Small Nucleolar RNAs (snoRNAs)-Based Risk Score Classifier Predicts Overall Survival in Bladder Carcinoma.

BACKGROUND Bladder carcinoma (BLCA) is a leading cause of cancer-related deaths worldwide. The aim of this work was to develop an accurate stratification in predicting the prognosis and directing the treatment of BLCA patients based on small nucleolar RNAs (snoRNAs). MATERIAL AND METHODS Expression profiles of snoRNAs were downloaded from the SNORic database. The expression profiles and clinical outcomes of BLCA patients were analyzed. Survival-associated snoRNAs were identified and used to develop a novel risk score classifier. Genes in the whole genome that were significantly correlated with the included prognostic snoRNAs were used for functional enrichment analysis. RESULTS The results showed that age, American Joint Committee on Cancer (AJCC) stage, and tumor status were significantly correlated with overall survival (OS) of BLCA patients. We selected 12 survival-associated snoRNAs to build a prognostic signature. Patients were separated into high- and low-risk groups based on the median value of the risk score. Patients in the high-risk group and low-risk group have distinct clinical outcomes. The AJCC TNM stage showed moderate utility as a prognostic indicator for clinical outcome prediction. Then, clinical parameters and risk scores were entered in multivariate Cox analysis. Notably, the prognostic signature remained an independent significant prognostic risk factor. The pathway analysis suggested that these genes were enriched in several types of cancer and "Focal adhesion" pathways. CONCLUSIONS The prognostic signature defined by expression profiles of 12 survival-associated snoRNAs appears to be an excellent predictor of the clinical outcome of BLCA patients.

Clinical value of miR-198-5p in lung squamous cell carcinoma assessed using microarray and RT-qPCR.

BACKGROUND: To examine the clinical value of miR-198-5p in lung squamous cell carcinoma (LUSC). METHODS: Gene Expression Omnibus (GEO) microarray datasets were used to explore the miR-198-5p expression and its diagnostic value in LUSC. Real-time reverse transcription quantitative polymerase chain reaction was used to evaluate the expression of miR-198-5p in 23 formalin-fixed, paraffin-embedded (FFPE) LUSC tissues and corresponding non-cancerous tissues. The correlation between miR-198-5p expression and clinic pathological features was assessed. Meanwhile, putative target messenger RNAs of miR-198-5p were identified based on the analysis of differentially expressed genes in the Cancer Genome Atlas (TCGA) and 12 miRNA prediction tools. Subsequently, the putative target genes were sent to Gene Ontology and Kyoto Encyclopedia of Genes and Genomes pathway analyses. RESULTS: MiR-198-5p was low expressed in LUSC tissues. The combined standard mean difference (SMD) values of miR-198-5p expression based on GEO datasets were - 0.30 (95% confidence interval (CI) - 0.54, - 0.06) and - 0.39 (95% CI - 0.83, 0.05) using fixed effect model and random effect model, respectively. The sensitivity and specificity were not sufficiently high, as the area under the curve (AUC) was 0.7749 (Q* = 0.7143) based on summarized receiver operating characteristic (SROC) curves constructed using GEO datasets. Based on the in-house RT-qPCR, miR-198-5p expression was 4.3826 ± 1.7660 in LUSC tissues and 4.4522 ± 1.8263 in adjacent normal tissues (P = 0.885). The expression of miR-198-5p was significantly higher in patients with early TNM stages (I-II) than that in cases with advanced TNM stages (III-IV) (5.4400 ± 1.5277 vs 3.5690 ± 1.5228, P = 0.008). Continuous variable-based meta-analysis of GEO and PCR data displayed the SMD values of - 0.26 (95% CI - 0.48, - 0.04) and - 0.34 (95% CI - 0.71, 0.04) based on fixed and random effect models, respectively. As for the diagnostic value of miR-198-5p, the AUC based on the SROC curve using GEO and PCR data was 0.7351 (Q* = 0.6812). In total, 542 genes were identified as the targets of miR-198-5p. The most enriched Gene Ontology terms were epidermis development among biological processes, cell junction among cellular components, and protein dimerization activity among molecule functions. The pathway of non-small cell lung cancer was the most significant pathway identified using Kyoto Encyclopedia of Genes and Genomes analysis. CONCLUSION: The expression of miR-198-5p is related to the TNM stage. Thus, miR-198-5p might play an important role via its target genes in LUSC.

