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1.
Nat Commun ; 11(1): 4284, 2020 08 27.
Artigo em Inglês | MEDLINE | ID: mdl-32855409

RESUMO

Cytokinins are mobile multifunctional plant hormones with roles in development and stress resilience. Although their Histidine Kinase receptors are substantially localised to the endoplasmic reticulum, cellular sites of cytokinin perception and importance of spatially heterogeneous cytokinin distribution continue to be debated. Here we show that cytokinin perception by plasma membrane receptors is an effective additional path for cytokinin response. Readout from a Two Component Signalling cytokinin-specific reporter (TCSn::GFP) closely matches intracellular cytokinin content in roots, yet we also find cytokinins in extracellular fluid, potentially enabling action at the cell surface. Cytokinins covalently linked to beads that could not pass the plasma membrane increased expression of both TCSn::GFP and Cytokinin Response Factors. Super-resolution microscopy of GFP-labelled receptors and diminished TCSn::GFP response to immobilised cytokinins in cytokinin receptor mutants, further indicate that receptors can function at the cell surface. We argue that dual intracellular and surface locations may augment flexibility of cytokinin responses.


Assuntos
Arabidopsis/metabolismo , Citocininas/metabolismo , Proteínas Recombinantes/metabolismo , Adenina/análogos & derivados , Adenina/farmacologia , Arabidopsis/citologia , Arabidopsis/efeitos dos fármacos , Arabidopsis/genética , Proteínas de Arabidopsis/genética , Proteínas de Arabidopsis/metabolismo , Líquido Extracelular/metabolismo , Proteínas de Fluorescência Verde/genética , Proteínas de Fluorescência Verde/metabolismo , Histidina Quinase/genética , Histidina Quinase/metabolismo , Mutação , Plantas Geneticamente Modificadas , Proteínas Recombinantes/genética , Transdução de Sinais
4.
BMC Infect Dis ; 20(1): 524, 2020 Jul 20.
Artigo em Inglês | MEDLINE | ID: mdl-32689975

RESUMO

BACKGROUND: Currently recommended boosted protease-inhibitor (bPI) regimens may be associated with increased risk of cardiovascular or chronic kidney diseases; in addition, boosted regimens are particularly associated with drug-drug interactions. Since both cardiovascular and renal disease, and polypharmacy, are common in ageing people with HIV, there is a need for alternative efficacious regimens. bPI-based regimens are often the treatment of choice for individuals with pre-treatment or treatment-acquired resistance but it is plausible that carefully selected HIV-positive individuals with drug resistance, who are virologically suppressed on their current bPI regimen, could maintain virological efficacy when switched to bictegravir, emtricitabine and tenofovir alafenamide (B/F/TAF) fixed dose combination (FDC). METHODS/DESIGN: A phase IV, investigator-initiated, multicentre, open label pilot, randomised two-arm study to assess the safety and efficacy of switching from bPI regimen to B/F/TAF single tablet regimen in integrase inhibitor-naïve, virologically suppressed adults with HIV-1 infection harbouring drug resistance mutations. Eligible individuals will either continue on their bPI regimen or switch to B/F/TAF FDC. After 24 weeks, all participants in the bPI arm will be switched to B/F/TAF and followed for a further 24 weeks and all participants will be followed for 48 weeks. The primary efficacy endpoint is the proportion of participants with HIV-1 RNA < 50 copies/mL at week 24 using pure virologic response whilst the secondary efficacy endpoint is the proportion of participants with HIV-1 RNA < 50 copies/mL at Week 48. Other secondary outcome measures include between arm comparisons of drug resistance at virological failure, safety and tolerability and patient-reported outcome measures. DISCUSSION: We aim to provide preliminary evidence of the efficacy of switching to B/F/TAF in patients with virological suppression on a bPI-based regimen who harbour select drug resistance mutations. TRIAL REGISTRATION: ISRCTN 44453201 , registered 19 June 2019 and EudraCT 2018-004732-30.


Assuntos
Adenina/análogos & derivados , Farmacorresistência Viral/genética , Emtricitabina/uso terapêutico , Infecções por HIV/tratamento farmacológico , Inibidores de Integrase de HIV/uso terapêutico , HIV-1/efeitos dos fármacos , Compostos Heterocíclicos de 4 ou mais Anéis/uso terapêutico , Inibidores de Proteases/uso terapêutico , Inibidores da Transcriptase Reversa/uso terapêutico , Adenina/uso terapêutico , Adulto , Combinação de Medicamentos , Emtricitabina/efeitos adversos , Feminino , Infecções por HIV/genética , Infecções por HIV/virologia , Inibidores de Integrase de HIV/efeitos adversos , Compostos Heterocíclicos de 4 ou mais Anéis/efeitos adversos , Humanos , Masculino , Mutação , Projetos Piloto , Estudos Prospectivos , Inibidores de Proteases/efeitos adversos , Inibidores da Transcriptase Reversa/efeitos adversos , Resultado do Tratamento
5.
Lancet ; 396(10246): 239-254, 2020 07 25.
Artigo em Inglês | MEDLINE | ID: mdl-32711800

