragment fraction (SFF). Methylation analyses evaluated the methylation levels of CG_16698623, a CG dinucleotide in the MGMT gene that we found hypermethylated in MTC tissues by analyzing public databases. The SFF ratio and methylation of CG_16698623 were significantly increased in plasma from MTC patients at diagnosis, and patients with clinical remission or stable disease at follow-up showed no significant SFF difference compared with healthy subjects. Our data support the diagnostic value of cfDNA traits that could enable better management of MTC patients.

Targeting FGFRs by pemigatinib induces G1 phase cell cycle arrest, cellular stress and upregulation of tumor suppressor microRNAs.

BACKGROUND: Fibroblast growth factor receptor (FGFR) gene family alterations are found in several cancers, indicating their importance as potential therapeutic targets. The FGFR-tyrosine kinase inhibitor (TKI) pemigatinib has been introduced in the treatment of advanced cholangiocarcinoma and more recently for relapsed or refractory myeloid/lymphoid neoplasms with FGFR2 and FGFR1 rearrangements, respectively. Several clinical trials are currently investigating the possible combination of pemigatinib with immunotherapy. In this study, we analyzed the biological and molecular effects of pemigatinib on different cancer cell models (lung, bladder, and gastric), which are currently objective of clinical trial investigations. METHODS: NCI-H1581 lung, KATO III gastric and RT-112 bladder cancer cell lines were evaluated for FGFR expression by qRT-PCR and Western blot. Cell lines were treated with Pem and then characterized for cell proliferation, apoptosis, production of intracellular reactive oxygen species (ROS), and induction of senescence. The expression of microRNAs with tumor suppressor functions was analyzed by qRT-PCR, while modulation of the proteins coded by their target genes was evaluated by Western blot and mRNA. Descriptive statistics was used to analyze the various data and student's t test to compare the analysis of two groups. RESULTS: Pemigatinib exposure triggered distinct signaling pathways and reduced the proliferative ability of all cancer cells, inducing G1 phase cell cycle arrest and strong intracellular stress resulting in ROS production, senescence and apoptosis. Pemigatinib treatment also caused the upregulation of microRNAs (miR-133b, miR-139, miR-186, miR-195) with tumor suppressor functions, along with the downregulation of validated protein targets with oncogenic roles (c-Myc, c-MET, CDK6, EGFR). CONCLUSIONS: These results contribute to clarifying the biological effects and molecular mechanisms mediated by the anti-FGFR TKI pemigatinib in distinct tumor settings and support its exploitation for combined therapies.

Molecular mechanisms of the tyrosine kinase inhibitor pralsetinib activity in in-vitro models of medullary thyroid carcinoma: Aberrant activation of the HH-Gli signaling pathway in acquired resistance.

Medullary thyroid carcinoma (MTC) is a malignant tumor with challenging management. Multi-targeted kinase inhibitors (MKI) and tyrosine-kinase inhibitors (TKI) with high specificity for RET protein are approved for advanced MTC treatment. However, their efficacy is hindered by evasion mechanisms of tumor cells. Thus, the aim of this study was the identification of an escape mechanism in MTC cells exposed to a highly selective RET TKI. TT cells were treated with TKI, MKI, and/or the HH-Gli inhibitors, GANT61 and Arsenic Trioxide (ATO), in the presence or absence of hypoxia. RET modifications, oncogenic signaling activation, proliferation and apoptosis were assessed. Additionally, cell modifications and HH-Gli activation were also evaluated in pralsetinib-resistant TT cells. Pralsetinib inhibited RET autophosphorylation and RET downstream pathways activation in normoxic and hypoxic conditions. Additionally, pralsetinib impaired proliferation, induced the activation of apoptosis and, in hypoxic cells, downregulated HIF-1α. Focusing on escape molecular mechanisms associated with therapy, we observed increased Gli1 levels in a subset of cells. Indeed, pralsetinib stimulated the re-localization of Gli1 into the cell nuclei. Treatment of TT cells with both pralsetinib and ATO resulted in Gli1 down-regulation and impaired cell viability. Moreover, pralsetinib-resistant cells confirmed Gli1 activation and up-regulation of its transcriptionally regulated target genes. Altogether, we showed that pralsetinib impairs MTC cell growth and induces cell death, also in hypoxic conditions. The HH-Gli pathway is a new molecular mechanism of escape to pralsetinib therapy that can be overcome through combined therapy.