Generation and differentiation of IL-17-producing CD4+ T cells in malignant pleural effusion.

IL-17-producing CD4(+) T (Th17) cells have been found to be increased in some human cancers; however, the possible implication of Th17 cells in regulating antitumor responses in malignant pleural effusion (MPE) remains to be elucidated. In the current study, distribution and phenotypic features of Th17 cells in both MPE and peripheral blood from patients with lung cancer were determined by flow cytometry or double immunofluorescence staining. The impacts of cytokines on Th17 cell generation and differentiation were explored. The chemoattractant activity of chemokines CCL20 and CCL22 for Th17 cells in vitro was also observed. It was found that the increased Th17 cells could be found in MPE compared with blood. The in vitro experiments showed that IL-1ß, IL-6, IL-23, or their various combinations could promote Th17 cell generation and differentiation from naive CD4(+) T cells. MPE was chemotactic for Th17 cells, and this activity was partly blocked by anti-CCL20 and/or CCL22 Abs. Our data also showed that the accumulation of Th17 cells in MPE predicted improved patient survival. It could be concluded that the overrepresentation of Th17 cells in MPE might be due to Th17 cell differentiation and expansion stimulated by pleural proinflammatory cytokines and to recruitment of Th17 cells from peripheral blood induced by pleural chemokines CCL20 and CCL22. Furthermore, the accumulation of Th17 cells in MPE predicted improved patient survival. These data provide the basis for developing immune-boosting strategies based on ridding the cancer patient of this cell population.

Downregulation of miRNA-205 Expression and Biological Mechanism in Prostate Cancer Tumorigenesis and Bone Metastasis.

BACKGROUND: The expression and mechanism of microRNA-205 (miRNA-205) in prostate cancer (PCa) and its bone metastasis remain controversial. MATERIALS AND METHODS: The expression and discriminating capability of miRNA-205 were assessed by drawing a forest plot and a summarized receiver operating characteristic (SROC) curve, using data available from 27 miRNA-array and miRNA-sequencing datasets. The miRNA-205 target genes were acquired from online prediction tools, differentially upregulated genes in PCa, and differentially expressed genes (DEGs) after miRNA-205 transfection into PCa cell lines. Functional enrichment analysis was conducted to explore the biological mechanism of miRNA-205 targets. Immunohistochemistry (IHC) was applied to verify the protein level of the hub gene. RESULTS: The expression of miRNA-205 in the PCa group (1,461 samples) was significantly lower than that in the noncancer group (510 samples), and the downregulation of miRNA-205 showed excellent sensitivity and specificity in differentiating between the two groups. In bone metastatic PCa, the miRNA-205 level was further reduced than in nonbone metastatic PCa, and it showed a good capability in distinguishing between the two groups. In total, 153 miRNA-205 targets were screened through the three aforementioned methods. Based on the results of functional enrichment analysis, the targets of miRNA-205 were mainly enriched during chromosome segregation and phospholipid-translocating ATPase activity and in the spindle microtubule and the p53 signaling pathway. CDK1 had the highest connectivity in the PPI network analysis and was screened as one of the hub genes. A statistically significant negative correlation between miRNA-205 and CDK1 was observed. The expression of CDK1 in PCa samples was pronouncedly upregulated in terms of both the mRNA level and the protein level when compared with noncancer samples. CONCLUSION: miRNA-205 may play a vital role in PCa tumorigenesis and bone metastasis by targeting CDK1.

Development and validation of an immune prognostic classifier for clear cell renal cell carcinoma.

BACKGROUND: Tumor-infiltrating immune cells are indispensable to the progression and prognosis of clear cell renal cell carcinoma (ccRCC). OBJECTIVE: The aim of this study was to explore the clinical implications of immune cell infiltrates in ccRCC. METHODS: The Cancer Genome Atlas (TCGA) database (N= 515) and E-MTAB-1980 cohort of patients (N= 101) were adopted to estimate the prognostic value of immune cell infiltration. Twenty-four types of immune cells were evaluated using single-sample gene set enrichment analysis. Cox regression analyses were conducted to develop an immune risk score. RESULTS: Survival analyses revealed that 13 genes significantly associated with the overall survival (OS). Furthermore, multivariate Cox analysis identified an immune risk score on the basis of mast cells, natural killer CD56bright cells, T helper 17 (Th17) cells, and Th2 cells. The immune risk score was associated with OS, with hazard ratios of 2.72 (95% CI 2.17-3.40) and 3.24 (95% CI 1.64-6.44) in TCGA and E-MTAB-1980 datasets, respectively. This immune risk score was significantly correlated with some immunotherapy-related biomarkers. CONCLUSIONS: We profiled a prognostic signature and established an immune risk score model for ccRCC, which could provide novel predictive markers for patients with ccRCC and an indicator for immunotherapy response measurement.

