Identification of Tissue miRNA Signatures for Pancreatic Ductal Adenocarcinoma.

Pancreatic ductal adenocarcinoma (PDAC), a neoplasm of the gastrointestinal tract, is the most common pancreatic malignancy (90%) and the fourth highest cause of cancer mortality worldwide. Surgery intervention is currently the only strategy able to offer an advantage in terms of overall survival, but prognosis remains poor even for operated patients. Therefore, the development of robust biomarkers for early diagnosis and prognostic stratification in clinical practice is urgently needed. In this work, we investigated deregulated microRNAs (miRNAs) in tissues from PDAC patients with high (G3) or low (G2) histological grade and with (N+) or without (N-) lymph node metastases. miRNA expression profiling was performed by a comprehensive PCR array and subsequent validation by RT-qPCR. The results showed a significant increase in miR-1-3p, miR-31-5p, and miR-205-5p expression in G3 compared to G2 patients (** p < 0.01; *** p < 0.001; *** p < 0.001). miR-518d-3p upregulation and miR-215-5p downregulation were observed in N+ compared to N- patients. A statistical analysis performed using OncomiR program showed the significant involvement (p < 0.05) of two miRNAs (miR-31 and miR-205) in the histological grade of PDAC patients. Also, an expression analysis in PDAC patients showed that miR-31 and miR-205 had the highest expression at grade 3 compared with normal and other tumor grades. Overall, survival plots confirmed that the overexpression of miR-31 and miR-205 was significantly correlated with decreased survival in TCGA PDAC clinical samples. A KEGG pathway analysis showed that all three miRNAs are involved in the regulation of multiple pathways, including the Hippo signaling, adherens junction and microRNAs in cancer, along with several target genes. Based on in silico analysis and experimental validation, our study suggests the potential role of miR-1-3p, miR-31-5p, and miR-205-5p as useful clinical biomarkers and putative therapeutic targets in PDAC, which should be further investigated to determine the specific molecular processes affected by their aberrant expression.

Genome editing without nucleases confers proliferative advantage to edited hepatocytes and corrects Wilson disease.

Application of classic liver-directed gene replacement strategies is limited in genetic diseases characterized by liver injury due to hepatocyte proliferation, resulting in decline of therapeutic transgene expression and potential genotoxic risk. Wilson disease (WD) is a life-threatening autosomal disorder of copper homeostasis caused by pathogenic variants in copper transporter ATP7B and characterized by toxic copper accumulation, resulting in severe liver and brain diseases. Genome editing holds promise for the treatment of WD; nevertheless, to rescue copper homeostasis, ATP7B function must be restored in at least 25% of the hepatocytes, which surpasses by far genome-editing correction rates. We applied a liver-directed, nuclease-free genome editing approach, based on adeno-associated viral vector-mediated (AAV-mediated) targeted integration of a promoterless mini-ATP7B cDNA into the albumin (Alb) locus. Administration of AAV-Alb-mini-ATP7B in 2 WD mouse models resulted in extensive liver repopulation by genome-edited hepatocytes holding a proliferative advantage over nonedited ones, and ameliorated liver injury and copper metabolism. Furthermore, combination of genome editing with a copper chelator, currently used for WD treatment, achieved greater disease improvement compared with chelation therapy alone. Nuclease-free genome editing provided therapeutic efficacy and may represent a safer and longer-lasting alternative to classic gene replacement strategies for WD.

Rare causes of acute non-variceal upper gastrointestinal bleeding: A comprehensive review.

Non-variceal upper gastrointestinal bleeding (NVUGIB) is a common gastroenterological emergency associated with significant morbidity and mortality. Gastroenterologists and other involved clinicians are generally assisted by international guidelines in its management. However, NVUGIB due to peptic ulcer disease only is mainly addressed by current guidelines, with upper gastrointestinal endoscopy being recommended as the gold standard modality for both diagnosis and treatment. Conversely, the management of rare and extraordinary rare causes of NVUGIB is not covered by current guidelines. Given they are frequently life-threatening conditions, all the involved clinicians, that is emergency physicians, diagnostic and interventional radiologists, surgeons, in addition obviously to gastroenterologists, should be aware of and familiar with their management. Indeed, they typically require a prompt diagnosis and treatment, engaging a dedicated, patient-tailored, multidisciplinary team approach. The aim of our review was to extensively summarize the current evidence with regard to the management of rare and extraordinary rare causes of NVUGIB.