A diagnostic circulating miRNA signature as orchestrator of cell invasion via TKS4/TKS5/EFHD2 modulation in human gliomas.

BACKGROUND: Altered microRNA profiles have been observed not only in tumour tissues but also in biofluids, where they circulate in a stable form thus representing interesting biomarker candidates. This study aimed to identify a microRNA signature as a non-invasive biomarker and to investigate its impact on glioma biology. METHODS: MicroRNAs were selected using a global expression profile in preoperative serum samples from 37 glioma patients. Comparison between serum samples from age and gender-matched controls was performed by using the droplet digital PCR. The ROC curve and Kaplan-Meier survival analyses were used to evaluate the diagnostic/prognostic values. The functional role of the identified signature was assessed by gain/loss of function strategies in glioma cells. RESULTS: A three-microRNA signature (miR-1-3p/-26a-1-3p/-487b-3p) was differentially expressed in the serum of patients according to the isocitrate dehydrogenase (IDH) genes mutation status and correlated with both patient Overall and Progression Free Survival. The identified signature was also downregulated in the serum of patients compared to controls. Consistent with these results, the signature expression and release in the conditioned medium of glioma cells was lower in IDH-wild type cells compared to the mutated counterpart. Furthermore, in silico analysis of glioma datasets showed a consistent deregulation of the signature according to the IDH mutation status in glioma tumour tissues. Ectopic expression of the signature negatively affects several glioma functions. Notably, it impacts the glioma invasive phenotype by directly targeting the invadopodia-related proteins TKS4, TKS5 and EFHD2. CONCLUSIONS: We identified a three microRNA signature as a promising complementary or even an independent non-invasive diagnostic/prognostic biomarker. The signature displays oncosuppressive functions in glioma cells and impacts on proteins crucial for migration and invasion, providing potential targets for therapeutic intervention.

ß-arrestin1-E2F1-ac axis regulates physiological apoptosis and cell cycle exit in cellular models of early postnatal cerebellum.

Development of the cerebellum is characterized by rapid proliferation of cerebellar granule cell precursors (GCPs) induced by paracrine stimulation of Sonic hedgehog (Shh) signaling from Purkinje cells, in the external granular layer (EGL). Then, granule cell precursors differentiate and migrate into the inner granular layer (IGL) of the cerebellum to form a terminally differentiated cell compartment. Aberrant activation of Sonic hedgehog signaling leads to granule cell precursors hyperproliferation and the onset of Sonic hedgehog medulloblastoma (MB), the most common embryonal brain tumor. ß-arrestin1 (ARRB1) protein plays an important role downstream of Smoothened, a component of the Sonic hedgehog pathway. In the medulloblastoma context, ß-arrestin1 is involved in a regulatory axis in association with the acetyltransferase P300, leading to the acetylated form of the transcription factor E2F1 (E2F1-ac) and redirecting its activity toward pro-apoptotic gene targets. This axis in the granule cell precursors physiological context has not been investigated yet. In this study, we demonstrate that ß-arrestin1 has antiproliferative and pro-apoptotic functions in cerebellar development. ß-arrestin1 silencing increases proliferation of Sonic hedgehog treated-cerebellar precursor cells while decreases the transcription of E2F1-ac pro-apoptotic targets genes, thus impairing apoptosis. Indeed, chromatin immunoprecipitation experiments show a direct interaction between ß-arrestin1 and the promoter regions of the pro-apoptotic E2F1 target gene and P27, indicating the double role of ß-arrestin1 in controlling apoptosis and cell cycle exit in a physiological context. Our data elucidate the role of ß-arrestin1 in the early postnatal stages of cerebellar development, in those cell compartments that give rise to medulloblastoma. This series of experiments suggests that the physiological function of ß-arrestin1 in neuronal progenitors is to directly control, cooperating with E2F1 acetylated form, transcription of pro-apoptotic genes.

