Biology Of Bladder Cancer Research Articles

New and contemporary markers of prognosis in nonmuscle invasive urothelial cancer.

Nonmuscle invasive (NMI) urothelial cancer (UC) is associated with varied biological potential. It is characterized by frequent recurrence and progression, which thus worsens the oncological outcome. Nearly three-quarters of NMI UCs recur within 5 years, whereas half can progress during follow-up. Progression is particularly seen in T1 and carcinoma in situ (CIS). Undoubtedly, NMI UC is one of the most expensive cancers to manage. The European Organisation for Research and Treatment of Cancer (EORTC) risk calculator is a commonly used tool for assessing the recurrence and progression potential of a newly diagnosed cancer. The parameters used in the assessment are tumor size and number, pathological stage and grade of the cancer, presence of CIS, and prior recurrence rate. The main advantages of the EORTC tool are its ease of use and the lack of need to run expensive molecular tests. However, reproducibility of pathologic stage and grade is modest, which is a concern to clinicians. Molecular markers have potential for predicting the clinical outcome of NMI UC, given that clinico-pathologic variables are not sufficient for prediction of prognosis in an individual. Significant work has been done in the past 2 decades in understanding the molecular biology of bladder cancer; however, the translational value of this knowledge remains poor. The role for molecular markers in predicting recurrence seems limited because multifocal disease and incomplete treatment are probably more important for recurrence than the molecular features of a resected tumor. Urinary markers have very limited value in prognostication of bladder cancer and are used (mainly as an adjunct to cytology) for detection and surveillance of urothelial cell cancer recurrence. Prediction of progression with molecular markers holds considerable promise. Nevertheless, the contemporary value of molecular markers over clinico-pathologic indexes is limited.

Molecular biology of bladder cancer: new insights into pathogenesis and clinical diversity.

Urothelial carcinoma of the bladder comprises two long-recognized disease entities with distinct molecular features and clinical outcome. Low-grade non-muscle-invasive tumours recur frequently but rarely progress to muscle invasion, whereas muscle-invasive tumours are usually diagnosed de novo and frequently metastasize. Recent genome-wide expression and sequencing studies identify genes and pathways that are key drivers of urothelial cancer and reveal a more complex picture with multiple molecular subclasses that traverse conventional grade and stage groupings. This improved understanding of molecular features, disease pathogenesis and heterogeneity provides new opportunities for prognostic application, disease monitoring and personalized therapy.

Regulation of inflammation and proliferation of human bladder carcinoma cells by type-1 and type-2 cannabinoid receptors.

Pro-inflammatory cytokines, growth and angiogenic factors released by leukocytes are involved in carcinogenesis and cancer progression, but they are also crucial for fighting tumour growth and spreading. We have previously demonstrated that endocannabinoids modulate cell-to-cell crosstalk during inflammation. Here, we investigated the inflammatory and tumourigenic properties of endocannabinoids in a human urinary bladder carcinoma cell line. Endocannabinoid-treated ECV304 cells were checked for tumour necrosis factor (TNF)-α secretion (by ELISA assay) and surface exposure of selectins (by in situ ELISA and FACS analysis). ECV304/Jurkat T cell interaction was assessed by adhesion and live imaging experiments. Proliferation rate, cell death and cell cycle were determined by FACS analysis. By binding to type-1 (CB1) and type-2 (CB2) cannabinoid receptors, the endocannabinoid 2-arachidonoylglycerol (2-AG) exacerbates the pro-inflammatory status surrounding bladder carcinoma ECV304 cells, by: (i) enhancing TNF-α release, (ii) increasing surface exposure of P- and E-selectins, and (iii) allowing Jurkat T lymphocytes to adhere to treated cancer cells. We also found that the CB1 inverse agonist AM281, unlike 2-AG, decreases cancer proliferation by delaying cell cycle progression. Our data suggest that 2-AG modulates the inflammatory milieu of cancer cells in vitro, while AM281 plays a more specific role in proliferation. Collectively, these findings suggest that CB receptors may play distinct roles in cancer biology, depending on the specific ligand employed. The in vivo assessment of the role of CB receptors in inflammation and cancer might be instrumental in broadening the understanding about bladder cancer biology.