Recurrence-Free Survival in Early and Locally Advanced Large Cell Neuroendocrine Carcinoma of the Lung after Complete Tumor Resection.

BACKGROUND: Large Cell Neuroendocrine Carcinoma (LCNEC) is a rare subtype of lung cancer with poor clinical outcomes. Data on recurrence-free survival (RFS) in early and locally advanced pure LCNEC after complete resection (R0) are lacking. This study aims to evaluate clinical outcomes in this subgroup of patients and to identify potential prognostic markers. METHODS: Retrospective multicenter study including patients with pure LCNEC stage I-III and R0 resection. Clinicopathological characteristics, RFS, and disease-specific survival (DSS) were evaluated. Univariate and multivariate analyses were performed. RESULTS: 39 patients (M:F = 26:13), with a median age of 64 years (44-83), were included. Lobectomy (69.2%), bilobectomy (5.1%), pneumonectomy (18%), and wedge resection (7.7%) were performed mostly associated with lymphadenectomy. Adjuvant therapy included platinum-based chemotherapy and/or radiotherapy in 58.9% of cases. After a median follow-up of 44 (4-169) months, the median RFS was 39 months with 1-, 2- and 5-year RFS rates of 60.0%, 54.6%, and 44.9%, respectively. Median DSS was 72 months with a 1-, 2- and 5-year rate of 86.8, 75.9, and 57.4%, respectively. At multivariate analysis, age (cut-off 65 years old) and pN status were independent prognostic factors for both RFS (HR = 4.19, 95%CI = 1.46-12.07, p = 0.008 and HR = 13.56, 95%CI 2.45-74.89, p = 0.003, respectively) and DSS (HR = 9.30, 95%CI 2.23-38.83, p = 0.002 and HR = 11.88, 95%CI 2.28-61.84, p = 0.003, respectively). CONCLUSION: After R0 resection of LCNEC, half of the patients recurred mostly within the first two years of follow-up. Age and lymph node metastasis could help to stratify patients for adjuvant therapy.

The prognostic value of histopathology in invasive lung adenocarcinoma: a comparative review of the main proposed grading systems.

INTRODUCTION: An accurate histological evaluation of invasive lung adenocarcinoma is essential for a correct clinical and pathological definition of the tumour. Different grading systems have been proposed to predict the prognosis of invasive lung adenocarcinoma. AREAS COVERED: Invasive non mucinous lung adenocarcinoma is often morphologically heterogeneous, consisting of complex combinations of architectural patterns with different proportions. Several grading systems for non-mucinous lung adenocarcinoma have been proposed, being the main based on architectural differentiation and the predominant growth pattern. Herein we perform a thorough review of the literature using PubMed, Scopus and Web of Science and we highlight the peculiarities and the differences between the main grading systems and compare the data about their prognostic value. In addition, we carried out an evaluation of the proposed grading systems for less common histological variants of lung adenocarcinoma, such as fetal adenocarcinoma and invasive mucinous adenocarcinoma. EXPERT OPINION: The current IASLC grading system, based on the combined score of predominant growth pattern plus high-grade histological pattern, shows the stronger prognostic significance than the previous grading systems in invasive non mucinous lung adenocarcinoma.

Neutrophil-to-Lymphocyte Ratio Is a Major Prognostic Factor in Non-small Cell Lung Carcinoma Patients Undergoing First Line Immunotherapy With Pembrolizumab.

