Virological response to nucleos(t)ide analogues treatment in chronic hepatitis B patients is associated with Bacteroides-dominant gut microbiome.

BACKGROUND: Gut dysbiosis is present in chronic hepatitis B virus (HBV) infection. In this study, we integrated microbiome and metabolome analysis to investigate the role of gut microbiome in virological response to nucleos(t)ide analogues (NAs) treatment. METHODS: Chronic HBV patients were prospectively recruited for steatosis and fibrosis assessments via liver elastography, with full-length 16S sequencing performed to identify the compositional gut microbiota differences. Fasting plasma bile acids were quantified by liquid chromatography-tandem mass spectrometry. FINDINGS: All patients (n = 110) were characterized into three distinct microbial clusters by their dominant genus: c-Bacteroides, c-Blautia, and c-Prevotella. Patients with c-Bacteroides had a higher plasma ursodeoxycholic acids (UDCA) level and an increase in 7-alpha-hydroxysteroid dehydrogenase (secondary bile acid biotransformation) than other clusters. In NAs-treated patients (n = 84), c-Bacteroides was associated with higher odds of plasma HBV-DNA undetectability when compared with non-c-Bacteroides clusters (OR 3.49, 95% CI 1.43-8.96, p = 0.01). c-Blautia was positively associated with advanced fibrosis (OR 2.74, 95% CI 1.09-7.31, p = 0.04). No such associations were found in treatment-naïve patients. Increased Escherichia coli relative abundance (0.21% vs. 0.03%, p = 0.035) was found in on-treatment patients (median treatment duration 98.1 months) with advanced fibrosis despite HBV DNA undetectability. An enrichment in l-tryptophan biosynthesis was observed in patients with advanced fibrosis, which exhibited a positive correlation with Escherichia coli. INTERPRETATION: Collectively, unique bacterial signatures, including c-Bacteroides and c-Blautia, were associated with virological undetectability and fibrosis evolution during NAs therapy in chronic HBV, setting up intriguing possibilities in optimizing HBV treatment. FUNDING: This study was supported by the Guangdong Natural Science Fund (2019A1515012003).

Combination of Tissue Microarray Profiling and Multiplexed IHC Approaches to Investigate Transport Mechanism of Nucleoside Analog Drug Resistance.

Nucleoside analogs (NAs) are an established class of anticancer agents being used clinically for the treatment of diverse cancers, either as monotherapy or in combination with other established anticancer or pharmacological agents. To date, nearly a dozen anticancer NAs are approved by the FDA, and several novel NAs are being tested in preclinical and clinical trials for future applications. However, improper delivery of NAs into tumor cells because of alterations in expression of one or more drug carrier proteins (e.g., solute carrier (SLC) transporters) within tumor cells or cells surrounding the tumor microenvironment stands as one of the primary reasons for therapeutic drug resistance. The combination of tissue microarray (TMA) and multiplexed immunohistochemistry (IHC) is an advanced, high-throughput approach over conventional IHC that enables researchers to effectively investigate alterations to numerous such chemosensitivity determinants simultaneously in hundreds of tumor tissues derived from patients. In this chapter, taking an example of a TMA from pancreatic cancer patients treated with gemcitabine (a NA chemotherapeutic agent), we describe the step-by-step procedure of performing multiplexed IHC, imaging of TMA slides, and quantification of expression of some relevant markers in these tissue sections as optimized in our laboratory and discuss considerations while designing and carrying out this experiment.

Differential response of HBV envelope-specific CD4 + T cells is related to HBsAg loss after stopping nucleos(t)ide analogue therapy.

