Expression of programmed cell death ligand 1 (PD-L1) on tumor cells contributes to cancer immune evasion by interacting with programmed cell death 1 on immune cells. γ-Interferon (IFN-γ) has been reported as a key extrinsic stimulator of PD-L1 expression, yet its mechanism of expression is poorly understood. This study analyzed the role of CD74 and its ligand macrophage migration inhibitory factor (MIF) on PD-L1 expression, by immunohistochemical analysis of melanoma tissue samples and in vitro analyses of melanoma cell lines treated with IFN-γ and inhibitors of the MIF-CD74 interaction. Immunohistochemical analyses of 97 melanoma tissue samples showed significant correlations between CD74 and the expression status of PD-L1 (P < .01). In vitro analysis of 2 melanoma cell lines, which are known to secrete MIF constitutively and express cell surface CD74 following IFN-γ stimulation, showed upregulation of PD-L1 levels by IFN-γ stimulation. This was suppressed by further treatment with the MIF-CD74 interaction inhibitor, 4-iodo-6-phenylpyrimidine. In the analysis of melanoma cell line WM1361A, which constitutively expresses PD-L1, CD74, and MIF in its non-treated state, treatment with 4-iodo-6-phenylpyrimidine and transfection of siRNAs targeting MIF and CD74 significantly suppressed the expression of PD-L1. Together, the results indicated that MIF-CD74 interaction directly regulated the expression of PD-L1 and helps tumor cells escape from antitumorigenic immune responses. In conclusion, the MIF-CD74 interaction could be a therapeutic target in the treatment of melanoma patients.

OBJECTIVE: S100A9 is a calcium- and zinc-binding molecule of S100 family. The aim of the study was to evaluate the role of S100A9 in osteosarcoma (OS) and its value as a diagnostic and therapeutic target in OS.
METHODS: Quantitative real-time polymerase chain reaction (qRT-PCR), immunohistochemistry and microdissection-based mRNA analysis were used to detect S100A9 mRNA and protein expression in OS and normal bone tissues and its potential as a diagnostic marker in OS. In vitro experiments with RNA interference were performed to evaluate the functional role of S100A9 and its potential as a therapeutic target in OS.
RESULTS: S100A9 mRNA levels were significantly higher in OS tissues than that of in normal bone tissues. Receiver operating characteristic curves showed that S100A9 could be a useful diagnostic marker in OS. In vitro data showed that inhibition of S100A9 decreased the proliferation and invasiveness of OS cells, and these findings were supported by microarray data.
CONCLUSIONS: Assessment of S100A9 mRNA expression is a promising tool for the diagnosis of OS, and S100A9 may be a promising therapeutic target in OS.

BACKGROUND: This study aimed to investigate the impact of miR-451 on the biological behaviours of colon cancer cells along with its targets interactions.
METHOD: The levels of miR-451 were tested in colon cancer cell lines (SW480 and SW48). Multiple functional and immunological assays were performed to analyse miR-451 induced growth changes in-vitro and downstream effects on target proteins.
RESULTS: Overexpression of miR-451 in colon cancer cells led to reduced cell proliferation, increased apoptosis and decrease accumulation of the cells at the G0/G1 phase of the cell cycle. In addition, a significant increase in the number of the cells was noted in the G2-M phase of cell cycle. Moreover, miR-451 reduced the expression of Oct-4, Sox-2 and Snail indicating its role in stem cell and epithelial-mesenchymal transition (EMT) regulation. An inverse correlation between miR-451 and macrophage migration inhibitory protein (MIF) protein expression occurred in colon cancer cells. Furthermore, restoration the level of miR-451 in colon cancer cells inhibits tumour spheres formation.
CONCLUSION: miR-451 has tumour suppressor effects in vitro, which can inhibit the cancer-related signalling pathways in colon cancer.

BACKGROUND: Glioma stem cells (GSCs) play important roles in the tumorigenesis of glioblastoma multiforme (GBM). Using a novel cellular bioinformatics pipeline, we aimed to characterize the differences in gene-expression profiles among GSCs, U251 (glioma cell line), and a human GBM tissue sample.
MATERIALS AND METHODS: Total RNA was extracted from GSCs, U251 and GBM and microarray analysis was performed; the data were then applied to the bioinformatics pipeline consisting of a principal component analysis (PCA) with factor loadings, an intracellular pathway analysis, and an immunopathway analysis.
RESULTS: The PCA clearly distinguished the three groups. The factor loadings of the PCA suggested that v-myc avian myelocytomatosis viral oncogene neuroblastoma derived homolog (MYCN), dipeptidyl-peptidase 4 (DPP4), and macrophage migration-inhibitory factor (MIF) contribute to the stemness of GSCs. The intracellular pathway and immunopathway analyses provided relevant information about the functions of representative genes in GSCs.
CONCLUSION: The newly-developed cellular bioinformatics pipeline was a useful method to clarify the similarities and differences among samples.

Oncologic relevant members of S100 proteins are described as promising biomarkers in molecular pathology for risk estimation in oral neoplasia exhibiting different stages of malignancy: gingiva as healthy tissue, irritation fibroma as benign, leukoplakia as precancerous, and oral squamous cell carcinoma as malignant entity. Gene expression levels of S100A4 (metastasin), S100A7 (psoriasin), S100A8 (calgranulin A), and S100A9 (calgranulin B) were analyzed using quantitative RT-PCR. In addition, immunohistochemistry-based microscopy was used to examine cellular localization and distribution of these biomarkers in tissue sections. The results indicate that S100 proteins represent promising biomarkers for early-stage diagnosis in oral lesions. The inclusion of expression profiles and ratios for each entity even improves their diagnostic validity.

