Pharmacologic inhibition of S-nitrosoglutathione reductase protects against experimental asthma in BALB/c mice through attenuation of both bronchoconstriction and inflammation.

BACKGROUND: S-nitrosoglutathione (GSNO) serves as a reservoir for nitric oxide (NO) and thus is a key homeostatic regulator of airway smooth muscle tone and inflammation. Decreased levels of GSNO in the lungs of asthmatics have been attributed to increased GSNO catabolism via GSNO reductase (GSNOR) leading to loss of GSNO- and NO- mediated bronchodilatory and anti-inflammatory actions. GSNOR inhibition with the novel small molecule, N6022, was explored as a therapeutic approach in an experimental model of asthma. METHODS: Female BALB/c mice were sensitized and subsequently challenged with ovalbumin (OVA). Efficacy was determined by measuring both airway hyper-responsiveness (AHR) upon methacholine (MCh) challenge using whole body plethysmography and pulmonary eosinophilia by quantifying the numbers of these cells in the bronchoalveolar lavage fluid (BALF). Several other potential biomarkers of GSNOR inhibition were measured including levels of nitrite, cyclic guanosine monophosphate (cGMP), and inflammatory cytokines, as well as DNA binding activity of nuclear factor kappa B (NFκB). The dose response, onset of action, and duration of action of a single intravenous dose of N6022 given from 30 min to 48 h prior to MCh challenge were determined and compared to effects in mice not sensitized to OVA. The direct effect of N6022 on airway smooth muscle tone also was assessed in isolated rat tracheal rings. RESULTS: N6022 attenuated AHR (ED50 of 0.015 ± 0.002 mg/kg; Mean ± SEM) and eosinophilia. Effects were observed from 30 min to 48 h after treatment and were comparable to those achieved with three inhaled doses of ipratropium plus albuterol used as the positive control. N6022 increased BALF nitrite and plasma cGMP, while restoring BALF and plasma inflammatory markers toward baseline values. N6022 treatment also attenuated the OVA-induced increase in NFκB activation. In rat tracheal rings, N6022 decreased contractile responses to MCh. CONCLUSIONS: The significant bronchodilatory and anti-inflammatory actions of N6022 in the airways are consistent with restoration of GSNO levels through GSNOR inhibition. GSNOR inhibition may offer a therapeutic approach for the treatment of asthma and other inflammatory lung diseases. N6022 is currently being evaluated in clinical trials for the treatment of inflammatory lung disease.

Mechanism of inhibition for N6022, a first-in-class drug targeting S-nitrosoglutathione reductase.

N6022 is a novel, first-in-class drug with potent inhibitory activity against S-nitrosoglutathione reductase (GSNOR), an enzyme important in the metabolism of S-nitrosoglutathione (GSNO) and in the maintenance of nitric oxide (NO) homeostasis. Inhibition of GSNOR by N6022 and related compounds has shown safety and efficacy in animal models of asthma, chronic obstructive pulmonary disease, and inflammatory bowel disease [Sun, X., et al. (2011) ACS Med. Chem. Lett. 2, 402-406]. N6022 is currently in early phase clinical studies in humans. We show here that N6022 is a tight-binding, specific, and fully reversible inhibitor of GSNOR with an IC(50) of 8 nM and a K(i) of 2.5 nM. We accounted for the fact that the NAD(+)- and NADH-dependent oxidation and reduction reactions, catalyzed by GSNOR are bisubstrate in nature in our calculations. N6022 binds in the GSNO substrate binding pocket like a competitive inhibitor, although in kinetic assays it behaves with a mixed uncompetitive mode of inhibition (MOI) toward the GSNO substrate and a mixed competitive MOI toward the formaldehyde adduct, S-hydroxymethylglutathione (HMGSH). N6022 is uncompetitive with cofactors NAD(+) and NADH. The potency, specificity, and MOI of related GSNOR inhibitor compounds are also reported.

Structure-activity relationship of pyrrole based S-nitrosoglutathione reductase inhibitors: carboxamide modification.

The enzyme S-nitrosoglutathione reductase (GSNOR) is a member of the alcohol dehydrogenase family (ADH) that regulates the levels of S-nitrosothiols (SNOs) through catabolism of S-nitrosoglutathione (GSNO). GSNO and SNOs are implicated in the pathogenesis of many diseases including those in respiratory, gastrointestinal, and cardiovascular systems. The pyrrole based N6022 was recently identified as a potent, selective, reversible, and efficacious GSNOR inhibitor which is currently in clinical development for acute asthma. We describe here the synthesis and structure-activity relationships (SAR) of novel pyrrole based analogs of N6022 focusing on carboxamide modifications on the pendant N-phenyl moiety. We have identified potent and novel GSNOR inhibitors that demonstrate efficacy in an ovalbumin (OVA) induced asthma model in mice.

