A copper(ii) peptide helicate selectively cleaves DNA replication foci in mammalian cells.

The use of copper-based artificial nucleases as potential anticancer agents has been hampered by their poor selectivity in the oxidative DNA cleavage process. An alternative strategy to solve this problem is to design systems capable of selectively damaging noncanonical DNA structures that play crucial roles in the cell cycle. We designed an oligocationic CuII peptide helicate that selectively binds and cleaves DNA three-way junctions (3WJs) and induces oxidative DNA damage via a ROS-mediated pathway both in vitro and in cellulo, specifically at DNA replication foci of the cell nucleus, where this DNA structure is transiently generated. To our knowledge, this is the first example of a targeted chemical nuclease that can discriminate with high selectivity 3WJs from other forms of DNA both in vitro and in mammalian cells. Since the DNA replication process is deregulated in cancer cells, this approach may pave the way for the development of a new class of anticancer agents based on copper-based artificial nucleases.

Nanoparticle- and Microparticle-Based Vaccines against Orbiviruses of Veterinary Importance.

Bluetongue virus (BTV) and African horse sickness virus (AHSV) are widespread arboviruses that cause important economic losses in the livestock and equine industries, respectively. In addition to these, another arthropod-transmitted orbivirus known as epizootic hemorrhagic disease virus (EHDV) entails a major threat as there is a conducive landscape that nurtures its emergence in non-endemic countries. To date, only vaccinations with live attenuated or inactivated vaccines permit the control of these three viral diseases, although important drawbacks, e.g., low safety profile and effectiveness, and lack of DIVA (differentiation of infected from vaccinated animals) properties, constrain their usage as prophylactic measures. Moreover, a substantial number of serotypes of BTV, AHSV and EHDV have been described, with poor induction of cross-protective immune responses among serotypes. In the context of next-generation vaccine development, antigen delivery systems based on nano- or microparticles have gathered significant attention during the last few decades. A diversity of technologies, such as virus-like particles or self-assembled protein complexes, have been implemented for vaccine design against these viruses. In this work, we offer a comprehensive review of the nano- and microparticulated vaccine candidates against these three relevant orbiviruses. Additionally, we also review an innovative technology for antigen delivery based on the avian reovirus nonstructural protein muNS and we explore the prospective functionality of the nonstructural protein NS1 nanotubules as a BTV-based delivery platform.

Production and Purification of Candidate Subunit Vaccines by IC-Tagging Protein Encapsulation.

Particulate material is more efficient in eliciting immune responses. Here we describe the production of micro- and nanospheres formed by protein muNS-Mi from avian reoviruses, loaded with foreign epitopes for their use as vaccines.

Chemical and thermal stabilization of CotA laccase via a novel one-step expression and immobilization in muNS-Mi nanospheres.

A methodology that programs eukaryotic or bacterial cells to encapsulate proteins of any kind inside micro/nanospheres formed by muNS-Mi viral protein was developed in our laboratory. In the present study such "in cellulo" encapsulation technology is utilized for immobilizing a protein with an enzymatic activity of industrial interest, CotA laccase. The encapsulation facilitates its purification, resulting in a cost-effective, one-step way of producing immobilized enzymes for industrial use. In addition to the ability to be recycled without activity loss, the encapsulated protein showed an increased pH working range and high resistance to chemical inactivation. Also, its activity was almost unaffected after 30 min incubation at 90 °C and 15 min at the almost-boiling temperature of 95 °C. Furthermore, the encapsulated laccase was able to efficiently decolorate the recalcitrant dye RB19 at room temperature.

Experiencia farmacoterapéutica de los pacientes con artritis reumatoide: el rol de la farmacia comunitaria / Exploring the medication experience of patients with rheumatoid arthritis: The role of community pharmacy

