An insight into the potential role of LINC00968 in luminal breast cancer: Case-control study and bioinformatics analysis.

Background: Luminal A and B subtypes of breast cancer (BC) comprises up to 70% of all BC patients. LncRNAs can affect many biological and pathological processes, and dysregulation of them is related to human cancers. The potential role of lncRNA LINC00968 in luminal BC is still unclear. Materials and methods: We analyzed the LINC00968 expression across 44 paired luminal BC tissues from the TCGA-BRCA RNA sequencing dataset. Besides, we used the GEPIA2 web server and GENEVESTIGATOR software, as well. Real-Time Quantitative Reverse Transcription PCR (qRT-PCR) assay was performed to confirm the LINC00968 expression in 71 paired luminal BC tissues and two luminal A cell lines (MCF7 and T47D). Moreover, to better understanding the potential role of LINC00968 in luminal BC, computational data analyses including co-expression analysis, functional annotation analysis, and genetic alteration analysis have been done. Results: The results of data analyses retrieved from BRCA dataset and databases revealed the significant downregulation of LINC00968 in luminal A and B BC. Also, the results of qRT-PCR in luminal BC tissues and cell lines confirmed the earlier data. LINC00968 expression was negatively associated with tumor stage and lymph node metastasis. Additionally, functional annotation analyses revealed that LINC00968 might be involved in vascular development and angiogenesis, extracellular matrix organization, and cell motility and migration. LINC00968 might play role in some cancer-related signaling pathways. Conclusion: Our study found that downregulation of LINC00968 might promote tumorigenesis, invasion, and metastasis of luminal BC.

A Proposed TUSC7/miR-211/Nurr1 ceRNET Might Potentially be Disturbed by a cer-SNP rs2615499 in Breast Cancer.

Evidence and in silico analyses showed that TUSC7, miR-211, and Nurr1 may be involved in BC pathogenesis by ceRNET signaling axis. This study aimed to investigate the potential role of TUSC7/miR-211/Nurr1 ceRNET and rs2615499 variant as a novel cer-SNP in BC subjects. The expression assays were conducted by qPCR on tumor tissues (n = 50), tumor-adjacent normal tissues (TANTs) (n = 50), and clinically healthy control tissues (n = 50). The expression of TUSC7 and Nurr1 significantly decreased, but the level of miR-211 significantly increased in tumor tissues compared to TANTs and healthy normal tissues. Altered expression of TUSC7 and miR-211 was associated with poor prognosis of patients. The Nurr1 exhibited a double-edged sword-like activity in BC. In addition, TUSC7, Nurr1, and miR-211 expressions were significantly related to a novel BC-associated rs2615499 (A > C) located in the miR-211 binding site on Nurr1 3'-UTR. In the second part of the study, a case-control study was performed on BC patients (n = 100) and matched healthy controls (n = 100). The genomic DNA was isolated and genotyping was performed using Tetra-Primer ARMS PCR. The CC and AC genotypes were associated with higher expression levels of Nurr1 and worse outcomes of the disease. Our findings revealed that TUSC7 functions as a tumor suppressor in BC potentially via miR-211/Nurr1, which might be disturbed by the cer-SNP rs2615499. However, functional studies are needed to validate these results.

Dysregulated Expression of miR-146a and Its Associated Immune Effectors in Peripheral Blood Mononuclear Cells of Esophageal Carcinoma Patients.

Esophageal cancer is one of the least studied aggressive tumors, with the squamous cell carcinoma (ESCC) being the most frequent histological type around the world. Growing evidence has shown that the abnormal expression of microRNAs (miRNAs) in peripheral blood mononuclear cells (PBMCs) is closely related to the pathogenesis of cancers. MiR-146a is a crucial regulator of inflammatory cascades. There is currently no data available regarding the possible role of miR-146a in PBMCs of ESCC patients. We evaluated the expression of miR-146a, as well as its target genes (IRAK1 and TRAF6) and its associated immune effectors (NF-κB1, IL1B, and IL6) in PBMCs of 40 ESCC patients and 50 control subjects. The geometric mean expression of five transcripts was used for normalizing expressions. The PBMC level of miR-146a, as measured by RT-qPCR, was upregulated, whereas levels of its target genes, IRAK1 and TRAF6, were downregulated in ESCC patients. NF-κB1 and IL6 was downregulated in PBMCs of ESCC patients. There was no difference in terms of the IL1B level between patients and the control group. Logistic regression and receiver operating characteristic curve analysis suggested that a model with PBMC levels of either NF-κB1+ IL6 or NF-κB1+ miR-146a as predictors may discriminate ESCC patients from subjects of the control group. Our findings, in the context of the current literature, may suggest a possible downregulatory mechanism of immune responses in PBMCs of ESCC patients.

