Schistosomiasis: Setting Routes for Drug Discovery

Schistosomiasis is the second most prevalent parasitic disease in the world. Currently, the treatment of this disease relies on a single drug, praziquantel, and due to the identification of resistant parasites, the development of new drugs is urged. The demand for the development of robust high‐throughput parasite screening techniques is increasing as drug discovery research in schistosomiasis gains significance. Here, we review the most common methods used for compound screening in the parasites life stages and also summarize some of the methods that have been recently developed. In addition, we reviewed the methods most commonly implemented to search for promising targets and how they have been used to validate new targets against the parasite Schistosoma mansoni. We also review some promising targets in this parasite and show the main approaches and the major advances that have been achieved by those studies. Moreover, we share our experiences in schistosomiasis drug discovery attained with our S. mansoni drug screening platform establishment

Exploring Host Factors of the Human Metabolism as Promising Targets for Dengue Treatment

The absence of specific therapy and the challenges posed by currently available palliative drugs, such as paracetamol, underscore the urgent need for targeting medications against dengue. Extensive research in the field of antiviral therapies has primarily focused on investigating viral proteins as potential targets. However, despite these efforts, finding an effective therapy for dengue fever remains a daunting task. Importantly, like all viruses, Dengue virus relies on human host proteins to enable infection. Recognizing this fact has prompted the consideration of host factors as viable targets for intervention strategies to combat the infection. This chapter aims to provide an overview of host-virus interactions during Dengue virus infection, emphasizing the importance of metabolic pathways, as well as molecular and cellular processes such as lipid metabolism, autophagy, apoptosis, and the immune system, which are critical for virus propagation. The main goal here is to expand the list of human factors that could serve as potential drug targets. Additionally, molecules that interact with these factors are explored for their therapeutic potential. This comprehensive exploration of host-virus interactions lays the groundwork for more effective dengue treatments. The molecules highlighted here hold promise as antiviral agents, and their inclusion in repurposing research could expedite the development of therapies for dengue fever

Use of BODIPY-Labeled ATP Analogues in the Development and Validation of a Fluorescence Polarization-Based Assay for Screening of Kinase Inhibitors

Copyright © 2020 American Chemical Society. The screening of compound libraries to identify small-molecule modulators of specific biological targets is crucial in the process for the discovery of novel therapeutics and molecular probes. Considering the need for simple single-tool assay technologies with which one could monitor "all" kinases, we developed a fluorescence polarization (FP)-based assay to monitor the binding capabilities of protein kinases to ATP. We used BODIPY ATP-y-S as a probe to measure the shift in the polarization of a light beam when passed through the sample. We were able to optimize the assay using commercial Protein Kinase A (PKA) and H7 efficiently inhibited the binding of the probe when added to the reaction. Furthermore, we were able to employ the assay in a high-throughput fashion and validate the screening of a set of small molecules predicted to dock into the ATP-binding site of PKA. This will be useful to screen larger libraries of compounds that may target protein kinases by blocking ATP binding

Pulmonary hemorrhage in dengue: differential diagnosis with acute viral respiratory syndromes including COVID-19

Clinical similarities among viral diseases become even more relevant considering the current scenario, especially in Brazil, where there is a high incidence of these diseases and overlapping seasonality. We report the case of a patient with acute clinical manifestations composed of predominant respiratory symptoms and alveolar hemorrhage in which three etiologies (dengue, influenza and COVID-19) were investigated concomitantly. Only the diagnosis of dengue was confirmed. Then, the patient’s immunological profile in response to stimulation of mononuclear cells with dengue virus antigen was analyzed in an attempt to identify specific characteristics that could be associated with the clinical manifestation

Schistosoma mansoni coactivator associated arginine methyltransferase 1 (SmCARM1) effect on parasite reproduction