RESUMO

BACKGROUND: Tenofovir alafenamide shows high antiviral efficacy and improved renal and bone safety compared with tenofovir disoproxil fumarate when used for HIV treatment. Here, we report primary results from a blinded phase 3 study evaluating the efficacy and safety of pre-exposure prophylaxis (PrEP) with emtricitabine and tenofovir alafenamide versus emtricitabine and tenofovir disoproxil fumarate for HIV prevention. METHODS: This study is an ongoing, randomised, double-blind, multicentre, active-controlled, phase 3, non-inferiority trial done at 94 community, public health, and hospital-associated clinics located in regions of Europe and North America, where there is a high incidence of HIV or prevalence of people living with HIV, or both. We enrolled adult cisgender men who have sex with men and transgender women who have sex with men, both with a high risk of acquiring HIV on the basis of their self-reported sexual behaviour in the past 12 weeks or their recent history (within 24 weeks of enrolment) of bacterial sexually transmitted infections. Participants with current or previous use of PrEP with emtricitabine and tenofovir disoproxil fumarate were not excluded. We used a computer-generated random allocation sequence to randomly assign (1:1) participants to receive either emtricitabine (200 mg) and tenofovir alafenamide (25 mg) tablets daily, with matched placebo tablets (emtricitabine and tenofovir alafenamide group), or emtricitabine (200 mg) and tenofovir disoproxil fumarate (300 mg) tablets daily, with matched placebo tablets (emtricitabine and tenofovir disoproxil fumarate group). As such, all participants were given two tablets. The trial sponsor, investigators, participants, and the study staff who provided the study drugs, assessed the outcomes, and collected the data were masked to group assignment. The primary efficacy outcome was incident HIV infection, which was assessed when all participants had completed 48 weeks of follow-up and half of all participants had completed 96 weeks of follow-up. This full analysis set included all randomly assigned participants who had received at least one dose of the assigned study drug and had at least one post-baseline HIV test. Non-inferiority of emtricitabine and tenofovir alafenamide to emtricitabine and tenofovir disoproxil fumarate was established if the upper bound of the 95·003% CI of the HIV incidence rate ratio (IRR) was less than the prespecified non-inferiority margin of 1·62. We prespecified six secondary bone mineral density and renal biomarker safety endpoints to evaluate using the safety analysis set. This analysis set included all randomly assigned participants who had received at least one dose of the assigned study drug. This trial is registered with ClinicalTrials.gov, NCT02842086, and is no longer recruiting. FINDINGS: Between Sept 13, 2016, and June 30, 2017, 5387 (92%) of 5857 participants were randomly assigned and received emtricitabine and tenofovir alafenamide (n=2694) or emtricitabine and tenofovir disoproxil fumarate (n=2693). At the time of the primary efficacy analysis (ie, when all participants had completed 48 weeks and 50% had completed 96 weeks) emtricitabine and tenofovir alafenamide was non-inferior to emtricitabine and tenofovir disoproxil fumarate for HIV prevention, as the upper limit of the 95% CI of the IRR, was less than the prespecified non-inferiority margin of 1·62 (IRR 0·47 [95% CI 0·19-1·15]). After 8756 person-years of follow-up, 22 participants were diagnosed with HIV, seven participants in the emtricitabine and tenofovir alafenamide group (0·16 infections per 100 person-years [95% CI 0·06-0·33]), and 15 participants in the emtricitabine and tenofovir disoproxil fumarate group (0·34 infections per 100 person-years [0·19-0·56]). Both regimens were well tolerated, with a low number of participants reporting adverse events that led to discontinuation of the study drug (36 [1%] of 2694 participants in the emtricitabine and tenofovir alafenamide group vs 49 [2%] of 2693 participants in the emtricitabine and tenofovir disoproxil fumarate group). Emtricitabine and tenofovir alafenamide was superior to emtricitabine and tenofovir disoproxil fumarate in all six prespecified bone mineral density and renal biomarker safety endpoints. INTERPRETATION: Daily emtricitabine and tenofovir alafenamide shows non-inferior efficacy to daily emtricitabine and tenofovir disoproxil fumarate for HIV prevention, and the number of adverse events for both regimens was low. Emtricitabine and tenofovir alafenamide had more favourable effects on bone mineral density and biomarkers of renal safety than emtricitabine and tenofovir disoproxil fumarate. FUNDING: Gilead Sciences.


Assuntos
Adenina/análogos & derivados , Fármacos Anti-HIV/uso terapêutico , Combinação Emtricitabina e Fumarato de Tenofovir Desoproxila/uso terapêutico , Emtricitabina/uso terapêutico , Infecções por HIV/tratamento farmacológico , Tenofovir/uso terapêutico , Adenina/efeitos adversos , Adenina/uso terapêutico , Adulto , Fármacos Anti-HIV/efeitos adversos , Método Duplo-Cego , Emtricitabina/efeitos adversos , Combinação Emtricitabina e Fumarato de Tenofovir Desoproxila/efeitos adversos , Europa (Continente)/epidemiologia , Feminino , Infecções por HIV/epidemiologia , Infecções por HIV/prevenção & controle , HIV-1/efeitos dos fármacos , Homossexualidade Masculina/etnologia , Humanos , Masculino , América do Norte/epidemiologia , Placebos/administração & dosagem , Profilaxia Pré-Exposição/métodos , Prevalência , Segurança , Minorias Sexuais e de Gênero , Tenofovir/efeitos adversos , Resultado do Tratamento
6.
J Nanosci Nanotechnol ; 20(12): 7311-7323, 2020 12 01.
Artigo em Inglês | MEDLINE | ID: mdl-32711596