BACKGROUND AND AIM: Long-term maintenance of viral control, even HBsAg loss, remains a challenge for chronic hepatitis B (CHB) patients undergoing nucleos(t)ide analogue (NA) discontinuation. This study aimed to investigate the relationship between HBV-specific T-cell responses targeting peptides spanning the whole proteome and clinical outcomes in CHB patients after NA discontinuation. APPROACH AND RESULTS: Eighty-eight CHB patients undergoing NA discontinuation were classified as responders (remained relapse-free up to 96 weeks) or relapsers (relapsed patients who underwent NA retreatment for up to 48 weeks and reachieved stable viral control). HBV-specific T-cell responses were detected at baseline and longitudinally throughout the follow-up. We found responders had a greater magnitude of HBV polymerase (Pol)-specific T-cell responses than relapsers at baseline. After long-term NA discontinuation, simultaneously enhanced HBV Core-induced and Pol-induced responses were observed in responders. Particularly, responders with HBsAg loss possessed enhanced HBV Envelope (Env)-induced responses after short-term and long-term follow-up. Notably, CD4 + T cells accounted for the predominance of HBV-specific T-cell responses. Correspondingly, CD4-deficient mice showed attenuated HBV-specific CD8 + T-cell responses, reduced HBsAb-producing B cells, and delayed HBsAg loss; in contrast, in vitro addition of CD4 + T cells promoted HBsAb production by B cells. Besides, IL-9, rather than PD-1 blockade, enhanced HBV Pol-specific CD4 + T-cell responses. CONCLUSION: HBV-specific CD4 + T-cell responses induced by the targeted peptide possess specificities for long-term viral control and HBsAg loss in CHB patients undergoing NA discontinuation, indicating that CD4 + T cells specific to distinct HBV antigens may endow with divergent antiviral potential.

Total synthesis of pseudouridimycin and its epimer via Ugi-type multicomponent reaction.

A total synthesis of pseudouridimycin (1) was accomplished featuring an unusual oxime Ugi-type multicomponent condensation to simultaneously construct the dipeptide moiety of this peptidyl nucleoside antibiotic. In this synthetic route 1 is readily accessible via a longest linear sequence of 9 synthetic steps from pseudouridine. This strategy can be applicable to a variety of pseudouridimycin analogues.

Assessment of nucleotide/nucleoside analog intervention in primer-dependent viral RNA-dependent RNA polymerases.

Nucleotide/nucleoside analogs (NAs) are important compounds used in antiviral drug development. To understand the action mode of NA drugs, we present an enzymology protocol to initially evaluate the intervention mechanism of the NTP forms of NAs on a coronaviral RNA-dependent RNA polymerase (RdRP). We describe the preparation of SARS-CoV-2 RdRP proteins and RNA constructs, followed by a primer-dependent RdRP assay to assess NTP forms of NAs. Two representative NA drugs, sofosbuvir and remdesivir, are used for demonstration of this protocol. For complete details on the use and execution of this protocol, please refer to Wu et al. (2021).

Nucleosides and emerging viruses: A new story.

With several US Food and Drug Administration (FDA)-approved drugs and high barriers to resistance, nucleoside and nucleotide analogs remain the cornerstone of antiviral therapies for not only herpesviruses, but also HIV and hepatitis viruses (B and C); however, with the exception of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), for which vaccines have been developed at unprecedented speed, there are no vaccines or small antivirals yet available for (re)emerging viruses, which are primarily RNA viruses. Thus, herein, we present an overview of ribonucleoside analogs recently developed and acting as inhibitors of the viral RNA-dependent RNA polymerase (RdRp). They are new lead structures that will be exploited for the discovery of new antiviral nucleosides.

Pyrimidine inhibitors synergize with nucleoside analogues to block SARS-CoV-2.

The SARS-CoV-2 virus has infected more than 261 million people and has led to more than 5 million deaths in the past year and a half1 ( https://www.who.org/ ). Individuals with SARS-CoV-2 infection typically develop mild-to-severe flu-like symptoms, whereas infection of a subset of individuals leads to severe-to-fatal clinical outcomes2. Although vaccines have been rapidly developed to combat SARS-CoV-2, there has been a dearth of antiviral therapeutics. There is an urgent need for therapeutics, which has been amplified by the emerging threats of variants that may evade vaccines. Large-scale efforts are underway to identify antiviral drugs. Here we screened approximately 18,000 drugs for antiviral activity using live virus infection in human respiratory cells and validated 122 drugs with antiviral activity and selectivity against SARS-CoV-2. Among these candidates are 16 nucleoside analogues, the largest category of clinically used antivirals. This included the antivirals remdesivir and molnupiravir, which have been approved for use in COVID-19. RNA viruses rely on a high supply of nucleoside triphosphates from the host to efficiently replicate, and we identified a panel of host nucleoside biosynthesis inhibitors as antiviral. Moreover, we found that combining pyrimidine biosynthesis inhibitors with antiviral nucleoside analogues synergistically inhibits SARS-CoV-2 infection in vitro and in vivo against emerging strains of SARS-CoV-2, suggesting a clinical path forward.