Expression profiles and functional prediction of long non-coding RNAs LINC01133, ZEB1-AS1 and ABHD11-AS1 in the luminal subtype of breast cancer.

BACKGROUND: Luminal breast cancer (BC) is the most frequent subtype accounting for more than 70% of BC. LncRNAs, a class of non-coding RNAs with more than 200 nucleotides, are involved in a variety of cellular processes and biological functions. Abberant expression is related to the development of various cancers, such as breast cancer. LINC01133, ZEB1-AS1, and ABHD11-AS1 were reported to be dysregulated in different cancers. However, their expression level in luminal BC remains poorly known. The aim of the present study was to evaluate the potential roles of these lncRNAs in BC, especially in luminal subtypes. METHODS: A comprehensive analysis was performed using the Lnc2Cancer database to identify novel cancer-associated lncRNA candidates. After conducting a literature review, three novel lncRNAs named LINC01133, ZEB1-AS1, and ABHD11-AS1 were chosen as target genes of the present study. Quantitative real-time polymerase chain reaction (qRT-PCR) was used to evaluate the expression level of the mentioned lncRNAs in both luminal BC tissues and cell lines. Then, the correlation of the three mentioned lncRNAs expression with clinicopathological characteristics of the patients was studied. Moreover, several datasets were used to discover the potential roles and functions of LINC01133, ZEB1-AS1 and ABHD11-AS1 in luminal subtype of BC. RESULTS: According to the qRT-PCR assay, the expression levels of LINC01133 and ZEB1-AS1 were decreased in luminal BC tissues and cell lines. On the other hand, ABHD11-AS1 was upregulated in the above-mentioned samples. The expression levels of LINC01133, ZEB1-AS1, and ABHD11-AS1 were not associated with any of the clinical features. Also, the results obtained from the bioinformatics analyses were consistent with qRT-PCR data. Functional annotation of the co-expressed genes with the target lncRNAs, protein-protein interactions and significantly enriched Kyoto Encyclopedia of Genes and Genomes (KEGG) pathways across luminal BC were also obtained using bioinformatics analysis. CONCLUSIONS: Taken together, our findings disclosed the dysregulation of LINC01133, ZEB1-AS1, and ABHD11-AS1 in luminal BC. It was revealed that LINC01133 and ZEB1-AS1 expression was significantly downregulated in luminal BC tissues and cell lines, while ABHD11-AS1 was upregulated considerably in the mentioned tissues and cell lines. Also, bioinformatics and systems biology analyses have helped to identify the possible role of these lncRNAs in luminal BC. However, further analysis is needed to confirm the current findings.

Young Breast Cancer: Novel Gene Methylation in WBC.

INTRODUCTION: Breast cancer is a highly diverse disease, and epigenomic alterations, as principle changes in the pathogenesis of breast cancer, have recently been noticed in epimarker research on peripheral blood. METHODS: In this study, DNA samples isolated from the white blood cells of 30 breast cancer patients were compared to 30 healthy controls using methylated DNA immunoprecipitation microarray (MeDIP-chip) to determine differentially methylated region as a potential epimarker in cancer and control cases. RESULTS: A total of 1799 differentially methylated regions were identified, including ZNF154, BCL9, and HOXD9, in which significant methylation differences were confirmed in breast cancer patients through a quantitative real-time polymerase chain reaction. Differential methylation of the mentioned genes has been reported in different cancer tissues and cell-free DNA, including breast cancer. Methylation of those genes listed in the white blood cells of our young patients not only relates to their importance in the pathogenesis of breast cancer but may also highlight their potential as primary epimarkers that can warrant further evaluation in large cohort studies. It is important to note that methylation alteration in WBC, as well as genetic mutation, can be identified years before cancer development, which emphasizes this issue as a potential screening marker.

Circular RNA hsa_circ_0044234 as distinct molecular signature of triple negative breast cancer: a potential regulator of GATA3.