Introducción: La gestión integral de la farmacoterapia de la AR es altamente compleja y la experiencia del paciente con la medicación condiciona su conducta. Objetivos. Explorar las dimensiones socioculturales de la experiencia farmacoterapéutica y determinar las necesidades a cubrir desde la farmacia comunitaria. Métodos: Investigación cualitativa, exploratoria, descriptiva y transversal con análisis temático secundario por categorías y unidades de significado emergentes. Resultados: La medicación pre-diagnóstico se percibe como paliativa no siempre efectiva. Tras el diagnóstico–a su vez alivio y sentencia-, se construye una relación con la medicación basada en las experiencias vividas, sensaciones físicas y emocionales, valorando continuamente su efectividad y seguridad a partir de sensaciones físicas y brotes. Los participantes reclaman mejor comunicación con los facultativos, mayor coordinación asistencial y que su experiencia sea considerada en la toma de decisiones. Destacan como positiva la gestión colectiva del dolor, tanto por el acompañamiento incondicional en el ámbito del hogar, como de las asociaciones de pacientes, claves para su empoderamiento. Conclusiones: La experiencia farmacoterapéutica redefine las fases de la enfermedad y se vertebra con el padecimiento y el conocimiento propio de la AR. La experiencia de los pacientes sobre la efectividad y seguridad de la medicación debería ser considerada en la toma de decisiones. La farmacia comunitaria tiene un rol importante a desarrollar para reducir el retraso diagnóstico, ofrecer información técnica sobre medicamentos, contribuir a la optimización de la farmacoterapia y al paciente a empoderase e integrarse con las redes sociales de cuidados.(AU)

Introduction: Comprehensive Medication Management in rheumatoid arthritis is highly complex and the patients experience with the medication determines the way in which it is used by them. Objectives: To explore sociocultural dimensions of the medication experience and determine the needs that should be covered by the pharmacy. Methods: It was carried out a qualitative, exploratory, descriptive, and cross-sectional research with secondary thematic analysis by categories and emerging units of meaning. Results: Prediagnostic medication is perceived as palliative and often ineffective. The diagnosis is bitter-sweet. After it, a relationship with the medication is built based on the lived experiences -both physical and emotional-assessing continually its effectiveness and safety according to physical sensations and outbreaks. Participants demand a better communication with doctors and a greater coordinated assistance. Thus, their experience should be considered in the decision-making process. They highlight the collective management of pain, the unconditional support at home as well as patients’ associations, which are key to their empowerment. Conclusions: Patients’ medication experience redefines the phases of the disease and is built with the suffering and the own knowledge of the rheumatoid arthritis. Patients’ experience with the effectiveness and safety of medication should be considered in the decision-making process. Community Pharmacy has an important role to reduce the diagnostic delay, to offer technical information on medications, to contribute to the optimization of pharmacotherapy and to empower the patient and get him or her integrated with the social care networks.(AU)

Incidencia de tuberculosis en pacientes que reciben fármacos anti-TNF( / Incidence of tuberculosis in patients receiving tumor necrosis factor-( blockers / Incidência de tuberculose em pacientes que recebem fármacos anti-TNF(

Resumen: Introducción: la tuberculosis (TB) es una complicación frecuente del uso de fármacos anti-TNF(. Ocurre por reactivación de una infección latente o por progresión de una infección reciente. Objetivos: conocer la incidencia de TB en la población que recibió fármacos anti- TNF(, analizar las formas de presentación y la realización de pesquisa de infección latente previo al inicio del tratamiento. Método: estudio de cohorte retrospectiva. Se incluyeron los pacientes que recibieron fármacos anti- TNF( entre 2010 y 2016. Los datos se obtuvieron de los sistemas informáticos del Fondo Nacional de Recursos y del Programa Nacional de Tuberculosis. Se calculó la incidencia de TB y se describieron los casos que desarrollaron TB. Resultados: se incluyeron 991 tratamientos para un total de 980 pacientes. Se reportaron nueve casos de TB. La incidencia global fue de 419,9 (IC 95% 191,9-591,2) por 100.000 personas/año. Solo hubo casos de TB en pacientes tratados con adalimumab. El cribado de infección tuberculosa latente (ITBL) previo al inicio del fármaco fue heterogéneo y predominaron las formas de TB diseminadas (6/9) sobre la afectación pulmonar aislada (3/9). En todos los casos se suspendió el anti- TNF( al diagnóstico de TB y en ningún caso se retomó. Conclusiones: la incidencia de TB en la población de pacientes bajo tratamiento con anti- TNF( fue 16,5 veces mayor que en la población general. Predominaron las formas de TB diseminadas y se dieron casos en sujetos que habían recibido tratamiento de ITBL previo al inicio del fármaco, sugiriendo que el riesgo persiste mientras exista exposición a éste.