Aptamers Entirely Built from Therapeutic Nucleoside Analogues for Targeted Cancer Therapy.

Owing to the specific and high binding affinity of aptamers to their targets, aptamer-drug conjugates (ApDCs) have emerged as a promising drug delivery system for targeted cancer therapy. However, in a conventional ApDC, the aptamer segment usually just serves as a targeting moiety, and only a limited number of drug molecules are sequentially conjugated to the oligonucleotide, giving a relatively low drug loading capacity. To address this challenge, herein we employ four clinically approved nucleoside analogues, including clofarabine (Clo), ara-guanosine (AraG), gemcitabine (Ge), and floxuridine (FdU), to replace all natural nucleosides in aptamer sequences, generating a series of whole drug-constituted DNA-like oligomers that are termed drugtamers. Similar to their parent aptamers, the obtained drugtamers maintain the targeting capability and can specifically bind to the target receptors overexpressed on the cancer cell surface. With 100% drug loading ratio, active targeting capability, and enzyme-mediated release of active therapeutics, our drugtamers can strongly induce the apoptosis of cancer cells and inhibit the tumor progression, which enables a new potential for a better targeted cancer therapy.

Total synthesis and chemical stability of pseudouridimycin.

We report the chemical synthesis of pseudouridimycin (1), an antimicrobial natural product that potently and selectively inhibits bacterial RNA polymerase. Chemical stability studies revealed intramolecular hydroxamate bond scission to be a major decomposition pathway for 1 in aqueous buffer. Replacement of the hydroxamate bond with a tertiary amide, as in 16, afforded a conformational isostere resistant to degradation. These studies pave the way for the design and synthesis of analogues with improved chemical stability and biological activity.

Total Synthesis of Pseudouridimycin.

Pseudouridimycin (1), a potent antibiotic against both Gram-positive and Gram-negative bacteria including multi-drug-resistant strains with a new mode of action isolated from Streptomyces sp., was synthesized by a convergent strategy from 5'-amino-pseudouridine 5 and N-hydroxy-dipeptide 26 in 23% total yield. The key intermediate 26 was synthesized by hydroxylaminolysis of the nitrone derived from glutamine and subsequent glycylation with glycine chloride. The synthetic method provides an efficient and practical way for the synthesis of N-hydroxylated peptidyl nucleoside.

Eficacia y seguridad de Molnupiravir para el tratamiento de COVID-19: actualización enero 2022 / Eficacia y seguridad de Molnupiravir para el tratamiento de COVID-19: nueva actualización 2022

INTRODUCCIÓN: El virus SARS-CoV-2, sigue presentando múltiples variantes, el espectro de la enfermedad es amplio e incluye desde cuadros leves y autolimitados hasta neumonía atípica severa y progresiva, falla multiorgánica y muerte. Actualmente, existen vacunas eficaces para prevenir COVID-19, y se están desarrollando otras más nuevas para la prevención del COVID-19 grave, asimismo algunos medicamentos muestran resultados promisorios para prevenir y combatir esta enfermedad. Sin embargo, se carece de fármacos antivirales eficaces para combatir esta enfermedad. Aunque algunos de los medicamentos han recibido recientemente una autorización de uso de emergencia (EUA) para COVID-19, todos estos agentes farmacéuticos deben administrarse en entornos hospitalarios. Este documento fue elaborado para actualizar la evidencia científica disponible respecto a la eficacia y seguridad de Molnupiravir en el tratamiento de COVID-19. OBJETIVO: El objetivo de este documento es actualizar la evidencia científica disponible respecto a la eficacia y seguridad de Molnupiravir en el tratamiento de COVID-19, mejorando los desenlaces de los pacientes, y/o previniendo la enfermedad COVID-19 causada por el virus SARS-CoV-2. MÉTODOS: En la SNT N° 08-2021: E

Off-Therapy Response After Nucleos(t)ide Analogue Withdrawal in Patients With Chronic Hepatitis B: An International, Multicenter, Multiethnic Cohort (RETRACT-B Study).

