Acyclic nucleoside phosphonates with 2-aminothiazole base as inhibitors of bacterial and mammalian adenylate cyclases.

A series of novel acyclic nucleoside phosphonates (ANPs) was synthesized as potential adenylate cyclase inhibitors, where the adenine nucleobase of adefovir (PMEA) was replaced with a 5-substituted 2-aminothiazole moiety. The design was based on the structure of MB05032, a potent and selective inhibitor of fructose 1,6-bisphosphatase and a good mimic of adenosine monophosphate (AMP). From the series of eighteen novel ANPs, which were prepared as phosphoroamidate prodrugs, fourteen compounds were potent (single digit micromolar or submicromolar) inhibitors of Bordetella pertussis adenylate cyclase toxin (ACT), mostly without observed cytotoxicity in J774A.1 macrophage cells. Selected phosphono diphosphates (nucleoside triphosphate analogues) were potent inhibitors of ACT (IC50 as low as 37 nM) and B. anthracis edema factor (IC50 as low as 235 nM) in enzymatic assays. Furthermore, several ANPs were found to be selective mammalian AC1 inhibitors in HEK293 cell-based assays (although with some associated cytotoxicity) and one compound exhibited selective inhibition of mammalian AC2 (only 12% of remaining adenylate cyclase activity) but no observed cytotoxicity. The mammalian AC1 inhibitors may represent potential leads in development of agents for treatment of human inflammatory and neuropathic pain.

Mechanisms of mitochondrial DNA escape and its relationship with different metabolic diseases.

It is well-known that mitochondrial DNA (mtDNA) can escape to intracellular or extracellular compartments under different stress conditions, yet understanding their escape mechanisms remains a challenge. Although Bax/Bak pores and VDAC oligomers are the strongest possibilities, other mechanisms may be involved. For example, mitochondria permeability transition, altered mitophagy, and mitochondrial dynamics are associated with intracellular mtDNA escape, while extracellular traps and extracellular vesicles can participate in extracellular mtDNA escape. The evidence suggests that mtDNA escape is a complex event with more than one mechanism involved. In addition, once the mtDNA is outside the mitochondria, the effects can be complex. Different danger signal sensors recognize the mtDNA as a damage-associated molecular pattern, triggering an innate immune inflammatory response that can be observed in multiple metabolic diseases characterized by chronic inflammation, including autoimmune diseases, diabetes, cancer, and cardiovascular disorders. For these reasons, we will review the most recent evidence regarding mtDNA escape mechanisms and their impact on different metabolic diseases.

Genome-Wide Small Interfering RNA Screening Reveals a Role for Cullin3-Really Interesting New Gene Ligase Signaling in Heterologous Sensitization of Adenylyl Cyclase.

Heterologous sensitization of adenylyl cyclase (AC) is revealed as enhanced or exaggerated AC/cAMP signaling that occurs following persistent activation of Gα i/o-coupled receptors. This paradoxical phenomenon was discovered more than 40 years ago and was proposed as a cellular mechanism to explain the adaptive changes that occur following chronic exposure to drugs of abuse. However, the underlying molecular mechanisms of heterologous sensitization of AC remain largely unknown. In the present study, we performed a genome-wide cell-based RNA interference screen as an unbiased approach to identify genes associated with heterologous sensitization of AC. Following a series of validation and confirmation assays, three genes that form an E3 ligase complex, cullin3 (CUL3), neural precursor-cell-expressed and developmentally downregulated 8 (NEDD8), and really interesting new gene (RING)-box protein 1 (RBX1), were identified as specific modulators of heterologous sensitization of AC. Furthermore, based on the downstream actions of these genes, we evaluated the activity of proteasome inhibitors as well as the specific NEDD8-activating enzyme inhibitor, MLN4924 (Pevonedistat), in AC sensitization. We demonstrate that MG-132 and bortezomib treatments could mimic the inhibitory effects observed with gene knockdown, and MLN4924 was potent and efficacious in blocking the development of heterologous sensitization of endogenous and recombinant AC isoforms, including AC1, AC2, AC5, and AC6. Together, by using genetic and pharmacological approaches, we identified, for the first time, cullin3-RING ligases and the protein degradation pathway as essential modulators for heterologous sensitization of AC. SIGNIFICANCE STATEMENT: Through a genome-wide cell-based RNA interference screening, we identified three genes that form an E3 ligase complex, cullin3, neural precursor-cell-expressed and developmentally downregulated 8 (NEDD8), and really interesting new gene-box protein 1, as specific modulators of heterologous sensitization of AC. The effect of cullin3, NEDD8, or really interesting new gene-box protein 1 small interfering RNAs on heterologous sensitization was recapitulated by proteasome inhibitors, MG132 and bortezomib, and the specific NEDD8-activating enzyme inhibitor, MLN4924. These results suggest a novel hypothesis in which protein degradation is involved in the sensitization of AC signaling that occurs following chronic activation of Gαi/o-coupled receptors.