RESUMO

We started a study on the molecular docking of six potential pharmacologically active inhibitors compounds that can be used clinically against the COVID-19 virus, in this case, remdesivir, ribavirin, favipiravir, galidesivir, hydroxychloroquine and chloroquine interacting with the main COVID-19 protease in complex with a COVID-19 N3 protease inhibitor. The highest values of affinity energy found in order from highest to lowest were chloroquine (CHL), hydroxychloroquine (HYC), favipiravir (FAV), galidesivir (GAL), remdesivir (REM) and ribavirin (RIB). The possible formation of hydrogen bonds, associations through London forces and permanent electric dipole were analyzed. The values of affinity energy obtained for the hydroxychloroquine ligands was -9.9 kcal/mol and for the chloroquine of -10.8 kcal/mol which indicate that the coupling contributes to an effective improvement of the affinity energies with the protease. Indicating that, the position chosen to make the substitutions may be a pharmacophoric group, and cause changes in the protease.


Assuntos
Antivirais/química , Antivirais/farmacologia , Betacoronavirus/efeitos dos fármacos , Betacoronavirus/enzimologia , Infecções por Coronavirus/tratamento farmacológico , Infecções por Coronavirus/virologia , Cisteína Endopeptidases/química , Pneumonia Viral/tratamento farmacológico , Pneumonia Viral/virologia , Inibidores de Proteases/química , Inibidores de Proteases/farmacologia , Proteínas não Estruturais Virais/antagonistas & inibidores , Proteínas não Estruturais Virais/química , Adenina/administração & dosagem , Adenina/análogos & derivados , Adenina/química , Adenina/farmacologia , Monofosfato de Adenosina/administração & dosagem , Monofosfato de Adenosina/análogos & derivados , Monofosfato de Adenosina/química , Monofosfato de Adenosina/farmacologia , Alanina/administração & dosagem , Alanina/análogos & derivados , Alanina/química , Alanina/farmacologia , Amidas/administração & dosagem , Amidas/química , Amidas/farmacologia , Antivirais/administração & dosagem , Sítios de Ligação , Cloroquina/administração & dosagem , Cloroquina/química , Cloroquina/farmacologia , Interações Medicamentosas , Humanos , Ligação de Hidrogênio , Hidroxicloroquina/administração & dosagem , Hidroxicloroquina/química , Hidroxicloroquina/farmacologia , Ligantes , Simulação de Acoplamento Molecular , Nanotecnologia , Pandemias , Inibidores de Proteases/administração & dosagem , Pirazinas/administração & dosagem , Pirazinas/química , Pirazinas/farmacologia , Pirrolidinas/administração & dosagem , Pirrolidinas/química , Pirrolidinas/farmacologia , Ribavirina/administração & dosagem , Ribavirina/química , Ribavirina/farmacologia , Eletricidade Estática
7.
Lancet HIV ; 7(6): e389-e400, 2020 06.
Artigo em Inglês | MEDLINE | ID: mdl-32504574

