Repositioning salicylanilide anthelmintic drugs to treat adenovirus infections

The repositioning of drugs already approved by regulatory agencies for other indications is an emerging alternative for the development of new antimicrobial therapies. The repositioning process involves lower risks and costs than the de novo development of novel antimicrobial drugs. Currently, infections by adenovirus show a steady increment with a high clinical impact in immunosuppressed and immunocompetent patients. The lack of a safe and efficacious drug to treat these infections supports the search for new antiviral drugs. Here we evaluated the anti-adenovirus activity of niclosanide, oxyclozanide, and rafoxanide, three salicylanilide anthelmintic drugs. Also, we carried out the cytotoxicity evaluation and partial characterization of the mechanism of action of these drugs. The salicylanilide anthelmintic drugs showed significant anti-adenovirus activity at low micromolar concentrations with little cytotoxicity. Moreover, our mechanistic assays suggest differences in the way the drugs exert anti-adenovirus activity. Niclosamide and rafoxanide target transport of the HAdV particle from the endosome to the nuclear envelope, whilst oxyclozanide specifically targets adenovirus immediately early gene E1A transcription. Data suggests that the studied salicylanilide anthelmintic drugs could be suitable for further clinical evaluation for the development of new antiviral drugs to treat infections by adenovirus in immunosuppressed patients and in immunocompetent individuals with community-acquired pneumonia.Supported by Plan Nacional de I + D + i 2013–2016 and Instituto de Salud Carlos III, Ministerio de Economía, Industria y Competitividad, Spanish Network for Research in Infectious Diseases (REIPI RD16/0016/0009) – co-financed by “A way to achieve Europe” ERDF, the Instituto de Salud Carlos III, Proyectos de Investigación en Salud (PI15/00489) and Proyectos de Desarrollo Tecnológico en Salud (DTS17/00130), and the Spanish Adenovirus Network (AdenoNet, BIO2015/68990-REDT). J.S.C. is supported by the “Contract to Access to the Spanish System of Research and Innovation of the Program of R + D + i of the University of Seville” (USE-13901-D) grant.Peer reviewe

In vitro co-infection by cytomegalovirus improves the antiviral activity of ganciclovir against human adenovirus

Human adenovirus (HAdV) infection has an important clinical impact in the immunosuppressed population and is associated with high morbidity and mortality rates. The lack of a specific, safe and effective antiviral treatment against HAdV makes necessary the search for new therapeutic options. The aim of this study was to evaluate the in vitro activity of ganciclovir (GCV) against HAdV in co-infection by human cytomegalovirus (HCMV) and HAdV in cellular cultures. Quantitative real-time polymerase chain reaction (qPCR) was used to measure HAdV and HCMV DNA replication efficiency in monocultures and in co-infection situations in the presence of both cidofovir (CDV) and GCV. The effects of GCV and CDV were also evaluated in a burst assay (used to measure the production of virus particles) for both viruses, alone and in combination. GCV decreased by 1-log the HAdV DNA replication efficiency in co-infection with HCMV compared with its activity in HAdV monoculture. The burst assay showed that the reductions in virus yield in the presence of GCV were higher for HCMV and co-infection than for HAdV in monoculture (145.2±35.5- vs. 116.4±27.3- vs. 23.0±10.0-fold, respectively, P<0.05). The improved anti-HAdV activity of GCV during co-infection may be because of the more efficient phosphorylation of GCV by the HCMV protein kinase, UL97. Patients treated with GCV as pre-emptive therapy for HCMV infection may be considered as low-risk for developing HAdV infections; however, further evaluations are required to confirm these results.Supported by Plan Nacional de I+D+i 2013‐2016 and Instituto de Salud Carlos III, Subdirección General de Redes y Centros de Investigación Cooperativa, Ministerio de Ciencia, Innovación y Universidades, Spanish Network for Research in Infectious Diseases (REIPI RD16/0016/0009) - co‐financed by European Development Regional Fund “A way to achieve Europe”, Operative program Intelligent Growth 2014‐2020, the Carlos III Health Institute (PI15/00489; DTS17/00130; PI18/01191) and the Spanish Adenovirus Network (AdenoNet, BIO2015/68990-REDT) grants

Marine Natural Products (MNPs) as sources of anti-adenovirus agents

Trabajo presentado en el 2nd AdenoNet Meeting of the Spanish Adenovirus Network, celebrado en Barcelona (España) del 5 al 6 de octubre de 2017.Using a library of 41 extracts generated from marine microorganisms we have identified nine extracts (9, 10, 17, 18, 21, 22, 25, 33 and 37) that showed significant inhibition of HAdV infection with no cytotoxicity at low concentration (concentrations ranging 0,5 and 5 µg/ml). The exposition to these extracts was associated with overall reductions in HAdV yield of two-fold. The impact of these extracts in the HAdV de novo synthesis of DNA was assayed by qPCR and showed significant reductions, ranging 2 to 15 fold.Peer Reviewe

T-cells immune response controls the high incidence of adenovirus infection in adult allogenic hematopoietic transplantation recipients

This work was supported by Plan Nacional de I+D+i 201

T-cells immune response controls the high incidence of adenovirus infection in adult allogenic hematopoietic transplantation recipients