Abstract 474: Changes in global chromatin landscape identify bladder cancer progression

Abstract Bladder cancer is the 4th most common cancer in men in the US and metastatic disease has a poor prognosis. To date, only few mutations in cancer driving genes have been identified in this tumor. Altered gene expression involving an estimate of >2,300 coding sequences is reported in high-grade bladder cancer. In addition to previously reported mutations in KRAS, FGFR3 and TPp53, recent data showed frequent deletions and mutations in genes involving several chromatin modifiers, such as UTX (KDM6A). These data indicate that chromatin organization may play an important role in bladder cancer development and progression. The process of in vivo passaging of a poorly tumorigenic bladder cells (T24) with subsequent injection into mice and selection for tumorigenic and metastatic phenotype specifically to the lung and liver, provides a unique opportunity for analysis of mechanisms associated with tumor development and progression. We compared whole exome sequencing and mRNA expression with analysis of DNase I hypersensitivity combined with deep sequencing (DHS-seq) on multiple cell lines developed by in vivo selection. Bioinformatics of DHS-seq was performed using algorithms developed at LRBGE. Analysis of microarray expression was performed using Ingenuity Pathways (IPA). Mutations in several genes, including TP53, RAS, TERT and chromatin remodeler UTX (KDM6A), were detected in parent cell line, T24. Only few additional mutations marked the progression to the metastatic phenotype. Changes in gene expression were already evident in progression from parental cells that were weakly tumorigenic (T24) to cells that were capable to form tumors in mice, but rarely formed metastasis (T24T). The highest score was assigned to genes involved in Cell-to-Cell interaction. Prominent changes included genes involved in cell adhesion and genes associated with EMT, followed by changes in genes associated with cancer. Thus, during the progression from weakly tumorigenic to fully tumorigenic but weakly metastatic cell type, bladder cancer cells acquired gene expression and chromatin landscape changes associated with tumor formation and metastasis. Gene expression profile for most genes correlated with nearby chromatin remodeling of regulatory regions. In addition, DHS analysis identified many novel areas of changes, termed “hot spots” which were located in upstream or downstream enhancer regions. Some of these changes were unique for each stage of tumor progression. Genome-wide analysis of DHS showed large-scale changes in chromatin landscape during tumorigenesis indicating massive reprogramming of regulatory networks during cancer development and progression. Thus, combination of exome sequencing with microarray gene expression and analysis of global unbiased chromatin landscape provides valuable new information on bladder cancer biology and tumor progression. Citation Format: Sohyoung Kim, Lyuba Varticovski, Lars Grontved, Songjoon Baek, Bethrice Thompson, Michael Dean, Daniel Theodoresco, Michael L. Nickerson, Gordon L. Hager. Changes in global chromatin landscape identify bladder cancer progression. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 474. doi:10.1158/1538-7445.AM2014-474

MP21-17 EFFECTS OF ADIPOSE TISSUE STROMAL CELLS ON APOPTOSIS, GROWTH AND INVASION OF SUPERFICIAL AND INVASIVE TYPE UROTHELIAL CARCINOMAS OF THE BLADDER

INTRODUCTION AND OBJECTIVES: Cancer-stromal interaction is critical for cancer biology. Bladder cancer develops within the mucosa, and invades into muscle layer surrounded by abundant adipose tissue. Adipose tissue has stromal cell types, such as preadipocytes and mesenchymal stem cells (MSCs). Thus, the adipose tissue stromal cells (ATSCs) seem critical for bladder cancer biology. The aim of this study is to address the effects of ATSCs on the apoptosis, growth and invasion of the superficial and invasive types. METHODS: RT4 and RT112 were used as human superficial urothelial carcinoma type of bladder, while EJ and HT1376 were done as invasive type. ATSCs were isolated from subcutaneous adipose tissue of 1-week old male Wister rats. We examined the apoptosis, growth and invasion of cancer cells, using collagen gel invasion assay system, in which the tumor cells were co-cultured on ATSC-embedded gel layer. The cellular behavior, and the apoptosis-, growthand ivasion-related molecules were analyzed by morphometry, immunohistochemistry, Western blot and real-time RT-PCR. RESULTS: ATSCs promoted and inhibited the apoptosis and growth of superficial type cancer cells, respectively. In contrast, ATSCs inhibited and promoted the apoptosis and growth of invasive type cancer cells. ATSCs had no effect on the invasion of superficial type, whereas they promoted the invasion of invasive type. ATSCs promoted the expression of MAPK proteins (raf-1, MEK-1 and ERK1/ERK2) in all cancer cell types. In turn, superficial type promoted [alpha]-SMA-positive myofibroblastic differentiation in ATSCs, while invasive type did lipid dropletand S-100 protein-positive adipogenic differentiation in ATSCs. CONCLUSIONS: The data suggest the following issues: 1) ATSCs suppress the progression of superficial type urothelial carcinoma through the growth inhibition and apoptosis promotion, whereas ATSCs promote that of invasive type through the enhancement of the growth and invasion, and the apoptosis prohibition; 2) ATSCinduced MAPK pathway activation plays differential roles in biological behavior of superficial and invasive types; and 3) Superficial type induces myofibroblastic differentiation in ATSCs as cancer-associated phenotype, while invasive type does adipogenic differentiation in ATSCs. Source of Funding: none