Genome-wide Analysis of the Alternative Splicing Profiles Revealed Novel Prognostic Index for Kidney Renal Cell Clear Cell Carcinoma.

Alternative splicing (AS) is a major mechanism that greatly enhanced the diversity of proteome. Mounting evidence demonstrated that aberration of AS are important steps for the initiation and progression of human cancers. Here, we comprehensively investigated the association between whole landscape of AS profiles and the survival outcome of renal cell carcinoma (RCC) patients using RNA-seq data from TCGA SpliceSeq. Because of the limited number size of deaths in kidney chromophobe renal cell carcinoma (KICH) and papillary renal cell carcinoma (KIRP) TCGA cohorts, we only conducted survival analysis in kidney clear renal cell carcinoma (KIRC). We further constructed prognostic index (PI) based on prognosis-related AS events and built correlation network for splicing factors and prognosis-related AS events. According to the results, a total of 5351 AS events in 3522 genes were significantly correlated with the overall survival (OS) of kidney clear cell renal cell carcinoma (KIRC) patients. Seven of the PI models exhibited preferable prognosis-predicting capacity for KIRC with PI-ALL reaching the highest area under curve value of 0.875. The splicing regulatory network between splicing factors and prognosis-related AS events depicted a tangled web of relationships between them. One of the splicing factors: KHDRBS3 was validated by immunohistochemistry to be down-regulated in KIRC tissues. In conclusion, the powerful efficiency of risk stratification of PI models indicated the potential of AS signature as promising prognostic markers for KIRC and the splicing regulation network provided possible genetic mechanism of KIRC.

Expression and clinical significance of neuropilin-1 in Epstein-Barr virus-associated lymphomas.

BACKGROUND: The expression of neuropilin-1 (NRP-1) in Epstein-Barr virus (EBV)-associated lymphomas and its relationships with clinicopathological parameters was investigated. METHODS: The researchers compared 111 cases of patients with lymphoma to 20 cases of reactive lymphoid hyperplasia. In situ hybridization was applied to observe the expression of EBV-encoded RNA (EBER) in lymphomas, and immunohistochemistry was used to detect the NRP-1 expression in lymphoma tissues and lymph node tissues with reactive hyperplasia. RESULTS: In these 111 cases, the EBER of 62 cases (55.9%) appeared positive. NRP-1 was relatively highly expressed in lymphomas (P= 0.019). Further, NRP-1 showed higher expression in lymphomas with positive EBER than in negative ones. A comprehensive analysis revealed that NRP-1 was differently expressed in NK/T-cell lymphoma, Hodgkin's lymphoma, diffuse large B-cell lymphoma, and anaplastic large cell lymphoma (P= 0.027). Moreover, highly expressed NRP-1 was found to be a useful independent prognostic factor in assessing overall survival and progression-free survival rates in cases of non-Hodgkin's lymphoma (NHL). CONCLUSIONS: NRP-1 exhibited higher expression in lymphomas, and it was positively expressed in EBV-positive lymphomas. Moreover, highly expressed NRP-1 can be used as an undesirable independent prognostic factor in NHL.

Erratum: Expression of exportin-1 in diffuse large B-cell lymphoma: immunohistochemistry and TCGA analyses.

[This corrects the article on p. 5547 in vol. 11, PMID: 31949642.].

Expression levels and co­targets of miRNA­126­3p and miRNA­126­5p in lung adenocarcinoma tissues: Αn exploration with RT­qPCR, microarray and bioinformatic analyses.

