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mRNA biogenesis in the eukaryotic nucleus is a highly complex process. The numerous RNA processing steps are tightly coordinated to ensure that only fully processed transcripts are released from chromatin for export from the nucleus. Here, we present the hypothesis that fission yeast Dbp2, a ribonucleoprotein complex (RNP) remodelling ATPase of the DEAD-box family, is the key enzyme in an RNP assembly checkpoint at the 3'-end of genes. We show that Dbp2 interacts with the cleavage and polyadenylation complex (CPAC) and localises to cleavage bodies, which are enriched for 3'-end processing factors and proteins involved in nuclear RNA surveillance. Upon loss of Dbp2, 3'-processed, polyadenylated RNAs accumulate on chromatin and in cleavage bodies, and CPAC components are depleted from the soluble pool. Under these conditions, cells display an increased likelihood to skip polyadenylation sites and a delayed transcription termination, suggesting that levels of free CPAC components are insufficient to maintain normal levels of 3'-end processing. Our data support a model in which Dbp2 is the active component of an mRNP remodelling checkpoint that licenses RNA export and is coupled to CPAC release.
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RNA Helicases DEAD-box , Ribonucleoproteínas , Proteínas de Schizosaccharomyces pombe , Schizosaccharomyces , Schizosaccharomyces/genética , Schizosaccharomyces/metabolismo , Ribonucleoproteínas/metabolismo , Ribonucleoproteínas/genética , RNA Helicases DEAD-box/metabolismo , RNA Helicases DEAD-box/genética , Proteínas de Schizosaccharomyces pombe/metabolismo , Proteínas de Schizosaccharomyces pombe/genética , Poliadenilação , RNA Mensageiro/metabolismo , RNA Mensageiro/genética , Fatores de Poliadenilação e Clivagem de mRNA/metabolismo , Fatores de Poliadenilação e Clivagem de mRNA/genética , Cromatina/metabolismo , RNA Fúngico/metabolismo , RNA Fúngico/genética , Núcleo Celular/metabolismoRESUMO
The use of Fpocket and virtual screening techniques enabled us to identify potential allosteric druggable pockets within the SARS-CoV-2 RNA-dependent RNA polymerase (RdRp). Of the compounds screened, compound 1 was identified as a promising inhibitor, lowering a SARS-CoV-2 RdRp activity to 57% in an enzymatic assay at 10 µM concentration. The structure of compound 1 was subsequently optimized in order to preserve or enhance inhibitory activity. This involved the substitution of problematic ester and aromatic nitro groups with more inert functionalities. The N,N'-diphenylurea scaffold with two NH groups was identified as essential for the compound's activity but also exhibited high toxicity in Calu-3 cells. To address this issue, a scaffold hopping approach was employed to replace the urea core with potentially less toxic urea isosteres. This approach yielded several structural analogues with notable activity, specifically 2,2'-bisimidazol (in compound 55 with residual activity RA = 42%) and (1H-imidazol-2-yl)urea (in compounds 59 and 60, with RA = 50 and 28%, respectively). Despite these advances, toxicity remained a major concern. These compounds represent a promising starting point for further structure-activity relationship studies of allosteric inhibitors of SARS-CoV-2 RdRp, with the goal of reducing their cytotoxicity and improving aqueous solubility.
