Gelsolin, an Actin-Binding Protein: Bioinformatic Analysis and Functional Significance in Urothelial Bladder Carcinoma.

The involvement of the actin-regulatory protein, gelsolin (GSN), in neoplastic transformation has been reported in different cancers including bladder cancer. However, the exact mechanism by which GSN influences bladder cancer development is not well understood. Here, we sought to reveal the functional significance of GSN in bladder cancer by undertaking a comprehensive bioinformatic analysis of TCGA datasets and through the assessment of multiple biological functions. GSN expression was knocked down in bladder cancer cell lines with two siRNA isoforms targeting GSN. Proliferation, migration, cell cycle and apoptosis assays were carried out. GSN expression, enrichment analysis, protein-protein interaction and immune infiltration analysis were verified through online TCGA tools. The data indicated that GSN expression is associated with bladder cancer proliferation, migration and enhanced cell apoptosis through regulation of NF-κB expression. GSN expression correlated with various inflammatory cells and may influence the immunity of the tumor microenvironment. Computational analysis identified several interacting partners which are associated with cancer progression and patient outcome. The present results demonstrate that GSN plays an important role in bladder cancer pathogenesis and may serve as a potential biomarker and therapeutic target for cancer therapy.

Interaction between HER2 and ATM predicts poor survival in bladder cancer patients.

Human Epidermal Growth Factor Receptor 2 (HER2) overexpression is considered one of the interesting prognostic biomarkers in bladder cancer. However, the mechanism of bladder cancer development in relation to HER2 status remains to be elucidated. In this study, we investigated HER2-Ataxia telangiectasia mutated (ATM) kinase interaction and their impact on patient survival and cancer aggressiveness. Using the Cancer Genome Atlas (TCGA) cohorts, we demonstrated that ATM expression (protein/mRNA) is increased in HER2 deficient compared with proficient HER2 patients. This finding was then validated using the Gene Expression Omnibus database (GEO). Correlation analysis (using low expression vs high expression as a discriminator) revealed a significant association of ATM low and HER2 high status with several clinicopathological variables such as high tumour grade, late disease stage and tumour shape. Kaplan-Meier survival analysis indicated that ATM low and HER2 high is a powerful prognosticator of both overall survival (OS) and disease-free survival (DFS). Furthermore, using bioinformatics and protein/protein interaction analyses, we identified 66 putative overlapping proteins with direct link between HER2 and ATM most of which are functionally involved in transcription regulation, apoptotic process and cell proliferation. Interestingly, the results showed that these proteins are strongly linked with PI3K-Akt pathway, p53 pathway and microRNAs in cancer. Altogether, our data pinpoint an important biological role of the interconnection between HER2 and ATM. The latter appear to be an independent prognostic biomarker and may serve as targets to develop novel combination therapies to improve the outcome of patients with bladder cancer.

The prognostic impact of GSTM1/GSTP1 genetic variants in bladder Cancer.

BACKGROUND: The glutathione S-transferases (GSTs) are a superfamily of phase II detoxifying enzymes that inactivates a wide variety of potential carcinogens through glutathione conjugation. Polymorphic changes in the GST genes have been reported to be associated with increased susceptibility to cancer development and anticancer drug resistance. In this study, we investigated the association between genetic variants in GSTM1 and GSTP1 and patients' clinicopathological parameters. The prognostic values of such associations were evaluated among bladder cancer patients. METHODS: Genotyping of GSTM1 and GSTP1 in bladder cancer patients was assessed using polymerase chain reaction followed by DNA sequencing. Overall survival was estimated using the Kaplan-Meier method and multiple logistic regression and correlation analysis were performed. RESULTS: The GSTM1 null genotype was significantly associated with poor overall survival compared with the wild-type GSTM1 genotype. There was a trend towards better overall survival in patients with wild-type GSTP1 allele (AA) compared with GSTP1 (AG/GG) genotype. Interestingly, Kaplan-meier survival curve for GSTM1 null patients adjusted for sub-cohort with amplified HER2 gene showed poor survival compared with the GSTM1 null/ non-amplified HER2 gene. Also the same population when adjusted with HER2 protein expression, data showed poor survival for patients harboring GSTM1 null/high HER2 protein expression compared with low protein expression. CONCLUSION: This study focuses on the impact of GSTM1 null genotype on bladder cancer patients' outcome. Further investigations are required to delineate the underlying mechanisms of combined GSTM-/- and HER2 status in bladder cancer.