This work was supported by Plan Nacional de I+D+i 2013‐2016 and Instituto de Salud Carlos III, Subdirección General de Redes y Centros de Investigación Cooperativa, Ministerio de Ciencia, Innovación y Universidades, Spanish Network for Research in Infectious Diseases (REIPI RD16/0016/0009) - cofinanced by European Development Regional Fund “A way to achieve Europe”, Operative program Intelligent Growth 2014‐2020, the Instituto de Salud Carlos III, Subdirección General de Evaluación y Fomento de la Investigación (PI15/00489), and the Spanish Adenovirus Network (AdenoNet, BIO2015/68990-REDT). JSC is a researcher belonging to the program “Nicolás Monardes” (C-0059-2018), Servicio Andaluz de Salud, Junta de Andalucía, Spain.Ye

Evaluación de la actividad anti-adenovirus de diferentes librerías químicas de pequeñas moléculas y caracterización de su mecanismo de acción

Motivación: El adenovirus humano (HAdV) es un virus que causa infecciones autolimitadas y leves en individuos inmunocompetentes, en cambio, en pacientes inmunodeprimidos las infecciones suelen ser de caracter grave, presentando elevada morbilidad y mortalidad. En la actualidad no existe un antiviral específico para tratar este tipo de infecciones y los antivirales de amplio espectro disponibles en la práctica clínica habitual no presentan una actividad satisfactoria(1). El objetivo de este proyecto es obtener, partir de diferentes librerías químicas de pequeñas moléculas, compuestos líderes capaces de bloquear la infección por HAdV como primer paso para el desarrollo de un antiviral específico frente HAdV. Métodos: Partimos de una librería de 50 moléculas de una quimioteca de derivados di- y trisustituidos de piperazina. Primero se realizaron ensayos en placa para evaluar su actividad anti-HAdV a una concentración de 10 µM utilizando células 293β5 y el HAdV-GFP no replicativo. Se seleccionaron aquellas moléculas que presentaron actividad inhibidora de la infección superior al 50% y un concentración citotóxica 50 (CC50) superior a 100 µM. Luego, se realizaron diferentes ensayos funcionales para poder identificar en qué punto del ciclo replicativo del virus actuaban dichos compuestos: Ensayo de entrada con el HAdV-GFP y células A549 para evaluar su actividad (50 µM) en etapas tempranas de la entrada del virus en la célula huésped, hasta la liberación de su ADN en el núcleo, y PCR a tiempo real cuantitativa (50 µM) para determinar si aquellos compuesto que no presentaron actividad en fases tempranas presentaban como diana el proceso de replicación del ADN del virus o si por el contrario estaban afectando pasos posteriores de su ciclo infectivo(2). Resultados: Se seleccionaron las moléculas 11, 12, 18 y 19 en base a los resultados del ensayo en placa (inhibición 81%- 96%). Las moléculas 11 y 12 presentaron así mismo inhibición en los ensayos de entrada. La molécula 19 mostró además actividad nhibitoria de la replicación del ADN viral. Conclusiones: Las moléculas 11, 12, 18 y 19 inhiben significativamente la infección por HAdV a bajas concentraciones sin presentar efectos citotóxicos. Además las moléculas 11 y 12 actuarían en fases tempranas del ciclo viral, la molécula 18 ejercería su actividad antiviral durante las fases tardías del ciclo viral y la molécula 19 tendría su diana de acción, al menos en parte, en el proceso de replicación del ADN viral

Optimization of piperazine-derived ureas privileged structures for effective antiadenovirus agents

In recent years, human adenovirus (HAdV) infections have shown a high clinical impact in both immunosuppressed and immunocompetent patients. The research into specific antiviral drugs for the treatment of HAdV infections in immunocompromised patients constitutes a principal objective for medicinal chemistry due to the lack of any specific secure drug to treat these infections. In this study, we report a small-molecule library (67 compounds) designed from an optimization process of piperazine-derived urea privileged structures and their biological evaluation: antiviral activity and cytotoxicity. The active compounds selected were further evaluated to gain mechanistic understanding for their inhibition. Twelve derivatives were identified that inhibited HAdV infections at nanomolar and low micromolar concentrations (IC50 from 0.6 to 5.1 μM) with low cytotoxicity. In addition, our mechanistic assays suggested differences in the way the derivatives exert their anti-HAdV activity targeting transcription, DNA replication and later steps in the HAdV replication cycle. Furthermore, eight of the 12 studied derivatives blocked human cytomegalovirus (HCMV) DNA replication at low micromolar concentrations. The data provided herein indicates that the 12 thiourea/urea piperazine derivatives studied may represent potential lead compounds for clinical evaluation and development of new anti-HAdV drugs.This work has been supported by Ministerio de Economía y Competitividad, Plan Estatal 2013–2016 Retos-Proyectos I + D + i (CTQ2016-78580-C2-2-R) and by Instituto de Salud Carlos III, Subdirección General de Redes y Centros de Investigación Cooperativa, Ministerio de Ciencia, Innovación y Universidades, Spanish Network for Research in Infectious Diseases (REIPI RD16/0016/0009) – co-financed by European Development Regional Fund “A way to achieve Europe”, Operative program Intelligent Growth 2014–2020, the Instituto de Salud Carlos III(DTS17/00130) and the Spanish Adenovirus Network (AdenoNet, BIO2015/68990-REDT) grants. MVH also thanks Ministerio de Economía y Competitividad, Plan Estatal 2013–2016 Excelencia I + D + i (CTQ2016-78703-P). JSC is a researcher belonging to the program “Nicolás Monardes” (C-0059-2018), Servicio Andaluz de Salud, Junta de Andalucía, Spain. FA thanks Dipartimento di Farmacia e Scienze della Salute e della Nutrizione-Dipartimento di Eccellenza MIUR L 232/2016". JAV thanks Junta de Andalucia (Contrato de Garantías Juveniles)