BACKGROUND: Circular RNAs (circRNAs) have been implicated in the initiation and development of breast cancer as functional non-coding RNAs (ncRNA). The roles of circRNAs as the competing endogenous RNAs (ceRNAs) to sponge microRNAs (miRNAs) have also been indicated. However, the functions of circRNAs in breast cancer have not been totally elucidated. This study aimed to explore the clinical implications and possible roles of circ_0044234 in carcinogenesis of the most problematic BC subtype, triple negative breast cancer (TNBC), which are in desperate need of biomarkers and targeted therapies. METHODS: The importance of circ_0044234 as one of the most dysregulated circRNAs in TNBC was discovered through microarray expression profile analysis. Reverse transcription-quantitative polymerase chain reaction (RT-qPCR) was performed to confirm the downregulation of circ_0044234 in triple negative tumors and cell lines versus non-triple negative ones. The bioinformatics prediction revealed that circ_0044234 could act as an upstream sponge in the miR-135b/GATA3 axis, two of the most dysregulated transcripts in TNBC. RESULTS: Our experimental investigation of circ_0044234 expressions in various BC subtypes as well as cell lines reveals that TNBC expresses circ_0044234 at a substantially lower level than non-TNBC. The ROC curve analysis indicates that it could be applied as a discriminative biomarker to identify TNBC from other BC subtypes. Moreover, circ_0044234 expression could be an independent prognostic biomarker in BC. Interestingly, a substantial inverse expression correlation was detected between circ_0044234 and miR-135b-5p as well as between miR-135b-5p and GATA3 in breast tumors. CONCLUSIONS: The possible clinical usefulness of circ_0044234 as a promising distinct biomarker and upcoming therapeutic target for TNBC have been indicated in this research. Our comprehensive approach revealed the potential circ_0044234/miR135b-5p/GATA3 ceRNA axis in TNBC.

Investigation of circRNA-miRNA-mRNA network in colorectal cancer using an integrative bioinformatics approach.

AIM: The aim of this study was to integrate both coding and non-coding available microarray data in the development of colorectal cancer (CRC) with bioinformatics analyses to attain a more inclusive pathobiologic map of their molecular interactions and functions. BACKGROUND: Identification of competing endogenous RNAs (ceRNAs), especially circRNAs, has become a new hotspot in cancer research, although their roles and underlying mechanisms in CRC development remain mostly unknown. METHODS: Microarray data was retrieved from the Gene Expression Omnibus (GEO) database and analyzed. Several bioinformatics tools and databases were applied for further elucidation. Principal component analysis (PCA) was run separately for four datasets. The dysregulated circRNA-miRNA-mRNA, co-expression, and protein-protein interaction (PPI) networks were established. RESULTS: PCA discloses colorectal tumors; normal tissue can be distinguished not only by mRNAs expression profile, but also by both circRNA and miRNA expression profiles. In this study, 14 DE mRNAs, 85 DE miRNAs, and 36 DE circRNAs were identified in CRC tissue and compared with normal tissue. Taking their potential interactions into account, a circRNA-miRNA-mRNA network was constructed. The results disclosed some DE circRNAs with potential oncogenic (circ_0014879) or tumor suppressive (circ_0001666 and circ_0000977) effects. Finally, the PPI network suggests pivotal roles for DOCK2 and PTPRC dysregulation in the progression of CRC, possibly by facilitating tumor escape from immune surveillance. CONCLUSION: The current study proposes a novel regulatory network consisting of DE circRNAs, miRNAs, and mRNAs in CRC development that highlights the roles of DE circRNAs at the upstream of oncotranscriptomic cascade in CRC development, suggesting their potential to be utilized as both prognostic and therapeutic biomarkers.

Bi-functionalized aminoguanidine-PEGylated periodic mesoporous organosilica nanoparticles: a promising nanocarrier for delivery of Cas9-sgRNA ribonucleoproteine.

BACKGROUND: There is a great interest in the efficient intracellular delivery of Cas9-sgRNA ribonucleoprotein complex (RNP) and its possible applications for in vivo CRISPR-based gene editing. In this study, a nanoporous mediated gene-editing approach has been successfully performed using a bi-functionalized aminoguanidine-PEGylated periodic mesoporous organosilica (PMO) nanoparticles (RNP@AGu@PEG1500-PMO) as a potent and biocompatible nanocarrier for RNP delivery. RESULTS: The bi-functionalized MSN-based nanomaterials have been fully characterized using electron microscopy (TEM and SEM), nitrogen adsorption measurements, thermogravimetric analysis (TGA), X-ray powder diffraction (XRD), Attenuated Total Reflectance-Fourier Transform Infrared Spectroscopy (ATR-FTIR), and dynamic light scattering (DLS). The results confirm that AGu@PEG1500-PMO can be applied for gene-editing with an efficiency of about 40% as measured by GFP gene knockdown of HT1080-GFP cells with no notable change in the morphology of the cells. CONCLUSIONS: Due to the high stability and biocompatibility, simple synthesis, and cost-effectiveness, the developed bi-functionalized PMO-based nano-network introduces a tailored nanocarrier that has remarkable potential as a promising trajectory for biomedical and RNP delivery applications.