Summary: Introduction: tuberculosis (TB) is a frequent complication in patients receiving tumor necrosis factor-alpha (TNF-() blockers. It occurs upon the reactivation of a latent infection or the progression of a recent infection. Objective: to learn about the incidence of TB in a population receiving tumor necrosis factor-alpha (TNF-() blockers, to analyze the presentation of this condition and to conduct a latent infection research prior to the initiation of therapy. Method: retrospective cohort study. Patients receiving tumor necrosis factor-alpha (TNF-() blockers between 2010 and 2016 were included in the study. Data were obtained from the IT systems of the National Resources Fund and the National Tuberculosis Program. The incidence of TB was calculated and the cases developing TB were described. Results: 991 treatments were included for 980 patients in total. 9 cases of TB were reported. Global incidence was 419.9 (IC 95% 191.9-591.2) out of 100,000 people per year. Cases of TB were only seen in patients treated with adalimumab. Screening for LTBI upon initiation of the drug was heterogeneous and the disseminated forms of TB prevailed (6/9) over isolated pulmonary affectation (3/9). In all cases anti- TNF( was suspended when TB was diagnosed, and it was not reinitiated. Conclusions: the incidence of TB in patients receiving tumor necrosis factor-alpha (TNF-() blockers was 16.5 times greater than in the general population. Disseminated forms of TB prevailed, and some cases occurred in individuals who had received LTBI therapy prior to the initiation of the drug, suggesting the risk persists as long as there is exposure to the drug.

Resumo: Introdução: a tuberculose (TB) é uma complicação frequente do uso de fármacos anti- TNF(. É causada pela reativação de uma infecção latente ou pela evolução de uma infecção recente. Objetivos: conhecer a incidência de TB na população que recebeu fármacos anti- TNF(, analisar as formas de apresentação e a realização de pesquisa de infecção latente prévia ao início do tratamento. Método: estudo de coorte retrospectivo. Foram incluídos todos os pacientes que receberam fármacos anti- TNF( no período 2010-2016. Os dados foram obtidos dos registros informatizados do Fondo Nacional de Recursos e do Programa Nacional de Tuberculosis. A incidência de TB foi calculada e foram descritos os casos que desenvolveram esta patologia. Resultados: foram incluídos 991 tratamentos para um total de 980 pacientes. Nove casos de TB foram registrados. A incidência global foi de 419,9 (IC 95% 191,9-591,2) por 100.000 pessoas/ano. Somente foram registrados casos de TB em pacientes tratados com adalimumab. A triagem de infecção tuberculosa latente (ITBL) prévia ao início do fármaco foi heterogênea e predominaram as formas de TB disseminadas (6/9) sobre a pulmonar isolada (3/9). Em todos os casos o uso de anti- TNF( foi suspenso definitivamente quando o diagnóstico de TB foi realizado. Conclusões: a incidência de TB na população de pacientes em tratamento com anti- TNF( foi 16,5 vezes maior que na população em geral. Predominaram as formas disseminadas de TB e foram registrados casos em pacientes que haviam recebido tratamento para ITBL prévio ao início do fármaco, sugerindo que o risco persiste enquanto houver exposição a este.

Cross-protective immune responses against African horse sickness virus after vaccination with protein NS1 delivered by avian reovirus muNS microspheres and modified vaccinia virus Ankara.

African horse sickness virus (AHSV) is an insect-borne pathogen that causes acute disease in horses and other equids. In an effort to improve the safety of currently available vaccines and to acquire new knowledge about the determinants of AHSV immunogenicity, new generation vaccines are being developed. In this work we have generated and tested a novel immunization approach comprised of nonstructural protein 1 (NS1) of AHSV serotype 4 (AHSV-4) incorporated into avian reovirus muNS protein microspheres (MS-NS1) and/or expressed using recombinant modified vaccinia virus Ankara vector (MVA-NS1). The protection conferred against AHSV by a homologous MS-NS1 or heterologous MS-NS1 and MVA-NS1 prime/boost was evaluated in IFNAR (-/-) mice. Our results indicate that immunization based on MS-NS1 and MVA-NS1 afforded complete protection against the infection with homologous AHSV-4. Moreover, priming with MS-NS1 and boost vaccination with MVA-NS1 (MS-MVA-NS1) triggered NS1 specific cytotoxic CD8 + T cells and prevented AHSV disease in IFNAR (-/-) mice after challenge with heterologous serotype AHSV-9. Cross-protective immune responses are highly important since AHS can be caused by nine different serotypes, which means that a universal polyvalent vaccination would need to induce protective immunity against all serotypes.