Lung adenocarcinoma (LUAD) is the most common histological subtype of lung cancer. Previous studies have found that many microRNAs (miRNAs), including miRNA­126­3p, may play a critical role in the development of LUAD. However, no study of LUAD has researched the synergistic effects and co­targets of both miRNA­126­3p and miRNA­126­5p. The present study used real­time quantitative polymerase chain reaction (RT­qPCR) to explore the expression values of miRNA­126­3p and miRNA­126­5p in 101 LUAD and 101 normal lung tissues. Ten relevant microarray datasets were screened to further validate the expression levels of miRNA­126­3p and ­5p in LUAD. Twelve prediction tools were employed to obtain potential targets of miRNA­126­3p and miRNA­126­5p. The results showed that both miRNA­126­3p and ­5p were expressed significantly lower in LUAD. A significant positive correlation was also present between miRNA­126­3p and ­5p expression in LUAD. In addition, lower expression of miRNA­126­3p and ­5p was indicative of vascular invasion, lymph node metastasis (LNM), and a later tumor/node/metastasis (TNM) stage of LUAD. The authors obtained 167 targets of miRNA­126­3p and 212 targets of miRNA­126­5p; 44 targets were co­targets of both. Eight co­target genes (IGF2BP1, TRPM8, DUSP4, SOX11, PLOD2, LIN28A, LIN28B and SLC7A11) were initially identified as key genes in LUAD. The results of the present study indicated that the co­regulation of miRNA­126­3p and miRNA­126­5p plays a key role in the development of LUAD, which also suggests a fail­proof mode between miRNA­3p and miRNA­126­5p.

Identification of potential drugs for diffuse large b-cell lymphoma based on bioinformatics and Connectivity Map database.

Diffuse large B-cell lymphoma (DLBCL) is the most main subtype in non-Hodgkin lymphoma. After chemotherapy, about 30% of patients with DLBCL develop resistance and relapse. This study was to identify potential therapeutic drugs for DLBCL using the bioinformatics method. The differentially expressed genes (DEGs) between DLBCL and non-cancer samples were downloaded from the Cancer Genome Atlas (TCGA) and the Gene Expression Omnibus (GEO). Gene ontology enrichment and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analysis of DEGs were analyzed using the Database for Annotation, Visualization, and Integrated Discovery. The R software package (SubpathwayMiner) was used to perform pathway analysis on DEGs affected by drugs found in the Connectivity Map (CMap) database. Protein-protein interaction (PPI) networks of DEGs were constructed using the Search Tool for the Retrieval of Interacting Genes online database and Cytoscape software. In order to identify potential novel drugs for DLBCL, the DLBCL-related pathways and drug-affected pathways were integrated. The results showed that 1927 DEGs were identified from TCGA and GEO. We found 54 significant pathways of DLBCL using KEGG pathway analysis. By integrating pathways, we identified five overlapping pathways and 47 drugs that affected these pathways. The PPI network analysis results showed that the CDK2 is closely associated with three overlapping pathways (cell cycle, p53 signaling pathway, and small cell lung cancer). The further literature verification results showed that etoposide, rinotecan, methotrexate, resveratrol, and irinotecan have been used as classic clinical drugs for DLBCL. Anisomycin, naproxen, gossypol, vorinostat, emetine, mycophenolic acid and daunorubicin also act on DLBCL. It was found through bioinformatics analysis that paclitaxel in the drug-pathway network can be used as a potential novel drug for DLBCL.

Survival associated alternative splicing events in diffuse large B-cell lymphoma.

Growing evidence has revealed that the initiation of various malignancies is closely associated with alternative splicing (AS) events in certain key oncogenes. However, in diffuse large B-cell lymphoma (DLBCL), there is still a great deal to learn about AS variants. In this study, 33,724 AS variant profiles were obtained from 16,278 genes in 48 DLBCL cases. A total of 10 AS variants were identified as overall survival (OS)- related events via multivariate Cox regression analysis. Notably, alternative donor (AD) sites in AS events in the low-risk group showed a significantly better outcome in DLBCL patients than in the high-risk group (P=0.0002). The area under the curve (AUC) of the receiver-operator characteristic curve (ROC) for ADs in DLBCL was 0.746. Furthermore, 66 related splicing factors were obtained to investigate their potential correlations with AS events. Factors SF1, HNRNPC, HNRNPD, and HNRNPH3 were significantly involved in different OS-related AS variants. Collectively, we constructed valuable prognostic predictors for DLBCL patients and mapped novel splicing networks for further investigation of the underlying mechanisms related to AS variants in DLBCLs.

Morphological characteristics of fatal pediatric hand, foot and mouth disease: A clinicopathological study with related receptors of EV71.