S100A8 and S100A9 proteins form part of a paracrine feedback loop between pancreatic cancer cells and monocytes.

BACKGROUND: The secretion of soluble factors enables communication between tumour cells and the surrounding microenvironment and plays an important role in oncogenesis. Pancreatic ductal adenocarcinoma (PDAC) is characterised by a highly reactive microenvironment, harbouring a variety of cell types, including S100A8/S100A9-expressing monocytes. S100A8/S100A9 proteins regulate the behaviour of cancer cells by inducing pre-metastatic cascades associated with cancer spread. The aim of this study was to examine how S100A8/A9 proteins mediate tumour-stroma crosstalk in PDAC. METHODS: Cytokine profiling of pancreatic cancer cell-derived conditioned media was performed using Bio-Plex Pro 27 Plex Human Cytokine assays. Protein expression and activation of downstream signalling effectors and NF-κB were assessed by western blotting analysis and reporter assays respectively. RESULTS: Stimulation of cultured pancreatic cancer cells with S100A8 and S100A9 increased the secretion of the pro-inflammatory cytokines IL-8, TNF-α, and FGF. S100A8, but not S100A9 induced PDGF secretion. Conversely, pancreatic cancer cell-derived conditioned media and the individual cytokines, TNF-α and TGF-ß induced the expression of S100A8 and S100A9 proteins in the HL-60 monocytic cell line and primary human monocytes, while FGF and IL-8 induced the expression of S100A9 only. S100A8 and S100A9 activated MAPK and NF-κB signalling in pancreatic cancer. This was partially mediated via activation of the receptor of advanced glycosylation end-product (RAGE). CONCLUSION: S100A8 and S100A9 proteins induce specific cytokine secretion from PDAC cells, which in turn enhances the expression of S100A8/A9. This paracrine crosstalk could have implications for PDAC invasiveness and metastatic potential.

Association of Interleukin-6 and Other Cytokines with Self-Reported Pain in Prostate Cancer Patients Receiving Chemotherapy.

Background: Pain is a common and dose-limiting side effect of many potentially curative cancer chemotherapeutic agents. This chemotherapy-induced pain (CIP) affects the quality of life of cancer patients and survivors and hampers the optimal clinical management of chemotherapy in cancer patients. The underlying mechanisms remain largely unknown, but changes in levels of cytokines/chemokines may contribute to the pathophysiology of CIP. Objective: This retrospective study was aimed at examining whether plasma levels of various cytokines change in prostate cancer patients after chemotherapy treatment and whether such changes (if any) are associated with their pain intensity. Methods: Using a Luminex assay, plasma levels of 27 cytokines/chemokines were measured in 78 men: 30 patients with metastatic prostate cancer who received chemotherapy (Docetaxel, 75 mg/m2 intravenously), 29 untreated patients with nonmetastatic prostate cancer, and 19 healthy controls. Subjective pain was assessed in chemotherapy-treated cancer patients using the 11-point numeric rating scale (NRS) pain scores. Results: Chemotherapy-treated patients with pain (NRS ≥ 3) exhibited significantly increased levels of the pro-inflammatory cytokines (IL-6, IL-8) and chemokines (Eotaxin, VEGF, and IP-10) compared with untreated cancer patients or with patients without pain (NRS = 0). Of the 27 cytokines examined, only IL-6 was positively correlated with pain intensity in the chemotherapy-treated patients with pain. Conclusions: These findings suggest that the cytokines, particularly IL-6, whose levels were elevated in the chemotherapy-treated patients may be involved in the pathophysiology of CIP, and that they might be potential new targets for pain control in cancer patients receiving chemotherapy.

UHRF1 regulation of the Keap1-Nrf2 pathway in pancreatic cancer contributes to oncogenesis.