Discovery of potent and novel S-nitrosoglutathione reductase inhibitors devoid of cytochrome P450 activities.

The pyrrole based N6022 was recently identified as a potent, selective, reversible, and efficacious S-nitrosoglutathione reductase (GSNOR) inhibitor and is currently undergoing clinical development for the treatment of acute asthma. GSNOR is a member of the alcohol dehydrogenase family (ADH) and regulates the levels of S-nitrosothiols (SNOs) through catabolism of S-nitrosoglutathione (GSNO). Reduced levels of GSNO, as well as other nitrosothiols (SNOs), have been implicated in the pathogenesis of many diseases including those of the respiratory, cardiovascular, and gastrointestinal systems. Preservation of endogenous SNOs through GSNOR inhibition presents a novel therapeutic approach with broad applicability. We describe here the synthesis and structure-activity relationships (SAR) of novel pyrrole based analogues of N6022 focusing on removal of cytochrome P450 inhibition activities. We identified potent and novel GSNOR inhibitors having reduced CYP inhibition activities and demonstrated efficacy in a mouse ovalbumin (OVA) model of asthma.

Structure-activity relationships of pyrrole based S-nitrosoglutathione reductase inhibitors: pyrrole regioisomers and propionic acid replacement.

S-Nitrosoglutathione reductase (GSNOR) is a member of the alcohol dehydrogenase family (ADH) that regulates the levels of S-nitrosothiols (SNOs) through catabolism of S-nitrosoglutathione (GSNO). GSNO and SNOs are implicated in the pathogenesis of many diseases including those in respiratory, cardiovascular, and gastrointestinal systems. The pyrrole based N6022 was recently identified as a potent, selective, reversible, and efficacious GSNOR inhibitor which is currently undergoing clinical development. We describe here the synthesis and structure-activity relationships (SAR) of novel pyrrole based analogues of N6022 focusing on scaffold modification and propionic acid replacement. We identified equally potent and novel GSNOR inhibitors having pyrrole regioisomers as scaffolds using a structure based approach.

Elimination of Onchocercia volvulus transmission in the Santa Rosa focus of Guatemala.

To eliminate transmission of Onchocerca volvulus, semiannual mass treatment with ivermectin (Mectizan; donated by Merck & Co) has been underway in Guatemala since 2000. We applied the 2001 World Health Organization (WHO) elimination criteria in the Santa Rosa focus of onchocerciasis transmission in Guatemala (10,923 persons at risk). No evidence of parasite DNA was found in 2,221 Simulium ochraceum vectors (one-sided 95% confidence interval [CI], 0-0.086%), and no IgG4 antibody positives to recombinant antigen OV16 were found in a sample of 3,232 school children (95% CI, 0-0.009%). We also found no evidence of microfilariae in the anterior segment of the eye in 363 area residents (95% CI, 0-0.08%). Our interpretation of these data, together with historical information, suggest that transmission of O. volvulus is permanently interrupted in Santa Rosa and that ivermectin treatments there can be halted.

Staff, associate specialist and specialty doctors' national audit on the management of gonorrhoea in the United Kingdom, 2015.

The British Association for Sexual Health and HIV (BASHH) revised United Kingdom national guideline for the management of gonorrhoea in adults, 2011, identified five auditable outcome measures, namely, that all patients should receive first-line treatment, be screened or treated for chlamydial infection, have a test of cure (TOC), be offered written information and have partner notification carried out. The UK National Guideline for Gonorrhoea Testing, Clinical Effectiveness Group, BASHH, 2012, recommended in addition that all reactive nucleic acid amplification tests (NAATs) from pharynx and rectum should be confirmed by supplementary testing, using a second NAAT which detects a different nucleic acid target, all those with a positive NAAT for gonorrhoea should have culture and antimicrobial susceptibility testing and that TOC should be done by two weeks. Staff, associate specialist and specialty doctors performed a national audit against these standards. Data from 3233 cases were submitted; 8% of cases of gonorrhoea diagnosed in England, Scotland and Wales over this period. We found that 83% patients received first-line treatment with a reason for not doing so provided for 11%. TOC was documented for 62% and written information was offered to 41%. Results about supplementary testing were inconsistent. The results for the other outcomes were satisfactory.

Good Laboratory Practice Preclinical Safety Studies for GSK2696273 (MLV Vector-Based Ex Vivo Gene Therapy for Adenosine Deaminase Deficiency Severe Combined Immunodeficiency) in NSG Mice.