OBJECTIVE: To investigate the pathological features of fatal pediatric hand foot and mouth disease (HFMD). METHODS: The histopathological features of HFMD were first summarized from literature, and then confirmed by in-house autopsies. Furthermore, immunohistochemistry was conducted to detect the distribution and expression level of two enterovirus 71 (EV71) receptors scavenger receptor class B, member 2 (SCARB2), and P-selectin glycoprotein ligand-1 (PSGL1) in the samples of autopsies. RESULTS: The main symptoms of HFMD included hand and foot rashes, as well as oral herpes. The fatal HFMD patients had typical histopathological change in the central nervous system, such as encephaledema and encephalitis. As for respiratory system, the fatal HFMD patients suffered acute pulmonary edema and congestion. SCARB2 positive signaling was distributed equally in bronchial and bronchiolar epithelial cells, alveolar epithelial cells and inflammatory cells of all HFMD patients, healthy children and adults without significant difference. PSGL-1 dispersed in bronchial and bronchiolar epithelial cells of healthy adults, but no PSGL-1 expression was detected in HFMD patients and healthy children. CONCLUSIONS: Both of the central nervous and respiratory systems may be involved in the fatal HFMD patients. The EV71 receptor PSGL-1 might play essential parts in the pathogenesis of fatal HFMD, however, the hypothesis needs to be further investigated.

Invasive cribriform carcinoma of the breast: a clinicopathological analysis of 12 cases with review of literature.

Invasive cribriform carcinoma (ICC) is a rare type of invasive breast cancer. We aim to investigate the clinicopathological features, immunophenotypes, diagnosis and differential diagnosis of ICC. Thus, clinicopathological data of 12 ICC patients were collected. All 12 cases were female, aged 38 to 75 years, with a median age of 53 years old. The maximum diameter of the tumor was 2 cm to 10 cm, in which the median tumor size was 2.54 cm in pure ICC and classical ICC. Microscopically, the cancer nests of ICC assumed an invasive, irregular island-shaped distribution, with an irregular mesh structure internally and fibrous reactions around most cancer nests. 67% (8/12) of cases were grade 1 and 33% (4/12) of cases were grade 2 tumors. Immunohistochemically, ER and PR were moderately to strongly positive with the positive tumor cell number accounting for 30% to 95% in all cases. HER-2 was negative in all cases except in one case which was positive (2+). Myoepithelial markers such as Calponin, p63, CK5/6 and CD10 were all negative in the cancer nests. 58% (7/12) of cases had a ki67 index of ≤ 14%. All follow-up patients were followed for 12 to 70 months (with a mean of 42 months), and were disease-free after treatment except for one patient whom we lost during the follow up. In conclusion, ICC, as a special type of breast cancer, has its unique clinicopathological and immunophenotypic characteristics, leading to a good prognosis.

High expression and frequently humoral immune response of melanoma-associated antigen D4 in glioma.

MAGE-D4 is a novel member of MAGE super-family. It has preliminarily been demonstrated that MAGE-D4 mRNA is not expressed in majority of normal tissues except for brain and ovary in which only trace amount of MAGE-D4 mRNA can be detected, but predominantly expressed in glioma. MAGE-D4 protein expression and its immunogenicity in glioma have not been elucidated well. This study was designed to analyze MAGE-D4 expression both at mRNA and protein level, characteristic of humoral immune response, and their relationships with glioma patients' clinicopathological parameters. Recombinant MAGE-D4 protein and antiserum were generated. Quantitative RT-PCR analysis revealed that MAGE-D4 mRNA expression was overall up-regulated in 41 glioma specimens compared with that in 14 normal brain tissues. Immunohistochemistry analysis showed that 78% (21/27) glioma tissues expressed MAGE-D4 protein, which was predominantly located in the cytoplasm of tumor cells, but absent in any neuroglia cell of normal brain tissues. ELISA analysis demonstrated that humoral response against MAGE-D4 was detected in 17% (7/41) of glioma patients' sera but not in 77 healthy donors. No apparent correlation was observed between the expression and immunogenicity of MAGE-D4 with clinicopathological parameters of glioma. In summary, these results indicate that MAGE-D4 is highly expressed in glioma and can develop specifically humoral response in glioma patients, which supports that it may be a promising biomarker for glioma diagnosis and immunotherapy.