BACKGROUND & AIMS: Hepatocellular carcinoma (HCC) typically arises in fibrotic or cirrhotic livers, which are characterized by pathogenic angiogenesis. Myeloid immune cells, specifically tumor-associated macrophages (TAMs), may represent potential novel therapeutic targets in HCC, complementing current ablative or immune therapies. However, the detailed functions of TAM subsets in hepatocarcinogenesis have remained obscure.
METHODS: TAM subsets were analyzed in-depth in human HCC samples and a combined fibrosis-HCC mouse model, established by i.p. injection with diethylnitrosamine after birth and repetitive carbon tetrachloride (CCl
RESULTS: We show that human CCR2
CONCLUSIONS: The HCC microenvironment in human patients and mice is characterized by functionally distinct macrophage populations, of which the CCR2

OBJECTIVES: The aim of the study is to explore the relationship between CCDC69 expression and resistance of ovarian cancer cells to cisplatin and reveal the underlying mechanism.
METHODS: One hundred thirty five ovarian cancer patients with intact chemo-response information from The Cancer Genome Atlas (TCGA) database were included and analyzed. Stable CCDC69 overexpressing 293 and ovarian cancer A2780 cell lines were established and subjected to examine cell apoptosis and cell cycle distribution using CCK-8 assay and flow cytometry. Cell cycle and apoptosis pathway were evaluated by immunoblots. Stability of p14
RESULTS: We found that CCDC69 expression was significantly higher in chemo-sensitive groups compared with chemo-resistant groups from TCGA database. High CCDC69 expression was associated longer survival. CCDC69 overexpressing 293 and A2780 cells with wildtype p53 and contributes to cisplatin sensitivity following treatment with cisplatin. We further found over-expression of CCDC69 activated p14
CONCLUSIONS: It is tempting to conclude that targeting CCDC69 may play a role in cisplatin resistance.

BACKGROUND: Numerous studies have investigated the associations between RARβ2, DAPK, hMLH1, p14, and p15 promoter hypermethylation and clinical progression of patients with breast cancer, however the results remained uncertain due to the small sample size. Therefore, we performed a meta-analysis to explore the role of RARβ2, DAPK, hMLH1, p14, and p15 promoter hypermethylation in the susceptibility and clinical progression of breast cancer.
METHODS: Eligible studies were obtained by searching Medicine, Embase, Web of knowledge, and Chinese National Knowledge Infrastructure (CNKI) databases. The odds ratios (OR) and 95% confidence intervals (CI) were calculated to evaluate the associations of RARβ2, DAPK, hMLH1, p14, and p15 promoter hypermethylation with breast cancer pathogenesis. Trial sequential analysis (TSA) was applied to observe the reliability of pooled results of RARβ2 gene, and obtain a conservative required information size (RIS).
RESULTS: In primary screened 445 articles, 39 literatures with 4492 breast cancer patients were finally enrolled in the final meta-analysis. The results indicated that the frequency of RARβ2 promoter hypermethylation in case group was significantly higher than the frequency of control group (OR = 7.21, 95% CI = 1.54-33.80, P < .05). The RARβ2 promoter hypermethylation had a significant association with lymph node metastasis of breast cancer (OR = 2.13, 95% CI = 1.04-4.47, P < .05). And, the RARβ2 promoter hypermethylation was more common in the breast cancer patients of TNM III-IV stage than those patients of TNM I-II stage (OR = 1.85, 95% CI = 1.33-2.57, P < .05). In addition, the promoter hypermethylation of DAPK, hMLH1, and p14 genes were significantly associated with the susceptibility of breast cancer (for DAPK, OR = 4.93, 95% CI = 3.17-7.65; for hMLH1, OR = 1.84, 95% CI = 1.26-1.29; for p14, OR = 22.52, 95% CI = 7.00-72.41; for p15, OR = 2.13, 95% CI = 0.30-15.07).
CONCLUSIONS: Our findings revealed that the RARβ2 promoter hypermethylation significantly increased the risk of breast cancer. In the meantime, the meta-analysis demonstrated that there were significant associations of RARβ2 promoter hypermethylation with lymph node metastasis and TNM-stage of breast cancer patients. In addition, DAPK, hMLH1, and p14 genes promoter hypermethylation were significantly associated with the susceptibility of breast cancer.

PURPOSE: We investigated the effects of alterations in the biological markers p14, p53, p21, and p16 in relation to tumour cell proliferation, T-category, N- category, lymphovascular invasion, and the ability to predict prognosis in patients with muscle-invasive bladder cancer (MIBC) treated with cystectomy and, if applicable, chemotherapy.
MATERIALS AND METHODS: We prospectively studied patients with urinary bladder cancer pathological stage pT1 to pT4 treated with cystectomy, pelvic lymph node dissection and postoperative chemotherapy. Tissue microarrays from paraffin-embedded cystectomy tumour samples were examined for expression of immunostaining of p14, p53, p21, p16 and Ki-67 in relation to other clinical and pathological factors as well as cancer-specific survival.
RESULTS: The median age of the 110 patients was 70 years (range 51-87 years), and 85 (77%) were male. Pathological staging was pT1 to pT2 (organ-confined) in 28 (25%) patients and pT3 to pT4 (non-organ-confined) in 82 (75%) patients. Lymph node metastases were found in 47 patients (43%). P14 expression was more common in tumours with higher T-stages (P = 0.05). The expression of p14 in p53 negative tumours was associated with a significantly shorter survival time (P=0.003). Independently of p53 expression, p14 expression was associated with an impaired response to chemotherapy (P=0.001). The expression of p21 in p53 negative tumours was associated with significantly decrease levels of tumour cell proliferation detected as Ki-67 expression (P=0.03).
CONCLUSIONS: The simultaneous expression of the senescence markers involved in the p53-pathway shows a more relevant correlation to the pathological outcome of MIBC than each protein separately. P14 expression in tumours with non-altered (p53-) tumours is associated with poor prognosis. P14 expression is associated with impaired response to chemotherapy. P21 expression is related to decreased tumour cell proliferation.