Health promotion before the HIV/AIDS epidemic in primary care in Punta Arenas / Promoção da saúde frente à epidemia de HIV/AIDS na atenção primária em Punta Arenas / Promoción de la salud frente a epidemia del VIH/AIDS en atención primaria en Punta Arenas

ABSTRACT Objective: to identify the attitudes and actions of health professionals in health promotion before the HIV/AIDS epidemic in Primary Health Care (PHC), Punta Arenas, Magallanes and Chilean Antartica Chilena region, Chile. Method: a qualitative, descriptive, and exploratory study carried out at PHC facilities in the city of Punta Arenas, Chile. Sixteen professionals who work in the five Family Health Centers participated. Data collection was carried out between March and May 2019. Data was obtained through a semi-structured interview. To organize data, ATLAS ti® was used, which were analyzed according to thematic content analysis. Results: two categories have emerged. The first category, Health promotion and prevention actions before HIV/AIDS, described the actions carried out by nurses in their daily work, such as counseling on STI/AIDS prevention, sexual/reproductive health and education for professionals and the community. The second category, Nurse-midwives' attitudes towards caring for people living with HIV/AIDS, includes caring without prejudice, active behavior and empathy due to high workload. Conclusion: health promotion and prevention actions carried out by PHC professionals are mainly related to counseling and education for professionals and the community. The need to integrate prevention measures with other professionals and vulnerable groups in the community stands out.

RESUMO Objetivo: identificar as atitudes e ações dos profissionais de saúde na promoção da saúde frente à epidemia de HIV/Aids na Atenção Primária de Saúde, Punta Arenas, região de Magalhães e Antártica Chilena, Chile. Método: estudo de enfoque qualitativo, descritivo, exploratório, realizado em estabelecimentos de Atenção Primária de Saúde da cidade de Punta Arenas, Chile. Participaram 16 profissionais que atuam nos cinco Centros de Saúde da Família. A coleta de dados foi realizada entre março e maio de 2019. Os dados foram obtidos através de entrevista semiestruturada. Foi utilizado o ATLAS-ti® para a organização dos dados, e estes foram analisados segundo a análise de conteúdo temática. Resultados: resultou em duas categorias. A primeira categoria, Ações de promoção da saúde e prevenção frente ao HIV/Aids descreve as ações realizadas pelas enfermeiras em seu trabalho diário, como o aconselhamento em prevenção de IST/Aids, saúde sexual/reprodutiva e a educação para profissionais e a comunidade. A segunda categoria, Atitudes das enfermeiras e parteiras frente à atenção de pessoas vivendo com HIV/Aids, abarca cuidar sem preconceitos, com escuta ativa e empatia diante da alta carga de trabalho. Conclusão: as ações de promoção da saúde e prevenção realizadas pelos profissionais de Atenção Primária de Saúde se relacionam principalmente com o aconselhamento e educação para profissionais e comunidade. Destaca a necessidade de transversalizar as medidas de prevenção a outros profissionais e grupos vulneráveis da comunidade.

RESUMEN Objetivo: identificar las actitudes y acciones de los profesionales de la salud en promoción de la salud frente a epidemia del VIH/SIDA en Atención Primaria de Salud, Punta Arenas, región de Magallanes y Antártica Chilena, Chile. Método: estudio de enfoque cualitativo, descriptivo, exploratorio, realizado en establecimientos de Atención Primaria de Salud de la ciudad de Punta Arenas, Chile. Participaron 16 professionales que desempeñan en los cinco Centros de Salud Familiar. La recolección de datos se realizó entre marzo y mayo de 2019. Los datos fueron obtenidos a través de entrevista semiestructurada. Para la organización de los datos, se utilizó ATLAS ti®, los cuáles fueron analizados según a análisis de contenido temática. Resultados: se obtuvo como resultados dos categorías. La primera categoría, Acciones de promoción de la salud y prevención frente al VIH/SIDA, describiu las acciones que realizan las enfermeras en su trabajo diario, como la consejería en la prevención de ITS/SIDA, salud sexual/reproductiva y la educación para profesionales y la comunidad. La segunda categoría, Actitudes de las enfermeras y matronas frente a la atención de personas viviendo con VIH/SIDA, inclue atender sin preconceitos, com escuta activa e empatía diante da alta carga laboral. Conclusión: las acciones de promoción de la salud y prevención realizadas por los profesionales de Atención Primaria de Salud se relacionan principalmente con la consejería y educación para profesionales y comunidad. Destaca la necesidad de transversalizar las medidas de prevención a otros profissionales y grupos vulnerables de la comunidad.