A Cell of Origin Gene Signature Indicates Human Bladder Cancer Has Distinct Cellular Progenitors

There are two distinct forms of urothelial (bladder) cancer: muscle-invasive (MI) and nonmuscle invasive (NMI) disease. Since it is currently believed that bladder cancer arises by transformation of urothelial cells of the basal layer, bladder cancer stem cells (CSCs) have been isolated based on expression markers found in such cells. However, these CSCs have only been identified in MI tumors raising the intriguing hypothesis that NMI tumor progenitors do not arise from the basal compartment. To test this hypothesis, we carried out genome-wide expression profiling of laser capture microdissected basal and umbrella cells, the two most histologically distinct cell types in normal urothelium and developed a cell of origin (COO) gene signature that distinguishes these. The COO signature was a better predictor of stage and survival than other bladder, generic, or breast CSC signatures and bladder cell differentiation markers in multiple patient cohorts. To assess whether NMI and MI tumors arise from a distinct progenitor cell (DPC) or common progenitor cell, we developed a novel statistical framework that predicts COO score as a function of known genetic alterations (TP53, HRAS, KDM6A, and FGFR3) that drive either MI or NMI bladder cancer and compared this to the observed COO score of the tumor. Analysis of 874 patients in five cohorts established the DPC model as the best fit to the available data. This observation supports distinct progenitor cells in NMI and MI tumors and provides a paradigm shift in our understanding of bladder cancer biology that has significant diagnostic and therapeutic implications.

Urinary Metabotyping of Bladder Cancer Using Two-Dimensional Gas Chromatography Time-of-Flight Mass Spectrometry

Cystoscopy is the gold standard clinical diagnosis of human bladder cancer (BC). As cystoscopy is expensive and invasive, it compromises patients' compliance toward surveillance screening and challenges the detection of recurrent BC. Therefore, the development of a noninvasive method for the diagnosis and surveillance of BC and the elucidation of BC progression become pertinent. In this study, urine samples from 38 BC patients and 61 non-BC controls were subjected to urinary metabotyping using two-dimensional gas chromatography time-of-flight mass spectrometry (GC×GC-TOFMS). Subsequent to data preprocessing and chemometric analysis, the orthogonal partial least-squares discriminant analysis (OPLS-DA, R2X=0.278, R2Y=0.904 and Q2Y (cumulative)=0.398) model was validated using permutation tests and receiver operating characteristic (ROC) analysis. Marker metabolites were further screened from the OPLS-DA model using statistical tests. GC×GC-TOFMS urinary metabotyping demonstrated 100% specificity and 71% sensitivity in detecting BC, while 100% specificity and 46% sensitivity were observed via cytology. In addition, the model revealed 46 metabolites that characterize human BC. Among the perturbed metabolic pathways, our clinical finding on the alteration of the tryptophan-quinolinic metabolic axis in BC suggested the potential roles of kynurenine in the malignancy and therapy of BC. In conclusion, global urinary metabotyping holds potential for the noninvasive diagnosis and surveillance of BC in clinics. In addition, perturbed metabolic pathways gleaned from urinary metabotyping shed new and established insights on the biology of human BC.