Frecuencia de consultas cardiológicas y paros cardíacos extrahospitalarios durante la pandemia COVID-19 en Uruguay / Frequency of cardiological consultations and out-of-hospital cardiac arrests during the COVID-19 pandemic / Frequência de consultas cardiológicas e paradas cardíacas fora do hospital durante a pandemia de COVID-19

Resumen: Introducción: la pandemia COVID-19 ha determinado la aplicación de medidas sanitarias de emergencia, tendientes a evitar la progresión a nivel nacional. Reportes internacionales han sugerido que dichas medidas determinaron secundariamente una desatención en otras patologías, principalmente cardiovasculares, y eventual aumento de paros cardíacos extrahospitalarios (PCEH). Objetivo: determinar la frecuencia de consultas en emergencia por dolor precordial a nivel nacional y de PCEH asistidos por los principales servicios de emergencia prehospitalaria (SEP) de Montevideo, durante la pandemia. Método: se solicitó información del número de consultas por dolor precordial y de consultas totales, a las instituciones médicas de Montevideo y del interior del país, realizadas en el período comprendido entre el 13 de marzo y el 30 de abril de los años 2018, 2019 y 2020. Se solicitó a los SEP información sobre la frecuencia de PCEH en Montevideo durante los mismos períodos. Los datos se expresan como frecuencia absoluta y tasa de incidencia (por 100.000 afiliados) con su intervalo de confianza estimado mediante Fisher. Resultados: el número de consultas por dolor precordial fue estable durante el período 2018-2019. Durante 2020, dichas consultas disminuyeron, representando entre 11,3% y 21,7% del total de consultas. Se evidenció un aumento no significativo en la tasa de PCEH en el 2020 (9,05, IC95%: 7,15-11,30) comparado con el 2019 (7,94, IC95%: 6,19-10,04) y 2018 (7,43, IC95%: 5,75-9,45). Conclusiones: los datos crudos presentados muestran que desde el 13 de marzo hasta el 30 de abril de 2020 hubo una disminución en las consultas en emergencia por dolor precordial, aumentando de forma no significativa el número de PCEH durante el mes de abril, respecto al mismo período de los dos años anteriores.

Summary: Introduction: the COVID-19 pandemic has determined the application of emergency health measures aimed at preventing progression at national level. International reports have suggested that these measures lead to a lack of care in other pathologies, mainly cardiovascular, and eventually increase out-of-hospital cardiac arrests. Objective: to determine the frequency of emergency consultation for chest pain and out-of-hospital cardiac arrests, assisted by the main pre-hospital emergency services of Montevideo, during the COVID-19 pandemic. Methods: information was requested to the medical institutions of Montevideo and all over the country, on the number of consultations for chest pain and total consultations, in the period March 13- April 30 of 2018, 2019 and 2020. The frequency of out-of-hospital cardiac arrests in Montevideo was requested to the pre-hospital emergency services during the same period. The data is expressed as absolute frequency and incidence rates (x 100,000) with its 95% CI calculated by Fisher. Results: the number of consultations for precordial pain was stable during the 2018-2019 period. During 2020, these consultations decreased and represented between 11.3% and 21.7% of the total number of consultations. Out-of-hospital cardiac arrests showed a non significant increase in its incidence rate in 2020 (9.05, 95%IC: 7.15-11.30) compared with 2019 (7.94, 95%IC: 6.19-10.04) and 2018 (7.43, 95%IC: 5.75-9.45). Conclusions: the raw data presented shows that from March 13 to April 30 of 2020, there was a decrease in emergency visits for precordial pain and a non-significant increase in the incidence rate of out-of-hospital cardiac arrests.