Up to 60% of untreated atypical hyperplastic endometrium will develop into endometrial carcinoma (EC), and for those who underwent a hysterectomy a coexisting EC is found in up to 50%. Gene promoter methylation might be related to the EC development. The aim of this study is to determine changes in gene promoter profiles in normal endometrium, atypical hyperplasia (AH) and EC in relation to K-Ras mutations. A retrospective study was conducted in patients diagnosed with endometrial hyperplasia with and without subsequent EC. Promoter methylation of APC, hMLh1, O6-MGMT, P14, P16, RASSF1, RUNX3 was analysed on pre-operative biopsies, and correlated to the final histological diagnosis, and related to the presence of K-Ras mutations. In the study cohort (n=98), differences in promoter methylation were observed for hMLH1, O6-MGMT, and P16. Promoter methylation of hMLH1 and O6-MGMT gradually increased from histologically normal endometrium to AH to EC; 27.3, 36.4% and 38.0% for hMLH1 and 8.3%, 18.2% and 31.4% for O6-MGMT, respectively. P16 promoter methylation was significantly different in AH (7.7%) compared to EC (38%). K-Ras mutations were observed in 12.1% of AH, and in 19.6% of EC cases. No association of K-Ras mutation with promoter methylation of any of the tested genes was found. In conclusion, hMLH1 and O6-MGMT promoter methylation are frequently present in AH, and thus considered to be early events in the carcinogenesis of EC, whereas P16 promoter methylation was mainly present in EC, and not in precursor lesions supporting a late event in the carcinogenesis.

BACKGROUND: The S100 protein family is implicated in tumor invasion and metastasis, but its prognostic roles in gastric cancer (GC) has not been elucidated.
MATERIALS AND METHODS: In the current study, Kaplan-Meier plotter (KM plotter) database integrated the expression data and survival information of 1065 GC patients were downloaded from the Gene Expression Omnibus (GEO) (GSE22377, GSE14210 and GSE51105) that published by the three major cancer centers (Berlin, Bethesda and Melbourne). Then this database was used to explore the prognostic values of mRNA expression of each individual S100 in GC patients. We further assessed the prognostic value of S100 in different Lauren classifications, clinicopathological features and clinical treatment of gastric cancer.
RESULTS: Expression of 12 members of the S100 family correlated with overall survival (OS) for all GC patients. Increased expression of S100A3, S100A5, S100A7, S100A7A, S100A11, S100A13, S100Z and S100 G were found to be strongly associated with worse survival, while S100A8, S100A9, S100B and S100 P were correlated with better prognosis in all GC patients. Further assessment of prognostic values of S100 in gastric cancer with different clinical features indicated that different S100 members may interact with different signaling pathways and exerted different functions in gastric cancer development.
CONCLUSIONS: Although the results should be further testified in clinical studies, our findings offer new insights into the contribution of S100 members to GC progression and might promote development of S100 targeted reagents for treating GC.

BACKGROUND: Gene fusions and fusion products have been proven to be ideal biomarkers and drug targets for cancer. Even though a comprehensive study of cervical cancer has been conducted as part of the Cancer Genome Atlas (TCGA) project, few recurrent gene fusions have been found, and none above 3% of frequency.
METHODS: We believe that chimeric fusion RNAs generated by intergenic splicing represent a new repertoire of biomarkers and/or therapeutic targets. However, they would be missed when only genome sequences and fusions at DNA level are considered. We performed extensive data mining for chimeric RNAs using both our and TCGA cervical cancer RNA-Seq datasets. Multiple criteria were applied. We analyzed the landscape of chimeric RNAs at various levels, and from different angles.
FINDINGS: The chimeric RNA landscape changed as different filters were applied. 15 highly frequent (>10%) chimeric RNAs were identified. LHX6-NDUFA8 was detected exclusively in cervical cancer tissues and Pap smears, but not in normal controls. Mechanistically, it is not due to interstitial deletion, but a product of cis-splicing between adjacent genes. Silencing of another recurrent chimera, SLC2A11-MIF, resulted in cell cycle arrest and reduced cellular proliferation. This effect is unique to the chimera, and not shared by the two parental genes.
INTERPRETATION: Highly frequent chimeric RNAs are present in cervical cancers. They can be formed by intergenic splicing. Some have clear implications as potential biomarkers, or for shedding new light on the biology of the disease. FUND: Stand Up To Cancer and the National Science Foundation of China.