RESUMO

BACKGROUND: In the primary week-48 analyses of two phase 3 studies, coformulated bictegravir, emtricitabine, and tenofovir alafenamide was non-inferior to a dolutegravir-containing regimen in treatment-naive people with HIV. We report week-144 efficacy and safety results from these studies. METHODS: We did two double-blind, active-controlled studies (now in open-label extension phase). Study 1 randomly assigned (1:1) HLA-B*5701-negative adults without hepatitis B virus co-infection to receive coformulated bictegravir 50 mg, emtricitabine 200 mg, and tenofovir alafenamide 25 mg, or coformulated dolutegravir 50 mg, abacavir 600 mg, and lamivudine 300 mg once daily. Study 2 randomly assigned (1:1) adults to bictegravir, emtricitabine, and tenofovir alafenamide, or dolutegravir 50 mg given with coformulated emtricitabine 200 mg and tenofovir alafenamide 25 mg. We previously reported non-inferiority at the primary endpoint. Here, we report the week-144 secondary outcome of proportion of participants with plasma HIV-1 RNA less than 50 copies per mL at week 144, by US Food and Drug Administration Snapshot algorithm, analysed in the same manner. These studies were registered with ClinicalTrials.gov, NCT02607930 and NCT02607956. FINDINGS: 629 participants were randomly assigned and treated in study 1 (314 to bictegravir, emtricitabine, and tenofovir alafenamide, and 315 to dolutegravir, abacavir, and lamivudine) and 645 in study 2 (327 to bictegravir, emtricitabine, and tenofovir alafenamide, 325 to dolutegravir, emtricitabine, tenofovir alafenamide). At week 144, bictegravir, emtricitabine, and tenofovir alafenamide was non-inferior to both dolutegravir-containing regimens for efficacy. In study 1, 256 (82%) of 314 participants had plasma HIV-1 RNA less than 50 copies per mL in the bictegravir, emtricitabine, and tenofovir alafenamide group and 265 (84%) of 315 in the dolutegravir, abacavir, and lamivudine group (difference -2·6%, 95% CI -8·5 to 3·4). In study 2, 262 (82%) of 320 participants had plasma HIV-1 RNA less than 50 copies per mL in the bictegravir, emtricitabine, and tenofovir alafenamide group and 273 (84%) of 325 in the dolutegravir, emtricitabine, and tenofovir alafenamide group (difference -1·9%, -7·8 to 3·9). In both studies, no participant had treatment-emergent resistance to study drugs up to week 144. All treatment regimens were well tolerated with additional exposure. Adverse events that led to study drug discontinuation were reported for no participants in the bictegravir, emtricitabine, and tenofovir alafenamide group versus five (2%) of 315 in the dolutegravir, abacavir, and lamivudine group (study 1), and six (2%) of 320 in the bictegravir, emtricitabine, and tenofovir alafenamide versus six (2%) of 325 in the dolutegravir, emtricitabine, and tenofovir alafenamide group (study 2). In study 1, statistically significant differences were observed in median changes from baseline in fasting total cholesterol (14 mg/dL vs 10 mg/dL; p=0·034), direct LDL (21 mg/dL vs 14 mg/dL; p=0·004), and total cholesterol to HDL ratio (-0·1 vs -0·3; p=0·007) at week 144; no differences were observed between groups in study 2. Weight gain was seen across all treatment groups in both studies, with no differences in median changes from baseline in weight at week 144 for either study. INTERPRETATION: These long-term data support the use of bictegravir, emtricitabine, and tenofovir alafenamide as a safe, well tolerated, and durable treatment for people with HIV, with no emergent resistance. FUNDING: Gilead Sciences.


Assuntos
Adenina/análogos & derivados , Didesoxinucleosídeos/administração & dosagem , Emtricitabina/administração & dosagem , Infecções por HIV/tratamento farmacológico , Compostos Heterocíclicos com 3 Anéis/administração & dosagem , Compostos Heterocíclicos de 4 ou mais Anéis/administração & dosagem , Lamivudina/administração & dosagem , Tenofovir/administração & dosagem , Adenina/administração & dosagem , Adenina/efeitos adversos , Adulto , Idoso , Didesoxinucleosídeos/efeitos adversos , Método Duplo-Cego , Esquema de Medicação , Combinação de Medicamentos , Emtricitabina/efeitos adversos , Feminino , Infecções por HIV/virologia , HIV-1/genética , Compostos Heterocíclicos com 3 Anéis/efeitos adversos , Compostos Heterocíclicos de 4 ou mais Anéis/efeitos adversos , Humanos , Lamivudina/efeitos adversos , Masculino , Pessoa de Meia-Idade , RNA Viral/sangue , Tenofovir/efeitos adversos , Resultado do Tratamento , Adulto Jovem
10.
Int. j. morphol ; 38(3): 530-535, June 2020. graf
Artigo em Inglês | LILACS | ID: biblio-1098283

RESUMO

Dysregulated autophagy, whether excessive or downregulated, has been thought to be associated with neurodegenerative disorders including Parkinson's disease. Accordingly, the present study was carried out to investigate whether 3-methyladenine, an autophagy inhibitor, can modulate the effects of rotenone on dopaminergic neurons in primary mesencephalic cell culture. Cultures were prepared from embryonic mouse mesencephala at gestation day 14. Four groups of cultures were treated on the 10th DIV for 48 h as follows: the first was kept as an untreated control, the second was treated with 3-methyladenine alone (1, 10, 100, 200 mM), the third was treated with 20 nM rotenone and the fourth was co-treated with 20 nM rotenone and 3-methyladenine (1, 10, 100, 200 mM). On the 12th DIV, cultured cells were stained immunohistochemically against tyrosine hydroxylase and culture media were used to measure the levels of lactate dehydrogenase. 3methyladenine had no effects on both the survival of dopaminergic neurons and the release of lactate dehydrogenase. Rotenone significantly decreased the number of dopaminergic neurons and increased the levels of lactate dehydrogenase in the culture media. When cultures concomitantly treated with 3-methyladenine and rotenone, 3-methyladenine had no effect against rotenone-induced dopaminergic cell damage and lactate dehydrogenase release into the culture medium. In conclusion, the autophagy inhibitor 3-methyladenine could not modulate rotenone-induced dopaminergic cell damage in primary mesencephalic cell culture.