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Infection control measures to prevent viral and bacterial infection spread are critical to maintaining a healthy environment. Pathogens such as viruses and pyogenic bacteria can cause infectious complications. Viruses such as SARS-CoV-2 are known to spread through the aerosol route and on fomite surfaces, lasting for a prolonged time in the environment. Developing technologies to mitigate the spread of pathogens through airborne routes and on surfaces is critical, especially for patients at high risk for infectious complications. Multifunctional coatings with a broad capacity to bind pathogens that result in inactivation can disrupt infectious spread through aerosol and inanimate surface spread. This study uses C-POLAR, a proprietary cationic, polyamine, organic polymer with a charged, dielectric property coated onto air filtration material and textiles. Using both SARS-CoV-2 live viral particles and bovine coronavirus models, C-POLAR-treated material shows a dramatic 2-log reduction in circulating viral inoculum. This reduction is consistent in a static room model, indicating simple airflow through a static C-POLAR hanging can capture significant airborne particles. Finally, Gram-positive and Gram-negative bacteria are applied to C-POLAR textiles using a viability indicator to demonstrate eradication on fomite surfaces. These data suggest that a cationic polymer surface can capture and eradicate human pathogens, potentially interrupting the infectious spread for a more resilient environment. IMPORTANCE: Infection control is critical for maintaining a healthy home, work, and hospital environment. We test a cationic polymer capable of capturing and eradicating viral and bacterial pathogens by applying the polymer to the air filtration material and textiles. The data suggest that the simple addition of cationic material can result in the improvement of an infectious resilient environment against viral and bacterial pathogens.
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BACKGROUND: Patients with congestive heart failure (CHF), chronic obstructive pulmonary disease (COPD) and dementia are underrepresented in specialist palliative home care (SPHC). However, the complexity of their conditions requires collaboration between general practitioners (GPs) and SPHC teams and timely integration into SPHC to effectively meet their needs. OBJECTIVE: To facilitate joint palliative care planning and the timely transfer of patients with advanced chronic non-malignant conditions to SPHC. METHODS: A two-arm, unblinded, cluster-randomised controlled trial. 49 GP practices in northern Germany were randomised using web-based block randomisation. We included patients with advanced CHF, COPD and/or dementia. The KOPAL intervention consisted of a SPHC nurse-patient consultation followed by an interprofessional telephone case conference between SPHC team and GP. The primary outcome was the number of hospital admissions 48 weeks after baseline. Secondary analyses examined the effects on health-related quality of life and self-rated health status, as measured by the EuroQol 5D scale. RESULTS: A total of 172 patients were included in the analyses. 80.4% of GP practices had worked with SHPC before, most of them exclusively for cancer patients. At baseline, patients reported a mean EQ-VAS of 48.4, a mean quality of life index (EQ-5D-5L) of 0.63 and an average of 0.80 hospital admissions in the previous year. The intervention did not significantly reduce hospital admissions (incidence rate ratio = 0.79, 95%CI: [0.49, 1.26], P = 0.31) or the number of days spent in hospital (incidence rate ratio = 0.65, 95%CI: [0.28, 1.49], P = 0.29). There was also no significant effect on quality of life (∆ = -0.02, 95%CI: [-0.09, 0.05], P = 0.53) or self-rated health (∆ = -2.48, 95%CI: [-9.95, 4.99], P = 0.51). CONCLUSIONS: The study did not show the hypothesised effect on hospitalisations and health-related quality of life. Future research should focus on refining this approach, with particular emphasis on optimising the timing of case conferences and implementing discussed changes to treatment plans, to improve collaboration between GPs and SPHC teams.
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Insuficiência Cardíaca , Cuidados Paliativos , Atenção Primária à Saúde , Doença Pulmonar Obstrutiva Crônica , Qualidade de Vida , Humanos , Cuidados Paliativos/métodos , Masculino , Feminino , Idoso , Alemanha , Doença Pulmonar Obstrutiva Crônica/terapia , Doença Pulmonar Obstrutiva Crônica/diagnóstico , Idoso de 80 Anos ou mais , Insuficiência Cardíaca/terapia , Insuficiência Cardíaca/diagnóstico , Demência/terapia , Doença Crônica , Serviços de Assistência Domiciliar , Equipe de Assistência ao Paciente , Fatores de Tempo , Comunicação Interdisciplinar , Prestação Integrada de Cuidados de Saúde/organização & administraçãoRESUMO
Nicotinamide adenine dinucleotide (NAD) is a critical component of the cellular metabolism and also serves as an alternative 5' cap on various RNAs. However, the function of the NAD RNA cap is still under investigation. We studied NAD capping of RNAs in HIV-1-infected cells because HIV-1 is responsible for the depletion of the NAD/NADH cellular pool and causing intracellular pellagra. By applying the NAD captureSeq protocol to HIV-1-infected and uninfected cells, we revealed that four snRNAs (e.g., U1) and four snoRNAs lost their NAD cap when infected with HIV-1. Here, we provide evidence that the presence of the NAD cap decreases the stability of the U1/HIV-1 pre-mRNA duplex. Additionally, we demonstrate that reducing the quantity of NAD-capped RNA by overexpressing the NAD RNA decapping enzyme DXO results in an increase in HIV-1 infectivity. This suggests that NAD capping is unfavorable for HIV-1 and plays a role in its infectivity.