A methylation signature at the CpG island promoter of estrogen receptor beta (ER-ß) in breasts of women may be an early footmark of lack of breastfeeding and nulliparity.

Although little is known regarding the mechanisms behind the onset of breast cancer (BC) through reproductive risk factors, new researches have highlighted some early tumor-related methylation footmarks in the breast tissue of apparently clinically healthy women as their potential epigenetic mechanism. Previous evidence supports that the estrogen receptor beta (ER-ß), whose anti-cancer roles had already been revealed in BC, is downregulated in the breasts of healthy nulliparous women. Nevertheless, data on such a link about its methylation alterations have not been reported. The goal of current study was to determine possible methylation alterations at CpG island promoter of the ER-ß gene, including promoter 0 N and exon 0 N, in relation to aspects of reproductive history in the healthy breasts. The DNA was extracted from the breasts of 120 subjects undergoing cosmetic mammoplasty. Thereafter, the methylation levels of targeted regions in ER-ß gene were determined by using MeDIP-qPCR assay. The results revealed that ER-ß exon 0 N had no methylation in 84.2 % of the women, whereas the rest, comprising 2.5 % and 13.3 % of the samples, showed a lower and higher of its methylation, respectively. Interestingly, nulliparous women were found to have an elevated methylation level of the ER-ß exon 0 N than parous women (P = 0.036). Moreover, we observed a high methylation of the ER-ß exon 0 N in the breasts of non-breastfeeding women compared to breastfeeding subgroup (P = 0.048). Likewise, the non-breastfeeding subgroup showed exon 0N high methylation in comparison to women with breastfeeding >24 months (P = 0.023). Finally, although we found that 6.67 % of the samples had a high methylation level at the promoter 0N, no any relationship was found between its methylation and reproductive history. These results may provide key clues to revealing the epigenetic mechanism through which the nulliparity and lack of breastfeeding influencing the risk factor of BC as well as introducing the potential new early prediction and prevention strategies. Although further investigations need to be done in order to gain a better understanding the roles of these epigenetic signatures.

Evaluation of the potential role of long non-coding RNA LINC00961 in luminal breast cancer: a case-control and systems biology study.

BACKGROUND: Luminal subtype is the most common subgroup of breast cancer (BC), accounting for more than 70% of this cancer. Long non-coding RNAs (lncRNAs) are a group of RNAs which play critical roles in diverse cellular processes. It is proved that dysregulation of them can contribute to the development of various cancers, including BC. LINC00961 was reported to be downregulated in several cancers, however, its expression level in BC remains largely unknown. The purpose of the present study was to investigate the possible role of LINC00961 in luminal A and B subtypes of BC. METHODS: To obtain novel lncRNAs associated with different cancers and differentially expressed lncRNAs (DElncRNAs) between BC tumor and normal tissues, Lnc2Cancer and GDC databases were used, respectively. After performing literature review, the expression level of the selected lncRNA (LINC00961) was evaluated in 79 luminal A and B BC specimens and adjacent non-cancerous tissues by Quantitative Reverse Transcription PCR (qRT-PCR). LINC00961 expression was also evaluated in two luminal A BC cell lines, compared to a normal breast cell line. The comparison of the differences between tumor and adjacent non-tumor samples was performed by paired sample t-test. Moreover, correlation analysis between LINC00961 expression and clinicopathological features was performed using the chi-square, fisher exact, and independent t-test. In order to investigate the possible roles of LINC00961 in luminal A and B BC, different bioinformatics analyses such as functional annotation of the LINC00961 co-expressed genes and protein-protein interaction (PPI) networks construction were also performed. RESULTS: LINC00961 was selected as a significant DElncRNA which had not been studied in BC. According to q-RT PCR assay, LINC00961 was downregulated in luminal BC tissues and cell lines. Its expression was correlated with smoking status and the age of menarche in luminal BC patients. Also, the results of the bioinformatics analysis were consistent with the data obtained from q-RT PCR assay. The final results indicated that LINC00961 might be involved in multiple cancer-associated pathways such as chemokine, Ras and PI3K-Akt signaling pathways, GPCR ligand binding, and signal transduction in luminal subtypes of BC. CDH5, GNG11, GNG8, SELL, S1PR1, CCL19, FYN, ACAN, CD3E, ACVRL1, CAV1, and PPARGC1A were identified as the top hub genes of the PPI networks across luminal subgroup. CONCLUSION: Our findings suggested that LINC00961 was significantly downregulated in luminal A and B subtypes of BC. Moreover, bioinformatics analysis provided a basis for better identification of the potential role of LINC00961 in luminal subtype of BC.