Se estima que la autofagia desregulada, ya sea excesiva o con baja regulación, está asociada con trastornos neurodegenerativos, incluyendo la enfermedad de Parkinson. En consecuencia, el se realizó este estudio para investigar si la 3metiladenina, un inhibidor de la autofagia,puede modular los efectos de la rotenona en las neuronas dopaminérgicas en el cultivo primario de células mesencefálicas. Los cultivos se prepararon a partir de mesencéfalo de ratón embrionario el día 14 de gestación. Cuatro grupos de cultivos se trataron en el 10º DIV durante 48 h de la siguiente manera: el primer grupo se mantuvo como un control no tratado, el segundo se trató con 3-metiladenina sola (1, 10, 100, 200 mM), el tercer grupo se trató con rotenona 20 nM y el cuarto se trató conjuntamente con rotenona 20 nM y 3-metiladenina (1, 10, 100, 200 mM). En el 12º DIV; las células cultivadas fueron tratadas mediante tinción inmunohistoquímica en tirosina hidroxilasa y se usaron medios de cultivo para medir los niveles de lactato deshidrogenasa. La 3-metiladenina no tuvo efectos tanto en la supervivencia de las neuronas dopaminérgicas como en la liberación de lactato deshidrogenasa. La rotenona disminuyó significativamente el número de neuronas dopaminérgicas y se observó un aumento de los niveles de lactato deshidrogenasa en los medios de cultivo. Cuando los cultivos tratados concomitantemente con 3-metiladenina y rotenona, la 3metiladenina no tuvo efecto contra el daño celular dopaminérgico inducido por la rotenona y la liberación de lactato deshidrogenasa en el medio de cultivo. En conclusión, el inhibidor de la autofagia 3-metiladenina no moduló el daño celular dopaminérgico inducido por la rotenona en el cultivo celular mesencefálico primario.


Assuntos
Animais , Camundongos , Doença de Parkinson , Rotenona/toxicidade , Adenina/análogos & derivados , Autofagia , Mesencéfalo , Adenina/farmacologia , Células Cultivadas , Morte Celular/efeitos dos fármacos , Neurônios Dopaminérgicos/efeitos dos fármacos , L-Lactato Desidrogenase/análise
11.
Ann Intern Med ; 173(7): 536-541, 2020 10 06.
Artigo em Inglês | MEDLINE | ID: mdl-32589451

RESUMO

BACKGROUND: The incidence and severity of coronavirus disease 2019 (COVID-19) among HIV-positive persons receiving antiretroviral therapy (ART) have not been characterized in large populations. OBJECTIVE: To describe the incidence and severity of COVID-19 by nucleos(t)ide reverse transcriptase inhibitor (NRTI) use among HIV-positive persons receiving ART. DESIGN: Cohort study. SETTING: HIV clinics in 60 Spanish hospitals between 1 February and 15 April 2020. PARTICIPANTS: 77 590 HIV-positive persons receiving ART. MEASUREMENTS: Estimated risks (cumulative incidences) per 10 000 persons and 95% CIs for polymerase chain reaction-confirmed COVID-19 diagnosis, hospitalization, intensive care unit (ICU) admission, and death. Risk and 95% CIs for COVID-19 diagnosis and hospital admission by use of the NRTIs tenofovir disoproxil fumarate (TDF)/emtricitabine (FTC), tenofovir alafenamide (TAF)/FTC, abacavir (ABC)/lamivudine (3TC), and others were estimated through Poisson regression models. RESULTS: Of 77 590 HIV-positive persons receiving ART, 236 were diagnosed with COVID-19, 151 were hospitalized, 15 were admitted to the ICU, and 20 died. The risks for COVID-19 diagnosis and hospitalization were greater in men and persons older than 70 years. The risk for COVID-19 hospitalization was 20.3 (95% CI, 15.2 to 26.7) among patients receiving TAF/FTC, 10.5 (CI, 5.6 to 17.9) among those receiving TDF/FTC, 23.4 (CI, 17.2 to 31.1) among those receiving ABC/3TC, and 20.0 (CI, 14.2 to 27.3) for those receiving other regimens. The corresponding risks for COVID-19 diagnosis were 39.1 (CI, 31.8 to 47.6), 16.9 (CI, 10.5 to 25.9), 28.3 (CI, 21.5 to 36.7), and 29.7 (CI, 22.6 to 38.4), respectively. No patient receiving TDF/FTC was admitted to the ICU or died. LIMITATION: Residual confounding by comorbid conditions cannot be completely excluded. CONCLUSION: HIV-positive patients receiving TDF/FTC have a lower risk for COVID-19 and related hospitalization than those receiving other therapies. These findings warrant further investigation in HIV preexposure prophylaxis studies and randomized trials in persons without HIV. PRIMARY FUNDING SOURCE: Instituto de Salud Carlos III and National Institutes of Health.