Toward a Molecular Pathologic Classification of Urothelial Carcinoma

We recently defined molecular subtypes of urothelial carcinomas according to whole genome gene expression. Herein we describe molecular pathologic characterization of the subtypes using 20 genes and IHC of 237 tumors. In addition to differences in expression levels, the subtypes show important differences in stratification of protein expression. The selected genes included biological features central to bladder cancer biology, eg, cell cycle activity, cellular architecture, cell-cell interactions, and key receptor tyrosine kinases. We show that the urobasal (Uro) A subtype shares features with normal urothelium such as keratin 5 (KRT5), P-cadherin (P-Cad), and epidermal growth factor receptor (EGFR) expression confined to basal cells, and cell cycle activity (CCNB1) restricted to the tumor-stroma interface. In contrast, the squamous cell cancer-like (SCCL) subtype uniformly expresses KRT5, P-Cad, EGFR, KRT14, and cell cycle genes throughout the tumor parenchyma. The genomically unstable subtype shows proliferation throughout the tumor parenchyma and high ERBB2 and E-Cad expression but absence of KRT5, P-Cad, and EGFR expression. UroB tumors demonstrate features shared by both UroA and SCCL subtypes. A major transition in tumor progression seems to be loss of dependency of stromal interaction for proliferation. We present a simple IHC/histology-based classifier that is easy to implement as a standard pathologic evaluation to differentiate the three major subtypes: urobasal, genomically unstable, and SCCL. These three major subtypes exhibit important prognostic differences.

Abstract 1922: Plasminogen activator inhibitor type-1 (PAI-1) enhances bladder cancer tumor progression.

Abstract Introduction: Bladder cancer (BCa) is among the five most common malignancies worldwide and has the highest recurrence rate of any tumor type. BCa biology is poorly understood and early detection remains one of the most urgent issues in BCa research. Previously we have identified a preliminary molecular signature of BCa in voided urine samples from BCa patients. Our results revealed aberrant overexpression of the serine protease inhibitor (serpin) plasminogen activator inhibitor type-1 (PAI-1), both at the genomic and protein level. Increasing evidence indicate that elevated levels of PAI-1 correlates to a poor prognosis in many tumor types including breast, colorectal, gastric, ovarian, and more recently bladder cancer. PAI-1 has emerged as a potential target for BCa treatment and the aim of this study is to provide a greater understanding of its role in bladder cancer progression. Methods: PAI-1 expression levels in UROtsa (benign urothelial cell), KU7 (low-grade papillary urothelial cancer cell), and UMUC14 and T24 (high-grade invasive urothelial cancer cell lines) was tested by real-time PCR (RTPCR) and Western blot analysis. Secreted PAI-1 levels were measured by ELISA. KU7 stable clones overexpressing PAI-1 was established by transfection of plasmid DNA encoding human PAI-1. T24 and UMUC14 stable clones with shRNA knockdown of PAI-1 were established by infection with retroviral particles. Cell proliferation, migration, invasion, and apoptosis assays were performed in stable clones. Results: RTPCR analysis revealed an increase of PAI-1 in T24 and UMUC14 cells (27.9 and 5.2 times, respectively) while KU7 cells exhibited significantly reduced levels as compared to the UROtsa control. Similar results were observed for PAI-1 protein concentrations in cell culture supernatants and western blot analysis. While overexpression of PAI-1 in KU7 cells promoted an increase in proliferation, migration and invasion, these were significantly reduced upon inactivation of PAI-1 in T24 and UMUC14 cells stably expressing shRNA PAI-1. Loss of PAI-1 function also led to a substantial increase in apoptotic cell death. Conclusions: These results suggest that PAI-1 overexpression in BCa promotes tumor progression and therapeutic strategies targeting PAI-1 may serve clinically beneficial outcomes. Funding provided by research grants from Florida Department of Health James and Esther King Team Science Award 10KT-01 (CJR) and National Cancer Institute RO1 CA116161 (SG). Citation Format: Evan Gomes, Steven Goodison, Makito Miyake, Shanti Ross, Ge Zhang, Charles Rosser. Plasminogen activator inhibitor type-1 (PAI-1) enhances bladder cancer tumor progression. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 1922. doi:10.1158/1538-7445.AM2013-1922

Abstract ADV09: Bladder cancer basics.

