Malignant gliomas, the most frequent primary brain tumors, are characterized by a dismal prognosis. Reliable biomarkers complementary to neuroradiology in the differential diagnosis of gliomas and monitoring for post-surgical progression are unmet needs. Altered expression of several microRNAs in tumour tissues from patients with gliomas compared to normal brain tissue have been described, thus supporting the rationale of using microRNA-based biomarkers. Although different circulating microRNAs were proposed in association with gliomas, they have not been introduced into clinical practice so far. Blood samples were collected from patients with high and low grade gliomas, both before and after surgical resection, and the expression of miR-21, miR-222 and miR-124-3p was measured in exosomes isolated from serum. The expression levels of miR-21, miR-222 and miR-124-3p in serum exosomes of patients with high grade gliomas were significantly higher than those of low grade gliomas and healthy controls and were sharply decreased in samples obtained after surgery. The analysis of miR-21, miR-222 and miR-124-3p in serum exosomes of patients affected by gliomas can provide a minimally invasive and innovative tool to help the differential diagnosis of gliomas at their onset in the brain and predict glioma grading and non glial metastases before surgery.

MicroRNAs are a family of small noncoding RNAs regulating gene expression by sequence-selective mRNA targeting, leading to a translational repression or mRNA degradation. The oncomiR miR-221 is highly expressed in human gliomas, as confirmed in this study in samples of low and high grade gliomas, as well in the cell lines U251, U373 and T98G. In order to alter the biological functions of miR-221, a peptide nucleic acid targeting miR-221 (R8-PNA-a221) was produced, bearing a oligoarginine peptide (R8) to facilitate uptake by glioma cells. The effects of R8-PNA-a221 were analyzed in U251, U373 and T98G glioma cells and found to strongly inhibit miR-221. In addition, the effects of R8-PNA-a221 on p27(Kip1) (a target of miR-221) were analyzed in U251 and T98G cells by RT-qPCR and by Western blotting. No change of p27(Kip1) mRNA content occurs in U251 cells in the presence of PNA-a221 (lacking the R8 peptide), whereas significant increase of p27(Kip1) mRNA was observed with the R8-PNA-a221. These data were confirmed by Western blot assay. A clear increment of p27(Kip1) protein expression in the samples treated with R8-PNA-a221 was detected. In addition, R8-PNA-a221 was found able to increase TIMP3 expression (another target of miR-221) in T98G cells. These results suggest that PNAs against oncomiRNA miR-221 might be proposed for experimental treatment of human gliomas.

Loss of chromosome 9p has been implicated in the progression of renal cell carcinoma. We evaluated the clinical utility of fluorescence in situ hybridization analysis of loss of chromosome 9p in 73 patients with clear cell renal cell carcinomas with varied stage, size, grade, necrosis (SSIGN) scores. Loss of chromosome 9p was observed in 13 tumors (18%). The 5-year cancer-specific survival of patients without loss of chromosome 9p was 88% and was 43% in those with loss of chromosome 9p (Po0.001). Local extension of the primary tumor according to the 2002 TNM staging system, lymph node involvement, the presence of distant metastases, and the SSIGN score were the other variables that predicted cancer-specific survival in univariate analysis. Loss of chromosome 9p was an independent prognostic factor in multivariate analysis. Our data indicate that the detection of chromosome 9p loss by fluorescence in situ hybridization analysis of clear cell renal cell carcinoma adds prognostic information beyond the pathological factors included in the current predictive models for renal cell carcinoma, such as SSIGN score.

We evaluated intratumoral heterogeneity of 30 ductal breast carcinomas with HER2/neu amplification, scored by the American Society of Clinical Oncology/ College of American Pathologists (ASCO/CAP) criteria, and 3+ immunoexpression. High-grade (ratio > or =4.0) vs low-grade amplification (ratio >2.2 to <4.0) and chromosome 17 polysomy were also evaluated. On whole tissue sections, 20 tumors (67%) showed high-grade and 10 (33%) showed low-grade HER2/ neu amplification. Of 20 tumors with high-grade amplification, 14 (70%) showed no intratumoral genotypic heterogeneity; 6 (30%) showed at least 1 core with low-grade amplification. Of 10 cases with low-grade amplification, 6 (60%) showed no intratumoral heterogeneity; 4 (40%) showed chromosome 17 polysomy without gene amplification in 2 of 3 cores per case. Of 30 cases with gene amplification, 4 (13%) showed a "not-amplified pattern" in other parts of the tumor. The routine assessment of HER2/neu amplification using the ASCO/CAP criteria on whole tissue sections is not significantly confounded by intratumoral heterogeneity in breast cancer with high-grade amplification; however, genetic heterogeneity exists in a subset of breast carcinomas with low-grade amplification. The clinical relevance and impact on treatment outcome of intratumoral heterogeneity in breast cancer with low-grade HER2/neu amplification or chromosome 17 polysomy need further investigation.