Multiple Myeloma (MM) is the second most common hematological malignancy with a median survival of 5-10 years. While current treatments initially cause remission, relapse almost always occurs, leading to the hypothesis that a chemotherapy-resistant cancer stem cell (CSC) remains dormant, and undergoes self-renewal and differentiation to reestablish disease. Our finding is that the mature cancer cell (CD138+, rapidly proliferating and chemosensitive) has developmental plasticity; namely, the ability to dedifferentiate back into its own chemoresistant CSC progenitor, the CD138-, quiescent pre-plasma cell. We observe multiple cycles of differentiation and dedifferentiation in the absence of niche or supportive accessory cells, suggesting that soluble cytokines secreted by the MM cells themselves are responsible for this bidirectional interconversion and that stemness and chemoresistance are dynamic characteristics that can be acquired or lost and thus may be targetable. By examining cytokine secretion of CD138- and CD138+ RPMI-8226 cells, we identified that concomitant with interconversion, Macrophage Migration Inhibitory Factor (MIF-1) is secreted. The addition of a small molecule MIF-1 inhibitor (4-IPP) or MIF-1 neutralizing antibodies to CD138+ cells accelerated dedifferentiation back into the CD138- progenitor, while addition of recombinant MIF-1 drove cells towards CD138+ differentiation. A similar increase in the CD138- population is seen when MM tumor cells isolated from primary bone marrow aspirates are cultured in the presence of 4-IPP. As the CD138+ MM cell is chemosensitive, targeting MIF-1 and/or the pathways that it regulates could be a viable way to modulate stemness and chemosensitivity, which could in turn transform the treatment of MM.

Oral squamous cell carcinoma (OSCC) is an extremely aggressive disease associated with a poor prognosis. Previous studies have established that cancer stem cells (CSCs) actively participate in OSCC development, progression and resistance to conventional treatments. Furthermore, CSCs frequently exhibit a deregulated expression of normal stem cell signalling pathways, thereby acquiring their distinctive abilities, of which self-renewal is an example. In this study, we examined the effects of GLI3 knockdown in OSCC, as well as the differentially expressed genes in CSC-like cells (CSCLCs) expressing high (CD44high) or low (CD44low) levels of CD44. The prognostic value of GLI3 in OSCC was also evaluated. The OSCC cell lines were sorted based on CD44 expression; gene expression was evaluated using a PCR array. Following this, we examined the effects of GLI3 knockdown on CD44 and ESA expression, colony and sphere formation capability, stem-related gene expression, proliferation and invasion. The overexpression of genes related to the Notch, transforming growth factor (TGF)β, FGF, Hedgehog, Wnt and pluripotency maintenance pathways was observed in the CD44high cells. GLI3 knockdown was associated with a significant decrease in different CSCLC fractions, spheres and colonies in addition to the downregulation of the CD44, Octamer-binding transcription factor 4 (OCT4; also known as POU5F1) and BMI1 genes. This downregulation was accompanied by an increase in the expression of the Involucrin (IVL) and S100A9 genes. Cellular proliferation and invasion were inhibited following GLI3 knockdown. In OSCC samples, a high GLI3 expression was associated with tumour size but not with prognosis. On the whole, the findings of this study demonstrate for the first time, at least to the best of our knowledge, that GLI3 contributes to OSCC stemness and malignant behaviour. These findings suggest the potential for the development of novel therapies, either in isolation or in combination with other drugs, based on CSCs in OSCC.

Long non-coding RNA MIF-AS1 (lncMIF-AS1) has been found to be upregulated in the tumor tissues of gastric cancer; however, its importance for the progression of gastric cancer remains unknown. Thus, the present study was designed to determine the role of the lncMIF-AS1-based signal transduction pathway in mediating the proliferation and apoptosis of gastric cancer cells. Differentially expressed lncRNAs and mRNAs were screened out using microarray analysis, based on the published data (GSE63288), and validated using quantitative RT-PCR. Target relationships between lncRNA-micro RNA (miRNA) and miRNA-mRNA were predicted by bioinformatics analysis and verified by dual-luciferase reporter assay. Protein expression of NDUFA4, COX6C and COX5B was detected by western blot. Cell proliferation, cell cycle and apoptosis were determined using colony formation assay and flow cytometry analysis. Oxidative phosphorylation in gastric cancer cells was assessed by levels of oxygen consumption and ATP synthase activity. Expression of lncMIF-AS1 and NDUFA4 were upregulated in gastric cancer tissues and cells as compared with non-cancerous gastric tissues and cells (P < .05). MiR-212-5p was identified as the most important miRNA linker between lncMIF-AS1 and NDUFA4, which was negatively regulated by lncMIF-AS1 and its depletion is the main cause of NDUFA4 overexpression (P < .01). The upregulated expression of NDUFA4 then greatly promoted the proliferation and decreased the apoptosis of gastric cancer cells through activation of the oxidative phosphorylation pathway. Taken together, the present study implies that inhibition of lncMIF-AS1/miR-212-5p/NDUFA4 signal transduction may provide a promising therapeutic target for the treatment of gastric cancer.

Macrophage migration inhibitory factor (MIF) is a pro‑inflammatory cytokine that serves important roles in cancer. MIF overexpression is frequently observed in numerous human cancer types, including pancreatic carcinoma. However, the prognostic value and function of MIF in pancreatic ductal adenocarcinoma (PDAC) have not been fully elucidated. In the present study, upregulation of MIF expression in PDAC tissue compared with adjacent normal tissue was observed. Furthermore, MIF overexpression was identified to be significantly associated with poor survival rates in patients with PDAC. Multivariate Cox regression analysis confirmed that MIF was an independent risk factor for poor survival. Functional analyses demonstrated that MIF knockdown significantly inhibited the proliferation and invasion of pancreatic cancer cells in vitro compared with control cells. IN addition, mechanistic investigations revealed that silencing MIF leads to inhibition of AKT serine/threonine kinase and extracellular‑signal‑regulated kinase activation, and suppression of cyclin D1 and matrix metalloproteinase‑2 expression, which may suppress tumor proliferation and invasion. These results highlight the importance of MIF overexpression in PDAC aggressiveness, and indicate that MIF may be a potential therapeutic target for pancreatic cancer.