Abstract The Bladder Cancer Advocacy Network's (BCAN) mission is to increase public awareness about bladder cancer, to advance bladder cancer research, and to provide educational and support services for the bladder cancer community. BCAN works to serve over 520,000 bladder cancer survivors and 73,000 newly diagnosed patients each year in the United States. BCAN engages in many initiatives and programs. BCAN's current initiatives include the Women's Outreach Initiative and Veterans’ Outreach Initiative, which are engaging national task forces to raise awareness of bladder cancer among these constituencies. In patient education and support, BCAN's website: www/bacan.org- is one of the most comprehensive bladder cancer resources on the internet. BCAN also provides free bladder cancer patient education materials including patient tip sheets and a patient handbook-Bladder Cancer Basics for the Newly Diagnosed, which was revised this year and is now available in Spanish. BCAN offers Patient Insight Webinars throughout the year. BCAN also provides a free clinical trial matching service in coordination with Emerging Med, to help people obtain specific information about participating in bladder cancer research trials. BCAN staff answers patient calls and e-mails for more information, resources, and support. BCAN has facilitated the development of in-person bladder cancer support groups around the country and offers access to an online bladder cancer support community. BCAN also offers the Young Investigator Awards, announced this year, for three awards of $100,000 over two years, to increase the understanding of bladder cancer risk, biology, prevention, and treatment while encouraging young investigators to pursue a career in bladder cancer research. BCAN is also engaged in advocacy with staff members and volunteers serving on panels and participating in coalitions. BCAN has coordinated Walks for Bladder Cancer to raise awareness of the disease since 2011, on the first Saturday in May. In 2012, the Walks for Bladder Cancer held on May 5 took place in 60 communities, had more than 5,000 participants and raised more than $275,000. Citation Format: Vanessa Hoffman. Bladder cancer basics. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr ADV09. doi:10.1158/1538-7445.AM2013-ADV09

TSP-1-1223 A/G Polymorphism as a Potential Predictor of the Recurrence Risk of Bladder Cancer in a Chinese Population

Backgrounds. TSP-1 is a glycoprotein that functions in the biology of bladder cancer. We investigated the relationship between the distribution of TSP-1-1223 A/G polymorphism (rs2169830) and the clinical characteristics of bladder cancer. Materials and Methods. TaqMan assay was performed to determine the genotype of 609 cases and 670 control subjects in a Chinese population. Logistic regression was used to assess the association between the polymorphism and the risk of bladder cancer. Quantitative real-time polymerase chain reaction was performed to determine TSP-1 mRNA expression. Survival curves were generated using the Kaplan-Meier method. Results. No significant differences were detected in the genotype frequencies of healthy control subjects and patients with bladder cancer. By contrast, the time until the first recurrence differed significantly between genotypes (P = 0.017). The expression of TSP-1 mRNA in bladder cancer tissues was lower in patients with an AG genotype than in those with an AA genotype. The lowest expression was observed in patients with a GG genotype. Conclusions. In conclusion, TSP-1-1223 A/G polymorphism may contribute to the recurrence of bladder cancer in Chinese population.

Integrated Epigenome Profiling of Repressive Histone Modifications, DNA Methylation and Gene Expression in Normal and Malignant Urothelial Cells

Epigenetic regulation of gene expression is commonly altered in human cancer. We have observed alterations of DNA methylation and microRNA expression that reflect the biology of bladder cancer. This common disease arises by distinct pathways with low and high-grade differentiation. We hypothesized that epigenetic gene regulation reflects an interaction between histone and DNA modifications, and differences between normal and malignant urothelial cells represent carcinogenic events within bladder cancer. To test this we profiled two repressive histone modifications (H3K9m3 and H3K27m3) using ChIP-Seq, cytosine methylation using MeDIP and mRNA expression in normal and malignant urothelial cell lines. In genes with low expression we identified H3K27m3 and DNA methylation each in 20–30% of genes and both marks in 5% of genes. H3K9m3 was detected in 5–10% of genes but was not associated with overall expression. DNA methylation was more closely related to gene expression in malignant than normal cells. H3K27m3 was the epigenetic mark most specifically correlated to gene silencing. Our data suggest that urothelial carcinogenesis is accompanied by a loss of control of both DNA methylation and H3k27 methylation. From our observations we identified a panel of genes with cancer specific-epigenetic mediated aberrant expression including those with reported carcinogenic functions and members potentially mediating a positive epigenetic feedback loop. Pathway enrichment analysis revealed genes marked by H3K9m3 were involved with cell homeostasis, those marked by H3K27m3 mediated pro-carcinogenic processes and those marked with cytosine methylation were mixed in function. In 150 normal and malignant urothelial samples, our gene panel correctly estimated expression in 65% of its members. Hierarchical clustering revealed that this gene panel stratified samples according to the presence and phenotype of bladder cancer.