BACKGROUND/AIM: Lung cancer is one of the most common malignant neoplastic diseases and by far the leading cause of cancer death worldwide. Recently, immune checkpoint inhibitors (ICIs) have received increasing attention for playing a crucial role in non-small cell lung cancer (NSCLC). Biomarkers, such as programmed cell death-ligand 1 (PD-L1) and tumor mutational burden (TMB), seemed to be helpful in selecting patients who are more likely to benefit from ICI treatment: however, their role has not yet been fully clarified. PATIENTS AND METHODS: In this retrospective study, we evaluated the relationship between pre-treatment peripheral blood neutrophil-to-lymphocyte ratio (NLR) and survival in 252 patients suffering from advanced NSCLC who had received pembrolizumab as their first-line immunotherapy. RESULTS: Compared to their NLR low counterparts who had a median overall survival (OS) of 34.8 months, patients with NLRs above 4.8 had a median OS of 7.6 months (HR=3.26, 95%Cl=2.3-4.6, p-value<0.0000001). In multivariate Cox regression analysis, alongside other variables, such as metastatic sites, age, and sex, NLR and PD-L1 predicted progression-free survival and OS; furthermore, a very high NLR - over 10 - seemed to forecast a very dismal prognosis in patients undergoing immunotherapy, with sudden deaths in the days immediately following therapy (median OS=3.8 months). CONCLUSION: NLR acts as a valuable and reliable prognostic factor in non-small cell lung carcinoma patients undergoing first line immunotherapy with pembrolizumab. Additional investigation is necessary to fully elucidate the underlying biological rationale, which can be found in myeloid derived suppressor cells, a heterogeneous population of cells with neutrophil-like immunophenotypic features.

Full-length ATP7B reconstituted through protein trans-splicing corrects Wilson disease in mice.

Wilson disease (WD) is a genetic disorder of copper homeostasis, caused by deficiency of the copper transporter ATP7B. Gene therapy with recombinant adeno-associated vectors (AAV) holds promises for WD treatment. However, the full-length human ATP7B gene exceeds the limited AAV cargo capacity, hampering the applicability of AAV in this disease context. To overcome this limitation, we designed a dual AAV vector approach using split intein technology. Split inteins catalyze seamless ligation of two separate polypeptides in a highly specific manner. We selected a DnaE intein from Nostoc punctiforme (Npu) that recognizes a specific tripeptide in the human ATP7B coding sequence. We generated two AAVs expressing either the 5'-half of a codon-optimized human ATP7B cDNA followed by the N-terminal Npu DnaE intein or the C-terminal Npu DnaE intein followed by the 3'-half of ATP7B cDNA, under the control of a liver-specific promoter. Intravenous co-injection of the two vectors in wild-type and Atp7b -/- mice resulted in efficient reconstitution of full-length ATP7B protein in the liver. Moreover, Atp7b -/- mice treated with intein-ATP7B vectors were protected from liver damage and showed improvements in copper homeostasis. Taken together, these data demonstrate the efficacy of split intein technology to drive the reconstitution of full-length human ATP7B and to rescue copper-mediated liver damage in Atp7b -/- mice, paving the way to the development of a new gene therapy approach for WD.

Stocky/Packed Pancreas: A Case of Focal Drug-Induced Acute Pancreatitis Mimicking Cancer.

Drug-induced acute pancreatitis (DIP) is a recognised but underreported entity in the literature. Immunotherapy drugs have been described as one possible emerging cause, although the pathogenic mechanism is still largely unclear. To date, only a few cases have been reported, even if in recent times there is an over-increasing awareness of this pathologic entity. The imaging-based diagnosis of DIP can be difficult to establish, representing a real challenge for a radiologist, especially when the inflammatory disease appears as a focal mass suspicious for a malignancy. Case report: We herein report the case of a 71-year-old man with a known history of partially responsive lung adenocarcinoma subtype with high programmed cell death ligand 1 (PD-L1) expression, who underwent positron emission tomography (PET)/computed tomography (CT) imaging follow-up after one year of immunotherapy. The exam revealed a stocky/packed lesion in the pancreatic body, with increased 18F-fluorodeoxyglucose (FDG) accumulation highly suggestive of pancreatic cancer, which finally was proven to be a DIP induced by immunotherapy. Conclusion: Distinguishing between focal DIP and pancreatic neoplasm is, therefore, crucial for timely therapeutic management and prognostic stratification. A deep knowledge of possible imaging pitfalls coupled with a comprehensive clinical and laboratory assessment is pivotal to avoid any delays in diagnosis.