Hedgehog-GLI and Notch Pathways Sustain Chemoresistance and Invasiveness in Colorectal Cancer and Their Inhibition Restores Chemotherapy Efficacy.

Colorectal cancer (CRC) is a leading cause of cancer-related mortality and chemoresistance is a major medical issue. The epithelial-to-mesenchymal transition (EMT) is the primary step in the emergence of the invasive phenotype and the Hedgehog-GLI (HH-GLI) and NOTCH signaling pathways are associated with poor prognosis and EMT in CRC. CRC cell lines harboring KRAS or BRAF mutations, grown as monolayers and organoids, were treated with the chemotherapeutic agent 5-Fluorouracil (5-FU) alone or combined with HH-GLI and NOTCH pathway inhibitors GANT61 and DAPT, or arsenic trioxide (ATO) to inhibit both pathways. Treatment with 5-FU led to the activation of HH-GLI and NOTCH pathways in both models. In KRAS mutant CRC, HH-GLI and NOTCH signaling activation co-operate to enhance chemoresistance and cell motility, while in BRAF mutant CRC, the HH-GLI pathway drives the chemoresistant and motile phenotype. We then showed that 5-FU promotes the mesenchymal and thus invasive phenotype in KRAS and BRAF mutant organoids and that chemosensitivity could be restored by targeting the HH-GLI pathway in BRAF mutant CRC or both HH-GLI and NOTCH pathways in KRAS mutant CRC. We suggest that in KRAS-driven CRC, the FDA-approved ATO acts as a chemotherapeutic sensitizer, whereas GANT61 is a promising chemotherapeutic sensitizer in BRAF-driven CRC.

Extra-Cellular Vesicles Derived from Thyroid Cancer Cells Promote the Epithelial to Mesenchymal Transition (EMT) and the Transfer of Malignant Phenotypes through Immune Mediated Mechanisms.

Thyroid cancer is the most common endocrine cancer, and its incidence is increasing in many countries around the world. Among thyroid cancers, the papillary thyroid cancer (PTC) histotype is particularly prevalent. A small percentage of papillary tumors is associated with metastases and aggressive behavior due to de-differentiation obtained through the epithelial-mesenchymal transition (EMT) by which epithelial thyroid cells acquire a fibroblast-like morphology, reduce cellular adhesion, increase motility and expression of mesenchymal proteins. The tumor microenvironment plays an important role in promoting an aggressive phenotype through hypoxia and the secretion of HMGB1 and other factors. Hypoxia has been shown to drastically change the tumor cell phenotype and has been associated with increasing metastatic and migratory behavior. Cells transfer information to neighboring cells or distant locations by releasing extracellular membrane vesicles (EVs) that contain key molecules, such as mRNAs, microRNAs (miRNAs), and proteins, that are able to modify protein expression in recipient cells. In this study, we investigated the potential role of EVs released by the anaplastic cancer cell line CAL-62 in inducing a malignant phenotype in a papillary cancer cell line (BCPAP).

A novel microRNA signature for the detection of melanoma by liquid biopsy.