Hypermethylation in bladder cancer: biological pathways and translational applications

A compelling body of evidences sustains the importance of epigenetic mechanisms in the development and progression of cancer. Assessing the epigenetic component of bladder tumors is strongly improving our understanding of their biology and clinical behavior. In terms of DNA methylation, cancer cells show genome-wide hypomethylation and site-specific CpG island promoter hypermethylation. In the context of other epigenetic alterations, this review will focus on the hypermethylation of CpG islands in promoter regions, as the most widely described epigenetic modification in bladder cancer. CpG islands hypermethylation is believed to be critical in the transcriptional silencing and regulation of tumor suppressor and crucial cancer genes involved in the major molecular pathways controlling bladder cancer development and progression. In particular, several biological pathways of frequently methylated genes include cell cycle, DNA repair, apoptosis, and invasion, among others. Furthermore, translational aspects of bladder cancer methylomes described to date will be discussed towards their potential application as bladder cancer biomarkers. Several tissue methylation signatures and individual candidates have been evidenced, that could potentially stratify tumors histopathologically, and discriminate patients in terms of their clinical outcome. Tumor methylation profiles could also be detected in urinary specimens showing a promising role as non-invasive markers for cancer diagnosis towards an early detection and potentially for the surveillance of bladder cancer patients in a near future. However, the epigenomic exploration of bladder cancer has only just begun. Genome-scale DNA methylation profiling studies will further highlight the relevance of the epigenetic component to gain knowledge of bladder cancer biology and identify those profiles and candidates better correlating with clinical behavior.

Progress in Personalizing Chemotherapy for Bladder Cancer

Platinum-based chemotherapy is commonly used for the treatment of locally advanced and metastatic bladder cancer. However, there are currently no methods to predict chemotherapy response in this disease setting. A better understanding of the biology of bladder cancer has led to developments of molecular biomarkers that may help guide clinical decision making. These biomarkers, while promising, have not yet been validated in prospective trials and are not ready for clinical applications. As alkylating agents, platinum drugs kill cancer cells mainly through induction of DNA damage. A microdosing approach is currently being tested to determine if chemoresistance can be identified by measuring platinum-induced DNA damage using highly sensitive accelerator mass spectrometry technology. The hope is that these emerging strategies will help pave the road towards personalized therapy in advanced bladder cancer.

Current therapeutic strategies for invasive and metastatic bladder cancer.

BackgroundBladder cancer is one of the most common cancers in Europe, the United States, and Northern African countries. Muscle-invasive bladder cancer is an aggressive epithelial tumor, with a high rate of early systemic dissemination. Superficial, noninvasive bladder cancer can most often be cured; a good proportion of invasive cases can also be cured by a combined modality approach of surgery, chemotherapy, and radiation. Recurrences are common and mostly manifest as metastatic disease. Those with distant metastatic disease can sometime achieve partial or complete remission with combination chemotherapy.Recent developmentsBetter understanding of the biology of the disease has led to the incorporation of molecular and genetic features along with factors such as tumor grade, lympho-vascular invasion, and aberrant histology, thereby allowing identification of ‘favorable’ and ‘unfavorable’ cancers which helps a more accurate informed and objective selection of patients who would benefit from neoadjuvant and adjuvant chemotherapy. Gene expression profiling has been used to find molecular signature patterns that can potentially be predictive of drug sensitivity and metastasis. Understanding the molecular pathways of invasive bladder cancer has led to clinical investigation of several targeted therapeutics such as anti-angiogenics, mTOR inhibitors, and anti-EGFR agents.ConclusionWith improvements in the understanding of the biology of bladder cancer, clinical trials studying novel and targeted agents alone or in combination with chemotherapy have increased the armamentarium for the treatment of bladder cancer. Although the novel biomarkers and gene expression profiles have been shown to provide important predictive and prognostic information and are anticipated to be incorporated in clinical decision-making, their exact utility and relevance calls for a larger prospective validation.