The cellular defence protein Nrf2 is a mediator of oncogenesis in pancreatic ductal adenocarcinoma (PDAC) and other cancers. However, the control of Nrf2 expression and activity in cancer is not fully understood. We previously reported the absence of Keap1, a pivotal regulator of Nrf2, in ∼70% of PDAC cases. Here we describe a novel mechanism whereby the epigenetic regulator UHRF1 suppresses Keap1 protein levels. UHRF1 expression was observed in 20% (5 of 25) of benign pancreatic ducts compared to 86% (114 of 132) of pancreatic tumours, and an inverse relationship between UHRF1 and Keap1 levels in PDAC tumours (n = 124) was apparent (p = 0.002). We also provide evidence that UHRF1-mediated regulation of the Nrf2 pathway contributes to the aggressive behaviour of PDAC. Depletion of UHRF1 from PDAC cells decreased growth and enhanced apoptosis and cell cycle arrest. UHRF1 depletion also led to reduced levels of Nrf2-regulated downstream proteins and was accompanied by heightened oxidative stress, in the form of lower glutathione levels and increased reactive oxygen species. Concomitant depletion of Keap1 and UHRF1 restored Nrf2 levels and reversed cell cycle arrest and the increase in reactive oxygen species. Mechanistically, depletion of UHRF1 reduced global and tumour suppressor promoter methylation in pancreatic cancer cell lines, and KEAP1 gene promoter methylation was reduced in one of three cell lines examined. Thus, methylation of the KEAP1 gene promoter may contribute to the suppression of Keap1 protein levels by UHRF1, although our data suggest that additional mechanisms need to be explored. Finally, we demonstrate that K-Ras drives UHRF1 expression, establishing a novel link between this oncogene and Nrf2-mediated cellular protection. Since UHRF1 over-expression occurs in other cancers, its ability to regulate the Keap1-Nrf2 pathway may be critically important to the malignant behaviour of these cancers.

Prognostic value of HER2 status in bladder transitional cell carcinoma revealed by both IHC and BDISH techniques.

BACKGROUND: Her2/neu is an oncogene that plays an important role in the pathogenesis of many cancer types. In bladder carcinoma (BC), the clinical significance of Her2/neu status remains under-investigated and poorly linked to the patients' clinic-pathological features and survival status. Thus, the current study was conducted to assess Her2/neu status in a cohort of patients' in Saudi Arabia, and to explore its prognostic value in BC. METHODS: A total of 160 consent patients of transitional cell carcinoma (TCC) of bladder were arranged on a tissue microarray (TMA) and stained by immunohistochemistry (IHC) and bright-field dual in situ hybridization (BDISH) methods. The intensity of Her2/neu protein receptor immunostaining was evaluated, correlated to Her2/neu gene amplification status in TCC and assessed for potential clinical value by correlation measures. RESULTS: IHC data demonstrated that Her2/neu protein is expressed in 60 % (2+ and 3+) of our TCC patient's cohort from Saudi Arabia. Her2/neu gene amplification is detected in 25 % by BDISH. There was a strong association between Her2/neu protein levels and lymph node invasion (p = 0.04), tumor stage (p = 0.002), vascular invasion and borderline significance with distant metastasis (p = 0.07). Amplification of Her2/neu gene was associated with tumor grade (p = 0.03) and poor disease-specific survival (p = 0.02), in that, patients with non-amplified Her2/neu gene live longer. Interestingly, there was a reasonable concordance rate (71 %) between IHC and BDISH data in the analyzed cohort. CONCLUSION: The study showed that 25 % of our patients' cohort has Her2/neu over-expression. This Her2/neu (over-expression/amplification) status was concordant using either IHC or BDISH and significantly associated with disease aggressiveness and poor outcome. These findings suggested a potential impact of anti-Her2 targeted therapy in the treatment of bladder cancer with amplified/overexpressed HER2 that needs further investigation.

Does Val/Val genotype of GSTP1 enzyme affects susceptibility to colorectal cancer in Saudi Arabia?.