BACKGROUND: Melanoma is the deadliest form of skin cancer and metastatic disease is associated with a significant survival rate drop. There is an urgent need for consistent tumor biomarkers to scale precision medicine and reduce cancer mortality. Here, we aimed to identify a melanoma-specific circulating microRNA signature and assess its value as a diagnostic tool. METHODS: The study consisted of a discovery phase and two validation phases. Circulating plasma extracellular vesicles (pEV) associated microRNA profiles were obtained from a discovery cohort of metastatic melanoma patients and normal subjects as controls. A pEV-microRNA signature was obtained using a LASSO penalized logistic regression model. The pEV-microRNA signature was subsequently validated both in a publicly available dataset and in an independent internal cohort. RESULTS: We identified and validated in three independent cohorts a panel of melanoma-specific circulating microRNAs that showed high accuracy in differentiating melanoma patients from healthy subjects with an area under the curve (AUC) of 1.00, 0.94 and 0.75 respectively. Investigation of the function of the pEV-microRNA signature evidenced their possible immune suppressive role in melanoma patients. CONCLUSIONS: We demonstrate that a blood test based on circulating microRNAs can non-invasively detect melanoma, offering a novel diagnostic tool for improving standard care. Moreover, we revealed an immune suppressive role for melanoma pEV-microRNAs.

Reduced miR-184-3p expression protects pancreatic ß-cells from lipotoxic and proinflammatory apoptosis in type 2 diabetes via CRTC1 upregulation.

The loss of functional ß-cell mass in type 2 diabetes (T2D) is associated with molecular events that include ß-cell apoptosis, dysfunction and/or dedifferentiation. MicroRNA miR-184-3p has been shown to be involved in several ß-cell functions, including insulin secretion, proliferation and survival. However, the downstream targets and upstream regulators of miR-184-3p have not been fully elucidated. Here, we show reduced miR-184-3p levels in human T2D pancreatic islets, whereas its direct target CREB regulated transcription coactivator 1 (CRTC1) was increased and protects ß-cells from lipotoxicity- and inflammation-induced apoptosis. Downregulation of miR-184-3p in ß-cells leads to upregulation of CRTC1 at both the mRNA and protein levels. Remarkably, the protective effect of miR-184-3p is dependent on CRTC1, as its silencing in human ß-cells abrogates the protective mechanism mediated by inhibition of miR-184-3p. Furthermore, in accordance with miR-184-3p downregulation, we also found that the ß-cell-specific transcription factor NKX6.1, DNA-binding sites of which are predicted in the promoter sequence of human and mouse MIR184 gene, is reduced in human pancreatic T2D islets. Using chromatin immunoprecipitation analysis and mRNA silencing experiments, we demonstrated that NKX6.1 directly controls both human and murine miR-184 expression. In summary, we provide evidence that the decrease in NKX6.1 expression is accompanied by a significant reduction in miR-184-3p expression and that reduction of miR-184-3p protects ß-cells from apoptosis through a CRTC1-dependent mechanism.

MiR-1248: a new prognostic biomarker able to identify supratentorial hemispheric pediatric low-grade gliomas patients associated with progression.

BACKGROUND: Pediatric low-grade gliomas (pLGGs), particularly incompletely resected supratentorial tumours, can undergo progression after surgery. However to date, there are no predictive biomarkers for progression. Here, we aimed to identify pLGG-specific microRNA signatures and evaluate their value as a prognostic tool. METHODS: We identified and validated supratentorial incompletey resected pLGG-specific microRNAs in independent cohorts from four European Pediatric Neuro-Oncology Centres. RESULTS: These microRNAs demonstrated high accuracy in differentiating patients with or without progression. Specifically, incompletely resected supratentorial pLGGs with disease progression showed significantly higher miR-1248 combined with lower miR-376a-3p and miR-888-5p levels than tumours without progression. A significant (p < 0.001) prognostic performance for miR-1248 was reported with an area under the curve (AUC) of 1.00. We also highlighted a critical oncogenic role for miR-1248 in gliomas tumours. Indeed, high miR-1248 levels maintain low its validated target genes (CDKN1A (p21)/FRK/SPOP/VHL/MTAP) and consequently sustain the activation of oncogenic pathways. CONCLUSIONS: Altogether, we provide a novel molecular biomarker able to successfully identify pLGG patients associated with disease progression that could support the clinicians in the decision-making strategy, advancing personalized medicine.

Circulating microRNAs as clinically useful biomarkers for Type 2 Diabetes Mellitus: miRNomics from bench to bedside.