Identification of dysregulated miRNAs-genes network in ovarian cancer: An integrative approach to uncover the molecular interactions and oncomechanisms.

BACKGROUND: Ovarian (OV) cancer is considered as one of the most deadly malignancies in women, since it is unfortunately diagnosed in advanced stages. Nowadays, the importance of bioinformatics tools and their frequent usage in tracking dysregulated cancer-related genes and pathways have been highlighted in researches. AIM: The aim of this study is to investigate dysregulated miRNAs-genes network and its function in OV tumors based on the integration of microarray data through a system biology approach. METHODS: Two microarray data (GSE119056 and GSE4122) were analyzed to explore the differentially expressed miRNAs (DEmiRs) and genes among OV tumors and normal tissues. Then, through the help of TargetScan, miRmap, and miRTarBase databases, the dysregulated miRNA-gene network in OV tumors was constructed by Cytoscape. In the next step, co-expression and protein-protein interaction networks were made using GEPIA and STRING databases. Moreover, the functional analysis of the hub genes was done by DAVID, KEGG, and Enrichr databases. Eventually, the regulatory network of TF-miRNA-gene was constructed. RESULTS: The potential dysregulated miRNAs-genes network in OV tumors has been constructed, including 109 differentially expressed genes (DEGs), 25 DEmiRs, and 213 interactions. Two down-regulated microRNAs, miR-660-3p and hsa-miR-4510, have the most interactions with up-expressed oncogenic DEGs. CDK1, PLK1, CCNB1, CCNA2, and EZH2 are involved in protein module, which show significant overexpression in OV tumors according to The Cancer Genome Atlas (TCGA) data. EZH2 shows amplification in OV tumors with remarkable percentage. The transcription factors TFAP2C and GATA4 have the pivotal regulatory functions in oncotranscriptomic profile of OV tumors. CONCLUSION: In current study, we have collected and integrated different data to uncover the complex molecular interactions and oncomechanisms in OV tumors. The DEmiRs-DEGs and TF-miRNA-gene networks reveal the potential interactions that could be a significant piece of the OV onco-puzzle.

Expression and clinicopathological significance of AOC4P, PRNCR1, and PCAT1 lncRNAs in breast cancer.

Long none coding RNAs (lncRNAs) AOC4P, PRNCR1, and PCAT-1 are dysregulated in various types of malignancies. However, their expression and clinicopathological significances are uncertain in breast cancer (BC). Quantitative real-time polymerase chain reaction (RT- qPCR) was used to measure the expression levels of the selected lncRNAs in tumor tissues obtained from 50 BC patients compared to the normal adjacent tissues (NATs) and 50 clinically healthy normal tissues. Our results revealed a significant downregulation of AOC4P, however, upregulated PRNCR1 and PCAT1 were found in tumor tissues compared to NATs and clinically healthy normal tissues (P < 0.05). Interestingly, remarkable decreased expression of AOC4P was observed in NATs than clinically healthy normal tissues. Dysregulation of the lncRNAs was correlated with worse outcomes of patients. Furthermore, our data showed that the altered expression levels of lncRNAs AOC4P, PRNCR1, and PCAT1 might be occurred through the function of demographic and reproductive variables. Taken together, the altered regulation of AOC4P, PRNCR1, and PCAT1 may highlight their crucial roles in BC development and pathogenesis. Our findings also proposed demographic and reproductive variables as risk factors in BC through the possible influence on the expression of the studied lncRNAs. Nevertheless, further explorations are required to elucidate the more detailed functions of these lncRNAs in BC.

Expression Analysis of GRHL3 and PHLDA3 in Head and Neck Squamous Cell Carcinoma.

