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The Lynch syndrome (LS) is an autosomal dominant disorder characterized by a strongly increased risk of developing colorectal cancer and several extra-colonic malignancies, such as carcinomas of the endometrium, ovary, ureter, stomach, and small intestine [1]. Lynch syndrome is caused by germline mutations in mismatch repair genes (MMR)[2], mainly in MLH1 and MSH2, rarely in MSH6 and PMS2 [3,4]. Tumors usually develop at a relatively young age (G mutation (rs267607760)in MLH1gene.

Introduction: Angiopoietin-like protein 3 (ANGPTL3) and 4 (ANGPTL4) are regulators of triglycerides storage and utilization. Bariatric surgery (BS) determines profound changes in adipose tissue composition and energy metabolism. We evaluated the impact of BS on plasma levels of ANGPTL3 and ANGPTL4.Method: Twenty-seven morbidly obese subjects with or without type 2 diabetes (T2D) underwent Roux-en-Y gastric bypass (RYGB) and 18 patients with advanced T2D received Biliopancreatic Diversion (BPD). Fasting ANGPTLs levels, insulin sensitivity (evaluated by euglycemic hyperinsulinemic clamp), total bile acids (TBA) and free fatty acids (FFA) were measured at baseline and 1year after surgery.Results: Both surgical procedures resulted in fat mass loss, improved glucose control, and a ~2-fold increase of insulin sensitivity. ANGPTL4 levels decreased significantly with both RYGB (26.6 ± 0.6 to 24.4 ± 0.3 ng/mL, p=0.001) and BPD (27.9 ± 1.5 to 24.0 ± 0.5 ng/mL, p=0.003). In contrast, ANGPTL3 concentrations did not change after RYGB but rose following BPD (225 ± 20 to 300 ± 15 ng/mL, p=0.003). By multiple regression analysis, changes in ANGPTL4 were independently associated with those of blood glucose, p=0.0169) whereas changes in ANGPTL3 after BPD were associated with variations in FFA (p=0.008) and insulin sensitivity (p=0.0427). Discussion: Circulating ANGPTL4 is reduced by BS probably due to the loss of fat mass and improved insulin sensitivity. Conversely, ANGPTL3 levels increased after BPD, presumably because of the metabolic changes induced by the malabsorptive effect of this surgical procedure.

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Context Approximately 60% of adults harbor 1 or more thyroid nodules. The possibility of cancer is the overriding concern, but only about 5% prove to be malignant. The widespread use of diagnostic imaging and improved access to health care favor the discovery of small, subclinical nodules and small papillary cancers. Overdiagnosis and overtreatment is associated with potentially excessive costs and nonnegligible morbidity for patients. Evidence Acquisition We conducted a PubMed search for the recent English-language articles dealing with thyroid nodule management. Evidence Synthesis The initial assessment includes an evaluation of clinical risk factors and sonographic examination of the neck. Sonographic risk-stratification systems (e.g., Thyroid Imaging Reporting and Data Systems) can be used to estimate the risk of malignancy and the need for biopsy based on nodule features and size. When cytology findings are indeterminate, molecular analysis of the aspirate may obviate the need for diagnostic surgery. Many nodules will not require biopsy. These nodules and those that are cytologically benign can be managed with long-term follow-up alone. If malignancy is suspected, options include surgery (increasingly less extensive), active surveillance or, in selected cases, minimally invasive techniques. Conclusion Thyroid nodule evaluation is no longer a 1-size-fits-all proposition. For most nodules, the likelihood of malignancy can be confidently estimated without resorting to cytology or molecular testing, and low-frequency surveillance is sufficient for most patients. When there are multiple options for diagnosis and/or treatment, they should be discussed with patients as frankly as possible to identify an approach that best meets their needs.

Purpose: Several studies have shown that different tumour types sharing a driver gene mutation do not respond uniformly to the same targeted agent. Our aim was to use an unbiased network-based approach to investigate this fundamental issue using BRAF mutant tumours and the BRAF inhibitor vemurafenib. Methods: We applied SWIM, a software able to identify putative regulatory (switch) genes involved in drastic changes to the cell phenotype, to gene expression profiles of different BRAF mutant cancers and their normal counterparts in order to identify the switch genes that could potentially explain the heterogeneity of these tumours' responses to vemurafenib. Results: We identified lung adenocarcinoma as the tumour with the highest number of switch genes (298) compared to its normal counterpart. By looking for switch genes encoding for kinases with homology sequences similar to known vemurafenib targets, we found that thyroid cancer and lung adenocarcinoma have a similar number of putative targetable switch gene kinases (5 and 6, respectively) whereas colorectal cancer has just one. Conclusions: We are persuaded that our network analysis may aid in the comprehension of molecular mechanisms underlying the different responses to vemurafenib in BRAF mutant tumours.

