Rapid developments are expected in the molecular pharmacology of both B1, and B2 types of kinin receptors, since the underlying genetic structures are now known and widely studied. The consequences of kinin receptor duality and physiopathological regulation have not yet been fully appreciated. Medicinal chemistry is also an active front of research in kinin pharmacology, as more effective drugs targeted at kinin receptors are regularly reported. Various complementary molecular approaches (the receptor binding, cloning, immunoreacting, mutagenesis, inactivation, the study of regulation, allelic polymorphisms, and so forth) are expanding our knowledge of the role of kinins in allergy, inflammation, and singularly, renal medicine.

BackgroundOvarian carcinoma is the most lethal gynecological malignancy due to early dissemination and acquired resistance to platinum-based chemotherapy. Reliable markers that are independent and complementary to clinical parameters are needed to improve the management of patients with this disease. The Canadian Ovarian Experimental Unified Resource (COEUR) provides researchers with biological material and associated clinical data to conduct biomarker validation studies. Using standards defined by the Canadian Tissue Repository Network (CTRNet), we have previously demonstrated the quality of the biological material from this resource. Here we describe the clinical characteristics of the COEUR cohort.MethodsWith support from 12 Canadian ovarian cancer biobanks in Canada, we created a central retrospective cohort comprised of more than 2000 patient tissue samples with associated clinical data, including 1246 high-grade serous, 102 low-grade serous, 295 endometrioid, 259 clear cell and 89 mucinous carcinoma histotypes. A two-step reclassification process was applied to assure contemporary histological classification (histotyping). For each histotypes individually, we evaluated the association between the known clinico-pathological parameters (stage, cytoreduction, chemotherapy treatment, BRCA1 and BRCA2 mutation) and patient outcome by using Kaplan-Meier and Cox proportional hazard regression analyses.ResultsThe median follow-up time of the cohort was 45 months and the 5-year survival rate for patients with high-grade serous carcinomas was 34%, in contrast to endometrioid carcinomas with 80% at 5 years. Survival profiles differed by histotype when stratified by stage or cytoreduction. Women with mucinous or clear cell carcinomas at advanced stage or with non-optimally debulked disease had the worst outcomes. In high-grade serous carcinoma, we observed significant association with longer survival in women harboring BRCA1 or BRCA2 mutation as compared to patients without detectable mutation.ConclusionsOur results show the expected survival rates, as compared with current literature, in each histotype suggesting that the cohort is an unbiased representation of the five major histotypes. COEUR, a one stop comprehensive biorepository, has collected mature outcome data and relevant clinical data in a comprehensive manner allowing stratified analysis.

Sorafenib, a multikinase inhibitor, recently received FDA approval for the treatment of advanced hepatocellular carcinoma (HCC). However, as the clinical application of sorafenib evolves, there is increasing interest in defining the mechanisms underlying its anti-tumor activity. Considering that this specific inhibitor could target unexpected molecules depending on the biologic context, a precise understanding of its mechanism of action could be critical to maximize its treatment efficacy, while minimizing adverse effects. Two human HCC cell lines (HepG2 and Huh7), carrying different biological and genetic characteristics, were used in this study to examine the intracellular events leading to sorafenib-induced HCC cell-growth inhibition. Sorafenib inhibited cell growth in both cell lines in a dose- and time-dependent manner and significantly altered expression levels of 826 and 2011 transcripts in HepG2 and Huh7 cells, respectively. Genes functionally involved in angiogenesis, apoptosis, transcription regulation, signal transduction, protein biosynthesis and modification were predominantly upregulated, while genes implicated in cell cycle control, DNA replication recombination and repair, cell adhesion, metabolism and transport were mainly downregulated upon treatment. However, each sorafenib-treated HCC cell line displayed specificity in the expression and activity of crucial factors involved in hepatocarcinogenesis. The altered expression of some of these genes was confirmed by semiquantitative and quantitative RT-PCR and by western blotting. Many novel genes emerged from our transcriptomics analysis that had not previously been reported to be effected by sorafenib. Further functional analyses may determine whether these genes can serve as potential molecular targets for more effective anti-HCC strategies.