IntroductionThe human blood fluke parasite Schistosoma mansoni relies on diverse mechanisms to adapt to its diverse environments and hosts. Epigenetic mechanisms play a central role in gene expression regulation, culminating in such adaptations. Protein arginine methyltransferases (PRMTs) promote posttranslational modifications, modulating the function of histones and non-histone targets. The coactivator-associated arginine methyltransferase 1 (CARM1/PRMT4) is one of the S. mansoni proteins with the PRMT core domain.MethodsWe carried out in silico analyses to verify the expression of SmPRMTs in public datasets from different infection stages, single-sex versus mixed-worms, and cell types. The SmCARM1 function was evaluated by RNA interference. Gene expression levels were assessed, and phenotypic alterations were analyzed in vitro, in vivo, and ex vivo.ResultsThe scRNAseq data showed that SmPRMTs expression is not enriched in any cell cluster in adult worms or schistosomula, except for Smcarm1 expression which is enriched in clusters of ambiguous cells and Smprmt1 in NDF+ neurons and stem/germinal cells from schistosomula. Smprmt1 is also enriched in S1 and late female germ cells from adult worms. After dsRNA exposure in vitro, we observed a Smcarm1 knockdown in schistosomula and adult worms, 83 and 69%, respectively. Smcarm1-knockdown resulted in reduced oviposition and no significant changes in the schistosomula or adult worm phenotypes. In vivo analysis after murine infection with Smcarm1 knocked-down schistosomula, showed no significant change in the number of worms recovered from mice, however, a significant reduction in the number of eggs recovered was detected. The ex vivo worms presented a significant decrease in the ovary area with a lower degree of cell differentiation, vitelline glands cell disorganization, and a decrease in the testicular lobe area. The worm tegument presented a lower number of tubercles, and the ventral sucker of the parasites presented a damaged tegument and points of detachment from the parasite body.DiscussionThis work brings the first functional characterization of SmCARM1 shedding light on its roles in S. mansoni biology and its potential as a drug target. Additional studies are necessary to investigate whether the reported effects of Smcarm1 knockdown are a consequence of the SmCARM1-mediated methylation of histone tails involved in DNA packaging or other non-histone proteins

Smp38 MAP Kinase Regulation in Schistosoma mansoni: Roles in Survival, Oviposition, and Protection Against Oxidative Stress

Eukaryotic protein kinases (ePKs) are good medical targets for drug development in different biological systems. ePKs participate in many cellular processes, including the p38 MAPK regulation of homeostasis upon oxidative stress. We propose to assess the role of Smp38 MAPK signaling pathway in Schistosoma mansoni development and protection against oxidative stress, parasite survival, and also to elucidate which target genes have their expression regulated by Smp38 MAPK. After a significant reduction of up to 84% in the transcription level by Smp38 MAPK gene knockdown, no visible phenotypic changes were reported in schistosomula in culture. The development of adult worms was tested in vivo in mice infected with the Smp38 knocked-down schistosomula. It was observed that Smp38 MAPK has an essential role in the transformation and survival of the parasites as a low number of adult worms was recovered. Smp38 knockdown also resulted in decreased egg production, damaged adult worm tegument, and underdeveloped ovaries in females. Furthermore, only ~13% of the eggs produced developed into mature eggs. Our results suggest that inhibition of the Smp38 MAPK activity interfere in parasites protection against reactive oxygen species. Smp38 knockdown in adult worms resulted in 80% reduction in transcription levels on the 10th day, with consequent reduction of 94.4% in oviposition in vitro. In order to search for Smp38 MAPK pathway regulated genes, we used an RNASeq approach and identified 1,154 DEGs in Smp38 knockdown schistosomula. A substantial proportion of DEGs encode proteins with unknown function. The results indicate that Smp38 regulates essential signaling pathways for the establishment of parasite homeostasis, including genes related to antioxidant defense, structural composition of ribosomes, spliceosomes, cytoskeleton, as well as, purine and pyrimidine metabolism pathways. Our data show that the Smp38 MAPK signaling pathway is a critical route for parasite development and may present attractive therapeutic targets for the treatment and control of schistosomiasis

Estabelecimento de uma plataforma de teste de compostos em Schistosoma mansoni utilizando inibidores de fatores de modificação epigenética