Impact of gender on bladder cancer incidence, staging, and prognosis

While patient gender is an important factor in the clinical decision-making for the management of bladder cancer, there are minimal evidence-based recommendations to guide health care professionals. Recent epidemiologic and translational research has shed some light on the complex relationship between gender and bladder cancer. Our aim was to review the literature on the effect of gender on bladder cancer incidence, biology, mortality, and treatment. Using MEDLINE, we performed a search of the literature between January 1975 and April 2011. Although men are nearly 3-4 times more likely to develop bladder cancer than women, women present with more advanced disease and have worse survival. Recently, a number of population-based and multicenter collaborative studies have shown that female gender is associated with a significantly higher rate of cancer-specific recurrence and mortality after radical cystectomy. The disparity between genders is proposed to be the result of a differences exposure to carcinogens (i.e., tobacco and chemicals) as well as reflective of genetic, anatomic, hormonal, societal, and environmental factors. Explanations for the differential behavior of bladder cancer between genders include sex steroids and their receptors as well as inferior quality of care for women (inpatient length of stay, referral patterns, and surgical outcomes). It is imperative that health care practitioners and researchers from disparate disciplines collectively focus efforts to appropriately develop gender-specific evidence-based guidelines for bladder cancer patients. We must strive to develop multidisciplinary collaborative efforts to provide tailored gender-specific care for bladder cancer patients.

Abstract 3910: An integrated genomic and epigenomic approach for identification of novel disease-related regions and genes in bladder cancer progression

Abstract Different pathways drive bladder cancer distinct presentations, such as superficial papillary (Ta) when compared to carcinoma in situ (Cis) or muscle invasive tumors. Although multiple chromosomal alterations have been identified, the specific genes and loci driving the distinct pathways remain largely unknown. In part this uncertainty is due to the limited resolution of the techniques used to detect these alterations. In this study, we applied a high-resolution genome-wide method, Affymetrix 250K Nsp1 SNP mapping arrays, to screen for somatic DNA copy number variation (CNV) among 49 bladder tumors [17 (35%) clinical stage 1 (15 Ta and 2 Cis), 11 (23%) stage 2, 15 (31%) stage 3 and 6 to be classified] samples and corresponding normals (lymphocytes from the same patient). GeneChip Genotyping analysis software (GTYPE v4.0), Partek Genomic SuiteTM and Hidden Markov model assuming diploid copy for normal samples were employed to infer the CNV estimates. We utilized a genomic segmentation algorithm (requiring a minimum of 10 probes) to characterize significantly higher or lower intensity ratios into cancer specific CNV regions. A threshold of > 3 (gains, amplifications) and < 1.25 (losses, deletions) was used to categorize altered regions. Genome Wide CNV analysis were determined in different groups of comparison (tumor versus normal, superficial vs muscle-invasive stages, Ta vs Cis). Furthermore, to identify the genes that are inactivated one allele by methylation and the other allele by deletion, we performed genome-wide methylation analysis on a subset of these samples by Illumina Methylation Bead Array containing over 27,000 CpG sites, focused on the promoter regions of 14,495 genes. Significant major alterations were seen on chromosome 6q, 8p, 8q, 9p, 9q, 5p, 12p, 12q, 18p, 18q and 20q. We also identified novel small deletions and amplifications throughout the genome. Amplification of loci 6q21-22 (NKAIN2 and FRK), 9q22 (PHF2) and 11p15 (OR51F2) and deletion of loci 11p15 (MRGPRX1), 16p23 (ADAMTS18), 11q15 (MCT2) and 14q11 (DAD1) were observed in superficial Ta compared to Cis. Interestingly, among the 366 CpG probes classified as significantly methylated in the bladder cancer samples, ADAMTS18 was also present. This gene is a disintegrin and metalloproteinase with thrombospondin motifs and has been reported to be epigenetically inactivated in tumors other than bladder cancer, including nasopharyngeal, esophageal, gastric, colorectal and pancreatic cancers. Genetic alterations have also being described: deletions in breast cancer as well as somatic mutations in melanoma. Our initial findings strengthen ADAMTS18 potential's of being a candidate gene related to bladder tumorigenesis. The integration of genetic and epigenetic alterations may lead to a comprehensive knowledge of the biology of bladder cancer and will open up fertile avenue for further research. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 3910. doi:10.1158/1538-7445.AM2011-3910