Genes in the S100 family are abnormally expressed in a variety of tumor cells and are associated with clinical pathology, but their prognostic value in melanoma patients has not yet been fully elucidated. In this study, we extracted and profiled S100 family mRNA expression data and corresponding clinical data from the Gene Expression Omnibus database to analyze how expression of these genes correlates with clinical pathology. Compared with normal skin, S100A1, S100A13, and S100B were expressed at significantly higher levels in melanoma samples. S100A2, S100A7, S100A8, S100A9, S100A10, S100A11, and S100P were all highly expressed in primary melanoma samples but were expressed at low levels in metastatic melanoma, and all of these genes were strongly correlated with each other (P<0.001). We found the expression of these S100 family genes to be significantly correlated with both lymphatic and distant melanoma metastasis, as well as with American Joint Committee on Cancer grade but not with Clark's grade, age, or sex. This suggests that expression of these genes may be related to the degree of tumor invasion. Although further validation through basic and clinical trials is needed, our results suggest that the S100 family genes have the potential to play an important role in the diagnosis of melanoma. S100 expression may be related to tumor invasion and may facilitate the early diagnosis of melanoma, allowing for a more accurate prognosis. Targeted S100 therapies are also potentially viable strategies in the context of melanoma.

BACKGROUND: Squamous cell lung cancer (SqCLC) is a distinct histologic subtype of non-small cell lung cancer (NSCLC). Although the discovery of driver mutations and their targeted drugs has remarkably improved the treatment outcomes for lung adenocarcinoma, currently no such molecular target is clinically available for SqCLC. The CDKN2A locus at 9p21 encodes two alternatively spliced proteins, p16
METHODS: We investigated whether the CDK inhibitors flavopiridol and dinaciclib may exhibit antitumor activity in CDKN2A-defective SqCLC cells compared to control cells. The cytotoxic effect of the CDK inhibitors was evaluated using cell viability assays, and the induction of apoptosis was assessed using TUNEL assays and Western blot analyses. Finally, anti-tumor effects of the CDK inhibitors on xenografted cells were investigated in vivo.
RESULTS: We found that flavopiridol and dinaciclib induced cytotoxicity by enhancing apoptosis in CDKN2A-defective SqCLC cells, and that epithelial to mesenchymal transition (EMT) decreased and autophagy increased during this process. In addition, we found that autophagy had a cytoprotective role.
CONCLUSION: Our data suggest a potential role of CDK inhibitors in managing CDKN2A-defective SqCLC.

EBV causes human B-cell lymphomas and transforms B cells in vitro. EBNA3C, an EBV protein expressed in latently-infected cells, is required for EBV transformation of B cells in vitro. While EBNA3C undoubtedly plays a key role in allowing EBV to successfully infect B cells, many EBV+ lymphomas do not express this protein, suggesting that cellular mutations and/or signaling pathways may obviate the need for EBNA3C in vivo under certain conditions. EBNA3C collaborates with EBNA3A to repress expression of the CDKN2A-encoded tumor suppressors, p16 and p14, and EBNA3C-deleted EBV transforms B cells containing a p16 germline mutation in vitro. Here we have examined the phenotype of an EBNAC-deleted virus (Δ3C EBV) in a cord blood-humanized mouse model (CBH). We found that the Δ3C virus induced fewer lymphomas (occurring with a delayed onset) in comparison to the wild-type (WT) control virus, although a subset (10/26) of Δ3C-infected CBH mice eventually developed invasive diffuse large B cell lymphomas with type III latency. Both WT and Δ3C viruses induced B-cell lymphomas with restricted B-cell populations and heterogeneous T-cell infiltration. In comparison to WT-infected tumors, Δ3C-infected tumors had greatly increased p16 levels, and RNA-seq analysis revealed a decrease in E2F target gene expression. However, we found that Δ3C-infected tumors expressed c-Myc and cyclin E at similar levels compared to WT-infected tumors, allowing cells to at least partially bypass p16-mediated cell cycle inhibition. The anti-apoptotic proteins, BCL2 and IRF4, were expressed in Δ3C-infected tumors, likely helping cells avoid c-Myc-induced apoptosis. Unexpectedly, Δ3C-infected tumors had increased T-cell infiltration, increased expression of T-cell chemokines (CCL5, CCL20 and CCL22) and enhanced type I interferon response in comparison to WT tumors. Together, these results reveal that EBNA3C contributes to, but is not essential for, EBV-induced lymphomagenesis in CBH mice, and suggest potentially important immunologic roles of EBNA3C in vivo.