Sorafenib, an oral multikinase inhibitor, is the only approved agent for the treatment of advanced hepatocellular carcinoma (HCC). However, its benefits are modest, and as its mechanisms of action remain elusive, a better understanding of its anticancer effects is needed. Based on our previous study results, we investigated here the implication of the nuclear protein 1 (NUPR1) in HCC and its role in sorafenib treatment. NUPR1 is a stress-inducible protein that is overexpressed in various malignancies, but its role in HCC is not yet fully understood. We found that NUPR1 expression was significantly higher in primary human HCC samples than in the normal liver. Knockdown of NUPR1 significantly increased cell sensitivity to sorafenib and inhibited the cell growth, migration and invasion of HCC cells, both in vitro and in vivo. Moreover, NUPR1 silencing influenced the expression of RELB and IER3 genes. Unsurprisingly, RELB and IER3 knockdown also inhibited HCC cell viability, growth and migration. Using gene expression profiling of HCC cells following stable NUPR1 knockdown, we found that genes functionally involved in cell death and survival, cellular response to therapies, lipid metabolism, cell growth and proliferation, molecular transport and cellular movement were mostly suppressed. Network analysis of dynamic gene expression identified NF-κB and ERK as downregulated gene nodes, and several HCC-related oncogenes were also suppressed. We identified Runt-related transcription factor 2 (RUNX2) gene as a NUPR1-regulated gene and demonstrated that RUNX2 gene silencing inhibits HCC cell viability, growth, migration and increased cell sensitivity to sorafenib. We propose that the NUPR1/RELB/IER3/RUNX2 pathway has a pivotal role in hepatocarcinogenesis. The identification of the NUPR1/RELB/IER3/RUNX2 pathway as a potential therapeutic target may contribute to the development of new treatment strategies for HCC management.

The kallikrein-kinin system, activated during inflammatory conditions and the regulation of specific cardiovascular and renal functions, includes two G protein-coupled receptors for bradykinin (BK)-related peptides. 9 -BK (1-100 nM) rapidly concentrated the receptor-associated fluorescence into multiple aggregates that remained associated with the plasma membrane (no significant internalization) and colocalized with caveolin-1. This reaction was slowly reversible upon agonist washing at 37°C and prevented pretreatment with a B 1 R antagonist. ␤-Cyclodextrin treatment, which extracts cholesterol from membranes and disrupts caveolae-related rafts, prevented agonist-induced redistribution of B 1 R-YFP but not the PLA 2 activation mediated by this receptor. The agonist radioligand copurified with caveolin-1 to a greater extent than the tritiated antagonist in buoyant fractions of HEK 293 cells treated with the ligands. Agonistinduced cellular translocation of the kinin B 1 R to caveolaerelated rafts without endocytosis is a novel variation on the theme of G protein-coupled receptor adaptation.