GSK2696273 (autologous CD34+ cells transduced with retroviral vector that encodes for the human adenosine deaminase [ADA] enzyme) is a gamma-retroviral ex vivo gene therapy of bone marrow-derived CD34+ cells for the treatment of adenosine deaminase deficiency severe combined immunodeficiency (ADA-SCID). ADA-SCID is a severe monogenic disease characterized by immunologic and nonimmunologic symptoms. Bone-marrow transplant from a matched related donor is the treatment of choice, but it is available for only a small proportion of patients. Ex vivo gene therapy of patient bone-marrow CD34+ cells is an alternative treatment. In order to prepare for a marketing authorization application in the European Union, preclinical safety studies in mice were requested by the European Medicines Agency (EMA). A pilot study and a main biodistribution study were performed according to Good Laboratory Practice (GLP) at the San Raffaele Telethon Institute for Gene Therapy test facility. In the main study, human umbilical cord blood (UCB)-derived CD34+ cells were transduced with gamma-retroviral vector used in the production of GSK2696273. Groups of 10 male and 10 female NOD-SCID gamma (NSG) mice were injected intravenously with a single dose of transduced- or mock-transduced UCB CD34+ cells, and they were observed for 4 months. Engraftment and multilineage differentiation of blood cells was observed in the majority of animals in both groups. There was no significant difference in the level of chimerism between the two groups. In the gene therapy group, vector was detectable in lymphohemopoietic and nonlymphohemopoietic tissues, consistent with the presence of gene-modified human hematopoietic donor cells. Given the absence of relevant safety concerns in the data, the nonclinical studies and the clinical experience with GSK2696273 supported a successful application for market authorization in the European Union for the treatment of ADA-SCID patients, for whom no suitable human leukocyte antigen-matched related donor is available.

Optimism in close relationships: How seeing things in a positive light makes them so.

Does expecting positive outcomes--especially in important life domains such as relationships--make these positive outcomes more likely? In a longitudinal study of dating couples, the authors tested whether optimists (who have a cognitive disposition to expect positive outcomes) and their romantic partners are more satisfied in their relationships, and if so, whether this is due to optimists perceiving greater support from their partners. In cross-sectional analyses, both optimists and their partners indicated greater relationship satisfaction, an effect that was mediated by optimists' greater perceived support. When the couples engaged in a conflict conversation, optimists and their partners saw each other as engaging more constructively during the conflict, which in turn led both partners to feel that the conflict was better resolved 1 week later. In a 1-year follow-up, men's optimism predicted relationship status. Effects of optimism were mediated by the optimists' perceived support, which appears to promote a variety of beneficial processes in romantic relationships.

Level 2 genitourinary medicine training and ongoing supervision -- how we can all benefit.

An innovative six-month, two sessions a week, funded clinical attachment programme was devised and introduced for practical training and experience to meet the demands and requirements of those wishing practical experience and training in genitourinary medicine. This has been generated by interest from previous theoretical teaching and as a desire to provide Level 2 services in the community. To date four doctors have completed this training with excellent mutual evaluation and a recognition by all involved, including commissioners, of the importance of maintaining a regular formal session while providing Level 2 community sexual health services. To be formally recognized improved evaluation, standardization and certification are required. However, if pursued, this package with continuing specialist links should prove beneficial to all involved and with the current epidemic of sexually transmitted infections will improve and enhance available services.

Lying words: predicting deception from linguistic styles.

Telling lies often requires creating a story about an experience or attitude that does not exist. As a result, false stories may be qualitatively different from true stories. The current project investigated the features of linguistic style that distinguish between true and false stories. In an analysis of five independent samples, a computer-based text analysis program correctly classified liars and truth-tellers at a rate of 67% when the topic was constant and a rate of 61% overall. Compared to truth-tellers, liars showed lower cognitive complexity, used fewer self-references and other-references, and used more negative emotion words.

ADH IB expression, but not ADH III, is decreased in human lung cancer.

Endogenous S-nitrosothiols, including S-nitrosoglutathione (GSNO), mediate nitric oxide (NO)-based signaling, inflammatory responses, and smooth muscle function. Reduced GSNO levels have been implicated in several respiratory diseases, and inhibition of GSNO reductase, (GSNOR) the primary enzyme that metabolizes GSNO, represents a novel approach to treating inflammatory lung diseases. Recently, an association between decreased GSNOR expression and human lung cancer risk was proposed in part based on immunohistochemical staining using a polyclonal GSNOR antibody. GSNOR is an isozyme of the alcohol dehydrogenase (ADH) family, and we demonstrate that the antibody used in those studies cross reacts substantially with other ADH proteins and may not be an appropriate reagent. We evaluated human lung cancer tissue arrays using monoclonal antibodies highly specific for human GSNOR with minimal cross reactivity to other ADH proteins. We verified the presence of GSNOR in ≥85% of specimens examined, and extensive analysis of these samples demonstrated no difference in GSNOR protein expression between cancerous and normal lung tissues. Additionally, GSNOR and other ADH mRNA levels were evaluated quantitatively in lung cancer cDNA arrays by qPCR. Consistent with our immunohistochemical findings, GSNOR mRNA levels were not changed in lung cancer tissues, however the expression levels of other ADH genes were decreased. ADH IB mRNA levels were reduced (>10-fold) in 65% of the lung cancer cDNA specimens. We conclude that the previously reported results showed an incorrect association of GSNOR and human lung cancer risk, and a decrease in ADH IB, rather than GSNOR, correlates with human lung cancer.