Assuntos
Terapia Antirretroviral de Alta Atividade , Infecções por Coronavirus/epidemiologia , Infecções por HIV/tratamento farmacológico , Pneumonia Viral/epidemiologia , Adenina/análogos & derivados , Adulto , Idoso , Betacoronavirus , Infecções por Coronavirus/mortalidade , Didesoxinucleosídeos , Combinação de Medicamentos , Emtricitabina , Feminino , Infecções por HIV/mortalidade , Hospitalização/estatística & dados numéricos , Humanos , Incidência , Unidades de Terapia Intensiva/estatística & dados numéricos , Lamivudina , Masculino , Pessoa de Meia-Idade , Pandemias , Pneumonia Viral/mortalidade , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Índice de Gravidade de Doença , Espanha/epidemiologia , Tenofovir
12.
J Phys Chem Lett ; 11(11): 4430-4435, 2020 Jun 04.
Artigo em Inglês | MEDLINE | ID: covidwho-233085

RESUMO

The pandemic outbreak of a new coronavirus (CoV), SARS-CoV-2, has captured the world's attention, demonstrating that CoVs represent a continuous global threat. As this is a highly contagious virus, it is imperative to understand RNA-dependent-RNA-polymerase (RdRp), the key component in virus replication. Although the SARS-CoV-2 genome shares 80% sequence identity with severe acute respiratory syndrome SARS-CoV, their RdRps and nucleotidyl-transferases (NiRAN) share 98.1% and 93.2% identity, respectively. Sequence alignment of six coronaviruses demonstrated higher identity among their RdRps (60.9%-98.1%) and lower identity among their Spike proteins (27%-77%). Thus, a 3D structural model of RdRp, NiRAN, non-structural protein 7 (nsp7), and nsp8 of SARS-CoV-2 was generated by modeling starting from the SARS counterpart structures. Furthermore, we demonstrate the binding poses of three viral RdRp inhibitors (Galidesivir, Favipiravir, and Penciclovir), which were recently reported to have clinical significance for SARS-CoV-2. The network of interactions established by these drug molecules affirms their efficacy to inhibit viral RNA replication and provides an insight into their structure-based rational optimization for SARS-CoV-2 inhibition.


Assuntos
Betacoronavirus/enzimologia , Nucleotidiltransferases/química , RNA Replicase/química , Adenina/análogos & derivados , Adenina/química , Adenina/metabolismo , Amidas/química , Amidas/metabolismo , Antivirais/química , Antivirais/metabolismo , Betacoronavirus/isolamento & purificação , Sítios de Ligação , Infecções por Coronavirus/epidemiologia , Infecções por Coronavirus/patologia , Infecções por Coronavirus/virologia , Humanos , Simulação de Acoplamento Molecular , Nucleotidiltransferases/metabolismo , Pandemias , Pneumonia Viral/epidemiologia , Pneumonia Viral/patologia , Pneumonia Viral/virologia , Estrutura Terciária de Proteína , Pirazinas/química , Pirazinas/metabolismo , Pirrolidinas/química , Pirrolidinas/metabolismo , RNA Replicase/metabolismo
13.
J Phys Chem Lett ; 11(11): 4430-4435, 2020 Jun 04.
Artigo em Inglês | MEDLINE | ID: mdl-32392072

RESUMO

The pandemic outbreak of a new coronavirus (CoV), SARS-CoV-2, has captured the world's attention, demonstrating that CoVs represent a continuous global threat. As this is a highly contagious virus, it is imperative to understand RNA-dependent-RNA-polymerase (RdRp), the key component in virus replication. Although the SARS-CoV-2 genome shares 80% sequence identity with severe acute respiratory syndrome SARS-CoV, their RdRps and nucleotidyl-transferases (NiRAN) share 98.1% and 93.2% identity, respectively. Sequence alignment of six coronaviruses demonstrated higher identity among their RdRps (60.9%-98.1%) and lower identity among their Spike proteins (27%-77%). Thus, a 3D structural model of RdRp, NiRAN, non-structural protein 7 (nsp7), and nsp8 of SARS-CoV-2 was generated by modeling starting from the SARS counterpart structures. Furthermore, we demonstrate the binding poses of three viral RdRp inhibitors (Galidesivir, Favipiravir, and Penciclovir), which were recently reported to have clinical significance for SARS-CoV-2. The network of interactions established by these drug molecules affirms their efficacy to inhibit viral RNA replication and provides an insight into their structure-based rational optimization for SARS-CoV-2 inhibition.


Assuntos
Betacoronavirus/enzimologia , Nucleotidiltransferases/química , RNA Replicase/química , Adenina/análogos & derivados , Adenina/química , Adenina/metabolismo , Amidas/química , Amidas/metabolismo , Antivirais/química , Antivirais/metabolismo , Betacoronavirus/isolamento & purificação , Sítios de Ligação , Infecções por Coronavirus/epidemiologia , Infecções por Coronavirus/patologia , Infecções por Coronavirus/virologia , Humanos , Simulação de Acoplamento Molecular , Nucleotidiltransferases/metabolismo , Pandemias , Pneumonia Viral/epidemiologia , Pneumonia Viral/patologia , Pneumonia Viral/virologia , Estrutura Terciária de Proteína , Pirazinas/química , Pirazinas/metabolismo , Pirrolidinas/química , Pirrolidinas/metabolismo , RNA Replicase/metabolismo
14.
Arterioscler Thromb Vasc Biol ; 40(6): 1510-1522, 2020 06.
Artigo em Inglês | MEDLINE | ID: mdl-32349535