BACKGROUND: Head and neck squamous cell carcinoma (HNSCC) includes a group of heterogeneous tumors with generally invasive behavior. The PI3K/AKT pathway plays an important role in the pathogenesis of HNSCC. METHODS: In the current study, we investigated the expression of two negative feedback regulators of the PI3K pathway, namely PHLDA3 and GRHL3, in 45 paired samples of HNSCC and adjacent non-cancerous tissues (ANCTs). RESULTS: While expression of GRHL3 was down-regulated in tumoral tissues compared with ANCTs by the factor 4.21, PHLDA3 expression levels were up-regulated by 5.99-times. Gender-based analysis revealed a significant down-regulation of GRHL3 gene expression level in male patients compared with the control samples and significant up-regulation of PHLDA3 gene expression level in both sexes compared with the control samples. Differences in the expressions of both genes were significant in patients aged more than 60 years, but not in the younger patients. Expression of GRHL3 was only down-regulated in patients with positive smoking history. Expression of GRHL3 was decreased in grades 2 and 3 samples compared with controls. There was a significant increase in transcript levels of PHLDA3 in stages II and III HNSCC samples compared with the controls group. ROC curve analysis indicated that the expression level of PHLDA3 could be a promising marker for the diagnosis of HNSCC patients with a sensitivity and specificity of 0.666 and 0.688, respectively. In addition, sensitivity and specificity of GRHL3 were 0.755 and 0.577, respectively. DISCUSSION: The current study indicates dysregulation of regulators of PI3K pathway in HNSCC and their potential application as putative biomarkers for this cancer.

The effect of cryopreservation on DNA methylation patterns of the chromosome 15q11-q13 region in human spermatozoa.

Human sperm cryopreservation is a common technique which is used in assisted reproductive technologies. Despite the existence of evidence supporting the production of ROS and DNA fragmentation during sperm cryopreservation, there is little and equivocal information about the cryopreservation effects on methylation of imprinted genes and imprinting control regions. In this study, we have investigated the effects of cryopreservation on DNA methylation in promoter regions of SNURF-SNRPN and UBE3A imprinted genes, PWS-ICR and AS-ICR in the chromosome 15q11-q13 region. Semen samples from 10 healthy normozoospermic men were collected and each sample was divided into four equal aliquots: fresh, cryoprotectant, cryopreservation, and H2O2. We measured the ROS levels and DNA fragmentation using DCFH-DA and TUNEL assay respectively by flow cytometry. DNA methylation in promoter regions of SNURF-SNRPN and UBE3A imprinted genes, PWS-ICR and AS-ICR in the chromosome 15q11-q13 region were evaluated by quantitative methylation-specific PCR technique. Intracellular levels of ROS and percentage of TUNEL-positive spermatozoa significantly increased in cryopreservation group compared to fresh group. Exposure to cryoprotectant had no significant effect on ROS levels and DNA fragmentation. Neither cryopreservation nor exposure to cryoprotectant significantly affected DNA methylation of the selected gene regions. However, DNA fragmentation had positive correlation with DNA methylation of AS-ICR. In conclusion, based on our study, clinical use of sperm cryopreservation for fertility treatments appear to be safe in regard to DNA methylation in the chromosome 15q11-q13 region.

Anti-tumor activity of cediranib, a pan-vascular endothelial growth factor receptor inhibitor, in pancreatic ductal adenocarcinoma cells.

PURPOSE: Pancreatic ductal adenocarcinoma (PDAC) is the most common and lethal subtype of pancreatic cancer, with a 5-year survival rate of < 3%. Early tumor dissemination, late diagnosis and insensitivity to conventional treatment are the major reasons for its high mortality rate. Members of the vascular endothelial growth factor (VEGF) family are overexpressed in PDAC and play important roles in its malignant progression, suggesting that VEGF-targeted therapies may interrupt the proliferation and motility of PDAC cells. Here, we evaluated the anti-tumor activity of cediranib, a pan-VEGF receptor inhibitor, on PDAC cells. METHODS: Anti-proliferative effects of cediranib were determined using cell proliferation and crystal violet staining assays. Annexin V/PI staining, radiation therapy, and cell migration and invasion assays were carried out to examine the effects of cediranib on apoptosis, radio-sensitivity and cell motility, respectively. Quantitative reverse transcription-PCR (qRT-PCR) and Western blot analyses were applied to elucidate the molecular mechanisms underlying the anti-tumor activity of cediranib. RESULTS: We found that cediranib decreased PDAC cell proliferation and clonogenic survival and induced apoptotic cell death through inhibition of the anti-apoptotic proteins cIAP1, XIAP, MCL-1 and survivin. Combination with cediranib synergistically increased the sensitivity of PDAC cells to chemotherapeutic agents such as gemcitabine and paclitaxel, and potentiated the effects of radiation therapy on PDAC cell growth inhibition and apoptosis induction. Furthermore, we found that treatment with cediranib impaired PDAC cell migration and invasion via expression reduction of the epithelial-to-mesenchymal transition (EMT) markers ZEB1, N-cadherin and Snail. CONCLUSIONS: Our data indicate that cediranib may exhibit anti-tumor activity in PDAC cells and provide a rationale for further investigation of the potential of VEGF receptor-targeted therapies for the treatment of PDAC.