OBJECTIVES: Glutathione S-transferase pi (GSTP1) is a candidate enzyme that may be involved in colorectal cancer susceptibility. Polymorphism of GSTP1 gene may cause changes in expression or structure which lead to alteration in the efficiency of catalytic function of the enzyme variants, i.e., deficient detoxification of carcinogens and consequently influences coloreActal cancer development. The present report examined the possible impact of GSTP1 (Ilel05Val) polymorphism and the risk of colorectal cancer. METHODS: Samples of paraffin embedded tissues from 83 patients with colorectal cancer as well as thirty five non-cancerous colon tissues were collected from the archive of the pathology department at King Abdulaziz University in Saudi Arabia. All cancer and control samples were subjects to DNA extraction then amplification. DNA genetic analyzer from Applied Biosystems was used to sequence the product of amplification for genotypes determination. RESULTS: None of the genotypes of GSTP1 was associated with the risk of colorectal cancer development. There were no statistical differences in the frequencies of GSTP1 genotypes between colorectal cancer cases and controls. CONCLUSION: The incidence of (Val/Val) genotype in colorectal cancer cases was three folds higher than controls. This finding is not statistically significant, but it could be of clinical consequence that it may increase the risk of colorectal cancer in Saudi Arabia.

Frequent methylation of the KLOTHO gene and overexpression of the FGFR4 receptor in invasive ductal carcinoma of the breast.

Invasive ductal carcinoma of the breast is the most common cancer affecting women worldwide. The marked heterogeneity of breast cancer is matched only with the heterogeneity in its associated or causative factors. Breast cancer in Saudi Arabia is apparently an early onset with many of the affected females diagnosed before they reach the age of 50 years. One possible rationale underlying this observation is that consanguinity, which is widely spread in the Saudi community, is causing the accumulation of yet undetermined cancer susceptibility mutations. Another factor could be the accumulation of epigenetic aberrations caused by the shift toward a Western-like lifestyle in the past two decades. In order to shed some light into the molecular mechanisms underlying breast cancer in the Saudi community, we identified KLOTHO (KL) as a tumor-specific methylated gene using genome-wide methylation analysis of primary breast tumors utilizing the MBD-seq approach. KL methylation was frequent as it was detected in 55.3 % of breast cancer cases from Saudi Arabia (n = 179) using MethyLight assay. Furthermore, KL is downregulated in breast tumors with its expression induced following treatment with 5-azacytidine. The involvement of KL in breast cancer led us to investigate its relationship in the context of breast cancer, with one of the protagonists of its function, fibroblast growth factor receptor 4 (FGFR4). Overexpression of FGFR4 in breast cancer is frequent in our cohort and this overexpression is associated with poor overall survival. Interestingly, FGFR4 expression is higher in the absence of KL methylation and lower when KL is methylated and presumably silenced, which is suggestive of an intricate relationship between the two factors. In conclusion, our findings further implicate "metabolic" genes or pathways in breast cancer that are disrupted by epigenetic mechanisms and could provide new avenues for understanding this disease in a new context.

Tackling dengue fever: Current status and challenges.

According to recent statistics, 96 million apparent dengue infections were estimated worldwide in 2010. This figure is by far greater than the WHO prediction which indicates the rapid spread of this disease posing a growing threat to the economy and a major challenge to clinicians and health care services across the globe particularly in the affected areas.This article aims at bringing to light the current epidemiological and clinical status of the dengue fever. The relationship between genetic mutations, single nucleotide polymorphism (SNP) and the pathophysiology of disease progression will be put into perspective. It will also highlight the recent advances in dengue vaccine development.Thus far, a significant progress has been made in unraveling the risk factors and understanding the molecular pathogenesis associated with the disease. However, further insights in molecular features of the disease and the development of animal models will enormously help improving the therapeutic interventions and potentially contribute to finding new preventive measures for population at risk.

High fibroblast growth factor 19 (FGF19) expression predicts worse prognosis in invasive ductal carcinoma of breast.

Metabolic diseases like diabetes and obesity are major risk factors for breast cancer. Aberrant expression of metabolic effectors such as fibroblast growth factor 19 (FGF19) could be therefore associated with the disease. The expression of FGF19 was examined in 193 archival breast tumor samples by immunohistochemistry and evaluated semi-quantitatively by determining the staining index and correlating it with clinicopathological parameters using Fisher's exact test. The correlation between FGF19 expression and 5-year disease-specific survival rate was determined using the univariate Kaplan-Meier analysis. The prognostic value of FGF19 expression was evaluated using the multivariate Cox regression analysis. Of the 193 tumors analyzed, 40% were classified with low FGF19 expression, whereas 60% were categorized as tumors with high FGF19 expression. There was a highly significant correlation between high FGF19 expression and patients' age (p = 0.008) as well as 5-year disease-specific survival (p = 0.001). However, FGF19 expression did not show any significant correlations with other clinicopathological parameters, including hormonal status, tumor grade, tumor size, or lymph node status. Univariate Kaplan-Meier log rank analysis showed that patients with high FGF19 expression exhibited a significantly shorter disease-specific 5-year survival (p = 0.007). This effect was exacerbated by lymph node metastasis (p = 0.001), negative estrogen receptor (ER) status (p = 0.002), or old age (p = 0.013). Multivariate analysis showed that high FGF19 expression could be an independent prognostic marker of disease-specific survival in breast cancer patients (p = 0.030). Quantification of FGF19 expression appears to provide valuable prognostic information in breast cancer, particularly in older patients with lymph node metastasis and negative ER status.