Submitted by Nuzia Santos ([email protected]) on 2017-12-14T16:11:00Z No. of bitstreams: 1 Naiara Clemente.pdf: 4922764 bytes, checksum: 6459865a412eab6469bbfb9a08f99d06 (MD5)Approved for entry into archive by Nuzia Santos ([email protected]) on 2017-12-14T16:17:24Z (GMT) No. of bitstreams: 1 Naiara Clemente.pdf: 4922764 bytes, checksum: 6459865a412eab6469bbfb9a08f99d06 (MD5)Made available in DSpace on 2017-12-14T16:17:24Z (GMT). No. of bitstreams: 1 Naiara Clemente.pdf: 4922764 bytes, checksum: 6459865a412eab6469bbfb9a08f99d06 (MD5) Previous issue date: 2016CNPqCAPESFAPEMIGFundação Oswaldo Cruz. Centro de Pesquisas René Rachou. Belo Horizonte, MG, Brasil.A esquistossomose é a segunda doença parasitária mais prevalente no mundo. Atualmente, seu tratamento depende de uma única droga , o Praziquantel; e devido a identificação de parasitos resistentes, o desenvolv imento de novas drogas é necessário. No Brasil, essa doença afeta cerca de 25 milhões de pessoas e não há uma plataforma de teste de novas drogas estabelecida . Com o intuito de estabelecer uma plataforma de testes de drogas anti- Schistosoma mansoni , foram padronizadas 3 metodologias para triagem de drogas em esquistossômul os, o ensaio de fluorescência utilizando alamarBlue® , a análise quan titativa de lactato e o ensaio visual por coloração com iodeto de propídio. A análi se quantitativa de lactato e o ensaio visual por coloração com iodeto de propídio demonstraram ser mais sensíveis e confiáveis que o ensaio de fluorescência com alamarBlue® e foram escolhidos para a triagem de drogas em esquistossômulo s. Para a triagem de drogas em vermes adultos foi utilizado o software para análise de movimento WormAssay. Compostos desenvolvidos para fatores de m odificação epigenética por colaboradores do consórcio AParaDDisE foram utilizado s para a validação da plataforma. Após a triagem de drogas em esquistossômulo s, os compostos ativos foram selecionados para a determinação do IC50 e ín dice de seletividade. Nesse trabalho um total de 252 compostos foram testados e 132 foram ativos em S. mansoni , desse, 72 foram ativos em esquistossômulose vermes adultos, 50 foram ativos somente em vermes adultos e 8 foram ativos s omente em esquistossômulos. Dentre os compostos ativos foi possível determinar o índice de seletividade de 29 inibidores. Os compostos JQ1, MC3031, MC2141, MC290 8, MC3668, MC2705, MC3073, C-646 e SPV-106 foram considerados potenciai s drogas para o tratamento da esquistossomose, uma vez que foram ativos nas duas fases do parasito e exibirem alto índice de seletividade. Esses resultad os mostram que os fatores de modificações epigenéticas podem apresentar important e papel na viabilidade e sobrevivência do S. mansoni , confirmando o potencial dessas enzimas como alvo de drogas.Schistosomiasis is the second most prevalent parasit ic disease in the world. Currently, the treatment of this disease rely on a single drug, Praziquantel; and due to the identification of resistant parasites, the d evelopment of new drugs is urged. In Brazil this disease affects around 25 million people and an anti- Schistosoma mansoni drug screening platform is inexistent. In order to s olve this problem, 3 in vitro methodologies were standardized for schistosomula, t he alamarBlue® fluorescence assay, lactate quantitative analysis and visual assay using propidium iodide staining. The lactate quantitative analysis an d visual assay using propidium iodide staining proved to be more sensitive and reli able than the alamarBlue® fluorescence assay, thus were chosen for the schistosomula drug s creening. For the drug screening in adult worms, the movement analysis software WormAssay was standardized and used. Compounds developed for epige netic modification factors by collaborators of A-ParaDDisE consortium were tested fo r platform validation. After the drug screening in schistosomula, the active compo unds were analyzed for its IC50 and selectivity index determination. In this wo rk, a total of 252 compounds were tested and 132 compounds were active in S. mansoni , among those, 73 were active in schistosomula and adult worms, 50 were active in a dult worms, only, and 8 were active in schistosomula only. Among the active compo unds, it was possible to determine the selectivity index of 29 inhibitors. T he compounds JQ1, MC3031, MC2141, MC2908, MC3668, MC2705, MC3073, C-646 e SPV- 106 were considered potential drugs for schistosomiasis therapy conside ring that they were active in both Schistosoma stages and exhibited a high selectivity index and/or low IC50. The results showed that epigenetic modifications factors play an important role in parasite viability and survival, confirming the potential of these enzymes as drug targets

Chapter Schistosomiasis: Setting Routes for Drug Discovery

Air quality sampling campaigns in three European subway systems (Barcelona, Athens and Oporto) were conducted in order to characterise particulate matter (PM) to better understand the main factors controlling it. PM mass concentrations varied among the European subway platforms, and also within the same underground system, this being mainly associated to differences in the design of the stations and tunnels, system age, train frequency, ventilation and air‐conditioning systems, commuter's density, rails geometry and outdoor air quality. PM concentrations displayed clear diurnal patterns, depending largely on the operation and frequency of the trains and the ventilation system. Chemically, subway PM2.5 on the platforms consisted of iron, carbonaceous material, crustal matter, secondary inorganic compounds, insoluble sulphate, halite and trace elements. Fe was the most abundant element, accounting for 19–46% of the bulk PM2.5, which is generated mainly from mechanical wear at rail‐wheel‐brake interfaces. A source apportionment analysis allowed the identification of outdoor (sea salt, fuel‐oil combustion and secondary aerosol) and subway sources on platforms. The use of air‐conditioning inside the trains was an effective approach to reduce exposure concentrations, being more efficient removing coarser particles. PM concentrations inside the trains were greatly affected by the surrounding (i.e. platforms and tunnels) air quality conditions

Iron oxide polyaniline-coated nanoparticles modulate tumor microenvironment in breast cancer: an in vitro study on the reprogramming of tumor-associated macrophages