PURPOSE: Chronic inflammation is an important factor in the etiology of prostate cancer. Macrophage migration inhibitory factor (MIF) plays an important regulatory role in inflammatory responses. The aim of this study was to investigate the potential association between MIF-173 G/C polymorphism, and both biological behavior and incidence of prostate cancer.
MATERIALS AND METHODS: Analysis of polymorphic variants for MIF was performed using the polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) method in 128 subjects with prostate cancer and 135 controls.
RESULTS: The frequency of MIF-173 *C allele was significantly (OR = 2.18, 95% CI = 1.32-3.61) higher in patients with prostate cancer (19.5%) than in healthy individuals (10%). Prostate cancer patients with Gleason scores ? 7 had higher frequency of MIF-173 *C allele than Gleason scores < 7 (86.1% vs. 27.1%, P = 0.003, OR = 3.18, 95%CI = 1.46-6.95). The frequency of MIF-173 *C allele was significantly different in patients with T1, T2 and ?T3 clinical stages of prostate cancer (15.2% vs. 42.6% and 47.8%, P = 0.003).
CONCLUSION: Our data suggest that MIF-173 polymorphisms may be associated with a higher incidence of prostate cancer compared to controls. We believe that MIF-173 GC+CC genotype can be used as a predictive factor for aggressive behavior of prostate cancer including pathological stage and Gleason scores as well as metastatic potential.

BACKGROUND: Breast cancer is the most prevalent tumor entity in Li-Fraumeni syndrome. Up to 80% of individuals with a Li-Fraumeni-like phenotype do not harbor detectable causative germline TP53 variants. Yet, no systematic panel analyses for a wide range of cancer predisposition genes have been conducted on cohorts of women with breast cancer fulfilling Li-Fraumeni(-like) clinical diagnostic criteria.
METHODS: To specifically help explain the diagnostic gap of TP53 wild-type Li-Fraumeni(-like) breast cancer cases, we performed array-based CGH (comparative genomic hybridization) and panel-based sequencing of 94 cancer predisposition genes on 83 breast cancer patients suggestive of Li-Fraumeni syndrome who had previously had negative test results for causative BRCA1, BRCA2, and TP53 germline variants.
RESULTS: We identified 13 pathogenic or likely pathogenic germline variants in ten patients and in nine genes, including four copy number aberrations and nine single-nucleotide variants or small indels. Three patients presented as double-mutation carriers involving two different genes each. In five patients (5 of 83; 6% of cohort), we detected causative pathogenic variants in established hereditary breast cancer susceptibility genes (i.e., PALB2, CHEK2, ATM). Five further patients (5 of 83; 6% of cohort) were found to harbor pathogenic variants in genes lacking a firm association with breast cancer susceptibility to date (i.e., Fanconi pathway genes, RECQ family genes, CDKN2A/p14
CONCLUSIONS: Our study details the mutational spectrum in breast cancer patients suggestive of Li-Fraumeni syndrome and indicates the need for intensified research on monoallelic variants in Fanconi pathway and RECQ family genes. Notably, this study further reveals a large portion of still unexplained Li-Fraumeni(-like) cases, warranting comprehensive investigation of recently described candidate genes as well as noncoding regions of the TP53 gene in patients with Li-Fraumeni(-like) syndrome lacking TP53 variants in coding regions.

Long non-coding RNAs (lncRNAs) are implicated to be involved in the pathogenesis of many cancers. Herein we report on our discovery of a novel lncRNA, ZFPM2 antisense RNA 1 (ZFPM2-AS1), and its critical role in gastric carcinogenesis. ZFPM2-AS1 expression in gastric cancer specimens was analyzed using Gene Expression Omnibus data set and validated in 73 paired gastric tumor and normal adjacent gastric tissue specimens using qRT-PCR. The effect of ZFPM2-AS1 expression on proliferation and apoptosis in gastric cancer cells was assessed by altering its expression in vitro and in vivo. Mechanistic investigation was carried out using cell and molecular biological approaches. ZFPM2-AS1 expression was higher in gastric tumors than in normal gastric tissue. Also, increased ZFPM2-AS1 expression in gastric cancer specimens was associated with tumor size, depth of tumor invasion, differentiation grade, and TNM stage. High ZFPM2-AS1 expression predicted markedly reduced overall and disease-free survival in gastric cancer patients. Functional experiments demonstrated that ZFPM2-AS1 expression promoted proliferation and suppressed apoptosis of gastric cancer cells in vitro and promoted tumor growth in vivo. This effect is associated with attenuated nuclear translocation of p53. Mechanistic experiments demonstrated that tumor-activated ZFPM2-AS1 could bind to and protect the degradation of macrophage migration inhibitory factor (MIF), a potent destabilizer of p53. Knockdown of MIF expression diminished ZFPM2-AS1's impact on p53 expression in gastric cancer cells. Our findings demonstrated that ZFPM2-AS1 regulates gastric cancer progression and revealed a novel ZFPM2-AS1/MIF/p53 signaling axis, shedding light on the molecular mechanisms underlying the tumorigenicity of certain malignant gastric cells.

PURPOSE: Dysregulation of the Wnt pathway is a crucial step in the tumorigenesis of colorectal cancer (CRC). This study aimed to determine whether DNA methylation of Wnt pathway genes helps predict treatment response and survival in patients with metastatic or recurrent CRC.
MATERIALS AND METHODS: We retrospectively collected primary tumor tissues from 194 patients with metastatic or recurrent CRC. Pyrosequencing was used to examine the methylation of 10 CpG island loci in DNA extracted from formalin-fixed paraffin-embedded specimens. To elucidate the predictive role of DNA methylation markers, Kaplan-Meier survival estimation and Cox regression were performed for progression-free survival and overall survival (OS).
RESULTS: The methylation frequencies of the 10 genes analyzed (p16, p14, MINT1, MINT2, MINT31, hMLH1, DKK3, WNT5A, AXIN2, and TFAP2E) were 47.9%, 10.8%, 21.1%, 16.0%, 20.6%, 0.5%, 53.1%, 32.0%, 2.6%, and 2.1%, respectively. We divided patients into three groups based on the number of methylated genes (group 1, no methylation n=38; group 2, 1-2 methylations n=92; group 3, 3 or more methylations n=64). Among patients treated with palliative chemotherapy (n=167), median OSs of groups 1, 2, and 3 were 39.1, 39.7, and 29.1 months, respectively (log rank p=0.013). After adjustment, number of methylations was identified as an independent poor prognostic factor (0-2 methylated vs. ≥3 methylated: hazard ratio, 1.72; 95% confidence interval, 1.16-2.56, p=0.007).
CONCLUSION: This study suggests that methylation of Wnt pathway genes, in addition to known CpG island methylator phenotype markers, may help predict treatment outcome and survival in patients with CRC.