Characterization of butyrate transport across the luminal membranes of equine large intestine.

The diet of the horse, pasture forage (grass), is fermented by the equine colonic microbiota to short-chain fatty acids, notably acetate, propionate and butyrate. Short-chain fatty acids provide a major source of energy for the horse and contribute to many vital physiological processes. We aimed to determine both the mechanism of butyrate uptake across the luminal membrane of equine colon and the nature of the protein involved. To this end, we isolated equine colonic luminal membrane vesicles. The abundance and activity of cysteine-sensitive alkaline phosphatase and villin, intestinal luminal membrane markers, were significantly enriched in membrane vesicles compared with the original homogenates. In contrast, the abundance of GLUT2 protein and the activity of Na(+)-K(+)-ATPase, known markers of the intestinal basolateral membrane, were hardly detectable. We demonstrated, by immunohistochemistry, that monocarboxylate transporter 1 (MCT1) protein is expressed on the luminal membrane of equine colonocytes. We showed that butyrate transport into luminal membrane vesicles is energized by a pH gradient (out < in) and is not Na(+) dependent. Moreover, butyrate uptake is time and concentration dependent, with a Michaelis-Menten constant of 5.6 ± 0.45 mm and maximal velocity of 614 ± 55 pmol s(-1) (mg protein)(-1). Butyrate transport is significantly inhibited by p-chloromercuribenzoate, phloretin and α-cyano-4-hydroxycinnamic acid, all potent inhibitors of MCT1. Moreover, acetate and propionate, as well as the monocarboxylates pyruvate and lactate, also inhibit butyrate uptake. Data presented here support the conclusion that transport of butyrate across the equine colonic luminal membrane is predominantly accomplished by MCT1.

Transcriptomics Demonstrates Significant Biological Effect of Growing Stem Cells on RGD-Cotton Scaffold.

Stem cell therapy provides a viable alternative treatment for degenerated or damaged tissue. Stem cells have been used either alone or in conjunction with an artificial scaffold. The latter provides a structural advantage by enabling the cells to thrive in three-dimensional (3D) settings, closely resembling the natural in vivo environments. Previously, we disclosed the development of a 3D scaffold made from cotton, which was conjugated with arginyl-glycyl-aspartic acid (RGD), to facilitate the growth and proliferation of mesenchymal stem cells (MSCs). This scaffold allowed the MSCs to adhere and proliferate without compromising their viability or their stem cell markers. A comprehensive analysis investigation of the molecular changes occurring in MSCs adhering to the cotton fibers will contribute to the advancement of therapy. The objective of this study is to analyze the molecular processes occurring in the growth of MSCs on a cotton-RGD conjugated-based scaffold by examining their gene expression profiles. To achieve this, we conducted an experiment where MSCs were seeded with and without the scaffold for a duration of 48 h. Subsequently, cells were collected for RNA extraction, cDNA synthesis, and whole-transcriptomic analysis performed on both populations. Our analysis revealed several upregulated and downregulated differently expressed genes in the MSCs adhering to the scaffold compared with the control cells. Through gene ontology analysis, we were able to identify enriched biological processes, molecular functions, pathways, and protein-protein interactions in these differentially expressed genes. Our data suggest that the scaffold may have the potential to enhance osteogenesis in the MSCs. Furthermore, our results indicate that the scaffold does not induce oxidative stress, inflammation, or aging in the MSCs. These findings provide valuable insights for the application of MSCs in tissue engineering and regenerative medicine.

Protective potential of mesenchymal stem cells against COVID-19 during pregnancy.