Type 2 diabetes (T2D), a chronic metabolic disease, has attained the status of a global epidemic with steadily increasing incidence worldwide. Improved diagnosis, stratification and prognosis of T2D patients and the development of more effective treatments are needed. In this era of personalized medicine, the discovery and evaluation of innovative circulating biomarkers can be an effective tool for better stratification, prognosis and therapeutic selection/management of T2D patients. MicroRNAs (miRNAs), a class of small non-coding RNAs that modulate gene expression, have been investigated as potential circulating biomarkers in T2D. Several studies have investigated the expression of circulating miRNAs in T2D patients from various biological fluids, including plasma and serum, and have demonstrated their potential as diagnostic and prognostic biomarkers, as well as biomarkers of response to therapy. In this review, we provide an overview of the current state of knowledge, focusing on circulating miRNAs that have been consistently expressed in at least two independent studies, in order to identify a set of consistent biomarker candidates in T2D. The expression levels of miRNAs, correlation with clinical parameters, functional roles of miRNAs and their potential as biomarkers are reported. A systematic literature search and assessment of studies led to the selection and review of 10 miRNAs (miR-126-3p, miR-223-3p, miR-21-5p, miR-15a-5p, miR-24-3p, miR-34a-5p, miR-146a-5p, miR-148a-3p, miR-30d-5p and miR-30c-5p). We also present technical challenges and our thoughts on the potential validation of circulating miRNAs and their application as biomarkers in the context of T2D.

HOXD8 hypermethylation as a fully sensitive and specific biomarker for biliary tract cancer detectable in tissue and bile samples.

BACKGROUND: Biliary tract cancers (BTC) are rare but highly aggressive tumours with poor prognosis, usually detected at advanced stages. Herein, we aimed at identifying BTC-specific DNA methylation alterations. METHODS: Study design included statistical power and sample size estimation. A genome-wide methylation study of an explorative cohort (50 BTC and ten matched non-tumoral tissue samples) has been performed. BTC-specific altered CpG islands were validated in over 180 samples (174 BTCs and 13 non-tumoral controls). The final biomarkers, selected by a machine-learning approach, were validated in independent tissue (18 BTCs, 14 matched non-tumoral samples) and bile (24 BTCs, five non-tumoral samples) replication series, using droplet digital PCR. RESULTS: We identified and successfully validated BTC-specific DNA methylation alterations in over 200 BTC samples. The two-biomarker panel, selected by an in-house algorithm, showed an AUC > 0.97. The best-performing biomarker (chr2:176993479-176995557), associated with HOXD8, a pivotal gene in cancer-related pathways, achieved 100% sensitivity and specificity in a new series of tissue and bile samples. CONCLUSIONS: We identified a novel fully efficient BTC biomarker, associated with HOXD8 gene, detectable both in tissue and bile by a standardised assay ready-to-use in clinical trials also including samples from non-invasive matrices.

Targeting cancer stem cells in medulloblastoma by inhibiting AMBRA1 dual function in autophagy and STAT3 signalling.

Medulloblastoma (MB) is a childhood malignant brain tumour comprising four main subgroups characterized by different genetic alterations and rate of mortality. Among MB subgroups, patients with enhanced levels of the c-MYC oncogene (MBGroup3) have the poorest prognosis. Here we identify a previously unrecognized role of the pro-autophagy factor AMBRA1 in regulating MB. We demonstrate that AMBRA1 expression depends on c-MYC levels and correlates with Group 3 patient poor prognosis; also, knockdown of AMBRA1 reduces MB stem potential, growth and migration of MBGroup3 stem cells. At a molecular level, AMBRA1 mediates these effects by suppressing SOCS3, an inhibitor of STAT3 activation. Importantly, pharmacological inhibition of autophagy profoundly affects both stem and invasion potential of MBGroup3 stem cells, and a combined anti-autophagy and anti-STAT3 approach impacts the MBGroup3 outcome. Taken together, our data support the c-MYC/AMBRA1/STAT3 axis as a strong oncogenic signalling pathway with significance for both patient stratification strategies and targeted treatments of MBGroup3.