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Infecções por HIV , HIV-1 , NAD , RNA Nuclear Pequeno , RNA Nucleolar Pequeno , Humanos , NAD/metabolismo , RNA Nucleolar Pequeno/metabolismo , RNA Nucleolar Pequeno/genética , RNA Nuclear Pequeno/metabolismo , Infecções por HIV/virologia , Infecções por HIV/metabolismo , Capuzes de RNA/metabolismoRESUMO
Stability and cytotoxicity of PEGylated Au NPs is crucial for biomedical application. In this study, we have focused on thermal stability of PEGylated Au NPs at 4 and 37 °C and after sterilization in autoclave. Gold nanoparticles were prepared by direct sputtering of gold into PEG and PEG-NH2. Transmission electron microscopy revealed that NPs exhibit a spherical shape with average dimensions 3.8 nm for both AuNP_PEG and AuNP_PEG-NH2. The single LSPR band at wavelength of 509 nm also confirmed presence of spherical Au NPs in both cases. Moreover, according to UV-Vis spectra, the Au NPs were overall stable during aging or thermal stressing and even after sterilization in autoclave. Based on gel electrophoresis results, the higher density of functionalizing ligands and the higher stability is assumed on AuNP_PEG-NH2. Changes in concentration of gold did not occur after thermal stress or with aging. pH values have to be adjusted to be suitable for bioapplications - original pH values are either too alkaline (AuNP_PEG-NH2, pH 10) or too acidic (AuNP_PEG, pH 5). Cytotoxicity was tested on human osteoblasts and fibroblasts. Overall, both Au NPs have shown good cytocompatibility either freshly prepared or even after Au NPs' sterilization in the autoclave. Prepared Au NP dispersions were also examined for their antiviral activity, however no significant effect was observed. We have synthesized highly stable, non-cytotoxic PEGylated Au NPs, which are ready for preclinical testing.
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Adhesion G protein-coupled receptors (aGPCRs) play an important role in neurodevelopment, immune defence and cancer; however, their role throughout viral infections is mostly unexplored. We have been searching for specific aGPCRs involved in SARS-CoV-2 infection of mammalian cells. In the present study, we infected human epithelial cell lines derived from lung adenocarcinoma (Calu-3) and colorectal carcinoma (Caco-2) with SARS-CoV-2 in order to analyse changes in the level of mRNA encoding individual aGPCRs at 6 and 12 h post infection. Based on significantly altered mRNA levels, we identified four aGPCR candidates-ADGRB3/BAI3, ADGRD1/GPR133, ADGRG7/GPR128 and ADGRV1/GPR98. Of these receptors, ADGRD1/GPR133 and ADGRG7/GPR128 showed the largest increase in mRNA levels in SARS-CoV-2-infected Calu-3 cells, whereas no increase was observed with heat-inactivated SARS-CoV-2 and virus-cleared conditioned media. Next, using specific siRNA, we downregulated the aGPCR candidates and analysed SARS-CoV-2 entry, replication and infectivity in both cell lines. We observed a significant decrease in the amount of SARS-CoV-2 newly released into the culture media by cells with downregulated ADGRD1/GPR133 and ADGRG7/GPR128. In addition, using a plaque assay, we observed a reduction in SARS-CoV-2 infectivity in Calu-3 cells. In summary, our data suggest that selected aGPCRs might play a role during SARS-CoV-2 infection of mammalian cells.