SARS-CoV-2 causes COVID-19. COVID-19 has led to severe clinical illnesses and an unprecedented death toll. The virus induces immune inflammatory responses specifically cytokine storm in lungs. Several published reports indicated that pregnant females are less likely to develop severe symptoms compared with non-pregnant. Putative protective role of maternal blood circulating fetal mesenchymal stem cells (MSCs) has emerged and have been put forward as an explanation to alleviated symptoms. MSCs with immune-modulatory, anti-inflammatory and anti-viral roles, hold great potential for the treatment of COVID-19. MSCs could be an alternative to treat infections resulting from the SARS-CoV-2 and potential future outbreaks. This review focuses on the MSCs putative protective roles against COVID-19 in pregnant females.

The COVID-19 outbreak is still posing a global health concern. Despite the herd immunity provided by vaccination programs, no efficient treatments are yet available especially against the severe forms of the disease. According to published reports, pregnant females are less likely to develop the severe form of the disease due to the protective effect of specialized cells named mesenchymal stem cells (MSCs). These cells are present in the placenta and amniotic fluid. They can migrate from these tissues to the mother's blood stream and are believed to confer protection to the pregnant females against severe form of COVID-19. Further investigations are on the way to uncover the potential role of MSC as an alternative therapy for COVID-19 and other diseases.

The virus SARS-CoV-2 results in COVID-19 infection. Several reports indicated that pregnant females are less likely to develop severe symptoms. Circulating fetal Mesenchymal stem cells in pregnant females might protect them.

Identification of SPP1 as a Prognostic Biomarker and Immune Cells Modulator in Urothelial Bladder Cancer: A Bioinformatics Analysis.

Secreted phosphoprotein-1 (SPP1) expression is differentially altered in many malignancies and could serve as a potential prognostic biomarker. Recent findings indicated that SPP1 possesses a broader role in bladder cancer (BC) pathogenesis than previously envisioned; however, the underlying mechanisms governing its expression, cellular localization, prognostic value and immune-related role in bladder cancer remain poorly understood. The expression and the prognosis value of SPP1 were assessed using immunohistochemistry (IHC) staining on a tissue microarray. SPP1 expression was correlated with the clinicopathological parameters, and survival analysis was calculated using a Kaplan-Meier plotter. Bioinformatics analysis of TCGA data was queried using UALCAN, CIBERSORT and TIMER datasets to decipher the biological processes enrichment pattern, protein-protein interactions and characterize tumor-infiltrating immune cells, respectively. IHC revealed that SPP1 expression is significantly associated with tumor type, stage, grade and smoking status. The Kaplan-Meier survival curve showed that low SPP1 expression is an unfavorable prognostic indicator in bladder cancer patients (p = 0.02, log-rank). The significant increased expression of the SPP1 level is associated with evident hypomethylation of the gene promoter in cancer compared to normal tissues in the TCGA-bladder dataset. Missense mutation is the most frequent genetic alteration of the SPP1 gene. Protein-protein interactions demonstrated that SPP1 shares the same network with many important genes and is involved in many signaling pathways and biological processes. TIMER reported a significant correlation between SPP1 expression and multiple immune cells infiltration. Furthermore, the expression of SPP1 was found to be positively correlated with a number of immune checkpoint genes such as PD-1 and CTLA4. The current investigation indicates that the SPP1 protein could serve as a prognostic biomarker and merit further investigation to validate its clinical usefulness in patients with bladder cancer.

Nrf2 is overexpressed in pancreatic cancer: implications for cell proliferation and therapy.