BACKGROUND: Data about the molecular pathogenesis of hepatitis C-related hepatocellular carcinoma (HCC) are still challenging.
OBJECTIVES: Therefore, we tried to investigate the epigenetic study of three nominated genes (APC, P14, and E-cadherin) in the pathogenesis of HCV-related HCC in Egyptian.
METHODS: Between March 2016 and March 2017, the DNA methylation, and quantification using (epigenetic ELISA kit) for E-cadherin, APC, and P14 genes were studied in three groups of patients: HCV related liver cirrhosis without HCC group (LC-group; n= 20), HCC on top of HCV-related cirrhosis (HCC-group; n= 20), and a third apparently healthy control group (control-group; n= 10).
RESULTS: E-cad methylation showed non-significant differences between groups. P14 methylation was occurred only in HCC-group (45%). APC methylation was the highest in HCC group (70%). Methylation level was high in HCC group in comparison to both LC and control groups (P< 0.001). DNA methylation at a cutoff point > 2.9 ng/ml predicts HCC in LC-group with 90% sensitivity and 80% specificity and at level > 2.3 ng/ml had 95% sensitivity and 90% specificity in control-group. The pooled sensitivity, specificity, positive and negative predictive values and accuracy were 90%, 60%, 69.2, 85.7 and 75% respectively.
CONCLUSION: Aberrant DNA methylation of multiple genes is associated with disease progression in HCV related cirrhosis. Moreover, early detection of promotor methylation of these may sever as good biomarker for early detection and therapeutic targets in high risk patients.

The Macrophage Migration Inhibitory Factor (MIF) is an inflammatory cytokine that is overexpressed in a number of cancer types, with increased MIF expression often correlating with tumor aggressiveness and poor patient outcomes. In this study, we aimed to better understand the link between primary tumor expression of MIF and increased tumor growth. Using the MMTV-PyMT murine model of breast cancer, we observed that elevated MIF expression promoted tumor appearance and growth. Supporting this, we confirmed our previous observation that higher MIF expression supported tumor growth in the 4T1 murine model of breast cancer. We subsequently discovered that loss of MIF expression in 4T1 cells led to decreased cell numbers and increased apoptosis in vitro under reduced serum culture conditions. We hypothesized that this increase in cell death would promote detection by the host immune system in vivo, which could explain the observed impairment in tumor growth. Supporting this, we demonstrated that loss of MIF expression in the primary tumor led to an increased abundance of intra-tumoral IFNgamma-producing CD4+ and CD8+ T cells, and that depletion of T cells from mice bearing MIF-deficient tumors restored growth to the level of MIF-expressing tumors. Furthermore, we found that MIF depletion from the tumor cells resulted in greater numbers of activated intra-tumoral dendritic cells (DCs). Lastly, we demonstrated that loss of MIF expression led to a robust induction of a specialized form of cell death, immunogenic cell death (ICD), in vitro. Together, our data suggests a model in which MIF expression in the primary tumor dampens the anti-tumor immune response, promoting tumor growth.

Cancer is world's second largest alarming disease, which involves abnormal cell growth and have potential to spread to other parts of the body. Most of the available anticancer drugs are designed to act on specific targets by altering the activity of involved transporters and genes. As cancer cells exhibit complex cellular machinery, the regeneration of cancer tissues and chemo resistance towards the therapy has been the main obstacle in cancer treatment. This fact encourages the researchers to explore the multitargeted use of existing medicines to overcome the shortcomings of chemotherapy for alternative and safer treatment strategies. Recent developments in genomics-proteomics and an understanding of the molecular pharmacology of cancer have also challenged researchers to come up with target-based drugs. The literature supports the evidence of natural compounds exhibiting antioxidant, antimitotic, anti-inflammatory, antibiotic as well as anticancer activity. In this review, we have selected marine sponges as a prolific source of bioactive compounds which can be explored for their possible use in cancer and have tried to link their role in cancer pathway. To prove this, we revisited the literature for the selection of cancer genes for the multitargeted use of existing drugs and natural products. We used Cytoscape network analysis and Search tool for retrieval of interacting genes/ proteins (STRING) to study the possible interactions to show the links between the antioxidants, antibiotics, anti-inflammatory and antimitotic agents and their targets for their possible use in cancer. We included total 78 pathways, their genes and natural compounds from the above four pharmacological classes used in cancer treatment for multitargeted approach. Based on the Cytoscape network analysis results, we shortlist 22 genes based on their average shortest path length connecting one node to all other nodes in a network. These selected genes are CDKN2A, FH, VHL, STK11, SUFU, RB1, MEN1, HRPT2, EXT1, 2, CDK4, p14, p16, TSC1, 2, AXIN2, SDBH C, D, NF1, 2, BHD, PTCH, GPC3, CYLD and WT1. The selected genes were analysed using STRING for their protein-protein interactions. Based on the above findings, we propose the selected genes to be considered as major targets and are suggested to be studied for discovering marine natural products as drug lead in cancer treatment.