RNA-based next-generation sequencing in non-small-cell lung cancer in a routine setting: an experience from an Italian referral center.

Aim: ALK, ROS1, NTRK and RET gene fusions and MET exon 14 skipping alterations represent novel predictive biomarkers for advanced non-small-cell lung cancer (NSCLC). Therefore, testing patients for these genetic variants is crucial for choosing the best selective treatment. Over the last couple of decades, next-generation sequencing (NGS) platforms have emerged as an extremely useful tool for detecting these variants. Materials & methods: In the present study, we report our NGS molecular records produced during a year of diagnostic activity. Results: Overall, our in-house developed NGS workflow successfully analyzed n = 116/131 (88.5%) NSCLC samples. Of these, eight (6.8%) and five (4.3%) out of 116 patients harbored ALK and RET gene rearrangements, respectively: one case harbored ROS1 gene fusion (0.7%). Conclusion: Our results highlight that an RNA-based NGS analysis can reliably detect gene fusion alterations, thereby playing a pivotal role in the management of NSCLC patients.

Pediatric Minimal Change Disease and AKI following the Pfizer-BioNTech COVID-19 Vaccine: causal or incidental correlation?

The global coronavirus 2019 (COVID-19) pandemic required vaccination even in children to reduce infection. We report on the development of acute kidney injury (AKI) and minimal change disease (MCD) nephrotic syndrome (NS), shortly after the first injection BNT162b2 COVID-19 vaccine (Pfizer-BioNTech). A 12-year-old previously healthy boy was referred to our hospital with complaints of peripheral edema and nephrotic range proteinuria. Nine days earlier he had received his first injection BNT162b2 COVID-19 vaccine (Pfizer-BioNTech). Seven days after injection, he developed leg edema, which rapidly progressed to anasarca with significant weight gain. On admission, serum creatinine was 1.3 mg/dL and 24-hour urinary protein excretion was 4 grams with fluid overload. As kidney function continued to decline over the next days, empirical steroid treatment and renal replacement therapy with ultrafiltration were started and kidney biopsy was performed. Seven days after steroid therapy, kidney function began to improve, gradually returning to normal. The association of MCD, nephrotic syndrome and AKI hasn't been previously described following the Pfizer-BioNTech COVID-19 vaccine in pediatric population, but this triad has been reported in adults. We need further similar case reports to establish the real incidence of this possible vaccine side effect.

Are Markers of Systemic Inflammatory Response Useful in the Management of Patients With Neuroendocrine Neoplasms?

Given the increasing incidence of neuroendocrine neoplasms (NENs) over the past few decades, a more comprehensive knowledge of their pathophysiological bases and the identification of innovative NEN biomarkers represents an urgent unmet need. There is still little advance in the early diagnosis and management of these tumors, due to the lack of sensible and specific markers with prognostic value and ability to early detect the response to treatment. Chronic systemic inflammation is a predisposing factor for multiple cancer hallmarks, as cancer proliferation, progression and immune-evading. Therefore, the relevance of inflammatory biomarkers has been identified as critical in several types of tumours, including NENs. A bidirectional relationship between chronic inflammation and development of NENs has been reported. Neuroendocrine cells can be over-stimulated by chronic inflammation, leading to hyperplasia and neoplastic transformation. As the modulation of inflammatory response represents a therapeutic target, inflammatory markers could represent a promising new key tool to be applied in the diagnosis, the prediction of response to treatment and also as prognostic biomarkers in NENs field. The present review provides an overview of the pre-clinical and clinical data relating the potentially usefulness of circulating inflammatory markers: neutrophil-lymphocyte ratio (NLR), platelet-lymphocyte ratio (PLR), cytokines and tissue inflammatory markers (PD-1/PD-L1), in the management of NENs. (1) NLR and PLR have both demonstrated to be promising and simple to acquire biomarkers in patients with advanced cancer, including NEN. To date, in the context of NENs, the prognostic role of NLR and PLR has been confirmed in 15 and 4 studies, respectively. However, the threshold value, both for NLR and PLR, still remains not defined. (2) Cytokines seem to play a central role in NENs tumorigenesis. In particular, IL-8 levels seems to be a good predictive marker of response to anti-angiogenic treatments. (3) PD-1 and PD-L1 expression on tumour cells and on TILs, have demonstrated to be promising predictive and prognostic biomarkers in NENs. Unfortunately, these two markers have not been validated so far and further studies are needed to establish their indications and utility.