BACKGROUND: Nrf2 is a key transcriptional regulator of a battery of genes that facilitate phase II/III drug metabolism and defence against oxidative stress. Nrf2 is largely regulated by Keap1, which directs Nrf2 for proteasomal degradation. The Nrf2/Keap1 system is dysregulated in lung, head and neck, and breast cancers and this affects cellular proliferation and response to therapy. Here, we have investigated the integrity of the Nrf2/Keap1 system in pancreatic cancer. RESULTS: Keap1, Nrf2 and the Nrf2 target genes AKR1c1 and GCLC were detected in a panel of five pancreatic cancer cell lines. Mutation analysis of NRF2 exon 2 and KEAP1 exons 2-6 in these cell lines identified no mutations in NRF2 and only synonomous mutations in KEAP1. RNAi depletion of Nrf2 caused a decrease in the proliferation of Suit-2, MiaPaca-2 and FAMPAC cells and enhanced sensitivity to gemcitabine (Suit-2), 5-flurouracil (FAMPAC), cisplatin (Suit-2 and FAMPAC) and gamma radiation (Suit-2). The expression of Nrf2 and Keap1 was also analysed in pancreatic ductal adenocarcinomas (n = 66 and 57, respectively) and matching normal benign epithelium (n = 21 cases). Whilst no significant correlation was seen between the expression levels of Keap1 and Nrf2 in the tumors, interestingly, Nrf2 staining was significantly greater in the cytoplasm of tumors compared to benign ducts (P < 0.001). CONCLUSIONS: Expression of Nrf2 is up-regulated in pancreatic cancer cell lines and ductal adenocarcinomas. This may reflect a greater intrinsic capacity of these cells to respond to stress signals and resist chemotherapeutic interventions. Nrf2 also appears to support proliferation in certain pancreatic adenocarinomas. Therefore, strategies to pharmacologically manipulate the levels and/or activity of Nrf2 may have the potential to reduce pancreatic tumor growth, and increase sensitivity to therapeutics.

Low molecular weight heat shock protein HSP27 is a prognostic indicator in rectal cancer but not colon cancer.

OBJECTIVE: There are currently no biomarkers in routine clinical use for determining prognosis in rectal cancer. In a preliminary proteomic study, variation in the levels of heat shock protein 27 (HSP27) in colorectal cancer samples was observed. The expression of HSP27 in a cohort of 404 patients with colorectal cancer with a predominantly poor prognosis was characterised and an investigation was undertaken of whether the differences were related to clinical outcome. HSP27 levels in diagnostic rectal biopsies were compared with matched surgical samples to determine whether changes in expression occurred in the time between biopsy and surgery and to investigate whether preoperative radiotherapy affected expression. Finally, the relationship between HSP27 expression and outcome was examined in an independent cohort of 315 patients with a predominantly good prognosis. METHODS: HSP27 levels were determined using combined two-dimensional gel electrophoresis and tandem mass spectrometry (12 cases) and by immunohistochemistry using tissue microarrays of colorectal cancers sampled at surgery and 80 diagnostic rectal biopsies. RESULTS: HSP27 overexpression was strongly associated with poor cancer-specific survival in rectal cancer (n=205, p=0.0063) but not colon cancer (n=199, p=0.7385) in the cohort with a poor prognosis. Multivariate Cox regression confirmed nodal metastases (p=0.0001) and HSP27 expression (p=0.0233) as independent markers of survival in rectal cancer. HSP27 levels remained unchanged in the majority of cases (65/80, 81%) between diagnostic biopsies and matched surgical samples, regardless of whether patients had undergone preoperative radiotherapy. In the cohort with a good prognosis the association between HSP27 and survival was not observed in patients with either rectal (n=115; p=0.308) or colon cancer (n=200; p=0.713). CONCLUSION: In a large cohort of patients with a poor prognosis, HSP27 is an independent marker of poor outcome in rectal cancer; its expression is not altered by neoadjuvant radiotherapy. This finding requires validation in an independent similar cohort of patients with rectal cancer. HSP27 levels merit evaluation as a stratification factor for treatment of rectal cancer.

Proteomic Profiling of Plasma-Derived Biomarkers in Patients with Bladder Cancer: A Step towards Clinical Translation.

BACKGROUND: Bladder cancer is a life-threatening disease and a major cause of cancer-associated complications. The main challenges confronted during the clinical management of bladder cancer are associated with recurrence and disease progression to the muscle-invasive phenotype. Improved early detection of the disease is of paramount importance to prevent disease progression and improve survival. Hence, novel clinically applicable biomarkers for early detection are warranted. METHODS: In the current study, a comparative proteomic approach was undertaken using plasma samples to identify protein biomarkers associated with the muscle-invasive phenotype of bladder carcinoma. Isolated plasma proteins were depleted, DIGE-labeled, then subjected to conventional 2D electrophoresis followed by mass spectrometry for identification of differentially expressed proteins. Western blot was used for data validation. RESULTS: Fourteen differentially expressed proteins with statistically significant changes in abundance between the cancer group and control group were identified. Three differentially expressed proteins were selected for validation, among which apolipoprotein A1 exhibited high specificity and sensitivity (AUC = 0.906). Ingenuity pathway analysis identified IFN-γ and TNF-α as the main signaling hub for the differentially regulated proteins. CONCLUSION: Our findings provide additional insight into understanding bladder cancer pathogenesis. Our data identified potential non-invasive plasma-derived biomarker proteins that merit additional investigation to validate its clinical usefulness to prevent bladder cancer progression.