An Association and Meta-Analysis of Esophageal Squamous Cell Carcinoma Risk Associated with PLCE1 rs2274223, C20orf54 rs13042395 and RUNX1 rs2014300 Polymorphisms.

One of the highest risk of esophageal squamous cell carcinoma (ESCC) in the world has been reported in Iran, which is located in the Asian esophageal cancer belt. ESCC constitutes 90% of the esophageal cancer cases in Iran. Genome wide association studies (GWASs) in Chinese have identified a number of candidate variants, of which PLCE1rs2274223, C20orf54rs13042395 and RUNX1rs2014300 are studied in high risk populations including Chinese, Caucasians and Africans. However, results are inconsistent and it is unknown whether similar associations exist in Iranian population. We evaluated association of three GWAS identified variants with risk of ESCC in an Iranian cohort consisted of 200 ESCC patients and 300 healthy controls and conducted meta-analysis of ESCC risk associated with rs2274223 (involving 9810 cases and 13,128 controls) and rs13042395 (involving 2363 cases and 5329 controls). Logistic regression analysis showed that rs2274223 was associated with ESCC under codominant [GG/AA, 2.47(1.17-5.23), P:0.021], dominant [AG + GG/AA, 1.57(1.09-2.27), P:0.016], recessive [GG/AA+AG, 2.18(1.04-4.56), P:0.036] and log-additive models [1.51(1.12-2.02), P:0.006]. C20orf54 rs13042395 was not associated with ESCC under any genetic model. RUNX1 rs2014300 was associated with risk of ESCC assuming codominant [AG/GG, 0.63(0.41-0.97), P:0.018], dominant [AG + AA/GG, 0.59 (0.39-0.89), P:0.010] and log-additive models [0.61 (0.42-0.87), P: 0.005]. Meta-analysis found significant associations between rs2274223 and ESCC under all analyzed genetic models. However, meta-analysis stratified by ethnicity showed a significant association in Asians but not non-Asian populations. No significant association was found for rs13042395 in meta-analysis. This study provided first evidence for association of GWAS-identified variants with risk of ESCC in an Iranian cohort.

Anti-tumor activity of neratinib, a pan-HER inhibitor, in gastric adenocarcinoma cells.

Gastric adenocarcinoma (GAC), the most common malignancy of the stomach, is the fourth most common and the second cause of cancer-related death worldwide. Although HER family plays a cardinal role in tumorigenesis of GAC, trastuzumab is the only approved anti-HER drug for this malignancy and development of resistance to trastuzumab is inevitable. Additionally, single-targeted HER inhibitors have demonstrated limited activity in GAC. Hence, there is a pressing need to devise more efficacious anti-HER therapeutic strategies. Here, we examined the anti-tumor activity of neratinb, a pan-HER inhibitor, on GAC cells. Anti-proliferative effects of neratinib were determined using a cell proliferation assay and crystal violet staining. Annexin V/PI staining, radiation therapy and anoikis resistance and wound healing assays were carried out to examine the effects of neratinib on apoptosis, radio-sensitivity and cell motility, respectively. Quantitative reverse transcription-PCR (qRT-PCR) analyses were applied to further investigate the anti-tumor activity of neratinib. We found that neratinib sensitized GAC cells to 5FU, carboplatin and oxaliplatin. Moreover, we found that neratinib was synergistic with trametinib (an approved MEK inhibitor) and foretinib (a c-MET inhibitor) and potentiated radio-sensitivity of GAC cells. Furthermore, we found that neratinib diminished GAC cell proliferation along with downregulation of FOXM1 and its targets. Additionally, neratinib induced apoptosis along with upregulation of pro-apoptotic and downregulation of anti-apoptotic genes. Treatment with neratinib attenuated invasive ability of GAC cells as shown by reduced anoikis resistance, downregulation of EMT markers, and reduced width in scratch assay. Our findings indicate that neratinib provides the therapeutic potential in the treatment of GAC.

Integrative analyses of triple negative dysregulated transcripts compared with non-triple negative tumors and their functional and molecular interactions.