Discovery of s-nitrosoglutathione reductase inhibitors: potential agents for the treatment of asthma and other inflammatory diseases.

S-Nitrosoglutathione reductase (GSNOR) regulates S-nitrosothiols (SNOs) and nitric oxide (NO) in vivo through catabolism of S-nitrosoglutathione (GSNO). GSNOR and the anti-inflammatory and smooth muscle relaxant activities of SNOs, GSNO, and NO play significant roles in pulmonary, cardiovascular, and gastrointestinal function. In GSNOR knockout mice, basal airway tone is reduced and the response to challenge with bronchoconstrictors or airway allergens is attenuated. Consequently, GSNOR has emerged as an attractive therapeutic target for several clinically important human diseases. As such, small molecule inhibitors of GSNOR were developed. These GSNOR inhibitors were potent, selective, and efficacious in animal models of inflammatory disease characterized by reduced levels of GSNO and bioavailable NO. N6022, a potent and reversible GSNOR inhibitor, reduced bronchoconstriction and pulmonary inflammation in a mouse model of asthma and demonstrated an acceptable safety profile. N6022 is currently in clinical development as a potential agent for the treatment of acute asthma.

Successful interruption of transmission of Onchocerca volvulus in the Escuintla-Guatemala focus, Guatemala.

BACKGROUND: Elimination of onchocerciasis (river blindness) through mass administration of ivermectin in the six countries in Latin America where it is endemic is considered feasible due to the relatively small size and geographic isolation of endemic foci. We evaluated whether transmission of onchocerciasis has been interrupted in the endemic focus of Escuintla-Guatemala in Guatemala, based on World Health Organization criteria for the certification of elimination of onchocerciasis. METHODOLOGY/PRINCIPAL FINDINGS: We conducted evaluations of ocular morbidity and past exposure to Onchocerca volvulus in the human population, while potential vectors (Simulium ochraceum) were captured and tested for O. volvulus DNA; all of the evaluations were carried out in potentially endemic communities (PEC; those with a history of actual or suspected transmission or those currently under semiannual mass treatment with ivermectin) within the focus. The prevalence of microfilariae in the anterior segment of the eye in 329 individuals (> or =7 years old, resident in the PEC for at least 5 years) was 0% (one-sided 95% confidence interval [CI] 0-0.9%). The prevalence of antibodies to a recombinant O. volvulus antigen (Ov-16) in 6,432 school children (aged 6 to 12 years old) was 0% (one-sided 95% IC 0-0.05%). Out of a total of 14,099 S. ochraceum tested for O. volvulus DNA, none was positive (95% CI 0-0.01%). The seasonal transmission potential was, therefore, 0 infective stage larvae per person per season. CONCLUSIONS/SIGNIFICANCE: Based on these evaluations, transmission of onchocerciasis in the Escuintla-Guatemala focus has been successfully interrupted. Although this is the second onchocerciasis focus in Latin America to have demonstrated interruption of transmission, it is the first focus with a well-documented history of intense transmission to have eliminated O. volvulus.

Exposure of seasonal migrant workers to Onchocerca volvulus on coffee plantations in Guatemala.

Onchocerciasis (river blindness), which is close to being eliminated from Guatemala through semiannual administration of ivermectin, is still transmitted in one area of the country that coincidentally receives an annual influx of migrant workers to harvest coffee. Migrant workers generally are not included in semiannual ivermectin treatments, but if infected could serve as a reservoir. We report on two studies undertaken to measure the exposure to onchocerciasis (presence of IgG4 antibodies to a recombinant Onchocerca volvulus antigen, OV-16) among migrant workers. During two coffee harvest seasons, 170 migrant workers with a history of working in the disease-endemic area were tested and 1 (0.6%, 95% confidence interval = 0-3.2%) was seropositive. This low rate of exposure in migrant workers indicates that they are unlikely to play a significant role in transmission of onchocerciasis and may indicate that transmission in the last remaining disease-endemic area of Guatemala is decreasing significantly.