Assessment of prognostic value of tissue inhibitors of metalloproteinase 3 (TIMP3) protein in ovarian cancer.

Background: TIMP3 is a multifunctional proteolytic enzyme belonging to TIMPs family and acts as a potent inhibitor of matrix metalloproteinases (MMPs). TIMP3 possesses a tumor suppresive function by directly promoting tumor cell apoptosis, preventing angiogenesis and extracellular matrix remodelling. The lower expression of TIMP3 was associated with poor prognosis and overall survival in various cancer types. The aim of this study was to evaluate the association of TIMP3 protein expression with ovarian cancer (OC) clinicopathological features and survival outcomes.Patients and Methods: One hundred forty four of OC FFPE samples were collected from King Abdulaziz University Hospital, Saudi Arabia and constructed in tissue microarray (TMA) slides. Automated Ventana immunostainer platform was used to evaluate TIMP3 protein expression patterns.Results: The study showed that TIMP3 exhibits cytoplasmic localisation. This TIMP3 protein expression was not associated with age, tumor size and the involvement of lymph nodes (p > 0.05). However, it was significantly correlated with tumor stage (p < 0.05) and borderline significant with endpoint status (p = 0.07). Interestingly, the Kaplan-Meier analysis of disease specific survival (DSS) outcomes showed a significant association (p = 0.02, log rank) between OC patients with higher TIMP3 expression compared to those with lower expression. In fact, OC patients with high TIMP3 expression had longer survivals. Multivariate Cox's regression analysis suggests that low TIMP3 protein expression pattern is an independent poor survival marker (p = 0.025).Conclusion: Cytoplasmic TIMP3 protein expression could be used as a good prognosticator to stratify poorly prognostic OC patients in order to personlaize their disease management.

Smad4 loss is associated with fewer S100A8-positive monocytes in colorectal tumors and attenuated response to S100A8 in colorectal and pancreatic cancer cells.

S100A8 and its dimerization partner S100A9 are emerging as important chemokines in cancer. We previously reported that Smad4-negative pancreatic tumors contain fewer stromal S100A8-positive monocytes than their Smad4-positive counterparts. Here, we studied S100A8/A9-expressing cells in colorectal tumors relating their presence to clinicopathological parameters and Smad4 status. Two-dimensional gel electrophoresis (n = 12) revealed variation in the levels of S100A8 protein in colorectal cancer tumors, whereas immunohistochemical analysis (n = 313) showed variation in the numbers of stromal S100A8-positive and S100A9-positive cells. Loss of Smad4 expression was observed in 42/304 (14%) colorectal tumors and was associated with reduced numbers of S100A8-positive (P = 0.03) but not S100A9-positive stromal cells (P = 0.26). High S100A9 cell counts were associated with large tumor sizes (P = 0.0006) and poor differentiation grade (P = 0.036). However, neither S100A8 nor S100A9 cell counts predicted poor survival, except for patients with Smad4-negative tumors (P = 0.02). To address the impact of environmental S100A8/A9 chemokines on tumor cells, we examined the effects of exogenously added S100A8 and S100A9 proteins on cellular migration and proliferation of colorectal and pancreatic cancer cells. S100A8 and S100A9 enhanced migration and proliferation in Smad4-positive and Smad4-negative cancer cells. However, transient depletion of Smad4 resulted in loss of responsiveness to exogenous S100A8, but not S100A9. S100A8 and S100A9 activated Smad4 signaling as evidenced by phosphorylation of Smad2/3; blockade of the receptor for the advanced glycation end products inhibited this response. In conclusion, Smad4 loss alters the tumor's interaction with stromal myeloid cells and the tumor cells' response to the stromal chemokine, S100A8.