BACKGROUND & AIMS: Functional cure, defined based on hepatitis B surface antigen (HBsAg) loss, is rare during nucleos(t)ide analogue (NA) therapy and guidelines on finite NA therapy have not been well established. We aim to analyze off-therapy outcomes after NA cessation in a large, international, multicenter, multiethnic cohort of patients with chronic hepatitis B (CHB). METHODS: This cohort study included patients with virally suppressed CHB who were hepatitis B e antigen (HBeAg)-negative and stopped NA therapy. Primary outcome was HBsAg loss after NA cessation, and secondary outcomes included virologic, biochemical, and clinical relapse, alanine aminotransferase flare, retreatment, and liver-related events after NA cessation. RESULTS: Among 1552 patients with CHB, cumulative probability of HBsAg loss was 3.2% at 12 months and 13.0% at 48 months of follow-up. HBsAg loss was higher among Whites (vs Asians: subdistribution hazard ratio, 6.8; 95% confidence interval, 2.7-16.8; P < .001) and among patients with HBsAg levels <100 IU/mL at end of therapy (vs ≥100 IU/mL: subdistribution hazard ratio, 22.5; 95% confidence interval, 13.1-38.7; P < .001). At 48 months of follow-up, Whites with HBsAg levels <1000 IU/mL and Asians with HBsAg levels <100 IU/mL at end of therapy had a high predicted probability of HBsAg loss (>30%). Incidence rate of hepatic decompensation and hepatocellular carcinoma was 0.48 per 1000 person-years and 0.29 per 1000 person-years, respectively. Death occurred in 7/19 decompensated patients and 2/14 patients with hepatocellular carcinoma. CONCLUSIONS: The best candidates for NA withdrawal are virally suppressed, HBeAg- negative, noncirrhotic patients with CHB with low HBsAg levels, particularly Whites with <1000 IU/mL and Asians with <100 IU/mL. However, strict surveillance is recommended to prevent deterioration.

Insufficient immunity led to virologic breakthrough in NAs-treated chronic hepatitis B patients switching to Peg-IFN-ɑ.

BACKGROUND: Virologic breakthrough (VBT) may occur in chronic hepatitis B (CHB) patients after switching from nucleos(t)ide analogues (NAs) to pegylated interferon alpha (Peg-IFN-ɑ). This study aimed to characterize the clinical and immunological features of VBT. METHODS: In NAs-treated patients switching to Peg-IFN-ɑ, innate and adaptive immune cell proportions were examined in peripheral blood and liver biopsy specimens. In vitro effect of IFN-ɑ on the expressions of toll-like receptors 2 (TLR2) and programmed cell death ligand 1 (PDL1) on monocytes, programmed cell death 1 (PD1) on CD8+T cells was examined. Peripheral blood mononuclear cells (PBMCs) were treated with TLR2 agonist and/or PDL1 blockade to evaluate their effect on HBV replication. RESULTS: 33 of 166 patients switching to Peg-IFN-ɑ experienced VBT after NA cessation, with majority being hepatitis B e antigen (HBeAg) positive or having higher hepatitis B core-related antigen (HBcrAg) levels. Patients with VBT exhibited lower proportions of TLR2+monocyte and increased PD1+HBV-specific CD8+T cell during the early phase of Peg-IFN-ɑ therapy after NA cessation in peripheral blood, as well as fewer TLR2+CD68+macrophages but more PDL1+CD68+macrophages and PD1+CD8+T cells in liver tissues. Simultaneous use of TLR2 agonist and PDL1 blockage ex vivo suppressed HBV replication by promoting cytokines production and CD8+T cells cytotoxicity. Upon in vitro IFN-ɑ stimulation, PDL1+monocytes and PD1+CD8+T cells were upregulated, whereas TLR2+monocytes were not increased in PBMC isolated from HBeAg-positive patients, or those with high HBcrAg titers. CONCLUSIONS: In NAs-treated patients, lower TLR2+monocyte and increased PD1+HBV-specific CD8+T cell proportions potentially contribute to VBT after switching to Peg-IFN-ɑ therapy. This insufficient immunity may be associated with the HBeAg status and HBcrAg levels.

Comparison of nucleoside and nucleotide analogs in the recurrence of hepatitis B virus-related hepatocellular carcinoma after surgical resection: A multicenter study.