PURPOSE: The purpose of this study was to identify novel plasma biomarkers for distinguishing nasopharyngeal carcinoma (NPC) patients from healthy individuals who have positive Epstein-Barr virus (EBV) viral capsid antigen (VCA-IgA).
Materials and Methods: One hundred seventy-four plasma cytokines were analyzed by a Cytokine Array in eight healthy individuals with positive EBV VCA-IgA and eight patients with NPC. Real-time polymerase chain reaction, Western blotting, enzyme-linked immunosorbent assay (ELISA), and immunohistochemistry were employed to detect the expression levels of macrophage migration inhibitory factor (MIF) and CC chemokine ligand 3 (CCL3) in NPC cell lines and tumor tissues. Plasma MIF and CCL3 were measured by ELISA in 138 NPC patients, 127 EBV VCA-IgA negative (VN) and 100 EBV VCA-IgA positive healthy donors (VP). Plasma EBV VCA-IgA was determined by immunoenzymatic techniques.
RESULTS: Thirty-four of the 174 cytokines varied significantly between the VP and NPC group. Plasma MIF and CCL3 were significantly elevated in NPC patients compared with VN and VP. Combination of MIF and CCL3 could be used for the differential diagnosis of NPC from VN cohort (area under the curve [AUC], 0.913; sensitivity, 90.00%; specificity, 80.30%), and combination of MIF, CCL3, and VCA-IgA could be used for the differential diagnosis of NPC from VP cohort (AUC, 0.920; sensitivity, 90.00%; specificity, 84.00%), from (VN+VP) cohort (AUC, 0.961; sensitivity, 90.00%; specificity, 92.00%). Overexpressions of MIF and CCL3 were observed in NPC plasma, NPC cell lines and NPC tissues.
CONCLUSION: Plasma MIF, CCL3, and VCA-IgA combination significantly improves the diagnostic specificity of NPC in high-risk individuals.

Lead is a widely existing environmental pollutant with potential carcinogenicity. To investigate the association of blood lead level (B-Pb) with potential chromosomal damage and cancer, we analyzed micronucleus (MN) frequency of peripheral blood lymphocytes (PBLs) and the methylation status of six human tumor suppressor genes (TSGs) post lead exposure. In the study, 147 lead-exposed workers were divided into two groups according to their B-Pb P

BACKGROUND: The purpose of this study was to evaluate the correlation between calcium-binding protein S100A8 and S100A9 expression in non-small cell lung cancer (NSCLC) and patients' clinical features.
METHODS: Fifty-two NSCLC patients who underwent surgery at Zhejiang Hospital from February 2014 to January 2016 were included in this study. Calcium-binding protein S100A8 and S100A9 expression patterns in cancer and para-cancer tissues were examined by immunohistochemistry assay. The correlation between calcium-binding protein S100A8 and S100A9 expression patterns and NSCLC patients' clinical characteristics, including age, gender, tumor node metastasis stage, and pathology type, were evaluated.
RESULTS: S100A8 and S100A9 were generally expressed on the cytoplasm and nucleus of NSCLC cells, mainly located in the cytoplasm, stained with brown particles, and distributed evenly. The positive expression rates of S100A8 and S100A9 in cancer tissues were 71.2% and 76.9%, respectively, which were significantly higher than in para-cancer tissues at 11.5% and 19.2%, respectively, with statistical significance (P < 0.05). S100A8 and S100A9 positive expression was associated with tumor differentiation degree (P < 0.05) but were not correlated with age, gender, smoking history, tumor diameter, pathology type, tumor node metastasis stage, or pleural effusion (P
CONCLUSION: S100A8 and S100A9 positive expression in cancer tissues was significantly higher than in para-cancer tissues and was correlated with tumor differentiation, which may be a potential marker for poor prognosis.

OBJECTIVE: To validate the gene expression profile obtained from the previous microarray analysis and to further study the biological functions of these genes in endometrial cancer. From our previous study, we identified 621 differentially expressed genes in laser-captured microdissected endometrioid endometrial cancer as compared to normal endometrial cells. Among these genes, 146 were significantly up-regulated in endometrial cancer.
MATERIALS AND METHODS: A total of 20 genes were selected from the list of up-regulated genes for the validation assay. The qPCR confirmed that 19 out of the 20 genes were up-regulated in endometrial cancer compared with normal endometrium. RNA interference (RNAi) was used to knockdown the expression of the upregulated genes in ECC-1 and HEC-1A endometrial cancer cell lines and its effect on proliferation, migration and invasion were examined.
RESULTS: Knockdown of MIF, SOD2, HIF1A and SLC7A5 by RNAi significantly decreased the proliferation of ECC-1 cells (p < 0.05). Our results also showed that the knockdown of MIF, SOD2 and SLC7A5 by RNAi significantly decreased the proliferation and migration abilities of HEC-1A cells (p < 0.05). Moreover, the knockdown of SLC38A1 and HIF1A by RNAi resulted in a significant decrease in the proliferation of HEC1A cells (p < 0.05).
CONCLUSION: We have identified the biological roles of SLC38A1, MIF, SOD2, HIF1A and SLC7A5 in endometrial cancer, which opens up the possibility of using the RNAi silencing approach to design therapeutic strategies for treatment of endometrial cancer.