Derecho a la salud en el trabajo: vulneración y fragmentación en su comprensión y materialización / Right to health at work: violation and fragmentation in its understanding and materialization / Dereito à saúde no trabalho: vulneração e fragmentação em sua comprensão e materialização

Objetivo: describir la problemática de hombres y mujeres trabajadores que al percibir vulnerado su derecho a la salud en el trabajo por sufrir presuntos o declarados ATEL, buscaron apoyo e interpusieron uno o más recursos constitucionales para lograr su restablecimiento efectivo. Materiales y métodos: estudio mixto. Se aplicó encuesta a 189 trabajadores que sufrieron un presunto o reconocido evento de ATEL y que buscaron apoyo en los servicios de extensión solidaria de la Universidad de Antioquia o en organizaciones de derechos humanos en Medellín. La encuesta captó datos sociodemográficos, de seguridad social, laborales, del SGSST, de los antecedentes, ocurrencia y consecuencias de los ATEL y los recursos interpuestos. Para el abordaje cualitativo se realizó: i) una entrevista en profundidad con un médico laboral y abogado independiente y ii) cuatro grupos focales con trabajadores que sufrieron ATEL, sindicalistas, abogados y médicos representantes de ARL, académicos. Resultados: los trabajadores más vulnerables fueron los más afectados en su derecho. Los agentes que integran el SGRL hacen un abordaje fragmentado de la salud y el trabajo que facilita la vulneración del derecho. Los trabajadores que sufren ATEL deben enfrentar barreras económicas, administrativas y de acceso a salud para restituir su derecho. Conclusiones: el trabajo decente y digno es un horizonte hacia el cual se deben apuntar los esfuerzos para que los trabajadores más vulnerados históricamente conquisten y sostengan una vida plena, digna y saludable. Los recursos como la acción de tutela contribuyen a lograr el cumplimiento del derecho a la salud en el trabajo.

Objective: to describe the problems of working men and women who, when perceiving their right to health at work violated due to alleged or declared workplace accidents or illnesses (ATEL for its acronym in Spanish) sought support and filed one or more constitutional resources to achieve their effective restoration. Materials and methods: Mixed study. A survey was applied to 189 workers who suffered an alleged ATEL event and who sought support in the solidarity extension services at Univesidad de Antioquia or in human rights organizations in the city of Medellin. The survey that captured sociodemographic data, social security data, labor data, the Occupational Health and Safety Management System (SGSST for its acronym in Spanish) data of the background, occurrence and consequences of ATELs and legal resources interposed. For the qualitative approach, an in-depth interview was conducted with an occupational physician and an independent lawyer and four focus groups with workers who suffered ATELs, trade unionists, lawyers and doctors representing the Occupational Risk Administrator (ARL for its acronym in Spanish), and academicians. Results: the most vulnerable workers were the most affected in their right. The agents that make up the General System of Occupational Risks (SGRL by its acronym in Spanish) make a fragmented approach to health and work that facilitates the violation of the right. Workers suffering from ATEL must face economic, administrative and access to health barriers to restore their right. Conclusions: Decent and respectable work is a horizon towards which efforts should be aimed so that the most historically vulnerable workers conquer and sustain full, dignified and healthy life. Resources such as guardianship action contribute to the fulfillment of the right to health at work.

Objetivo: descrever a problemática de homens e mulheres trabalhadores que ao perceber vulnerado seu direito à saúde no trabalho por sofrer presuntos ou declarados ATEL, procuraram apoio e interpuseram um ou mais recursos constitucionais para lograr seu restabelecimento efetivo. Materiais e métodos: estudo misto. Aplicou se enquete a 189 trabalhadores que sofreram um presunto ou reconhecido evento de ATEL e que buscaram apoio nos serviços de extensão solidaria da Universidade de Antioquia ou em organizações de diretos humanos em Medellín. A enquete captou dados sócios demográficos, de segurança social, laborais, do SGSST, dos antecedentes, ocorrência e consequências dos ATEL e os recursos interpostos. Para a abordagem qualitativa se realizou: i) uma entrevista em profundidade com um médico laboral e advogado independente e ii) quatro grupos focais com trabalhadores que sofreram ATEL, sindicalistas, advogados e médicos representantes de ARL, acadêmicos. Resultados: os trabalhadores mais vulneráveis foram os mais afetados em seu direito. Os agentes que integram o SGRL fazem uma abordagem fragmentada da saúde e o trabalho que facilita a vulneração do direito. Os trabalhadores que sofrem ATEL devem enfrentar barreiras econômicas, administrativas e de aceso a saúde para restituir seu direito. Conclusões: o trabalho decente e digno é um horizonte para o qual se devem apontar os esforços para que os trabalhadores mais vulnerados historicamente conquistem e sustenham uma vida plena, digna e saudável. Os recursos como a ação de mandato de segurança que contribui a obter o cumprimento do direito à saúde no trabalho.