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Adenocarcinoma de Pulmão , COVID-19 , RNA Mensageiro , Receptores Acoplados a Proteínas G , SARS-CoV-2 , Regulação para Cima , Humanos , Receptores Acoplados a Proteínas G/metabolismo , Receptores Acoplados a Proteínas G/genética , SARS-CoV-2/genética , SARS-CoV-2/fisiologia , SARS-CoV-2/metabolismo , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , COVID-19/genética , COVID-19/virologia , COVID-19/metabolismo , Adenocarcinoma de Pulmão/genética , Adenocarcinoma de Pulmão/virologia , Adenocarcinoma de Pulmão/patologia , Adenocarcinoma de Pulmão/metabolismo , Regulação para Cima/genética , Linhagem Celular Tumoral , Neoplasias Pulmonares/genética , Neoplasias Pulmonares/virologia , Neoplasias Pulmonares/patologia , Neoplasias Pulmonares/metabolismo , Células CACO-2RESUMO
The evolutionary processes that underlie the marked sensitivity of small cell lung cancer (SCLC) to chemotherapy and rapid relapse are unknown1-3. Here we determined tumour phylogenies at diagnosis and throughout chemotherapy and immunotherapy by multiregion sequencing of 160 tumours from 65 patients. Treatment-naive SCLC exhibited clonal homogeneity at distinct tumour sites, whereas first-line platinum-based chemotherapy led to a burst in genomic intratumour heterogeneity and spatial clonal diversity. We observed branched evolution and a shift to ancestral clones underlying tumour relapse. Effective radio- or immunotherapy induced a re-expansion of founder clones with acquired genomic damage from first-line chemotherapy. Whereas TP53 and RB1 alterations were exclusively part of the common ancestor, MYC family amplifications were frequently not constituents of the founder clone. At relapse, emerging subclonal mutations affected key genes associated with SCLC biology, and tumours harbouring clonal CREBBP/EP300 alterations underwent genome duplications. Gene-damaging TP53 alterations and co-alterations of TP53 missense mutations with TP73, CREBBP/EP300 or FMN2 were significantly associated with shorter disease relapse following chemotherapy. In summary, we uncover key processes of the genomic evolution of SCLC under therapy, identify the common ancestor as the source of clonal diversity at relapse and show central genomic patterns associated with sensitivity and resistance to chemotherapy.
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Evolução Molecular , Imunoterapia , Neoplasias Pulmonares , Platina , Carcinoma de Pequenas Células do Pulmão , Animais , Feminino , Humanos , Masculino , Camundongos , Pessoa de Meia-Idade , Células Clonais/efeitos dos fármacos , Células Clonais/metabolismo , Células Clonais/patologia , Resistencia a Medicamentos Antineoplásicos/efeitos dos fármacos , Resistencia a Medicamentos Antineoplásicos/genética , Genes myc/genética , Neoplasias Pulmonares/genética , Neoplasias Pulmonares/imunologia , Neoplasias Pulmonares/patologia , Neoplasias Pulmonares/terapia , Mutação , Recidiva Local de Neoplasia/genética , Recidiva Local de Neoplasia/patologia , Platina/farmacologia , Platina/uso terapêutico , Recidiva , Carcinoma de Pequenas Células do Pulmão/genética , Carcinoma de Pequenas Células do Pulmão/imunologia , Carcinoma de Pequenas Células do Pulmão/patologia , Carcinoma de Pequenas Células do Pulmão/terapiaRESUMO