BACKGROUND: Antiviral therapy should reduce the recurrence of hepatitis B virus-related hepatocellular carcinoma (HBV-related HCC) after surgical resection. However, there is little research on whether various antiviral drugs have different prognostic effects in patients with HBV-related HCC after curative liver resection. The present study compared the effects of nucleotide analog (NtA) and nucleoside analog (NsA) antiviral therapies after surgical resection on the prognosis of HBV-related HCC. METHODS: A total of 1303 patients with HBV-related HCC who received curative hepatectomy at five institutes between April 2014 and April 2019 were retrospectively enrolled and analyzed. Propensity matching analysis was used to compare the outcomes of HCC patients given NsA versus NtA therapy. Subgroup analysis of patients treated with entecavir (ETV) and tenofovir disoproxil fumarate (TDF) was also performed. RESULTS: Among 1303 patients, 759 (58.2%) patients developed recurrence, and 460 (35.3%) patients died. Multivariable analyses revealed that NtA therapy significantly decreased the risk of HCC recurrence (hazard ratio [HR], 0.64; 95% confidence interval [CI], 0.51-0.80; p < 0.001) and HCC-related death (HR, 0.52; 95% CI, 0.36-0.76; p = 0.001) compared to that with NsA therapy. Subgroup analysis showed that TDF treatment was associated with significantly lower rates of HCC recurrence (HR, 0.64; 95% CI, 0.49-0.83; p = 0.001) and death (HR, 0.32; 95% CI, 0.20-0.50; p < 0.001) than ETV treatment. CONCLUSIONS: Nucleotide analog treatment, but not NsA treatment, significantly reduced the risk of HCC recurrence in patients with HBV-related HCC and improved overall survival after curative hepatic resection.

Alerta monitoramento do horizonte tecnológico: Molnupiravir para o tratamento de COVID-19 / Technology horizon monitoring alert: Molnupiravir for the treatment of COVID-19