RNA-Based Assay for Next-Generation Sequencing of Clinically Relevant Gene Fusions in Non-Small Cell Lung Cancer.

Gene fusions represent novel predictive biomarkers for advanced non-small cell lung cancer (NSCLC). In this study, we validated a narrow NGS gene panel able to cover therapeutically-relevant gene fusions and splicing events in advanced-stage NSCLC patients. To this aim, we first assessed minimal complementary DNA (cDNA) input and the limit of detection (LoD) in different cell lines. Then, to evaluate the feasibility of applying our panel to routine clinical samples, we retrospectively selected archived lung adenocarcinoma histological and cytological (cell blocks) samples. Overall, our SiRe RNA fusion panel was able to detect all fusions and a splicing event harbored in a RNA pool diluted up to 2 ng/µL. It also successfully analyzed 46 (95.8%) out of 48 samples. Among these, 43 (93.5%) out of 46 samples reproduced the same results as those obtained with conventional techniques. Intriguingly, the three discordant results were confirmed by a CE-IVD automated real-time polymerase chain reaction (RT-PCR) analysis (Easy PGX platform, Diatech Pharmacogenetics, Jesi, Italy). Based on these findings, we conclude that our new SiRe RNA fusion panel is a valid and robust tool for the detection of clinically relevant gene fusions and splicing events in advanced NSCLC.

A case of haemoptysis and bilateral areas of lung consolidation sparing the right lower lobe.

Multiple primary lung cancers (MPLC) are often neglected. Obtaining pre-operative specimens through bronchoscopy could play a role. It is important to distinguish aerogenous metastasis from MPLC in the adenocarcinoma spectrum due to the different prognosis. https://bit.ly/3zbdVrw.

From microbiota toward gastro-enteropancreatic neuroendocrine neoplasms: Are we on the highway to hell?

Gut microbiota is represented by different microorganisms that colonize the intestinal tract, mostly the large intestine, such as bacteria, fungi, archaea and viruses. The gut microbial balance has a key role in several functions. It modulates the host's metabolism, maintains the gut barrier integrity, participates in the xenobiotics and drug metabolism, and acts as protection against gastro-intestinal pathogens through the host's immune system modulation. The impaired gut microbiota, called dysbiosis, may be the result of an imbalance in this equilibrium and is linked with different diseases, including cancer. While most of the studies have focused on the association between microbiota and gastrointestinal adenocarcinomas, very little is known about gastroenteropancreatic (GEP) neuroendocrine neoplasms (NENs). In this review, we provide an overview concerning the complex interplay between gut microbiota and GEP NENs, focusing on the potential role in tumorigenesis and progression in these tumors.

CHOP and c-JUN up-regulate the mutant Z α1-antitrypsin, exacerbating its aggregation and liver proteotoxicity.

α1-Antitrypsin (AAT) encoded by the SERPINA1 gene is an acute-phase protein synthesized in the liver and secreted into the circulation. Its primary role is to protect lung tissue by inhibiting neutrophil elastase. The Z allele of SERPINA1 encodes a mutant AAT, named ATZ, that changes the protein structure and leads to its misfolding and polymerization, which cause endoplasmic reticulum (ER) stress and liver disease through a gain-of-function toxic mechanism. Hepatic retention of ATZ results in deficiency of one of the most important circulating proteinase inhibitors and predisposes to early-onset emphysema through a loss-of-function mechanism. The pathogenetic mechanisms underlying the liver disease are not completely understood. C/EBP-homologous protein (CHOP), a transcription factor induced by ER stress, was found among the most up-regulated genes in livers of PiZ mice that express ATZ and in human livers of patients homozygous for the Z allele. Compared with controls, juvenile PiZ/Chop-/- mice showed reduced hepatic ATZ and a transcriptional response indicative of decreased ER stress by RNA-Seq analysis. Livers of PiZ/Chop-/- mice also showed reduced SERPINA1 mRNA levels. By chromatin immunoprecipitations and luciferase reporter-based transfection assays, CHOP was found to up-regulate SERPINA1 cooperating with c-JUN, which was previously shown to up-regulate SERPINA1, thus aggravating hepatic accumulation of ATZ. Increased CHOP levels were detected in diseased livers of children homozygous for the Z allele. In summary, CHOP and c-JUN up-regulate SERPINA1 transcription and play an important role in hepatic disease by increasing the burden of proteotoxic ATZ, particularly in the pediatric population.