Role of tissue and circulating microRNAs and DNA as biomarkers in medullary thyroid cancer.

Medullary thyroid carcinoma (MTC) is a rare neuroendocrine tumor comprising hereditary or sporadic form with frequent mutations in the rearranged during transfection (RET) or RAS genes. Diagnosis is based on the presence of thyroid tumor mass with altered levels of calcitonin (Ctn) and carcinoembryonal antigen (CEA) in the serum and/or in the cytological smears from fine needle aspiration biopsies. Treatment consists of total thyroidectomy, followed by tyrosine kinase inhibitors (TKi) in case of disease persistence. During TKi treatment, Ctn and CEA levels can fluctuate regardless of tumor volume, metastasis or response to therapy. Research for more reliable non-invasive biomarkers in MTC is still underway. In this context, circulating nucleic acids, namely circulating microRNAs (miRNAs) and cell free DNA (cfDNA), have been evaluated by different research groups. Aiming to shed light on whether miRNAs and cfDNA are suitable as MTC biomarkers we searched three different databases, PubMed, Scopus, WOS and reviewed the literature. We classified 83 publications fulfilling our search criteria and summarized the results. We report data on miRNAs and cfDNA that can be evaluated for validation in independent studies and clinical application.

Downregulation of miR-326 and its host gene ß-arrestin1 induces pro-survival activity of E2F1 and promotes medulloblastoma growth.

Persistent mortality rates of medulloblastoma (MB) and severe side effects of the current therapies require the definition of the molecular mechanisms that contribute to tumor progression. Using cultured MB cancer stem cells and xenograft tumors generated in mice, we show that low expression of miR-326 and its host gene ß-arrestin1 (ARRB1) promotes tumor growth enhancing the E2F1 pro-survival function. Our models revealed that miR-326 and ARRB1 are controlled by a bivalent domain, since the H3K27me3 repressive mark is found at their regulatory region together with the activation-associated H3K4me3 mark. High levels of EZH2, a feature of MB, are responsible for the presence of H3K27me3. Ectopic expression of miR-326 and ARRB1 provides hints into how their low levels regulate E2F1 activity. MiR-326 targets E2F1 mRNA, thereby reducing its protein levels; ARRB1, triggering E2F1 acetylation, reverses its function into pro-apoptotic activity. Similar to miR-326 and ARRB1 overexpression, we also show that EZH2 inhibition restores miR-326/ARRB1 expression, limiting E2F1 pro-proliferative activity. Our results reveal a new regulatory molecular axis critical for MB progression.

Upfront treatment with mTOR inhibitor everolimus in pediatric low-grade gliomas: A single-center experience.

Pediatric low-grade gliomas (pLGGs) are the most frequent brain tumor in children. Adjuvant treatment, consisting in chemotherapy and radiotherapy, is often necessary if a complete surgical resection cannot be obtained. Traditional treatment approaches result in a significant long-term morbidity, with a detrimental impact on quality of life. Dysregulation of the mitogen-activated protein kinase (MAPK) pathway is the molecular hallmark of pLGGs and hyperactivation of the downstream mammalian target of rapamycin (mTOR) pathway is frequently observed. We report clinical and radiological results of front-line treatment with everolimus in 10 consecutive patients diagnosed with m-TOR positive pLGGs at the Bambino Gesù Children's Hospital in Rome, Italy. Median duration of treatment was 19 months (range from 13-60). Brain magnetic resonance imaging showed stable disease in 7 patients, partial response in 1 and disease progression in 2. Therapy-related adverse events were always reversible after dose reduction or temporary treatment interruption. To the best of our knowledge, this is the first report of everolimus treatment for chemo- and radiotherapy-naïve children with pLGG. Our results provide preliminary support, despite low sample size, for the use of everolimus as target therapy in pLGG showing lack of progression with a manageable toxicity profile.