The SARS-CoV-2 pandemic is a worldwide public health emergency. Despite the beginning of a vaccination campaign, the search for new drugs to appropriately treat COVID-19 patients remains a priority. Drug repurposing represents a faster and cheaper method than de novo drug discovery. In this study, we examined three different network-based approaches to identify potentially repurposable drugs to treat COVID-19. We analyzed transcriptomic data from whole blood cells of patients with COVID-19 and 21 other related conditions, as compared with those of healthy subjects. In addition to conventionally used drugs (e.g., anticoagulants, antihistaminics, anti-TNFα antibodies, corticosteroids), unconventional candidate compounds, such as SCN5A inhibitors and drugs active in the central nervous system, were identified. Clinical judgment and validation through clinical trials are always mandatory before use of the identified drugs in a clinical setting., Graphical Abstract ga1

Angiopoietin-like proteins, namely ANGPTL3-4-8, are known as regulators of lipid metabolism. However, recent evidence points towards their involvement in the regulation of adipose tissue function. Alteration of adipose tissue functions (also called adiposopathy) is considered the main inducer of metabolic syndrome (MS) and its related complications. In this review, we intended to analyze available evidence derived from experimental and human investigations highlighting the contribution of ANGPTLs in the regulation of adipocyte metabolism, as well as their potential role in common cardiometabolic alterations associated with adiposopathy. We finally propose a model of ANGPTLs-based adipose tissue dysfunction, possibly linking abnormalities in the angiopoietins to the induction of adiposopathy and its related disorders.

Optimal management of patients with thyroid cancer requires the use of sensitive and specific biomarkers. For early diagnosis and effective follow-up, the currently available cytological and serum biomarkers, thyroglobulin and calcitonin, present severe limitations. Research on microRNA expression in thyroid tumors is providing new insights for the development of novel biomarkers that can be used to diagnose thyroid cancer and optimize its management. In this review, we will examine some of the methods commonly used to detect and quantify microRNA in biospecimens from patients with thyroid tumor, as well as the potential applications of these techniques for developing microRNA-based biomarkers for the diagnosis and prognostic evaluation of thyroid cancers.

H&uuml, rthle cell carcinomas (HCC) are rare differentiated thyroid cancers that display low avidity for radioactive iodine and respond poorly to kinase inhibitors. Here, using next-generation sequencing, we analyzed the mutational status of primary tissue and poorly differentiated metastatic tissue from two HCC patients. In both cases, metastatic tissues harbored a mutation of SETD2, each resulting in loss of the SRI and WW domains of SETD2, a methyltransferase that trimethylates H3K36 (H3K36me3) and also interacts with p53 to promote its stability. Functional studies of the novel p.D1890fs6* mutation (case 1) revealed significantly reduced H3K36me3 levels in SETD2-mutated tissue and primary cell cultures and decreased levels of the active form of p53. Restoration of SETD2-wildtype expression in the SETD2-mutant cells significantly reduced the expression of four well-known stemness markers (OCT-4, SOX2, IPF1, Goosecoid). These findings suggest potential roles for SETD2 loss-of-function mutations in HCC progression, possibly involving p53 destabilization and promotion of stemness. Their prevalence and potential treatment implications in thyroid cancer, especially HCC, require further study.

Background. Obesity has been hypothesized to contribute to the aggressiveness of thyroid cancer through the production of abnormal levels of serum adipokines. Leptin receptor (OB-R) expression has also been documented in papillary thyroid cancer (PTC). Aim. In this translational study, we analyzed in vitro the effects of leptin on the growth and migration of thyroid cancer cells (TPC-1 and K1), the molecular mechanisms underlying leptin’s action, and the influence of prolonged leptin exposure on cell response to a protein kinase inhibitor lenvatinib. The expression levels of OB-R mRNA and protein were also investigated in vivo in a series of aggressive PTCs divided into two groups based on the presence of the BRAF mutation. Results. In TPC-1 and K1 cells, prolonged treatment with leptin (500 ng/ml for 96 h) resulted in a mild increase in the proliferation (about 20% over control only in K1 cells, p<0.05) and in the migration of both cancer cell lines. Immunoblot analysis revealed a slight increase in the phosphorylation of AKT, but no effect on β-catenin and phospho-ERK expressions. The inhibitory effects of lenvatinib on the viability of both cell lines were not influenced by the leptin treatment. OB-R transcript (in fresh tissues) and proteins (in formalin-fixed and paraffin-embedded specimens) were expressed in all PTC tissues examined, with no significant differences between BRAF-mutated and BRAF-wild-type tumors. Conclusions. These results demonstrate leptin’s role in mildly increasing the aggressive phenotype of PTC cells but without influencing the action of lenvatinib. Further studies will clarify whether it is possible to target OB-R, expressed in all aggressive PTCs, as an adjuvant treatment approach for these malignancies.