Identification of Novel Adenylyl Cyclase 5 (AC5) Signaling Networks in D1 and D2 Medium Spiny Neurons using Bimolecular Fluorescence Complementation Screening.

Adenylyl cyclase type 5 (AC5), as the principal isoform expressed in striatal medium spiny neurons (MSNs), is essential for the integration of both stimulatory and inhibitory midbrain signals that initiate from dopaminergic G protein-coupled receptor (GPCR) activation. The spatial and temporal control of cAMP signaling is dependent upon the composition of local regulatory protein networks. However, there is little understanding of how adenylyl cyclase protein interaction networks adapt to the multifarious pressures of integrating acute versus chronic and inhibitory vs. stimulatory receptor signaling in striatal MSNs. Here, we presented the development of a novel bimolecular fluorescence complementation (BiFC)-based protein-protein interaction screening methodology to further identify and characterize elements important for homeostatic control of dopamine-modulated AC5 signaling in a neuronal model cell line and striatal MSNs. We identified two novel AC5 modulators: the protein phosphatase 2A (PP2A) catalytic subunit (PPP2CB) and the intracellular trafficking associated protein-NSF (N-ethylmaleimide-sensitive factor) attachment protein alpha (NAPA). The effects of genetic knockdown (KD) of each gene were evaluated in several cellular models, including D1- and D2-dopamine receptor-expressing MSNs from CAMPER mice. The knockdown of PPP2CB was associated with a reduction in acute and sensitized adenylyl cyclase activity, implicating PP2A is an important and persistent regulator of adenylyl cyclase activity. In contrast, the effects of NAPA knockdown were more nuanced and appeared to involve an activity-dependent protein interaction network. Taken together, these data represent a novel screening method and workflow for the identification and validation of adenylyl cyclase protein-protein interaction networks under diverse cAMP signaling paradigms.

2 year outcome for 8 year old female managed with partial cystectomy for primary bladder clear cell carcinoma.

Bladder cancer is rare in the pediatric population, and clear cell carcinoma is extremely rare with one other pediatric case reported. Here we report the clinical outcome for a medically complicated pediatric patient with muscle invasive clear cell carcinoma treated with partial cystectomy without neoadjuvant or adjuvant therapy. Final pathology was stage T2bN0M0 with negative margins. At 2 years, there is no disease recurrence by cystoscopy, chest and abdominal imaging. Postoperative issues have been related to reduced bladder capacity and compliance and the patient is currently managed with continuous urinary diversion and will require future definitive lower tract reconstruction.

Optimization of a 1,3,4-oxadiazole series for inhibition of Ca2+/calmodulin-stimulated activity of adenylyl cyclases 1 and 8 for the treatment of chronic pain.

Adenylyl cyclases type 1 (AC1) and 8 (AC8) are group 1 transmembrane adenylyl cyclases (AC) that are stimulated by Ca2+/calmodulin. Studies have shown that mice depleted of AC1 have attenuated inflammatory pain response, while AC1/AC8 double-knockout mice display both attenuated pain response and opioid dependence. Thus, AC1 has emerged as a promising new target for treating chronic pain and opioid abuse. We discovered that the 1,3,4-oxadiazole scaffold inhibits Ca2+/calmodulin-stimulated cyclic adenosine 3',5'-monophosphate (cAMP) production in cells stably expressing either AC1 or AC8. We then carried out structure-activity relationship studies, in which we designed and synthesized 65 analogs, to modulate potency and selectivity versus each AC isoform in cells. Furthermore, molecular docking of the analogs into an AC1 homology model suggests the molecules may bind at the ATP binding site. Finally, a prioritized analog was tested in a mouse model of inflammatory pain and exhibited modest analgesic properties. In summary, our data indicate the 1,3,4-oxadiazoles represent a novel scaffold for the cellular inhibition of Ca2+/calmodulin-stimulated AC1- and AC8 cAMP and warrant further exploration as potential lead compounds for the treatment of chronic inflammatory pain.

A Novel CRISPR/Cas9-Based Cellular Model to Explore Adenylyl Cyclase and cAMP Signaling.