Abstract Background Breast cancer is the neoplastic disease with the highest incidence and mortality in the female population worldwide. Treatment remains challenging due to various factors. Therefore, it is of great importance to develop new therapeutic strategies that promote the safe destruction of neoplastic cells without compromising patients' quality of life. Among advances in the treatment of breast cancer, immunotherapy stands out as a promising trend. Recent studies have demonstrated the potential of iron oxide nanoparticles in promoting the reprogramming of M2 macrophages (pro-tumor phenotype) into M1 macrophages (anti-tumor phenotype) within the tumor microenvironment, resulting in potent antitumor effects. In this study, the effect of polyaniline-coated iron oxide nanoparticles (Pani/y-Fe2O3) on macrophage polarization and breast cancer cell death was investigated. Methods The non-cytotoxic concentration of nanoparticles was determined using the MTT assay. For in vitro co-culture experiments, breast cancer cell lines MCF -7 and MDA-MB -231 and macrophages THP-1 were co-cultured in a Transwell system and then the effects of Pani/y-Fe2O3 on cell viability, gene expression, cytokine profile, and oxidative stress markers were investigated. Results The results showed that Pani/y-Fe2O3 nanoparticles induced M2-to-M1 macrophage polarization in both cell lines through different pathways. In MCF -7 and THP-1 macrophage co-culture, the study showed a decrease in cytokine levels IL -1β, upregulation of M1-associated genes (IL-12, TNF-α) in macrophages, resulting in increased MCF -7 cell death by apoptosis (caspase 3/7+). In MDA-MB -231 co-cultures, increases in cytokines IL -6, IL -1β, and oxidative stress markers were observed, as well as upregulation of the inducible nitric oxide synthase (iNOS) gene in macrophages, leading to tumor cell death via apoptosis-independent pathways (Sytox+). Conclusions These findings highlight the potential of Pani/y-Fe2O3 as a promising therapeutic approach in the context of breast cancer treatment by effectively reprogramming M2 macrophages into an anti-tumor M1 phenotype, Pani/y-Fe2O3 nanoparticles demonstrated the ability to elicit antitumor effects in both MCF-7 and MDA-MB-231 breast cancer cell lines

Whole-genome sequencing reveals host factors underlying critical COVID-19

Altres ajuts: Department of Health and Social Care (DHSC); Illumina; LifeArc; Medical Research Council (MRC); UKRI; Sepsis Research (the Fiona Elizabeth Agnew Trust); the Intensive Care Society, Wellcome Trust Senior Research Fellowship (223164/Z/21/Z); BBSRC Institute Program Support Grant to the Roslin Institute (BBS/E/D/20002172, BBS/E/D/10002070, BBS/E/D/30002275); UKRI grants (MC_PC_20004, MC_PC_19025, MC_PC_1905, MRNO2995X/1); UK Research and Innovation (MC_PC_20029); the Wellcome PhD training fellowship for clinicians (204979/Z/16/Z); the Edinburgh Clinical Academic Track (ECAT) programme; the National Institute for Health Research, the Wellcome Trust; the MRC; Cancer Research UK; the DHSC; NHS England; the Smilow family; the National Center for Advancing Translational Sciences of the National Institutes of Health (CTSA award number UL1TR001878); the Perelman School of Medicine at the University of Pennsylvania; National Institute on Aging (NIA U01AG009740); the National Institute on Aging (RC2 AG036495, RC4 AG039029); the Common Fund of the Office of the Director of the National Institutes of Health; NCI; NHGRI; NHLBI; NIDA; NIMH; NINDS.Critical COVID-19 is caused by immune-mediated inflammatory lung injury. Host genetic variation influences the development of illness requiring critical care or hospitalization after infection with SARS-CoV-2. The GenOMICC (Genetics of Mortality in Critical Care) study enables the comparison of genomes from individuals who are critically ill with those of population controls to find underlying disease mechanisms. Here we use whole-genome sequencing in 7,491 critically ill individuals compared with 48,400 controls to discover and replicate 23 independent variants that significantly predispose to critical COVID-19. We identify 16 new independent associations, including variants within genes that are involved in interferon signalling (IL10RB and PLSCR1), leucocyte differentiation (BCL11A) and blood-type antigen secretor status (FUT2). Using transcriptome-wide association and colocalization to infer the effect of gene expression on disease severity, we find evidence that implicates multiple genes-including reduced expression of a membrane flippase (ATP11A), and increased expression of a mucin (MUC1)-in critical disease. Mendelian randomization provides evidence in support of causal roles for myeloid cell adhesion molecules (SELE, ICAM5 and CD209) and the coagulation factor F8, all of which are potentially druggable targets. Our results are broadly consistent with a multi-component model of COVID-19 pathophysiology, in which at least two distinct mechanisms can predispose to life-threatening disease: failure to control viral replication; or an enhanced tendency towards pulmonary inflammation and intravascular coagulation. We show that comparison between cases of critical illness and population controls is highly efficient for the detection of therapeutically relevant mechanisms of disease