Recent studies have revealed in normal thyroid tissue the presence of the transcript of several phosphodiesterases (PDEs), enzymes responsible for the hydrolysis of cyclic nucleotides. In this work, we analyzed the expression of PDE5 in a series of human papillary thyroid carcinomas (PTCs) presenting or not BRAF V600E mutation and classified according to ATA risk criteria. Furthermore, we tested the effects of two PDE5 inhibitors (sildenafil, tadalafil) against human thyroid cancer cells. PDE5 gene and protein expression were analyzed in two different cohorts of PTCs by real-time PCR using a TaqMan micro-fluid card system and Western blot. MTT and migration assay were used to evaluate the effects of PDE5 inhibitors on proliferation and migration of TPC-1, BCPAP, and 8505C cells. In a first series of 36 PTCs, we found higher expression levels of PDE5A in tumors versus non-tumor (normal) tissues. PTCs with BRAF mutation showed higher levels of mRNA compared with those without mutation. No significant differences were detected between subgroups with low and intermediate ATA risk. Upregulation of PDE5 was also detected in tumor tissue proteins. Similar results were obtained analyzing the second cohort of 50 PTCs. Moreover, all tumor tissues with high PDE5 levels showed reduction of Thyroglobulin, TSH receptor, Thyroperoxidase, and NIS transcripts. In thyroid cancer cells in vitro, sildenafil and tadalafil determined a reduction of proliferation and cellular migration. Our findings demonstrate for the first time an overexpression of PDE5 in PTCs, and the ability of PDE5 inhibitors to block the proliferation of thyroid cancer cells in culture, therefore, suggesting that specific inhibition of PDE5 may be proposed for the treatment of these tumors.

Synonymous mutations continue to be filtered out from most large-scale cancer genome studies, but several lines of evidence suggest they can play driver roles in neoplastic disease. We investigated a case of an aggressive, apparently sporadic medullary thyroid carcinoma (MTC) harboring a somatic RET p.Cys634Arg mutation (a known MTC driver). A germ-line RET substitution (p.Cys630=) had also been found but was considered clinically irrelevant because of its synonymous nature. Next generation sequencing (NGS) of the tumor tissues revealed that the RET mutations were in cis. There was no evidence of gene amplification. Expression analysis found an increase of RET transcript in p.Cys630=;p.Cys634Arg patient compared with that found in 7 MTCs harboring p.Cys634 mutations. Minigene expression assays demonstrated that the presence of the synonymous RET mutation was sufficient to explain the increased RET mRNA level. In silico analyses and RNA immunoprecipitation experiments showed that the p.Cys630 = variant created new exonic splicing enhancer motifs that enhanced SRp55 recruitment to the mutant allele, leading to more efficient maturation of its pre-mRNA and an increased abundance of mature mRNA encoding a constitutively active RET receptor. These findings document a novel mechanism by which synonymous mutations can contribute to cancer progression., Author summary Synonymous mutations—once considered “silent” because they do not alter the gene product’s amino-acid sequence—are now emerging as potential drivers of cancer. Our recent investigation of an aggressive medullary thyroid carcinoma (MTC) revealed a novel mechanism that could underlie such effects. The MTC analyzed harbored a somatic p.Cys634Arg mutation of the RET protooncogene (a well-known MTC driver). A second RET substitution (p.Cys630=) discovered in the germline had been considered clinically irrelevant because of its synonymous nature. Our next-generation sequencing analysis of the patient’s cancer tissues revealed that the RET mutations were in cis. We also found that RET mRNA levels in patient’s MTC were significantly higher than those found in seven other MTCs with various amino-acid substitutions at position 634 in RET. Subsequent experiments demonstrated that the presence of the synonymous mutation created new exonic splicing enhancer motifs in the mutant allele, which led to more efficient maturation of its transcript and increased expression of a constitutively active RET receptor.