Functional characterization of adenylyl cyclase (AC) isoforms has proven challenging in mammalian cells because of the endogenous expression of multiple AC isoforms and the high background cAMP levels induced by nonselective AC activators. To simplify the characterization of individual transmembrane AC (mAC) isoforms, we generated a human embryonic kidney cell line 293 (HEK293) with low cAMP levels by knocking out two highly expressed ACs, AC3 and AC6, using CRISPR/Cas9 technology. Stable HEK293 cell lines lacking either AC6 (HEK-ACΔ6) or both AC3 and AC6 (HEK-ACΔ3/6) were generated. Knockout was confirmed genetically and by comparing cAMP responses of the knockout cells to the parental cell line. HEK-ACΔ6 and HEK-ACΔ3/6 cells revealed an 85% and 95% reduction in the forskolin-stimulated cAMP response, respectively. Forskolin- and Gαs-coupled receptor-induced activation was examined for the nine recombinant mAC isoforms in the HEK-ACΔ3/6 cells. Forskolin-mediated cAMP accumulation for AC1-6 and AC8 revealed 10- to 250-fold increases over the basal cAMP levels. All nine mAC isoforms, except AC8, also exhibited significantly higher cAMP levels than the control cells after Gαs-coupled receptor activation. Isoform-specific AC regulation by protein kinases and Ca2+/calmodulin was also recapitulated in the knockout cells. Furthermore, the utility of the HEK-ACΔ3/6 cell line was demonstrated by characterizing the activity of novel AC1 forskolin binding-site mutants. Hence, we have developed a HEK293 cell line deficient of endogenous AC3 and AC6 with low cAMP background levels for studies of cAMP signaling and AC isoform regulation.

Synthesis of α-Branched Acyclic Nucleoside Phosphonates as Potential Inhibitors of Bacterial Adenylate Cyclases.

Inhibition of Bordetella pertussis adenylate cyclase toxin (ACT) and Bacillus anthracis edema factor (EF), key virulence factors with adenylate cyclase activity, represents a potential method for treating or preventing toxemia related to whooping cough and anthrax, respectively. Novel α-branched acyclic nucleoside phosphonates (ANPs) having a hemiaminal ether moiety were synthesized as potential inhibitors of bacterial adenylate cyclases. ANPs prepared as bisamidates were not cytotoxic, but did not exhibit any profound activity (IC50 >10 µm) toward ACT in J774A.1 macrophages. The apparent lack of activity of the bisamidates is speculated to be due to the inefficient formation of the biologically active species (ANPpp) in the cells. Conversely, two 5-haloanthraniloyl-substituted ANPs in the form of diphosphates were shown to be potent ACT and EF inhibitors with IC50 values ranging from 55 to 362 nm.

Identification of FDA-Approved Small Molecules Capable of Disrupting the Calmodulin-Adenylyl Cyclase 8 Interaction through Direct Binding to Calmodulin.

Adenylyl cyclases (AC) catalyze the formation of cyclic AMP (cAMP) from ATP and are involved in a number of disease states, making them attractive potential drug targets. AC8, in particular, has been implicated in several neurological disorders. While development of small molecule AC inhibitors has generated some chemical leads, the lack of inhibitor specificity among AC family members has limited the identification of successful drug candidates. Therefore, finding alternative novel methods to suppress AC activity are needed. Because only AC1 and AC8 are robustly stimulated by calmodulin (CaM), we set out to explore the mechanism of disrupting the AC/CaM interaction as a way to selectively inhibit AC8. Through the development and implementation of a novel biochemical high-throughput-screening paradigm, we identified six small molecules from an FDA-approved compound library that are capable of disrupting the AC8/CaM interaction. These compounds were also shown to be able disrupt formation of this complex in cells, ultimately leading to decreased AC8 activity. Interestingly, further mechanistic analysis determined that these compounds functioned by binding to CaM and blocking its interaction with AC8. While these particular compounds could inhibit CaM interaction with both AC1 and AC8, they provide significant proof of concept for inhibition of ACs through disruption of CaM binding. These compounds, as dual AC1/AC8 inhibitors, provide important tools for probing pathological conditions where AC1/AC8 activity are enhanced, such as chronic pain and ethanol consumption. Furthermore, unlike tools such as genetic deletion, these compounds can be used in a dose-dependent fashion to determine the role of AC/CaM interactions in these pathologies.

Identification of a selective small-molecule inhibitor of type 1 adenylyl cyclase activity with analgesic properties.

Adenylyl cyclase 1 (AC1) belongs to a group of adenylyl cyclases (ACs) that are stimulated by calcium in a calmodulin-dependent manner. Studies with AC1 knockout mice suggest that inhibitors of AC1 may be useful for treating pain and opioid dependence. However, nonselective inhibition of AC isoforms could result in substantial adverse effects. We used chemical library screening to identify a selective AC1 inhibitor with a chromone core structure that may represent a new analgesic agent. After demonstrating that the compound (ST034307) inhibited Ca2+-stimulated adenosine 3',5'-monophosphate (cAMP) accumulation in human embryonic kidney (HEK) cells stably transfected with AC1 (HEK-AC1 cells), we confirmed selectivity for AC1 by testing against all isoforms of membrane-bound ACs. ST034307 also inhibited AC1 activity stimulated by forskolin- and Gαs-coupled receptors in HEK-AC1 cells and showed inhibitory activity in multiple AC1-containing membrane preparations and mouse hippocampal homogenates. ST034307 enhanced µ-opioid receptor (MOR)-mediated inhibition of AC1 in short-term inhibition assays in HEK-AC1 cells stably transfected with MOR; however, the compound blocked heterologous sensitization of AC1 caused by chronic MOR activation in these cells. ST034307 reduced pain responses in a mouse model of inflammatory pain. Our data indicate that ST034307 is a selective small-molecule inhibitor of AC1 and suggest that selective AC1 inhibitors may be useful for managing pain.