Resumo: Introdução: a pandemia do COVID-19 determinou a aplicação de medidas emergenciais de saúde destinadas a impedir a progressão em nível nacional. Relatórios internacionais sugerem que essas medidas levam à falta de atendimento em outras patologias, principalmente cardiovasculares, e eventualmente aumentam as paradas cardíacas fora do hospital. Objetivo: determinar a frequência da consulta de emergência para dor no peito e paradas cardíacas fora do hospital, assistidas pelos principais serviços de emergência pré-hospitalar de Montevidéu, durante a pandemia do COVID-19. Métodos: foram solicitadas informações às instituições médicas de Montevidéu e de todo o país sobre o número de consultas para dor no peito e total de consultas, no período de 13 de março a 30 de abril de 2018, 2019 e 2020. A frequência de -as paradas cardíacas no hospital foram solicitadas aos serviços de emergência pré-hospitalares durante o mesmo período. Os dados são expressos como frequência absoluta e taxa de incidência (x 100.000) com seu intervalo de confiança estimado por Fisher. Resultados: o número de consultas para dor precordial permaneceu estável no período 2018-2019. Durante 2020, essas consultas representaram entre 11,3% e 21,7% do número total de consultas. Um aumento não significativo da taxa de paradas cardíacas fora do hospital foi evidente em 2020 (9,05, IC 95%: 7,15-11,30) em comparação com 2019 (7,94, IC 95%: 6,19-10,04) e 2018 (7,43, 95%IC: 5,75-9,45), o que não é significativo. Conclusões: os dados brutos apresentados mostram que de 13 de março a 30 de abril de 2020 houve uma diminuição nas visitas de emergência por dor precordial, não aumentando significativamente o número de paradas cardíacas fora do hospital em comparação aos dois anos anteriores.

IC-Tagging methodology applied to the expression of viral glycoproteins and the difficult-to-express membrane-bound IGRP autoantigen.

We have previously developed a methodology to produce protein microspheres (MS) that can be loaded with proteins of interest in living cells through their C or N-terminal tagging with the so-called IC-Tag. The IC-Tagging method has many applications ranging from the production of immobilized enzymes for industrial use to the production of subunit vaccines due to its intrinsic adjuvancy. Here we show the adaptation of the IC-Tagging to work inside the endoplasmic reticulum and bacteria, allowing us to produce properly modified viral glycoproteins. Additionally, we were able to express the Islet-specific glucose-6-phosphatase catalytic subunit-related protein (IGRP), whose expression remained elusive to date possibly due to its toxicity when over-expressed. IGRP is an antigen of enormous pharmaceutical interest as it is specifically targeted during the autoimmune response taking place in both the Non-Obese Diabetic (NOD) mice and type 1 diabetes (T1D) patients leading to the destruction of insulin-producing beta cells.

Using IC-Tagging Methodology for Production and Purification of Epitope-Loaded Protein Microspheres for Vaccination.

Particulate material is more efficient in eliciting immune responses. Here we describe the production of microspheres formed by protein muNS-Mi from avian reoviruses, loaded with foreign epitopes by means of IC-Tagging, for their use as vaccines.

Estudio de una cohorte uruguaya de pacientes portadores de Glioblastoma tratados con radioterapia y Temozolamida / Analysis of a Uruguayan cohort of patients with Glioblastoma treated with radiotherapy and Temozalamide

Objetivos: conocer la sobrevida a dos años de pacientes con diagnóstico de Glioblastoma multiforme (GBM) tratados con Temozolamida con cobertura del Fondo Nacional de Recursos (FNR), evaluar la seguridad del tratamiento y los factores pronósticos. Metodología: cohorte de pacientes portadores de GBM, tratados con Temozolamida entre mayo de 2009 y diciembre 2011. Fuente de información: base de datos del FNR. Resultados: Se incluyeron 81 pacientes. La mediana de sobrevida global fue de 18 meses. Vivos a dos años: 33 de los pacientes. El análisis multivariado encontró a la edad y al peor estado funcional del paciente como los factores de riesgo para mortalidad. 8 pacientes tuvieron efectos adversos severos. Discusión: Se presentan los primeros resultados nacionales vinculados al tratamiento combinado de los GBM. La sobrevida fue similar a la reportada en los estudios de referencia internacionales y mayor que la descripta para el tratamiento radiante exclusivo. El beneficio es mayor en aquellos pacientes jóvenes con buena capacidad funcional previa al inicio del tratamiento. Su seguridad fue considerada como aceptable.