The regulation of protein turnover by the ubiquitin proteasome system (UPS) is a major posttranslational mechanism in eukaryotes. One of the key components of the UPS, the COP9 signalosome (CSN), regulates ‘cullin–ring' E3 ubiquitin ligases. In plants, CSN participates in diverse cellular and developmental processes, ranging from light signaling to cell cycle control. In this work, we isolated a new plant-specific CSN-interacting F-box protein, which we denominated CFK1 (COP9 INTERACTING F-BOX KELCH 1). We show that, in Arabidopsis thaliana , CFK1 is a component of a functional ubiquitin ligase complex. We also show that CFK1 stability is regulated by CSN and by proteasome-dependent proteolysis, and that light induces accumulation of the CFK1 transcript in the hypocotyl. Analysis of CFK1 knockdown, mutant, and overexpressing seedlings indicates that CFK1 promotes hypocotyl elongation by increasing cell size. Reduction of CSN levels enhances the short hypocotyl phenotype of CFK1 -depleted seedlings, while complete loss of CSN activity suppresses the long-hypocotyl phenotype of CFK1-overexpressing seedlings. We propose that CFK1 (and its regulation by CSN) is a novel component of the cellular mechanisms controlling hypocotyl elongation. SUMMARY The CSN is a multiprotein complex required for proper plant development. Here, we show the isolation of a new plant-specific CSN-interacting F-box protein, denominated CFK1, that is regulated by the CSN and promotes hypocotyl elongation.

MicroRNA (miRNA) expression is dysregulated in many human malignancies, and a growing number of studies are focused on their potential use as tumor biomarkers. To identify a miRNA signature for papillary thyroid carcinomas (PTC), we investigated miRNA expression profiles in two independent cohorts of PTCs, which included major histological subtypes [classical-type (PTC‑CT), follicular-variant (PTC‑FV), and tall-cell variant (PTC‑TCV)] and cases with low or intermediate risk of recurrence. Using TaqMan® Array Human MicroRNA A+B Cards v3.0, we first performed microRNA profiling of normal and neoplastic thyroid tissues from 29 PTC patients. Promising candidates were then investigated in a second, independent cohort of 76 PTCs using Custom TaqMan® Array MicroRNA Cards. We identified a molecular signature of 11 miRNAs that were significantly upregulated (miR‑146b-5p, miR‑146b-3p, miR‑221-3p, miR‑222‑5p, miR‑222‑3p) or downregulated (miR‑1179, miR‑486‑5p, miR‑204-5p, miR‑7-2-3p, miR‑144-5p, miR‑140-3p) in PTC tissues vs. normal thyroid tissue. Upregulation of miR‑146b-5p and miR‑222‑3p was also significantly associated with an increased risk of recurrence. Higher than normal expression of miR‑146b-5p and miR‑146b-3p characterized PTC‑CT and PTC‑TCV but not PTC‑FV, whereas miR‑21-5p was significantly upregulated only in PTC‑TCV. When PTC‑FV were subclassified as encapsulated (PTC‑EFV) or infiltrative (PTC‑IFV), miR‑204-5p was downregulated in all histological subtypes except PTC‑EFV, which displayed expression levels similar to those of normal thyroid tissues. These findings provide new insights into the molecular classification of PTC, showing that different miRNA expression profiles are associated with different histological types of PTC and different risks of recurrence.

Context: Trends toward more conservative management of papillary thyroid cancer (PTC) diminish the primacy of serum thyroglobulin (Tg) assays as a posttreatment surveillance tool. Objective: To identify thyroid tumor-associated microRNAs (miRNAs) in the serum with potential for development as unique biomarkers of PTC recurrence. Methods: We measured expression of 754 miRNAs in serum samples collected from 11 patients with PTC before and 30 days after thyroidectomy. Major candidates were then re-evaluated by absolute quantitative polymerase chain reaction analysis in an independent cohort of patients with PTC (n = 44) or benign nodules and 20 healthy controls (HCs). The 2 miRNAs most significantly associated with thyroid tumors were then assessed in matched serum samples (before and 30 days and 1 to 2 years after surgery) from the 20 PTC patients with complete follow-up datasets and results correlated with American Thyroid Association (ATA) responses to therapy. Results: Eight miRNAs (miR-221-3p, miR-222-3p, miR-146a-5p, miR-24-3p, miR-146b-5p, miR-191-5p, miR-103a-3p, and miR-28-3p) displayed levels in prethyroidectomy serum samples from patients with PTC that significantly exceeded those measured after thyroidectomy and those found in samples from HCs. The 2 most promising candidates—miR-146a-5p and miR-221-3p —were further analyzed in the 20 PTC patients mentioned earlier. Serum levels of both miRNAs after 1 to 2 years of follow-up were consistent with ATA responses to therapy in all patients, including 2 with structural evidence of disease whose Tg assays remained negative (, Precis: Analysis of miRNA levels in pre- and postoperative serum samples from PTC patients reveals possible associations between postoperative changes in miR-146a-5p and miR-221-3p and clinical outcome.

