GPCR Compound Library

Many genetic alterations have been reported at this locus including HhAntag691 structure deletions, duplications, nonsense, and missense mutations. Genotype/phenotype correlations have been described, with some variation. In general, mutations that abolish enzyme function are associated with a relatively uniform clinical and biochemical presentation. Animal models of OTCD that have been characterized and are readily available include the spf mouse and the spf-ash mouse. These models are currently maintained on a mixed background, B6EiC3Sn, which may limit certain issues of experimental design. Herein, we describe a new spontaneous hypomorphic missense mutation in the C57BL/6J-Otcspf-J/J that produces a new model of mild OTCD. Spf-J mice display normal plasma ammonia and plasma orotic acid at baseline and milder amino acid perturbations when compared to their predecessors. Despite this mild plasma biochemical phenotype, cerebral amino acid concentrations were elevated at baseline and, unlike WT, were depleted during a systemic immune response suggesting altered cerebral amino acid metabolism or transport. Overall, this mouse model is not only useful for further characterization of functional domains of the OTC enzyme, but is also useful for investigating the pathophysiology of cerebral amino acid metabolism in UCD. Due to the inheritance of OTCD, the phenotypic expression of the disease severity in patients is dependent upon the nature of the mutation, genetic background and in females, Xinactivation in the liver. In males, this disorder classically manifests with symptomatic hyperammonemia in infancy, although milder alleles have been described. In addition, phenotypic variability can be seen in males within the same family with the same mutation suggesting modifier alleles may play a role. Late-onset symptoms of OTCD, due to milder enzyme defects, may manifest in men as late as the fifth or sixth decade given the right precipitant, such as the Atkins’ diet. Galloway et al. described a previously healthy male who experienced his first documented hyperammonemic episode at 13 years of age. The precipitant of this episode was unknown. On admission to hospital, his ammonia was 750 μmol/L and hemodialysis was initiated. Urine orotate was 813 μmol/mmol creatinine, suggesting a diagnosis of OTCD. A subsequent liver biopsy demonstrated OTC enzyme activity to be 11% of normal. Molecular investigation revealed a lysine substitution for asparagine in the OTC protein. This amino acid substitution in the OTC enzyme is identical to the mutation described here in the spf-J mouse. Similar to the reported patient, the K80N mutation results in a reduction in enzyme activity to *11–12% in spf-J. The mutation lies outside the substrate binding and catalytic regions of the enzyme and may be related to homo-oligomerization or stability of the enzyme. In our studies, we demonstrated normal levels of OTC mRNA and OTC protein that was below the limit of detection by immunoblot. These findings lead us to suggest that the K80N mutation may destabilize the enzyme, leading to its early degradation.

In CF patients, S. aureus predominates in the lung of children and teenagers, while P. aeruginosa prevails in adults. Exopolysaccharide-enriched biofilms produced by P. aeruginosa increase the mucus viscosity, resistance to antibiotics and host immune effectors. Chronic bacterial infections are common in CF patients and facilitates lung inflammation, mucous obstruction and tissue remodeling, resulting in fatal loss of function. CF lungs display excessive inflammatory response, especially with increased neutrophil recruitment, the mechanism of this phenomenon is not adequately explained. However, intervention in this process likely will benefit CF patients. The role of the pro-inflammatory signaling cytokine Interleukin 1b in CF lung disease has been reported before. P. aeruginosa induces IL-1b or IL-18 production through NLRC4 inflammasome activation. P. aeruginosa flagellin and highly acylated LPS is recognized by TLR5 and TLR4 respectively. Human polymorphisms observed in the IL1B gene were associated with CF disease. CFTR deficient mice were found to be more susceptible to acute and chronic P. aeruginosa infection and display an exacerbated inflammatory response to LPS and P. aeruginosa activated alveolar macrophages from F508del mutant mice have enhanced expression of IL-1b. Huaux et al recently showed a deregulated inflammatory and fibrotic response in F508del mutant mice to bleomycin, which is IL-1R1 signaling dependent. Here we revisited the role of IL-1b in the resolution of P. aeruginosa infection, in a murine model based on mice carrying the most common CF mutation F508del CFTR. In this study, we show that excessive activation of IL-1b correlates with increased bacterial load, Epoxomicin inflammation and lung damage in F508del CFTR mice. Further, we show that IL-1b antibody neutralization attenuates the inflammatory response to P. aeruginosa infection. CF patients display increased susceptibility to chronic infections with opportunistic bacteria, excessive lung inflammation and fibrosis leading to fatal loss of function eventually. P. aeruginosa has been shown to induce IL-1b through NLRC4 inflammasome activation. CFTR deficient mice were found to be more susceptible to P. aeruginosa infection and have an exacerbated inflammatory response to LPS and P. aeruginosa. Furthermore, F508del CFTR mutant alveolar macrophages display an enhanced IL-1b production in response to LPS stimulation. Thus, we were interested in clarifying the role of IL-1b in the resolution of P. aeruginosa infection, in a murine model with the most common CF mutation F508del CFTR. Here, we show that, after P. aeruginosa infection, excessive activation of IL-1b in F508del CFTR mice compared to WT was accompanied by increased CFU in BALF, inflammation and lung damage. Further we show that a therapeutic antibody administration attenuates inflammatory response in an acute model of infection using WT mice. In CF patients bacterial infections persist and their chronicity facilitates lung inflammation, and subsequent tissue remodeling with severe loss of function.