Introduction: Immune checkpoint inhibitors have advanced the outcomes of many different types of cancer. A rare but extraordinarily severe complication of these agents resembles immune checkpoint inhibitor-related myocarditis, which typically occurs within the first few weeks after treatment initiation with a mortality of 25%-50%. Case report: A 57-year-old woman had uneventfully received pembrolizumab for metastatic non-small cell lung cancer for over 2.5 years and was admitted after an out-of-hospital cardiac arrest due to ventricular fibrillation. After successful cardiopulmonary resuscitation, the initial diagnostic work-up showed elevated cardiac enzymes and a limited left-ventricular ejection fraction, while coronary angiography did not show relevant stenosis. Despite cardiac MRI being unsuggestive of myocarditis, myocardial biopsies were obtained and histologically confirmed anti-PD-1 antibody-associated myocarditis. After the initiation of prednisone at 1â mg/kg body weight, the patient gradually recovered and was discharged three weeks later with markedly improved cardiac function. Conclusion: This case resembles the first description of a very late onset irMyocarditis, occurring over 2.5 years after the start of treatment. It demonstrates the importance of contemplating that severe immune-related toxicities with a sudden onset clinical presentation may occur even after long uneventful periods of anti-PD-1 immune checkpoint inhibitor treatment. Furthermore, it underlines the critical importance of myocardial biopsies in this setting, especially when cardiac MRI remains inconclusive. Moreover, it demonstrates the necessity and benefits of early immunosuppressive treatment if immune-related myocarditis is considered a differential diagnosis.
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Highly sulfated malto-oligomers, similar to heparin and heparan-sulfate, have good antiviral, antimetastatic, anti-inflammatory and cell growth inhibitory effects. Due to their broad biological activities and simple structure, sulfated malto-oligomer derivatives have a great therapeutic potential, therefore, the development of efficient synthesis methods for their production is of utmost importance. In this work, preparation of α-(1â4)-linked oligoglucosides containing a sulfonatomethyl moiety at position C-6 of each glucose unit was studied by different approaches. Malto-oligomeric sulfonic acid derivatives up to dodecasaccharides were prepared by polymerization using different protecting groups, and the composition of the product mixtures was analyzed by MALDI-MS methods and size-exclusion chromatography. Synthesis of lower oligomers was also accomplished by stepwise and block synthetic methods, and then the oligosaccharide products were persulfated. The antiviral, anti-inflammatory and cell growth inhibitory activity of the fully sulfated malto-oligosaccharide sulfonic acids were determined by in vitro tests. Four tested di- and trisaccharide sulfonic acids effectively inhibited the activation of the TNF-α-mediated inflammatory pathway without showing cytotoxicity.
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Oligossacarídeos , Sulfatos , Polimerização , Oligossacarídeos/farmacologia , Ácidos Sulfônicos , Anti-Inflamatórios/farmacologia , Antivirais/farmacologiaRESUMO
Contextual cues and prior evidence guide human goal-directed behavior. The neurophysiological mechanisms that implement contextual priors to guide subsequent actions in the human brain remain unclear. Using intracranial electroencephalography (iEEG), we demonstrate that increasing uncertainty introduces a shift from a purely oscillatory to a mixed processing regime with an additional ramping component. Oscillatory and ramping dynamics reflect dissociable signatures, which likely differentially contribute to the encoding and transfer of different cognitive variables in a cue-guided motor task. The results support the idea that prefrontal activity encodes rules and ensuing actions in distinct coding subspaces, while theta oscillations synchronize the prefrontal-motor network, possibly to guide action execution. Collectively, our results reveal how two key features of large-scale neural population activity, namely continuous ramping dynamics and oscillatory synchrony, jointly support rule-guided human behavior.