A TECNOLOGIA: Descrição da tecnologia: Molnupiravir (MK-4482 ou EIDD-2801) é um antiviral experimental de amplo espectro, atualmente em avaliação para uso na COVID-19. Este medicamento constitui-se de uma pró-droga do análogo de nucleosídeo N4-hidroxicitidina (NHC). No plasma, o molnupiravir é rapidamente convertido a NHC e, então, para a forma ativa da 5'-trifosfato. Esse último atua como um substrato competitivo para a RNA polimerase codificada pelo vírus, sendo incorporado e introduzindo mutações no RNA viral, as quais se acumulam a cada ciclo de replicação3 . Dessa forma, seu mecanismo de ação antiviral se dá pela introdução de erros de cópia durante a replicação do RNA do vírus . O molnupiravir tem alta barreia à resistência viral. Condição clínica: A COVID-19 é uma doença infecciosa respiratória aguda, causada pelo SARS-CoV-2 ou novo coronavírus, cujos primeiros relatos ocorreram em 2019 na cidade de Wuhan na China. No Brasil, o Ministério da Saúde (MS) recebeu a primeira notificação de um caso confirmado de COVID-19 em 26 de fevereiro de 2020. Em 11 de março de 2020, a Organização Mundial de Saúde (OMS) declarou pandemia de COVID-19. INFORMAÇÕES REGULATÓRIAS: Informações sobre registro: Até a última atualização deste alerta, o molnupiravir ainda não possuía registro definitivo ou autorização de uso emergencial na Agência Nacional de Vigilância Sanitária (Anvisa), U. S. Food and Drug Administration (FDA) e European Medicine Agency (EMA) para o tratamento na COVID-19. PANORAMA DE DESENVOLVIMENTO: Estratégia de busca: A estratégia de busca foi realizada em duas etapas. A primeira teve como objetivo identificar ensaios clínicos registrados sobre o uso do molnupiravir para tratamento da COVID-19. Para isso, foi consultada a base de dados ClinicalTrials.gov em 13 de outubro de 2021, usando-se o nome do medicamento (molnupiravir) e os seus códigos de pesquisa (EIDD-2801, MK-4482). Foram incluídos ensaios clínicos que avaliavam o uso de molnupiravir em pacientes com COVID-19, com qualquer tipo de desenho. A segunda etapa da busca objetivou identificar na literatura os resultados referentes aos ensaios clínicos sobre o molnupiravir no tratamento da COVID19. Para tal, no dia 14 de outubro de 2021, foram realizadas buscas nas bases de dados Medline (via Pubmed), Embase e Biblioteca Virtual em Saúde (BVS), utilizando-se termos controlados e seus respectivos sinônimos. Os ensaios clínicos são estudos realizados com humanos para medir os parâmetros de segurança e eficácia de novos medicamentos, sendo etapa essencial para a chegada de novas alternativas terapêuticas no mercado. Esses ensaios são divididos nas fases 1, 2, 3 e 4, de acordo com a quantidade de participantes e os objetivos específicos de cada etapa. CONSIDERAÇÕES FINAIS: O molnupiravir é um antiviral experimental análogo de ribonucleosídeo, que inibe a replicação a partir da indução de mutação no RNA viral. Está sendo investigado para o tratamento oral da COVID-19, especialmente em pessoas com sintomas precoces. Foram identificados cinco ensaios clínicos (um completo e quatro em andamento) com a tecnologia. Até a última atualização deste alerta, havia resultados disponíveis para dois desses estudos, sendo um deles sobre a fase 1 (AGILE/NCT04746183) de um ensaio fase 1/2 e o outro sobre um estudo de fase 2 (NCT04405570), esse último ainda em nota de imprensa. Os resultados da fase 1 apontam para um perfil de segurança favorável do uso de molnupiravir (800 mg por via oral duas vezes ao dia por cinco dias) em adultos com infecção por SARS-CoV-2 confirmada por PCR, início de sintomas em até cinco dias e não hospitalizados. Os resultados do estudo de fase 2 apontam para a redução da carga de RNA viral e de vírus infeccioso na nasofaringe com o uso de molnupiravir (especialmente na dose de 800 mg por via oral duas vezes ao dia por cinco dias) em adultos com infecção por SARS-CoV-2 confirmada e início de sintomas em até sete dias. Nesse estudo, o perfil de segurança também foi favorável. Também foi identificado uma nota de imprensa sobre os resultados interinos da fase 3 de um ensaio clínico fase 2/3 (NCT04575597/MOVe-OUT). Segundo informações da nota, o uso de molnupiravir reduziu em 50% o risco de hospitalização ou morte em adultos com COVID-19 leve a moderada, confirmada por teste laboratorial, início de sintomas em até cinco dias, não-hospitalizados e com pelo menos um fator de risco para desfechos desfavoráveis da doença. Diante do exposto, os resultados disponíveis apontam para um benefício da tecnologia no tratamento precoce de adultos com COVID-19 não hospitalizados. Entretanto, a evidência ainda é incipiente, referindo-se principalmente a resultados de ensaios clínicos de fase 1 e 2 (um deles ainda não publicado e não avaliado por pares), cujo objetivo principal foi avaliar a segurança e a eficácia virológica do molnupiravir. Além disso, os estudos apresentam limitações metodológicas importantes, a exemplo do pequeno tamanho amostral, pequena proporção de indivíduos com fatores de risco para desenvolvimento de doença grave e uso de desfechos primários que não são relevantes para demonstração de benefícios clínico do molnupiravir. Os resultados do ensaio clínico fase 3 que avalia desfechos clínicos relevantes, até a última atualização deste alerta, estava disponível apenas em forma de nota de imprensa, sem a descrição adequada da população incluída bem como dos detalhes metodológicos quanto à condução do estudo. Portanto, ainda não existem evidências substanciais para concluir sobre a eficácia e segurança do uso de molnupiravir no tratamento ou profilaxia de COVID-19. Além disso, para que ocorra a oferta do medicamento no SUS, é necessária sua análise pela Conitec, conforme disposto na Lei nº 12.401/2011, que alterou a Lei nº 8.080/1990. Os relatórios de recomendação da Conitec levam em consideração as evidências científicas sobre eficácia, a acurácia, a efetividade e a segurança do medicamento, e, também, a avaliação econômica comparativa dos benefícios e dos custos em relação às tecnologias já incorporadas e o impacto da incorporação da tecnologia no SUS.

An update on the progress of galidesivir (BCX4430), a broad-spectrum antiviral.