Though, no evidence was reported regarding their role in oral cancer. Perlecan is a large proteoglycan harboring five distinct structural domains, to which long chains of heparan sulfate and/or WZ4002 chondroitin sulfate are attached. This molecule is present in all vascularized tissues with a distribution that is primarily confined to basement membranes. Also, other studies have also identified perlecan in the stromal spaces of various pathophysiological conditions. Agrin shares a rather intriguing multimodular organization with perlecan, but more complexity to agrin can be added due to at least four sites of alternative splicing. The amino acid sequence of agrin encodes a protein with a molecular size of 220 kDa, but the observed molecular weight is around 400 kDa due to the long heparan sulfate and chondroitin sulfate glycosaminoglycans attached to the core protein. Although originally discovered in the neuromuscular junctions, agrin has been observed in numerous other tissues, and it is described as highly expressed in hepatocellular carcinomas and cholangiocellular carcinomas. Nevertheless, little is known about its role at locations other than the neuromuscular junctions, and even less information is known about its role in tumor tissues. In the present study, we focused on understanding the role of the proteoglycans agrin and perlecan in oral cancer. First, we sought to validated the overexpression of agrin and perlecan in oral cancer tissues compared to normal tissues and in cell lines with different site of origin: oral squamous carcinoma originated from human tongue, oral squamous carcinoma SCC-9 isolated from lymph nodes and a skin-derived squamous carcinoma. Next, we showed that oral squamous carcinoma cell line had a reduced ability to adhere to extracellular matrix proteins and increased sensibility to cisplatin when treated with chondroitinase. By specific target agrin and perlecan protein levels with siRNA, we showed that OSCC cells have decreased cell adhesion and migration and increased sensibility to cisplatin treatment. Overall, our findings opened new avenues to better understand the role of agrin and perlecan, as well as their involvement in carcinogenesis, which may offer a novel approach to cancer therapy by targeting the tumor microenvironment. Proteoglycans, essential macromolecules of the tumor microenvironment, have their expression altered during malignant transformation and tumor progression. Agrin and perlecan are two of the major HSPG identified in the basement membrane, and their functional roles in modulation of cancer growth have been reviewed elsewhere. In this study, we showed for the first time that agrin and perlecan are highly expressed in OSCCs, and the function of these proteins in oral cancer associated processes was investigated. Not only the expression of agrin and perlecan was shown to be higher in OSCC tissues compared to control tissues, but also their expression might be associated with different sites of origin, where higher expression of agrin was found in cell line originated from primary site.