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Encéfalo , Sinais (Psicologia) , Humanos , Encéfalo/fisiologia , Ritmo Teta/fisiologia , EletroencefalografiaRESUMO
The COVID-19 pandemic, caused by the SARS-CoV-2 virus, has led to significant global morbidity and mortality. A crucial viral protein, the non-structural protein 14 (nsp14), catalyzes the methylation of viral RNA and plays a critical role in viral genome replication and transcription. Due to the low mutation rate in the nsp region among various SARS-CoV-2 variants, nsp14 has emerged as a promising therapeutic target. However, discovering potential inhibitors remains a challenge. In this work, we introduce a computational pipeline for the rapid and efficient identification of potential nsp14 inhibitors by leveraging virtual screening and the NCI open compound collection, which contains 250,000 freely available molecules for researchers worldwide. The introduced pipeline provides a cost-effective and efficient approach for early-stage drug discovery by allowing researchers to evaluate promising molecules without incurring synthesis expenses. Our pipeline successfully identified seven promising candidates after experimentally validating only 40 compounds. Notably, we discovered NSC620333, a compound that exhibits a strong binding affinity to nsp14 with a dissociation constant of 427 ± 84 nM. In addition, we gained new insights into the structure and function of this protein through molecular dynamics simulations. We identified new conformational states of the protein and determined that residues Phe367, Tyr368, and Gln354 within the binding pocket serve as stabilizing residues for novel ligand interactions. We also found that metal coordination complexes are crucial for the overall function of the binding pocket. Lastly, we present the solved crystal structure of the nsp14-MTase complexed with SS148 (PDB:8BWU), a potent inhibitor of methyltransferase activity at the nanomolar level (IC50 value of 70 ± 6 nM). Our computational pipeline accurately predicted the binding pose of SS148, demonstrating its effectiveness and potential in accelerating drug discovery efforts against SARS-CoV-2 and other emerging viruses.
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(-)-Cannabidiol (CBD) and (-)-cannabigerol (CBG) are two major non-psychotropic phytocannabinoids that have many beneficial biological properties. However, due to their low water solubility and prominent first-pass metabolism, their oral bioavailability is moderate, which is unfavorable for medicinal use. Therefore, there is a great need for appropriate chemical modifications to improve their physicochemical and biological properties. In this study, Mannich-type reaction was used for the synthetic modification of CBD and CBG for the first time, and thus fifteen new cannabinoid derivatives containing one or two tertiary amino groups were prepared. Thereafter the antiviral, antiproliferative and antibacterial properties of the derivatives and their effects on certain skin cells were investigated. Some modified CBD derivatives showed remarkable antiviral activity against SARS-CoV-2 without cytotoxic effect, while synthetic modifications on CBG resulted in a significant increase in antiproliferative activity in some cases compared to the parent compound.
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Canabidiol , Canabinoides , Canabidiol/farmacologia , Canabinoides/farmacologia , Disponibilidade Biológica , Antivirais/farmacologiaRESUMO
Aim: The realities of emergency care and resuscitation research involving nursing home (NH) residents suggest an overuse of resuscitation attempts in NHs. A complete analysis of all NH resident deaths is needed to provide a complementary perspective of potential underuse. The present research investigated whether residents of different NH homes died at the NH during attempted resuscitation or after transfer to hospital. Methods: A full survey of resuscitation attempts and deaths among NH residents, via retrospective analysis of data from the death registry and the German Resuscitation Registry for the years 2018 to 2021. Results: Over the 4-year study period, 14,598 individuals died, of whom 3,288 (22.5%) were residents of 31 different NHs. The mean age of the deceased NH residents was 87 years (±8.6); 2,196 (66.8%) were female, 118 (3.6%) underwent a resuscitation attempt, and 58.5% died at the NH. NH averages were as follows: deaths per NH: 106 (±51; min-max: 36-292); number of beds: 102 (±39; 34-210); deaths per bed per year 0.27 (±0.07; 0.15-0.51); resuscitation attempts per 1,000 beds per year: 9.5 (±5.5; 0-21.1); and ratio of futile resuscitation attempts to deaths: 6.0% (0-12.5%). Considering the entire study region before and during the COVID-19 pandemic, a slight underuse of resuscitation attempts with female NH residents emerged. On a facility level, substantial disparities and opposing trends were found. The incidence of deaths and resuscitation attempts, as well as the place of death and the ratio of futile resuscitation attempts to deaths, varied considerably. Conclusion: Resuscitation attempts are rarely administered to dying NH residents. However, their frequency varies considerably between NHs.