Histone deacetylases (HDACs) regulate fundamental biological processes such as cellular proliferation, differentiation, and survival via genomic and nongenomic effects. This study examined the importance of HDAC activity in the regulation of gene expression and differentiation of the developing mouse kidney. Class I HDAC1-3 and class II HDAC4, -7, and -9 genes are developmentally regulated. Moreover, HDAC1-3 are highly expressed in nephron precursors. Short term treatment of cultured mouse embryonic kidneys with HDAC inhibitors (HDACi) induced global histone H3 and H4 hyperacetylation and H3K4 hypermethylation. However, genome-wide profiling revealed that the HDAC-regulated transcriptome is restricted and encompasses regulators of the cell cycle, Wnt/␤-catenin, TGF-␤/Smad, and PI3K-AKT pathways. Further analysis demonstrated that base-line expression of key developmental renal regulators, including Osr1, Eya1, Pax2/8, WT1, Gdnf, Wnt9b, Sfrp1/2, and Emx2, is dependent on intact HDAC activity. Treatment of cultured embryonic kidney cells with HDACi recapitulated these gene expression changes, and chromatin immunoprecipitation assays revealed that HDACi is associated with histone hyperacetylation of Pax2/Pax8, Gdnf, Sfrp1, and p21. Gene knockdown studies demonstrated that HDAC1 and HDAC2 play a redundant role in regulation of Pax2/8 and Sfrp1 but not Gdnf. Long term treatment of embryonic kidneys with HDACi impairs the ureteric bud branching morphogenesis program and provokes growth arrest and apoptosis. We conclude that HDAC activity is critical for normal embryonic kidney homeostasis, and we implicate class I HDACs in the regulation of early nephron gene expression, differentiation, and survival.Kidney development depends on reciprocal inductive interactions between the metanephric mesenchyme (MM), 4 a specified region in the caudal intermediate mesoderm, and the ureteric bud (UB), an epithelial outgrowth from the Wolffian (nephric) duct (1-3). Recent years have witnessed significant progress in our understanding of the gene regulatory networks of early kidney development (3-6). For example, the Osr1/ Eya1/Pax2/Six/Sall/WT1/Hoxd11 gene regulatory network specifies the MM and is absolutely required for expression of glia-derived neurotrophic factor (Gdnf) (7,8). Gdnf, in turn, is essential for UB outgrowth and subsequent branching (9 -11).Gdnf acts via activation of a c-Ret/PI3K/ERK-dependent gene network (Wnt11, Spry1, Etv4, Etv5, Cxcr4, Myb, Met, and Mmp14) in UB tip cells to control the branching morphogenesis program (12-14). Various growth factor/receptor signaling pathways, including FGFs, bone morphogenic proteins, VEGF, semaphorins, hepatocyte growth factor, EGF, among others, share signaling components with the c-Ret pathway and are required for optimal metanephric growth and patterning (15)(16)(17)(18)(19)(20)(21)(22). Following induction of the MM, activation of the Wnt/␤-catenin signaling pathway plays a key role in nephrogenesis. Release of Wnt9b from the UB branches triggers a ␤-catenindependent mo...

YY1 is a zinc finger DNA-binding transcription factor that influences expression of a wide variety of cellular and viral genes. YY1 is essential for the development of mammalian embryos. It regulates the expression of genes with important functions in DNA replication, protein synthesis, and cellular response to external stimuli during cell growth and differentiation. How YY1 accomplishes such a variety of functions is unknown. Here, we show that a subset of the nuclear YY1 appears to be O-GlcNAcylated regardless of the differentiation status of the cells. We found that glucose strongly stimulates O-linked N-acetylglucosaminylation (O-GlcNAcylation) on YY1. Glycosylated YY1 no longer binds the retinoblastoma protein (Rb). Upon dissociation from Rb, the glycosylated YY1 is free to bind DNA. The ability of the O-glycosylation on YY1 to disrupt the complex with Rb leads us to propose that O-glycosylation might have a profound effect on cell cycle transitions that regulate the YY1-Rb heterodimerization and promote the activity of YY1. Our observations provide strong evidence that YY1-regulated transcription is very likely connected to the pathway of glucose metabolism that culminates in the O-GlcNAcylation on YY1, changing its function in transcription.

The RNA polymerase I transcription factor UBF has been identified in human, mouse, rat and Xenopus and the primary structure of the human protein has been determined. Human UBF was shown to contain four tandem homologies to the folding domains of the HMG1 and 2 proteins and hence to belong to a previously unrecognised family of 'HMG-box' transcription factors. Here, cDNA clones encoding the Xenopus laevis UBF (xUBF) have been isolated and sequenced. Northern and Southern blots revealed that in tissue culture cells, xUBF is coded on a single major mRNA size species by a small number of genes. The deduced primary structure of xUBF is highly homologous with the human protein except for a central deletion which removes most of one HMG-box. This explains the major size difference between the X. laevis and human proteins and may well explain their different transcriptional specificities. It is shown that xUBF contains 5 tandemly repeated HMG-boxes and that by analogy the human protein contains 6.