Molecular Modeling Evaluation of the Enantiomers of a Novel Adenylyl Cyclase 2 Inhibitor.

Adenylyl cyclase 2 (AC2) is one of nine membrane-bound isoforms of adenylyl cyclase that converts ATP into cyclic AMP (cAMP), an important second messenger molecule. Upregulation of AC2 is linked to cancers like pancreatic and small intestinal neuroendocrine tumors (NETs). The structures of the various isoforms of adenylyl cyclases are highly homologous, posing a significant challenge to drug discovery efforts for an effective, isoform-selective modulator of AC2. In a previous study, a screen identified a potential isoform-selective and noncompetitive inhibitor of AC2, SKF83566. In the present study, molecular modeling is used to explore the mode of inhibition of AC2 by SKF83566 and to investigate the active enantiomer of SKF83566. Homology models of hAC2 were built based on canine AC5-C1a and rat AC2-C2a templates. With these models, a combination of flexible docking, molecular dynamics simulations, and free energy calculations using the MM/GBSA methodology suggested an allosteric mechanism in which (S)-SKF83566 binds to an allosteric site near ATP and alters the protein conformation of the ATP binding site, potentially preventing the adenosine moiety of ATP from forming an archlike shape to form cAMP. The predicted binding preference for the (S)-SKF83566 enantiomer and the predicted free energy are consistent with the experimental data.

Latin American Clinical Epidemiology Network Series - Paper 8: Ticagrelor was cost-effective vs. clopidogrel in acute coronary syndrome in Chile.

OBJECTIVE: To evaluate the incremental cost-effectiveness ratio (ICER) of the use of ticagrelor as a substitute for clopidogrel for secondary prevention of acute coronary syndrome in Chile. STUDY DESIGN AND SETTING: Cost-effectiveness analysis based on a Markov model: Safety and effectiveness data of ticagrelor were obtained from a systematic review of the literature. Costs are expressed in Chilean pesos (CLP) as of 2013. The evaluation was conducted from the payer standpoint. A probabilistic sensitivity analysis comprising discount rates and national cost variability was done. A budget impact analysis estimated for 2015 was conducted to calculate the total cost for both treatments. RESULTS: The ICER with a discount rate of 6% for ticagrelor vs. clopidogrel was CLP 4,893,126 per quality-adjusted life-year (QALY) gained (=9,689 US$). In the budget impact analysis for the baseline scenario, considering 100% of treatment, coverage, and adherence, ticagrelor represented an additional cost of CLP 5,233,854,272, for 979 QALYs gained compared with clopidogrel. CONCLUSIONS: Ticagrelor is cost-effective in comparison with clopidogrel for the secondary prevention of acute coronary syndrome. These findings are similar to those reported in other international cost-effectiveness studies.

Design and Synthesis of Fluorescent Acyclic Nucleoside Phosphonates as Potent Inhibitors of Bacterial Adenylate Cyclases.

Bordetella pertussis adenylate cyclase toxin (ACT) and Bacillus anthracis edema factor (EF) are key virulence factors with adenylate cyclase (AC) activity that substantially contribute to the pathogenesis of whooping cough and anthrax, respectively. There is an urgent need to develop potent and selective inhibitors of bacterial ACs with prospects for the development of potential antibacterial therapeutics and to study their molecular interactions with the target enzymes. Novel fluorescent 5-chloroanthraniloyl-substituted acyclic nucleoside phosphonates (Cl-ANT-ANPs) were designed and synthesized in the form of their diphosphates (Cl-ANT-ANPpp) as competitive ACT and EF inhibitors with sub-micromolar potency (IC50 values: 11-622 nm). Fluorescence experiments indicated that Cl-ANT-ANPpp analogues bind to the ACT active site, and docking studies suggested that the Cl-ANT group interacts with Phe306 and Leu60. Interestingly, the increase in direct fluorescence with Cl-ANT-ANPpp having an ester linker was strictly calmodulin (CaM)-dependent, whereas Cl-ANT-ANPpp analogues with an amide linker, upon binding to ACT, increased the fluorescence even in the absence of CaM. Such a dependence of binding on structural modification could be exploited in the future design of potent inhibitors of bacterial ACs. Furthermore, one Cl-ANT-ANP in the form of a bisamidate prodrug was able to inhibit B. pertussis ACT activity in macrophage cells with IC50 =12 µm.