Objectives: find out the two-year survival of patients diagnosed with Multiform Glioblastoma (MGB) treated with Temozolamide funded by the National Resource Fund (FNR), and evaluate the safety of therapy and prognostic factors. Methodology: cohort of patients with MGB, treated with Temozolamide between May 2009 and December 2011. Data source: database of the FNR. Results: the 81 patients included had a median overall survival of 18 months; 33 of them were still alive at two years. The multivariate analysis found that the patient’s age and functional status were the risk factors for mortality. Eight patients had severe adverse effects. Discussion: Presentation of the first national results related to the combined therapy of MGB. Survival was similar to that reported in the international reference studies and greater than that described for radiation therapy alone. Benefits were greater in young patients presenting with a good functional capacity before starting therapy. The safety of therapy was deemed acceptable.

Vacunación antigripal en personal de salud: cobertura, actitudes y barreras contra la vacunación en dos servicios de un hospital general / Influenza immunization in health care professionals: coverage, attitudes and barriers against immunization in two departments of a general hospital

Con el objetivo de conocer la cobertura y las barreras contra la vacunación antigripal, cuya aceptación estimábamos baja entre el personal de salud(PDS) se realizó un estudio transversal, descriptivo, con la metodología de encuesta, autoadministrada con opciones cerradas para su respuesta. Se utilizó una muestra de 142 PDS integrantes del Hospital Pasteur (MSP-ASSE), aleatorizada y adecuada para extraer conclusiones estadísticamente válidas. Los resultados mostraron que 87/142 (61,3%) de los encuestados conocía la campaña de vacunación antigripal en el Hospital, reconocieron tener indicación de vacunación antigripal 124/142 (87,3%) a pesar de lo cual se vacunaron en 2011 79/142 (55,6%) y en 2012 53/142 (37,3%). Los vacunados en 2012 respondieron como motivos principales de adherencia a la campaña protegerse a sí mismo de la enfermedad, pertenecer al PDS y proteger a su familia. De los no vacunados en 2012 respondieron como motivos principales: percepción de nunca haber contraído gripe, miedo a efectos adversos y descrédito al efecto inmunizador de las vacunas. Se confirmó la baja tasa de vacunación del PDS (37,3% en año 2012) a pesar de reconocer la indicación y tener accesibilidad a la misma; se identificaron como barreras las percepciones erróneas acerca de la vacunación...

Tratamiento de la artritis reumatoidea con AntiTNF. Experiencia en Uruguay / Treating rheumatoid arthritis with Anti-TNF. Experience in Uruguay

La artritis reumatoidea (AR) es una enfermedad sistémica autoinmune, que se asocia a afectación en la calidad de vida y a un incremento de la morbimortalidad. Los tratamientos remisivos de la AR incluyen fármacos antiartríticos modificadores de le enfermedad (FAME) y terapias con biológicos. En marzo 2010 se incorpora a las prestaciones del Fondo Nacional de Recursos (FNR) el tratamiento con Antifactor de necrosis tumoral (Anti-TNF) se realiza evaluación de la cohorte de pacientes que iniciaron tratamiento antes del 01/05/2011. Objetivos: caracterizar la población y evaluar resultado de la eficacia del tratamiento, según evolución de puntuación de DAS 28. Resultados: se incluyeron 69 pacientes. Las medias de DAs 28 pre y postratamiento evidenciaron una mejoría (p < 0,05). Criterio de mejoría en 76,4% (bajo grado de actividad 37,3% y remisión 14,9%). Efectos adversos en 20 pacientes (en 3 casos motivó suspensión de la terapia). Cambio de droga, principal causa: falla de tratamiento (8/9). Conclusiones: más de 70% obtiene criterio de mejoría por lo cual los resultados en esta primera evaluación de respuesta pueden calificarse como buenos, aunque deberán monitorizarse en el tiempo.