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BACKGROUND: Worldwide, progressive chronic, non-malignant diseases are highly prevalent. Especially with increasing age, they are characterised by high hospitalisation rates and high healthcare costs. Improved interprofessional collaboration between general practitioners (GPs) and specialist palliative home care (SPHC) teams might reduce hospitalisation while improving symptoms and quality of life, or preventing them from deterioration. The aim of this study was to examine the cost-effectiveness of a newly developed intervention in patients with advanced chronic, non-malignant diseases consisting of a structured palliative care nurse-patient consultation followed by an interprofessional telephone case conference. METHODS: The analysis was based on data from 172 participants of the KOPAL multi-centre, cluster randomised controlled trial. Patients with advanced congestive heart failure (CHF), chronic obstructive pulmonary disease (COPD), or dementia were randomised into intervention group (IG) and control group (CG, usual care). Cost-effectiveness was examined over 48 weeks from a societal and healthcare payer's perspective. Effects were quantified as quality-adjusted life years (QALYs, EQ-5D-5L). Incremental cost-effectiveness ratios were calculated and cost-effectiveness acceptability curves were constructed. RESULTS: Baseline imbalances in costs and effects could be observed between IG and CG. After adjusting for these imbalances and compared to the CG, mean costs in the IG were non-significantly higher from a societal and lower from a payer's perspective. On the effect side, the IG had marginally lower mean QALYs. The results were characterized by high statistical uncertainty, indicated by large confidence intervals for the cost and effect differences between groups and probabilities of cost-effectiveness between 18% and 65%, depending on the perspective and willingness-to-pay. CONCLUSIONS: Based on the results of this study, the cost-effectiveness of the KOPAL intervention was uncertain. The results highlighted (methodological) challenges of economic evaluations in patients with chronic, non-malignant diseases related to sample size, heterogeneity of participants, and the way the intervention effectiveness is typically captured in economic evaluations.
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Cuidados Paliativos , Qualidade de Vida , Humanos , Análise Custo-Benefício , Doença Crônica , Encaminhamento e Consulta , Anos de Vida Ajustados por Qualidade de VidaRESUMO
The RNA 2'-O methyltransferase (MTase) VP39 of the monkeypox virus (MpxV) participates in RNA capping within poxviruses. Sub-micromolar inhibitors targeting this enzyme were already reported. However, these 7-deaza analogs of S-adenosyl methionine (SAH) had not been tested in cellular assays until now. In this study, we employed plaque assays and cytopathic effect-based assays to evaluate the effectiveness of these compounds. All tested compounds demonstrated antiviral activity against MpxV, with EC50 values ranging from 0.06 to 2.7 µM. Nevertheless, some of these compounds also exhibited cytotoxicity in HeLa cells, while others showed no toxicity. Notably, the non-toxic compounds featured a large aromatic substituent at the 7-deaza position, whereas the toxic compounds had a small substituent at the same position. These findings suggest that VP39 represents a bona fide target for the development of antiviral drugs against MpxV.
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The search for new drugs against COVID-19 and its causative agent, SARS-CoV-2, is one of the major trends in the current medicinal chemistry. Targeting capping machinery could be one of the therapeutic concepts based on a unique mechanism of action. Viral RNA cap synthesis involves two methylation steps, the first of which is mediated by the nsp14 protein. Here, we rationally designed and synthesized a series of compounds capable of binding to both the S-adenosyl-l-methionine and the RNA-binding site of SARS-CoV-2 nsp14 N7-methyltransferase. These hybrid molecules showed excellent potency, high selectivity toward various human methyltransferases, nontoxicity, and high cell permeability. Despite the outstanding activity against the enzyme, our compounds showed poor antiviral performance in vitro. This suggests that the activity of this viral methyltransferase has no significant effect on virus transcription and replication at the cellular level. Therefore, our compounds represent unique tools to further explore the role of the SARS-CoV-2 nsp14 methyltransferase in the viral life cycle and the pathogenesis of COVID-19.