Expression of genes related to apoptosis, cell cycle and signaling pathways are independent of TP53 status in urinary bladder cancer cells

Urinary bladder cancer is the fourth most common malignancy in the Western world. Transitional cell carcinoma (TCC) is the most common subtype, accounting for about 90% of all bladder cancers. The TP53 gene plays an essential role in the regulation of the cell cycle and apoptosis and therefore contributes to cellular transformation and malignancy; however, little is known about the differential gene expression patterns in human tumors that present with the wild-type or mutated TP53 gene. Therefore, because gene profiling can provide new insights into the molecular biology of bladder cancer, the present study aimed to compare the molecular profiles of bladder cancer cell lines with different TP53 alleles, including the wild type (RT4) and two mutants (5637, with mutations in codons 280 and 72; and T24, a TP53 allele encoding an in-frame deletion of tyrosine 126). Unsupervised hierarchical clustering and gene networks were constructed based on data generated by cDNA microarrays using mRNA from the three cell lines. Differentially expressed genes related to the cell cycle, cell division, cell death, and cell proliferation were observed in the three cell lines. However, the cDNA microarray data did not cluster cell lines based on their TP53 allele. The gene profiles of the RT4 cells were more similar to those of T24 than to those of the 5637 cells. While the deregulation of both the cell cycle and the apoptotic pathways was particularly related to TCC, these alterations were not associated with the TP53 status.

Metabolic footprinting of tumorigenic and nontumorigenic uroepithelial cells using two-dimensional gas chromatography time-of-flight mass spectrometry

In this study, gas chromatography mass spectrometry (GC-MS) and two-dimensional gas chromatography time-of-flight mass spectrometry (GCxGC-TOFMS) were employed for the metabolic footprinting of a pair of immortalized human uroepithelial cells namely HUC-1 (nontumorigenic) and HUC T-2 (tumorigenic). Both HUC-1 and HUC T-2 cell lines were cultivated in 1 mL of Ham's F-12 media. Subsequent to 48 h of incubation, 200 microL of cell culture supernatant was protein-precipitated using 1.7 mL of methanol and an aliquot of 1.5 mL of the mixture was separated, dried, trimethylsilyl-derivatized, and analyzed using GC-MS and GCxGC-TOFMS. Metabolic profiles were analyzed using multivariate data analysis techniques to evaluate the changes of the metabolomes. Both GC-MS and GCxGC-TOFMS analyses showed distinct differences in metabolic phenotypes of the normal and tumorigenic human bladder cells (partial least squares-discriminant analysis (PLS-DA) of GCxGC-TOFMS data; two latent variables, R (2) X = 0.418, R (2) Y = 0.977 and Q (2) (cumulative) = 0.852). Twenty metabolites were identified as being statistically different between the two cell types. These metabolites revealed that several key metabolic pathways were perturbed in tumorigenic urothelial cells as compared to the normal cells. Application of GCxGC-TOFMS offered several advantages compared to classical one-dimensional GC-MS which include enhanced chromatographic resolution (without increase in analytical run time), increase in sensitivity, improved identification of metabolites, and also separation of reagent artifacts from the metabolite peaks. Our results reinforced the advantages of GCxGC-TOFMS and the role of metabolomics in characterizing bladder cancer biology using in vitro cell culture models.

Nouvelles molécules dans le cancer de la vessie

Cisplatin-based chemotherapy (MVAC : cisplatin, methotrexate, adriamycin, vinblastine ; or GC : cisplatin, gemcitabine) has been the standard of care for patients with advanced urothelial tumor during the last twenty years. Greater knowledge in the molecular biology of bladder cancer lead to the identification of promising target such as EGFR, HER2, or VEGF-VEGFR pathways. The role of targeted therapies as monotherapy, in combination with chemotheray or as maintenance post-chemotherapy is currently under study.