RESUMO

OBJECTIVE: Endothelial Cav-1 (caveolin-1) expression plays a relevant role during atherogenesis by controlling NO production, vascular inflammation, LDL (low-density lipoprotein) transcytosis, and extracellular matrix remodeling. Additional studies have identified cholesterol-rich membrane domains as important regulators of autophagy by recruiting ATGs (autophagy-related proteins) to the plasma membrane. Here, we investigate how the expression of Cav-1 in the aortic endothelium influences autophagy and whether enhanced autophagy contributes to the atheroprotective phenotype observed in Cav-1-deficient mice. Approach and Results: To analyze the impact of Cav-1 deficiency on regulation of autophagy in the aortic endothelium during the progression of atherosclerosis, we fed Ldlr-/- and Cav-1-/-Ldlr-/- mice a Western diet and assessed autophagy in the vasculature. We observe that the absence of Cav-1 promotes autophagy activation in athero-prone areas of the aortic endothelium by enhancing autophagic flux. Mechanistically, we found that Cav-1 interacts with the ATG5-ATG12 complex and influences the cellular localization of autophagosome components in lipid rafts, which controls the autophagosome formation and autophagic flux. Pharmacological inhibition of autophagy attenuates the atheroprotection observed in Cav-1-/- mice by increasing endothelial inflammation and macrophage recruitment, identifying a novel molecular mechanism by which Cav-1 deficiency protects against the progression of atherosclerosis. CONCLUSIONS: These results identify Cav-1 as a relevant regulator of autophagy in the aortic endothelium and demonstrate that pharmacological suppression of autophagic flux in Cav-1-deficient mice attenuates the atheroprotection observed in Cav-1-/- mice. Additionally, these findings suggest that activation of endothelial autophagy by blocking Cav-1 might provide a potential therapeutic strategy for cardiovascular diseases including atherosclerosis.


Assuntos
Aterosclerose/prevenção & controle , Autofagia/fisiologia , Caveolina 1/deficiência , Endotélio Vascular/fisiopatologia , Vasculite/prevenção & controle , Adenina/análogos & derivados , Adenina/farmacologia , Animais , Aorta/patologia , Aorta/fisiopatologia , Aorta/ultraestrutura , Aterosclerose/etiologia , Autofagia/efeitos dos fármacos , Caveolina 1/análise , Caveolina 1/fisiologia , Dieta Ocidental , Células Endoteliais/química , Células Endoteliais/fisiologia , Células Endoteliais/ultraestrutura , Endotélio Vascular/química , Endotélio Vascular/ultraestrutura , Feminino , Humanos , Masculino , Microdomínios da Membrana/química , Microdomínios da Membrana/fisiologia , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Células NIH 3T3 , Receptores de LDL/deficiência
15.
Gastroenterol Clin North Am ; 49(2): 239-252, 2020 06.
Artigo em Inglês | MEDLINE | ID: mdl-32389361

RESUMO

Half a century after its discovery, hepatitis delta remains a pertinent global health issue with a major clinical impact in endemic regions and an underestimated prevalence worldwide. Hepatitis delta virus infection follows a challenging clinical course and is responsible for significant liver-related morbidity. Although the only currently available treatment (pegylated interferon) does not provide consistent results, emerging therapeutic options are promising. This article explores the epidemiology, natural history, as well as current and potential therapeutic options for hepatitis delta virus infection.


Assuntos
Hepatite D/diagnóstico , Hepatite D/terapia , Adenina/análogos & derivados , Adenina/uso terapêutico , Anticorpos Antivirais/sangue , Antivirais/uso terapêutico , Biomarcadores/sangue , Carcinoma Hepatocelular/etiologia , Saúde Global , Hepatite D/epidemiologia , Hepatite D/virologia , Vírus Delta da Hepatite/genética , Vírus Delta da Hepatite/imunologia , Antígenos da Hepatite delta/imunologia , Humanos , Interferon alfa-2/uso terapêutico , Lamivudina/uso terapêutico , Cirrose Hepática/etiologia , Neoplasias Hepáticas/etiologia , Transplante de Fígado , Organofosfonatos/uso terapêutico , Prevalência , RNA Viral , Risco , Testes Sorológicos/métodos
16.
Nucleic Acids Res ; 48(9): 5094-5105, 2020 05 21.
Artigo em Inglês | MEDLINE | ID: mdl-32297938

RESUMO

Box C/D RNA protein complexes (RNPs) catalyze site-specific 2'-O-methylation of RNA with specificity determined by guide RNAs. In eukaryotic C/D RNP, the paralogous Nop58 and Nop56 proteins specifically associate with terminal C/D and internal C'/D' motifs of guide RNAs, respectively. We have reconstituted active C/D RNPs with recombinant proteins of the thermophilic yeast Chaetomium thermophilum. Nop58 and Nop56 could not distinguish between the two C/D motifs in the reconstituted enzyme, suggesting that the assembly specificity is imposed by trans-acting factors in vivo. The two C/D motifs are functionally independent and halfmer C/D RNAs can also guide site-specific methylation. Extensive pairing between C/D RNA and substrate is inhibitory to modification for both yeast and archaeal C/D RNPs. N6-methylated adenine at box D/D' interferes with the function of the coupled guide. Our data show that all C/D RNPs share the same functional organization and mechanism of action and provide insight into the assembly specificity of eukaryotic C/D RNPs.