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Molybdenum (Mo) as essential micronutrient for plants, acts as active component of molybdenum cofactor (Moco). Core metabolic processes like nitrate assimilation or abscisic-acid biosynthesis rely on Moco-dependent enzymes. Although a family of molybdate transport proteins (MOT1) is known to date in Arabidopsis, molybdate homeostasis remained unclear. Here we report a second family of molybdate transporters (MOT2) playing key roles in molybdate distribution and usage. KO phenotype-analyses, cellular and organ-specific localization, and connection to Moco-biosynthesis enzymes via protein-protein interaction suggest involvement in cellular import of molybdate in leaves and reproductive organs. Furthermore, we detected a glutathione-molybdate complex, which reveals how vacuolar storage is maintained. A putative Golgi S-adenosyl-methionine transport function was reported recently for the MOT2-family. Here, we propose a moonlighting function, since clear evidence of molybdate transport was found in a yeast-system. Our characterization of the MOT2-family and the detection of a glutathione-molybdate complex unveil the plant-wide way of molybdate.
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Arabidopsis , Arabidopsis/genética , Arabidopsis/metabolismo , Molibdênio/metabolismo , Proteínas de Membrana Transportadoras/genética , Proteínas de Membrana Transportadoras/metabolismo , Pteridinas , HomeostaseRESUMO
The human prefrontal cortex (PFC) constitutes the structural basis underlying flexible cognitive control, where mixed-selective neural populations encode multiple task features to guide subsequent behavior. The mechanisms by which the brain simultaneously encodes multiple task-relevant variables while minimizing interference from task-irrelevant features remain unknown. Leveraging intracranial recordings from the human PFC, we first demonstrate that competition between coexisting representations of past and present task variables incurs a behavioral switch cost. Our results reveal that this interference between past and present states in the PFC is resolved through coding partitioning into distinct low-dimensional neural states; thereby strongly attenuating behavioral switch costs. In sum, these findings uncover a fundamental coding mechanism that constitutes a central building block of flexible cognitive control.
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Cognição , Córtex Pré-Frontal , HumanosRESUMO
Proteases encoded by SARS-CoV-2 constitute a promising target for new therapies against COVID-19. SARS-CoV-2 main protease (Mpro, 3CLpro) and papain-like protease (PLpro) are responsible for viral polyprotein cleavage-a process crucial for viral survival and replication. Recently it was shown that 2-phenylbenzisoselenazol-3(2H)-one (ebselen), an organoselenium anti-inflammatory small-molecule drug, is a potent, covalent inhibitor of both the proteases and its potency was evaluated in enzymatic and antiviral assays. In this study, we screened a collection of 34 ebselen and ebselen diselenide derivatives for SARS-CoV-2 PLpro and Mpro inhibitors. Our studies revealed that ebselen derivatives are potent inhibitors of both the proteases. We identified three PLpro and four Mpro inhibitors superior to ebselen. Independently, ebselen was shown to inhibit the N7-methyltransferase activity of SARS-CoV-2 nsp14 protein involved in viral RNA cap modification. Hence, selected compounds were also evaluated as nsp14 inhibitors. In the second part of our work, we employed 11 ebselen analogues-bis(2-carbamoylaryl)phenyl diselenides-in biological assays to evaluate their anti-SARS-CoV-2 activity in Vero E6 cells. We present their antiviral and cytoprotective activity and also low cytotoxicity. Our work shows that ebselen, its derivatives, and diselenide analogues constitute a promising platform for development of new antivirals targeting the SARS-CoV-2 virus.