Galidesivir (BCX4430) is an adenosine nucleoside analog that is broadly active in cell culture against several RNA viruses of various families. This activity has also been shown in animal models of viral disease associated with Ebola, Marburg, yellow fever, Zika, and Rift Valley fever viruses. In many cases, the compound is more efficacious in animal models than cell culture activity would predict. Based on favorable data from in vivo animal studies, galidesivir has recently undergone evaluation in several phase I clinical trials, including against severe acute respiratory syndrome coronavirus 2, and as a medical countermeasure for the treatment of Marburg virus disease.

Eficacia y Seguridad de Molnupiravir para el tratamiento de COVID-19 / Efficacy and Safety of Molnupiravir for the treatment of COVID-19

ANTECEDENTES: El presente informe se realiza a solicitud del Viceministerio de Salud Pública del Ministerio de Salud, y tiene como objetivo identificar la evidencia disponible respecto a la eficacia y seguridad de Molnupiravir en el tratamiento de COVID-19. MÉTODOS: Se realizó una búsqueda en la plataforma Living Overview of the Evidence (L·OVE) de la Fundación Epistemonikos que consulta 10 bases de datos incluyendo Medline, EMBASE, LILACS, medRxiv (un servidor de distribución de manuscritos aún no publicados, sin certificación de revisión por pares) entre otros. La búsqueda se realizó el 13/08/2021. Los detalles de la metodología de búsqueda utilizada por L.OVE se encuentra disponible en: https://app.iloveevidence.com/loves/5e6fdb9669c00e4ac072701d?population=5e7fce7e3d05156b5f5e 032a§ion=methods&classification=all. Adicionalmente se consultó el Registro Cochrane Central de Ensayos Controlados (CENTRAL) y Clinicaltrials.gov de los Institutos de Salud Pública de Estados Unidos (NIH) para identificar los ensayos en curso. La pregunta PICO abordada es: ¿Cuál es la eficacia y seguridad de Molnupiravir para el tratamiento de COVID-19? Ensayos clínicos controlados que reporten la eficacia y seguridad para al menos uno de los desenlaces de interés. Idioma: inglés o español. Se excluyeron estudios preclínicos, series de casos, reportes de casos, reportes breves y cartas al editor. RESULTADOS: La búsqueda sistemática identificó 34 artículos, a la lectura de título y resumen fueron seleccionados 11, solo 3 respondían a la pregunta PICO. Características de los estudios incluidos: Se identificaron 03 estudios que cumplieron con los criterios de selección: 2 ensayos clínicos en fase I y uno en fase II. Dentro de los resultados, 2 de los reportes corresponden a manuscritos no publicados ni certificados por una revisión por pares, lo que implica que éstos podrían modificar sus resultados y/o conclusiones en sucesivas versiones hasta su publicación y no es posible garantizar que respondan satisfactoriamente la revisión por pares y sean finalmente publicados. CONCLUSIONES: Se identificaron 3 estudios, dos ensayos clínicos de fase I y uno de fase II. Dos ensayos evalúan la seguridad y tolerabilidad de una dosis de Molnupiravir comparada con placebo o cuidado estándar para ser usada en otra fase. Los estudios de fase I, reportan la incidencia de eventos adversos evaluados para el uso de Molnupiravir en dosis desde 200 hasta 800 mg vía oral. Sin embargo, recomiendan la dosis de 800 para futuros estudios. Un estudio de fase II, sugiere el uso de Molnupiravir para reducir la replicación del virus en etapas tempranas de la enfermedad, siendo que en el grupo en el que se presentaron mejores resultados fue con la dosis de 800 mg en solo 55 participantes comparado con 62 participantes en el grupo de 400 mg, 23 en el grupo de 200 mg y 62 en el grupo placebo. No existe evidencia sobre la eficacia o efectividad del uso de Molnupiravir para el tratamiento profiláctico, o terapéutico contra COVID-19, pues los ensayos clínicos desarrollados aún se encuentran en fases iniciales para determinar la dosis adecuada con mejores resultados clínicos.

Targeting SARS-CoV-2 Polymerase with New Nucleoside Analogues.