These findings were in contrast to previous studies, which have shown that an important characteristic of clostridial myonecrosis is an absence of vascular permeability due to the formation of platelet-leucocyte and platelet-platelet aggregates within the vasculature and the up-regulation of adhesion molecules on endothelial cells, leading to an occlusion of blood flow into infected regions.. Studies using an in vivo microvascular perfusion model also have shown that treatment of mouse cremaster muscle tissue with culture supernatants from wild-type C. perfringens strains led to a decreased functional capillary density, which was reversed when cells were pretreated with anti-Gr-1, a granulocyte-specific antibody, and antiplatelet serum. These data confirmed the importance of neutrophil-platelet aggregates in blocking vascular perfusion during C. perfringens infection. Using a mouse myonecrosis infection model, we have now shown that Regorafenib a-clostripain is not essential for C. perfringens-mediated disease. Mice infected with an a-clostripain mutant did not show any alteration in disease progression or in the development of vascular leukostasis, a hallmark of C. perfringens infection. Mutation of the ccp gene also had no affect on bacterial growth or on the extracellular level of the two major toxins, a-toxin and perfringolysin O, which are implicated in clostridial myonecrosis. In agreement with previous studies our results provide good evidence that a-clostripain is the major protease produced by C. perfringens strain 13. Other bacterial cysteine proteases, which unlike a-clostripain have been shown to be essential for virulence, also are involved in influencing the levels of other virulence factors, many of which have nonredundant functions. For example, mutation of both P. gingivalis gingipains not only reduces total cysteine protease activity, but also affects the growth and expression of cell surface structures such as fimbriae and vesicles, both of which are thought to assist in colonization and evasion of the immune response. Like a-clostripain other potential C. perfringens-encoded spreading factors, such as collagenase and two different sialidases, are not to be essential for disease in the mouse myonecrosis model. These enzymes may still have a role during infection. The breakdown of host tissues is important for the release of nutrients such as amino acids, sugars and essential minerals such as sequestered iron, which are required for the growth of C. perfringens cells in the lesion.

The high risk of metastasis accounts for the need of adjuvant therapy in early tumor stages. To date, IFN-a therapy represents the only effective adjuvant therapeutic approach against malignant melanoma as demonstrated in several studies. In malignant diseases, induction of tolerance by tumor-derived factors is one critical mechanism involved in tumor progression. The production of the immunosuppressive cytokine IL-10 by tumor cells themselves or by tumor-infiltrating immune cells is well known for malignant melanoma and other tumor entities. IL-10 modulates the biologic function of antigen presenting cells and of T cells. In previous studies, we and others demonstrated that IL-10 induces a tolerogenic phenotype of DC with impaired T cell stimulatory properties. Furthermore, IL-10 DC have been shown to induce anergic regulatory CD4+ and CD8+ T cells, which inhibit activated cytotoxic and helper T cells, INCB18424 JAK inhibitor resulting in a failure to kill melanoma cells. Human DC comprise a heterogeneous cell population and their functional potential depends on their origin, the cytokine microenvironment and cell/cell interactions. They are central in inducing immunity and in mediating immune tolerance in their role as professional antigen-presenting cells. Tolerogenic DC control the immune homeostasis and prevent the development of autoimmune diseases but are also involved in tumor development and progression. We hypothesized that IFN-a may interfere with tumorassociated tolerance mechanisms. In the present study, we demonstrate that IFN-a abrogates the tolerogenic function of human IL-10 DC and thereby prevents the induction of T cell anergy and, subsequently the differentiation into suppressive iTregs. The efficacy of IFN-a treatment in cancer and infectious diseases may therefore be related to its capacity to break tumorassociated tolerance induction. In our study, we demonstrate that IFN-a abolishes the tolerogenic phenotype of human IL-10 DC and, thereby, prevents the induction of T cell anergy and regulatory T cells. Tolerogenic DC can be induced by tumor- or immune cellderived factors like IL-10 and are critically involved in tumor progression. Previously, we have shown that addition of IL-10 during the generation of human DC induced a tolerogenic phenotype of DC which provoked antigen-specific tolerance in CD4+ and CD8+ T cells. The induced anergic iTregs inhibited activated effector T cell responses in an antigenspecific fashion and induced a melanoma antigen-specific anergy in CD8+ cytotoxic T cells resulting in failure of tumor lysis.