Revisión Sistemática sobre los Efectos Adversos de la Fluoración del Agua / Systematic Review of the Adverse Effects of Water Fluoridation

Durante las últimas décadas, una considerable atención científica ha sido puesta en la seguridad de los fluoruros, dada la amplia variedad de fuentes de ingestión a la que la población se encuentra expuesta y los riesgos a la salud de las personas que esto puede acarrear. El objetivo de esta investigación fue determinar si la fluoración del agua a concentraciones de 0,6 a 1 ppm se asocian a una mayor proporción de efectos adversos en la población general al compararlo con concentraciones subóptimas. Se realizó una revisión sistemática de la literatura en MEDLINE, EMBASE, COCHRANE, SCIELO, LILACS, CRD, BBO, PAHO y WHOLIS, limitada desde el 2002 al 2012. Se incluyeron estudios primarios y secundarios en español, inglés y portugués con al menos dos poblaciones comparadas, una con niveles óptimos de flúor en agua (0,6­1 ppm) y otra sin fluoración del agua (<0,3 ppm) o con niveles subóptimos (>0,3 < 0,6 ppm). Dos investigadores de forma independiente realizaron evaluación de la calidad de los artículos seleccionados y que cumplieron los criterios de inclusión. La búsqueda arrojó 1024 artículos de los cuales 24 cumplieron los criterios de inclusión y 10 fueron incluidos como evidencia. Con excepción de fluorosis dental, no hay asociación entre fluoración del agua con fracturas óseas, cáncer u otro efecto adverso. A pesar de la mayor prevalencia de fluorosis en zonas fluoradas, esta fue principalmente del tipo cuestionable a leve y la proporción de fluorosis con daño estético no difiere significativamente de la presente en zonas sin fluoración del agua.

During the last decades, considerable scientific attention has been paid to the safety of fluoride, given the wide variety of sources of intake at which the population is exposed and the risks to the health of people this may produce. The aim was to determine whether water fluoridation at concentrations from 0.6 to 1 ppm is associated with a higher proportion of adverse effects in the general population when comparing them to suboptimal concentrations. A systematic review was conducted of the literature in MEDLINE, EMBASE, COCHRANE, SCIELO, LILACS, CRD, BBO, PAHO and WHOLIS, limited to 2002 to 2012. Included were primary and secondary studies in Spanish, English and Portuguese with at least two compared populations, one with optimal fluoride levels in the water (0.6­1 ppm) and another without water fluoridation (<0.3 ppm) or with suboptimal levels (>0.3 < 0.6ppm). Two researchers independently evaluated the quality of the articles selected and which met the inclusion criteria. The search revealed 1024 articles, of which 24 met the inclusion criteria and 10 were included as evidence. With the exception of dental fluorosis, there is no association between any other adverse effect and water fluoridation. Despite the greater prevalence of fluorosis in fluoride than in non-fluoride zones, this was mainly questionable to slight and the proportion of fluorosis with esthetic damage does not differ significantly from this in zones without water fluoridation.

Principales aspectos metodológicos en la Evaluación de Tecnologías Sanitarias / The main methodological aspects in Health Technology Assessment

This article reviews the most relevant methodological aspects involved in Health Technology Assessment (HTA). Firstly, it addresses the process of defining the research problem (or scoping). Then it explains some specific aspects of systematic reviews of evidence, as well as indirect and mixed comparisons of the effectiveness of interventions. It covers also the methods for economic evaluation in healthcare and the budget impact analysis of interventions. Finally, the paper provides an empirical insight on the methodological emphasis used by HTA agencies around the world, and reflects on the available capacities in our country in the topics discussed.

Surgical management of male infertility: an update.

Male factor infertility is common, affecting 7% of the total population and up to half of couples who are trying to conceive. Various surgical and reconstructive options allow biological paternity depending on the etiology of the male factor issues. This article describes historical treatments and newer approaches, discussing the role for traditional open surgery, microsurgery and robotic surgery, as well as interventional radiologic procedures in the management of male infertility.