Epithelial-Mesenchymal Transition Proteins in Neuroendocrine Neoplasms: Differential Immunohistochemical Expression in Different Sites and Correlation with Clinico-Pathological Features.

The first step leading to metastasis, or for the acquisition of local invasiveness, involves changes in the phenotype of neoplastic cells in the primary tumor. The epithelial-mesenchymal transition (EMT) is a process that determines the acquisition of a form and a transcriptional program that are characteristic of mesenchymal cells, in epithelial cells. The factors involved in this process are E-cadherin and N-cadherin adhesion proteins and some transcription factors such as Slug and Twist. EMT is a site-specific mechanism that is also active in embryogenesis-embryonic cells are affected if invested in certain points, probably due to the signals emanating from the cells or groups of surrounding cells. It is known that neuroendocrine neoplasms have a biological behavior that differs in grading, staging, and site. The aim of our study was to investigate the immunohistochemical expression of EMT factors (Twist, Slug, and E-cadherin) in the neuroendocrine neoplasms of the gastrointestinal tract, the pancreas, and lungs, in 65 cases retrieved from the archives of the Department of Pathology, of three hospitals. The immunoscores were compared in each site and correlated with the clinico-pathological parameters. Statistical evaluation revealed an association between the higher Twist immunoscore and higher grading (p value < 0.0001) and staging (p value = 0.0055). Slug was detected only in pancreatic cases where its reduced expression was associated with a higher grading (p value = 0.0033). This data could be of diagnostic utility in the case of metastases from neuroendocrine neoplasm, to define the site of the primitive tumor when the traditional immunohistochemical panel is not sufficient. In summary, our results indicated, first that the EMT is also an active process in neuroendocrine neoplasms. To the best of our knowledge, this was the first study that evaluated the expression of EMT factors in neuroendocrine neoplasms of different districts.

PD-L1 expression in cell-blocks of non-small cell lung cancer: The impact of prolonged fixation.

INTRODUCTION: In the selection of non-small cell lung cancer (NSCLC) patients for immunotherapy, specimen processed as cell blocks (CBs) may be the only available material to assess PD-L1 expression. Therefore, optimal CB preparation becomes paramount. In this context, here we assessed whether inadequate fixation time might be one of the pre-analytical factors affecting PD-L1 expression. METHODS: Ex vivo CBs from placental (n = 3) and NSCLC (n = 8) resection specimens were obtained. PD-L1 staining was performed on CBs prepared at increasing fixation times (12 hours, 48 hours, 72 hours, 96 hours, 168 hours and 504 hours) using the companion diagnostic SP263 Assay and a validated 22C3 laboratory developed test (LDT). Staining intensity and percentage of positive cells were evaluated. RESULTS: All placental CBs showed moderate to strong PD-L1 positivity in most cells, regardless of the fixation time. Likewise, the percentage of SP263-stained NSCLC cells was similar at all fixation times except for one case, which showed less intense SP263 staining at 168 hours. Conversely, in 5/8 cases, the 22C3 LDT percentage of positive cells and staining intensity decreased at 168 hours and 504 hours. CONCLUSIONS: Our results show that fixation time influences the performance of 22C3 LDT on CBs. Thus, we recommend that the fixation time of cytological materials be carefully checked, especially when PD-L1 testing is delayed until the oncology request. Indeed, delays in tissue processing and paraffin embedding may lead to sub-optimal performance of PD-L1 staining on CBs.

Rapid On-site Molecular Evaluation in thyroid cytopathology: A same-day cytological and molecular diagnosis.