Assuntos
Metiltransferases/química , Metiltransferases/metabolismo , RNA Nucleolar Pequeno/química , RNA Nucleolar Pequeno/metabolismo , Ribonucleoproteínas/metabolismo , Adenina/análogos & derivados , Chaetomium/genética , Humanos , Metilação , Conformação de Ácido Nucleico , RNA/metabolismo , Ribonucleoproteínas/química , Sulfolobus solfataricus
17.
Nucleic Acids Res ; 48(9): 5119-5134, 2020 05 21.
Artigo em Inglês | MEDLINE | ID: mdl-32282906

RESUMO

Reactive oxygen species generate the genotoxic 8-oxoguanine (oxoG) and 8-oxoadenine (oxoA) as major oxidative lesions. The mutagenicity of oxoG is attributed to the lesion's ability to evade the geometric discrimination of DNA polymerases by adopting Hoogsteen base pairing with adenine in a Watson-Crick-like geometry. Compared with oxoG, the mutagenesis mechanism of oxoA, which preferentially induces A-to-C mutations, is poorly understood. In the absence of protein contacts, oxoA:G forms a wobble conformation, the formation of which is suppressed in the catalytic site of most DNA polymerases. Interestingly, human DNA polymerase η (polη) proficiently incorporates dGTP opposite oxoA, suggesting the nascent oxoA:dGTP overcomes the geometric discrimination of polη. To gain insights into oxoA-mediated mutagenesis, we determined crystal structures of polη bypassing oxoA. When paired with dGTP, oxoA adopted a syn-conformation and formed Hoogsteen pairing while in a wobble geometry, which was stabilized by Gln38-mediated minor groove contacts to oxoA:dGTP. Gln38Ala mutation reduced misinsertion efficiency ∼55-fold, indicating oxoA:dGTP misincorporation was promoted by minor groove interactions. Also, the efficiency of oxoA:dGTP insertion by the X-family polß decreased ∼380-fold when Asn279-mediated minor groove contact to dGTP was abolished. Overall, these results suggest that, unlike oxoG, oxoA-mediated mutagenesis is greatly induced by minor groove interactions.


Assuntos
Adenina/análogos & derivados , DNA Polimerase Dirigida por DNA/química , Mutagênese , Adenina/química , Pareamento de Bases , DNA Polimerase beta/química , DNA Polimerase beta/genética , DNA Polimerase Dirigida por DNA/genética , DNA Polimerase Dirigida por DNA/metabolismo , Nucleotídeos de Desoxiguanina/química , Nucleotídeos de Desoxiguanina/metabolismo , Humanos , Cinética , Mutação , Nucleotídeos de Timina/metabolismo
18.
Scand J Immunol ; 91(6): e12880, 2020 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-32219875

RESUMO

Synthetic Toll-like receptor (TLR) 7 agonists have been suggested as immune modulators in a range of conditions. In contrast, self-derived TLR7 activators, such as RNA-containing immune complexes (RNA-IC), can contribute to autoimmune diseases due to endogenous immune activation. The exact difference in immune cell response between synthetic and endogenous TLR7 triggers is only partly known. An understanding of these differences could aid in the development of new therapeutic agents and provide insights into autoimmune disease mechanisms. We therefore compared the stimulatory capacity of two TLR7 agonists, RNA-IC and a synthetic small molecule DSR-6434, on blood leucocytes, plasmacytoid dendritic cells (pDCs) and B cells from healthy individuals. IFN-α, IL-6, IL-8 and TNF levels were measured by immunoassays, and gene expression in pDCs was analysed by an expression array. DSR-6434 triggered 20-fold lower levels of IFN-α by pDCs, but higher production of IL-6, IL-8 and TNF, compared to RNA-IC. Furthermore, IFN-α and TNF production were increased with exogenous IFN-α2b priming, whereas IL-8 synthesis by B cells was reduced for both stimuli. Cocultivation of pDCs and B cells increased the RNA-IC-stimulated IFN-α and TNF levels, while only IL-6 production was enhanced in the DSR-6434-stimulated cocultures. When comparing pDCs stimulated with RNA-IC and DSR-6434, twelve genes were differentially expressed (log2 fold change >2, adjusted P-value <.05). In conclusion, RNA-IC, which mimics an endogenous TLR7 stimulator, and the synthetic TLR7 agonist DSR-6434 trigger distinct inflammatory profiles in immune cells. This demonstrates the importance of using relevant stimuli when targeting the TLR7 pathway for therapeutic purposes.


Assuntos
Adenina/farmacologia , Complexo Antígeno-Anticorpo/farmacologia , Linfócitos B/imunologia , Células Dendríticas/imunologia , Complexos Multiproteicos/farmacologia , RNA/farmacologia , Receptor 7 Toll-Like/metabolismo , Adenina/análogos & derivados , Adenina/química , Complexo Antígeno-Anticorpo/química , Diferenciação Celular , Células Cultivadas , Citocinas/metabolismo , Perfilação da Expressão Gênica , Humanos , Mediadores da Inflamação/metabolismo , Ativação Linfocitária , Estrutura Molecular , Complexos Multiproteicos/química , RNA/química , Receptor 7 Toll-Like/agonistas
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