Despite the fact that COVID-19 vaccines are already available on the market, there have not been any effective FDA-approved drugs to treat this disease. There are several already known drugs that through drug repositioning have shown an inhibitory activity against SARS-CoV-2 RNA-dependent RNA polymerase. These drugs are included in the family of nucleoside analogues. In our efforts, we synthesized a group of new nucleoside analogues, which are modified at the sugar moiety that is replaced by a quinazoline entity. Different nucleobase derivatives are used in order to increase the inhibition. Five new nucleoside analogues were evaluated with in vitro assays for targeting polymerase of SARS-CoV-2.

Synthesis and Antiviral Evaluation of Nucleoside Analogues Bearing One Pyrimidine Moiety and Two D-Ribofuranosyl Residues.

A series of 1,2,3-triazolyl nucleoside analogues in which 1,2,3-triazol-4-yl-ß-d-ribofuranosyl fragments are attached via polymethylene linkers to both nitrogen atoms of the heterocycle moiety (uracil, 6-methyluracil, thymine, quinazoline-2,4-dione, alloxazine) or to the C-5 and N-3 atoms of the 6-methyluracil moiety was synthesized. All compounds synthesized were evaluated for antiviral activity against influenza virus A/PR/8/34/(H1N1) and coxsackievirus B3. Antiviral assays revealed three compounds, 2i, 5i, 11c, which showed moderate activity against influenza virus A H1N1 with IC50 values of 57.5 µM, 24.3 µM, and 29.2 µM, respectively. In the first two nucleoside analogues, 1,2,3-triazol-4-yl-ß-d-ribofuranosyl fragments are attached via butylene linkers to N-1 and N-3 atoms of the heterocycle moiety (6-methyluracil and alloxazine, respectively). In nucleoside analogue 11c, two 1,2,3-triazol-4-yl-2',3',5'-tri-O-acetyl-ß-d-ribofuranose fragments are attached via propylene linkers to the C-5 and N-3 atoms of the 6-methyluracil moiety. Almost all synthesized 1,2,3-triazolyl nucleoside analogues showed no antiviral activity against the coxsackie B3 virus. Two exceptions are 1,2,3-triazolyl nucleoside analogs 2f and 5f, in which 1,2,3-triazol-4-yl-2',3',5'-tri-O-acetyl-ß-d-ribofuranose fragments are attached to the C-5 and N-3 atoms of the heterocycle moiety (6-methyluracil and alloxazine respectively). These compounds exhibited high antiviral potency against the coxsackie B3 virus with IC50 values of 12.4 and 11.3 µM, respectively, although both were inactive against influenza virus A H1N1. According to theoretical calculations, the antiviral activity of the 1,2,3-triazolyl nucleoside analogues 2i, 5i, and 11c against the H1N1 (A/PR/8/34) influenza virus can be explained by their influence on the functioning of the polymerase acidic protein (PA) of RNA-dependent RNA polymerase (RdRp). As to the antiviral activity of nucleoside analogs 2f and 5f against coxsackievirus B3, it can be explained by their interaction with the coat proteins VP1 and VP2.

Nucleoside Analogues Are Potent Inducers of Pol V-mediated Mutagenesis.

Drugs targeting DNA and RNA in mammalian cells or viruses can also affect bacteria present in the host and thereby induce the bacterial SOS system. This has the potential to increase mutagenesis and the development of antimicrobial resistance (AMR). Here, we have examined nucleoside analogues (NAs) commonly used in anti-viral and anti-cancer therapies for potential effects on mutagenesis in Escherichia coli, using the rifampicin mutagenicity assay. To further explore the mode of action of the NAs, we applied E. coli deletion mutants, a peptide inhibiting Pol V (APIM-peptide) and metabolome and proteome analyses. Five out of the thirteen NAs examined, including three nucleoside reverse transcriptase inhibitors (NRTIs) and two anti-cancer drugs, increased the mutation frequency in E. coli by more than 25-fold at doses that were within reported plasma concentration range (Pl.CR), but that did not affect bacterial growth. We show that the SOS response is induced and that the increase in mutation frequency is mediated by the TLS polymerase Pol V. Quantitative mass spectrometry-based metabolite profiling did not reveal large changes in nucleoside phosphate or other central carbon metabolite pools, which suggests that the SOS induction is an effect of increased replicative stress. Our results suggest that NAs/NRTIs can contribute to the development of AMR and that drugs inhibiting Pol V can reverse this mutagenesis.