Triple-negative (TN) tumors are a subtype of breast cancer with aggressive behaviors and limited targeted therapies. Microarray studies were not concerned with interactions and functional relations of dysregulated transcripts. Here, we aimed to conduct integrative strategy to analyze gene and miRNA available microarray data as well as bioinformatic analyses to catch a more inclusive picture of pivotal dysregulated transcripts and their interactions in TN tumors. Several online datasets and offline bioinformatic tools were used to detect differentially expressed (DE) transcripts, both protein and nonprotein coding, in TN compared with non-TN tumors and their functional and molecular interactions. Sixteen upregulated and 58 downregulated genes with a log fold change higher or equal to | 2 | were identified, including nine transcription factors. Coexpression network revealed EN1 as a hub gene, moreover Kaplan-Meier plotter survival analysis indicated that it was an appropriate prognostic marker for TN patients with breast cancer. Functional annotation analysis of protein-protein interaction network showed FOXM1 as an upexpressed and ESR1 as a downexpressed hub genes are suitable targets as far as antitumor protein therapy is concerned in TN breast cancers. The consensus analysis of two microRNA datasets revealed seven DE miRNAs. The gene-transcriptional factor (TF)-miRNA network revealed mir-135b and mir-29b are the hub nodes and involved in feedback loops with GATA3. This study suggests that dysregulated TFs and miRNAs have pivotal roles in regulation of TN oncotranscriptomic profile and might become both biomarkers and therapeutic targets.

The ERBB receptor inhibitor dacomitinib suppresses proliferation and invasion of pancreatic ductal adenocarcinoma cells.

PURPOSE: Pancreatic ductal adenocarcinoma (PDAC), the most common malignancy of the pancreas, is the fourth most common cause of cancer-related death in the USA. Local progression, early tumor dissemination and low efficacy of current treatments are the major reasons for its high mortality rate. The ERBB family is over-expressed in PDAC and plays essential roles in its tumorigenesis; however, single-targeted ERBB inhibitors have shown limited activity in this disease. Here, we examined the anti-tumor activity of dacomitinib, a pan-ERBB receptor inhibitor, on PDAC cells. METHODS: Anti-proliferative effects of dacomitinib were determined using a cell proliferation assay and crystal violet staining. Annexin V/PI staining, radiation therapy and cell migration and invasion assays were carried out to examine the effects of dacomitinib on apoptosis, radio-sensitivity and cell motility, respectively. Quantitative reverse transcription-PCR (qRT-PCR) and Western blot analyses were applied to elucidate the molecular mechanisms underlying the anti-tumor activity of dacomitinib. RESULTS: We found that dacomitinib diminished PDAC cell proliferation via inhibition of FOXM1 and its targets Aurora kinase B and cyclin B1. Moreover, we found that dacomitinib induced apoptosis and potentiated radio-sensitivity via inhibition of the anti-apoptotic proteins survivin and MCL1. Treatment with dacomitinib attenuated cell migration and invasion through inhibition of the epithelial-to-mesenchymal transition (EMT) markers ZEB1, Snail and N-cadherin. In contrast, we found that the anti-tumor activity of single-targeted ERBB agents including cetuximab (anti-EGFR mAb), trastuzumab (anti-HER2 mAb), H3.105.5 (anti-HER3 mAb) and erlotinib (EGFR small molecule inhibitor) were marginal. CONCLUSIONS: Our findings indicate that dacomitinib-mediated blockade of the ERBB receptors yields advantages over single-targeted ERBB inhibition and provide a rationale for further investigation of the therapeutic potential of dacomitinib in the treatment of ERBB-driven PDAC.

Molecular investigation of mutations in androgen receptor and 5-alpha-reductase-2 genes in 46,XY Disorders of Sex Development with normal testicular development.

In this study, we aimed to determine androgen receptor (AR) and SRD5A2 gene mutations in 45 patients characterised by 46,XY Disorders of Sex Differentiation (DSD) signs with normal testicular development referred to the Children's Medical Center from February 2015 to September 2017. Karyotype and sex hormone analyses were performed. Cytogenetic investigation showed that seven patients were 46,XX DSD, six patients were chromosomal DSD and 32 patients were 46,XY DSD. Eight exons of the AR gene and five exons of the SRD5A2 gene were amplified. Two cases were affected with androgen insensitivity syndrome (AIS) (missense mutation on exon 7, position c.3637 G>A: p.R841H and position c.3610 G>A: p.R832Q), one case was affected with 5-alpha-reductase deficiency type 2 (missense mutation at c.578A>G: p.N193S on exon 4), and 22 cases (88%) did not demonstrate AIS or 5α-RD2 gene abnormality. Due to the great impact of these disorders on human lifestyle, evaluation of genes involved can improve genetic counselling and therapeutic management. We focused on the AR and SRD5A2 genes in patients with 46,XY DSDs with normal testicular development referred to the Children's Medical Center from all over the country to eventually culminate in a reliable prenatal diagnosis protocol at this major referral centre giving service to a great number of families with consanguineous marriages.