Many studies have examined the diagnostic concordance of whole slide imaging (WSI) and light microscopy (LM) for surgical pathology. In cytopathology, WSI use has been more limited, mainly because of technical issues. The aim of this study was to review the literature and determine the overall diagnostic concordance of WSI and LM in cytopathology. A systematic search of PubMed, Scopus, and the Cochrane Library was performed, with data extracted from the included articles. A quality assessment of studies was performed with a modified Quality Assessment of Diagnostic Accuracy Studies 2 tool. The primary outcome was concordance for the diagnoses rendered by WSI and LM as shown by the concordance rate with the original diagnosis, intra‐observer and interobserver concordance with the κ coefficient, or a percentage. Secondary outcomes included the time taken to reach a diagnosis and the quality and perception of WSI. A descriptive survey was provided. Among 1867 publications, a total of 19 studies (1%) were included. Overall, the concordance between WSI and the original diagnosis was 84.1%, the intra‐observer concordance between WSI and LM was 92.5% with a κ coefficient of 0.66, and the interobserver κ coefficient was 0.69. The time to reach a diagnosis was longer with WSI in all studies. The quality of WSI was good, but diagnostic confidence and cytologist preference were higher for LM. In conclusion, the concordance of WSI with LM is acceptable and in line with systematic reviews in surgical pathology. However, the time required for scanning and technical issues represent barriers to complete adoption. It is foreseeable that technical advances and rigorous validation study design will help to improve the diagnostic concordance of WSI with LM in cytopathology.

The clinical significance and management (surgical excision vs. follow-up) of the patients with the diagnosis of flat epithelial atypia (FEA) on core needle biopsy (CNB) are actually under discussion. Using standardized criteria and precise terminology, we analyzed retrospectively our CNB diagnosis of FEA, dividing patients with pure FEA as the most advanced pathologic lesion from patients with FEA associated to atypical ductal hyperplasia (FEA+ADH). Both the categories were correlated with radiologic data and findings on subsequent surgery. We evaluated 875 core needle biopsies (11-gauge stereotactic vacuum-assisted procedure), performed over a 5-year period. A CNB diagnosis of pure FEA was made in 33/875 (3.7%) cases; in other 11 (1.2%) cases we observed the coexistence of FEA and ADH. Subsequent surgical excisions were available in 20/33 pure FEA and in 10/11 FEA+ADH: of the 20 patients with pure FEA on CNB, none had either ductal carcinoma in situ or invasive carcinoma in their excisional biopsy, whereas 3/10 (30%) FEA+ADH on CNB showed, at subsequent surgery, more advanced lesions (2 ductal carcinoma in situ, 1 invasive carcinoma). Our results suggest that patients with an 11-gauge vacuum-assisted CNB diagnosis of pure FEA (especially if related to a small radiologic target, completely or almost completely removed by the needle biopsy procedure) could be spared surgical excision and managed with close radiologic follow-up.

The application of next generation sequencing (NGS) technology to cytological samples has significantly modified molecular cytopathology practice. Cytological samples represent a valid source of high‐quality DNA for NGS analysis, especially for predicting patients' response to targeted treatments and for refining the risk of malignancy in indeterminate cytological diagnoses. However, several pre‐analytical factors may influence the reliability of NGS clinical analysis. Here, we briefly review the challenges of NGS in cytology practice, focusing on those pre‐analytical factors that may negatively affect NGS success rates and routine diagnostic applications. Finally, we address the future directions of the field.

Immune evasion is a key strategy adopted by tumor cells to escape the immune system while promoting their survival and metastatic spreading. Indeed, several mechanisms have been developed by tumors to inhibit immune responses. PD-1 is a cell surface inhibitory receptor, which plays a major physiological role in the maintenance of peripheral tolerance. In pathological conditions, activation of the PD-1/PD-Ls signaling pathway may block immune cell activation, a mechanism exploited by tumor cells to evade the antitumor immune control. Targeting the PD-1/PD-L1 axis has represented a major breakthrough in cancer treatment. Indeed, the success of PD-1 blockade immunotherapies represents an unprecedented success in the treatment of different cancer types. To improve the therapeutic efficacy, a deeper understanding of the mechanisms regulating PD-1 expression and signaling in the tumor context is required. We provide an overview of the current knowledge of PD-1 expression on both tumor-infiltrating T and NK cells, summarizing the recent evidence on the stimuli regulating its expression. We also highlight perspectives and limitations of the role of PD-L1 expression as a predictive marker, discuss well-established and novel potential approaches to improve patient selection and clinical outcome and summarize current indications for anti-PD1/PD-L1 immunotherapy.