Advanced medullary thyroid cancers (MTCs) are now being treated with drugs that inhibit receptor tyrosine kinases, many of which involved in angiogenesis. Response rates vary widely, and toxic effects are common, so treatment should be reserved for MTCs likely to be responsive to these drugs.RETmutations are common in MTCs, but it is unclear how they influence the microvascularization of these tumors. We examined 45 MTCs with germ-line or somaticRETmutations (RETmut group) and 34 with wild-typeRET(RETwt). Taqman Low-Density Arrays were used to assess proangiogenic gene expression. Immunohistochemistry was used to assess intratumoral, peritumoral and nontumoral expression levels of VEGFR1, R2, R3, PDGFRa, PDGFB and NOTCH3. We also assessed microvessel density (MVD) and lymphatic vessel density (LVD) based on CD31-positive and podoplanin-positive vessel counts, respectively, and vascular pericyte density based on staining for a-smooth muscle actin (a-SMA), a pericyte marker. Compared withRETwt tumors,RETmut tumors exhibited upregulated expression of proangiogenic genes (mRNA and protein), especially VEGFR1, PDGFB and NOTCH3. MVDs and LVDs were similar in the two groups. However, microvessels inRETmut tumors were more likely to be a-SMA positive, indicating enhanced coverage by pericytes, which play key roles in vessel sprouting, maturation and stabilization. These data suggest that angiogenesis inRETmut MTCs may be more intense and complete than that found inRETwt tumors, a feature that might increase their susceptibility to antiangiogenic therapy. Given their increased vascular pericyte density,RETmut MTCs might also benefit from combined or preliminary treatment with PDGF inhibitors.

In this study we analyzed the expression levels of markers of epithelial-to-mesenchymal transition (EMT) in several papillary thyroid carcinomas (PTCs) and the relation with tumor genotypes and clinicopathological characteristics. The role of fibronectin-1 (FN1) was investigated by analyzing the effects of FN1 silencing in two human thyroid cancer cell lines. Most of EMT markers were significantly over-expressed in a group of 36 PTCs. In particular, FN1 mRNA levels were higher in tumor vs non-tumor tissue (117.3, p 0.001) and also in aggressive and BRAF(V600E) samples. Similar results were observed (and confirmed at the protein level) when FN1 expression was analyzed in a validation group of 50 PTCs and six lymph node (LN) metastases. Silencing of FN1 in TPC-1 and BCPAP thyroid cancer cells significantly reduced proliferation, adhesion, migration, and invasion in both cell lines. Collectively, our data indicate that FN1 overexpression is an important determinant of thyroid cancer aggressiveness.

Abnormal expression of non-coding micro RNA (miRNA) has been described in medullary thyroid carcinoma (MTC). Expression of genes encoding factors involved in miRNA biogenesis results often deregulated in human cancer and correlates with aggressive clinical behavior. In this study, expression of four genes involved in miRNA biogenesis (DICER, DROSHA, DCGR8, and XPO5) was investigated in 54 specimens of MTC. Among them, 33 and 13 harbored RET and RAS mutations, respectively. DICER, DGCR8, and XPO5 mRNA levels were significantly overexpressed in MTC harboring RET mutations, in particular, in the presence of RET634 mutation. When MTCs with RET and RAS mutations were compared, only DGCR8 displayed a significant difference, while MTCs with RAS mutations did not show significant differences with respect to non-mutated tumors. We then attempted to correlate expression of miRNA biogenesis genes with tumor aggressiveness. According to the TNM status, MTCs were divided in two groups and compared (N0 M0 vs. N1 and/or M1): for all four genes no significant difference was detected. Cell line experiments, in which expression of a RET mutation is silenced by siRNA, suggest the existence of a causal relationship between RET mutation and overexpression of DICER, DGCR8, and XPO5 genes. These findings demonstrate that RET- but not RAS-driven tumorigenic alterations include abnormalities in the expression of some important genes involved in miRNA biogenesis that could represent new potential markers for targeted therapies in the treatment of RET-mutated MTCs aimed to restore the normal miRNA expression profile.