BACKGROUND: Thyroid fine-needle aspirates (FNAs) with undetermined morphology can be outsourced to centralized laboratories for comprehensive molecular profiling. When a local, rapid screening rules out easily detectable BRAF and NRAS mutations outsourcing is minimized, leading to cost savings. The fully automated Idylla technology, that does not require trained staff, is an emerging option. However, Idylla platform has only been validated to process formalin fixed paraffin embedded (FFPE) sections. Here we investigate whether also the FNA needle rinse could be genotyped by the same cytopathologist who performs the FNA, a procedure that can be termed rapid on site molecular evaluation (ROME). METHODS: To validate this approach, the Idylla BRAF and NRAS Test was performed on the rinses from 25 simulated (bench-top) FNAs, in a first part of the study. Genotyping data were compared with those obtained on matched histological FFPE blocks. The second part of the study was carried out on 25 prospectively collected routine FNAs to assess the performance of the Idylla BRAF and NRAS assay against a gold standard real time polymerase chain reaction method. RESULTS: Idylla NRAS-BRAF Mutation Test was performed on needle rinse as well as histological FFPE blocks. A sensitivity of 88.9%, a specificity of 100.0% were obtained comparing the Idylla NRAS-BRAF Mutation Test on needle rinse to the reference method. CONCLUSIONS: The FNA needle rinse can be directly genotyped. This obviates the need of cell block preparation, making possible a rapid combined morphological and molecular evaluation. Since DNA extraction is no longer necessary, the cytopathologist can perform ROME him/herself.

Primary Neuroendocrine Neoplasms of the Breast: Still Open Issues.

Neuroendocrine breast tumors represent a rare subtype of breast cancer, accounting for less than 1% of all neuroendocrine neoplasms. Starting from their pathology definition, and going through their prevalence, prognosis and treatment, our knowledge is still really uncertain. In the present short review of the medical literature on this topic, we have evaluated in details their epidemiology, risk factors, pathogenesis, pathology, clinical presentation, radiographic aspects, prognosis, and therapy. We have thus been able to identify a number of open issues regarding primary neuroendocrine neoplasms of the breast that need to be clarified. Our ultimate aim was actually to try to understand whether neuroendocrine neoplasms of the breast can be considered a definite clinical entity and if neuroendocrine differentiation of breast tumors has a really clinical relevance.

Analysis of CCDC6 as a novel biomarker for the clinical use of PARP1 inhibitors in malignant pleural mesothelioma.

OBJECTIVES: CCDC6 (coiled-coil domain containing 6) is a player of the HR response to DNA damage and has been predicted to interact with BAP1, another HR-DNA repair gene highly mutated in Malignant Pleural Mesothelioma (MPM), an aggressive cancer with poor prognosis. CCDC6 levels are modulated by the deubiquitinase USP7, and CCDC6 defects have been reported in several tumors determining PARP-inhibitors sensitivity. Our aim was to investigate the functional role of CCDC6 in MPM carcinogenesis and response to PARP-inhibitors. MATERIALS AND METHODS: The interaction between CCDC6 and BAP1 was confirmed in MPM cells, by co-immunoprecipitation. Upon USP7 inhibition, that induces CCDC6 degradation, the ability to repair the DSBs and the sensitivity to PARP inhibitors, was explored by HR reporter and by cells viability assays, respectively. A TMA including 34 MPM cores was immunostained for CCDC6, USP7 and BAP1 and the results correlated by statistical analysis. RESULTS: MPM cells depleted of CCDC6 showed defects in DSBs repair and sensitivity to PARP inhibitors. The silencing of CCDC6 when combined with the overexpression of BAP1-mutant (Δ221-238) enhanced the HR-DNA repair defects and the PARP inhibitors sensitivity. In the TMA of MPM primary samples, the staining of CCDC6 and of its de-ubiquitinase USP7 showed a significant correlation in the tested primary samples (p = 0.01). CCDC6 was barely detected in 30% of the tumors that also carried BAP1 defects. CONCLUSION: The combination of CCDC6 and BAP1 staining may indicate therapeutic options for DDR targeting, acting in synergism with cisplatinum.