Correlation between the response to Mycobacterium tuberculosis antigens and the tuberculin skin test in patients with rheumatoid arthritis in Colombia / Correlación de la respuesta a antígenos de Mycobacterium tuberculosis y la prueba cutánea de tuberculina en pacientes con artritis reumatoide

Introduction. Rheumatoid arthritis patients under treatment with anti-TNF-α are at a high risk of developing active tuberculosis, and therefore, screening for latent tuberculosis infection is recommended before anti-TNF-α therapy. Objective. To compare the tuberculin test and IFNγ production induced by culture filtrate proteins(CFPs) and Mycobacterium tuberculosis-specific CFP-10 antigens in rheumatoid arthritis patients. Materials and methods. An analytic transversal study was conducted in rheumatoid arthritis patients treated at Hospital Universitario San Vicente Fundación between January and December 2007. IFNγ production in response to CFPs and CFP-10 was measured in the supernatants of whole blood cultures and evaluated for correlations with tuberculin reactivity. The degree of concordance between both tests was also established. Results. Forty-five patients were included, of which 14 (31.1%) had a tuberculin reaction of ≥10 mm of induration, 9 (20%) produced IFNγ in response to CFP-10, and 7 were positive for both tests. The correlation between tests was r=0.53 (IC 95%:0.28-0.72), and the global concordance between tests was80%, with a Kappa coefficient of 0.48 (IC95%:0.20-0.76). Conclusions. Only two tuberculin (-)/CFP-10+ "anergic" patients were observed. By contrast, six tuberculin +/CFP-10(-) "tuberculin false-positive" patients were observed. These data suggest that the tuberculin test is not an appropriate tool for determining the need for tuberculosis prophylaxis.

Introducción. Los pacientes con artritis reumatoide bajo tratamiento con anti-TNFα están en alto riesgo de desarrollar tuberculosis activa, por lo cual se recomienda hacer la tamización para infección latente de tuberculosis, antes de iniciar el tratamiento. Objetivo. Comparar la prueba de tuberculina y la producción de IFNγ inducida por antígenos CFP (Culture Filtrate Protein) y antígenos específicos de Mycobacterium tuberculosis (CFP-10) para el diagnóstico de infección latente de tuberculosis en pacientes con artritis reumatoide. Materiales y métodos. Se llevó a cabo un estudio transversal analítico en pacientes con artritis reumatoide atendidos en el Hospital Universitario San Vicente Fundación, entre enero y diciembre de 2007, a los cuales se les determinó la producción de IFNγ en respuesta a CFP y CFP-10 en sobrenadantes de cultivos de sangre total, y se correlacionó con la reacción en la prueba de tuberculina. Además, se estableció el grado de concordancia entre ambas pruebas. Resultados. Se incluyeron 45 pacientes, de los cuales, 14 (31,1 %) tuvieron un diámetro de induración ≥10 mm (tuberculina positiva), nueve (20 %) produjeron IFNγ en respuesta a CFP-10, y siete fueron positivos para ambas pruebas. La correlación entre las pruebas fue de r=0,53 (IC95%: 0,28-0,72) y la concordancia global entre pruebas fue de 80 %, con un coeficiente kappa de 0,48 (IC95%: 0,20-0,76). Conclusiones. Solo se observaron dos pacientes con tuberculina positiva y CFP-10 positivo "anérgicos" y se encontraron seis pacientes con tuberculina positiva y CFP-10 negativa "falsos positivos para tuberculina", lo cual sugiere que la prueba de la tuberculina no es la más adecuada para indicar profilaxis para tuberculosis.

Correlation between the response to Mycobacterium tuberculosis antigens and the tuberculin skin test in patients with rheumatoid arthritis in Colombia.

INTRODUCTION: Rheumatoid arthritis patients under treatment with anti-TNF-α are at a high risk of developing active tuberculosis, and therefore, screening for latent tuberculosis infection is recommended before anti-TNF-α therapy. OBJECTIVE: To compare the tuberculin test and IFNγ production induced by culture filtrate proteins(CFPs) and Mycobacterium tuberculosis-specific CFP-10 antigens in rheumatoid arthritis patients. MATERIALS AND METHODS: An analytic transversal study was conducted in rheumatoid arthritis patients treated at Hospital Universitario San Vicente Fundación between January and December 2007. IFNγ production in response to CFPs and CFP-10 was measured in the supernatants of whole blood cultures and evaluated for correlations with tuberculin reactivity. The degree of concordance between both tests was also established. RESULTS: Forty-five patients were included, of which 14 (31.1%) had a tuberculin reaction of ≥10 mm of induration, 9 (20%) produced IFNγ in response to CFP-10, and 7 were positive for both tests. The correlation between tests was r=0.53 (IC 95%:0.28-0.72), and the global concordance between tests was80%, with a Kappa coefficient of 0.48 (IC95%:0.20-0.76). CONCLUSIONS: Only two tuberculin (-)/CFP-10+ "anergic" patients were observed. By contrast, six tuberculin +/CFP-10(-) "tuberculin false-positive" patients were observed. These data suggest that the tuberculin test is not an appropriate tool for determining the need for tuberculosis prophylaxis.