RESUMEN
ETHNOPHARMACOLOGY RELEVANCE: Lepidium meyenii Walp. (maca) has been traditionally used for centuries in the Central Andes region both as food and as medicine. In the last decades, its fertility enhancer properties have gained importance, with the majority of the scientific literature related to this topic. However, other traditional uses are less known as metabolic or infectious diseases. AIM OF THE STUDY: The main purpose of this study is to investigate the anti-infectious activity of L. meyenii, specifically in HIV-1 infection. There are previous reports of the transcriptional related activity of L. meyenii extracts in human T lymphocytes via transcription factors as NF-κB. Since T lymphocytes are the main target of HIV-1 infection and NF-κB is strongly involved in HIV-1 transcription, L. meyenii could display antiviral activity. MATERIAL AND METHODS: Chromatography and spectroscopy techniques were used to isolate and identify the compounds in the active extracts. An antiviral assay system based on recombinant viruses was used to evaluate the anti-HIV activity. Cell toxicity was tested for all the extracts and compounds. Viral entry was studied using VSV-HIV chimera viruses and reverse transcription and viral integration were studied by qPCR of viral DNA in infected cells. Finally, viral transcription was studied in primary lymphocytes transfected with HIV-1 or NF-κB luciferase reporter plasmids. RESULTS: n-Hexane extracts of purple maca displayed anti-HIV activity in an in vitro assay. A bioassay-guided fractionation led to the identification of three thiadiazole alkaloids with antiviral activity. All the compounds were able to inhibit HIV infection of MT-2 cell lines and primary lymphocytes (PBMCs) with IC50 values in the low micromolar range. The mechanism of action differs between the three compounds: one of them showed activity on viral entry, and all the three compounds inhibited viral integration at low concentrations. Remarkably, none of the compounds inhibited reverse transcription or viral transcription. CONCLUSIONS: n-Hexane extracts of the purple ecotype of L. meyenii inhibit HIV-1 infection in vitro and three active thiadiazole alkaloids were isolated acting mainly on viral integration and viral entry.
Asunto(s)
Alcaloides , Fármacos Anti-VIH , Infecciones por VIH , VIH-1 , Lepidium , Tiadiazoles , Integración Viral , Humanos , Alcaloides/aislamiento & purificación , Alcaloides/farmacología , Línea Celular , Etnofarmacología , Regulación Viral de la Expresión Génica/efectos de los fármacos , Infecciones por VIH/tratamiento farmacológico , Infecciones por VIH/virología , VIH-1/efectos de los fármacos , VIH-1/genética , VIH-1/crecimiento & desarrollo , VIH-1/fisiología , Hipocótilo/química , Lepidium/química , Linfocitos/efectos de los fármacos , Linfocitos/virología , Extractos Vegetales/química , Extractos Vegetales/farmacología , Transcripción Reversa/efectos de los fármacos , Tiadiazoles/aislamiento & purificación , Tiadiazoles/farmacología , Transcripción Genética/efectos de los fármacos , Integración Viral/efectos de los fármacos , Internalización del Virus/efectos de los fármacos , Replicación Viral/efectos de los fármacos , Fármacos Anti-VIH/química , Fármacos Anti-VIH/aislamiento & purificación , Fármacos Anti-VIH/farmacologíaRESUMEN
Epstein-Barr virus (EBV) is linked to lymphoma and epithelioma but lacks drugs specifically targeting EBV-positive tumors. BamHI A Rightward Transcript (BART) miRNAs are expressed in all EBV-positive tumors, suppressing both lytic infection and host cell apoptosis. We identified suberoylanilide hydroxamic acid (SAHA), an inhibitor of histone deacetylase enzymes, as an agent that suppresses BART promoter activity and transcription of BART miRNAs. SAHA treatment demonstrated a more pronounced inhibition of cell proliferation in EBV-positive cells compared to EBV-negative cells, affecting both p53 wild-type and mutant gastric epithelial cells. SAHA treatment enhanced lytic infection in wild-type EBV-infected cells, while also enhancing cell death in BZLF1-deficient EBV-infected cells. It reduced BART gene expression by 85% and increased the expression of proapoptotic factors targeted by BART miRNAs. These findings suggest that SAHA not only induces lytic infection but also leads to cell death by suppressing BART miRNA transcription and promoting the apoptotic program.
Asunto(s)
Apoptosis , Herpesvirus Humano 4 , Ácidos Hidroxámicos , MicroARNs , Vorinostat , Vorinostat/farmacología , Apoptosis/efectos de los fármacos , Humanos , MicroARNs/genética , MicroARNs/metabolismo , Herpesvirus Humano 4/genética , Herpesvirus Humano 4/fisiología , Herpesvirus Humano 4/efectos de los fármacos , Ácidos Hidroxámicos/farmacología , Regulación Viral de la Expresión Génica/efectos de los fármacos , Línea Celular , Inhibidores de Histona Desacetilasas/farmacología , Regiones Promotoras Genéticas , Proliferación Celular/efectos de los fármacosRESUMEN
Repression of human cytomegalovirus (HCMV) immediate-early (IE) gene expression is a key regulatory step in the establishment and maintenance of latent reservoirs. Viral IE transcription and protein accumulation can be elevated during latency by treatment with histone deacetylase inhibitors such as valproic acid (VPA), rendering infected cells visible to adaptive immune responses. However, the latency-associated viral protein UL138 inhibits the ability of VPA to enhance IE gene expression during infection of incompletely differentiated myeloid cells that support latency. UL138 also limits the accumulation of IFNß transcripts by inhibiting the cGAS-STING-TBK1 DNA-sensing pathway. Here, we show that, in the absence of UL138, the cGAS-STING-TBK1 pathway promotes both IFNß accumulation and VPA-responsive IE gene expression in incompletely differentiated myeloid cells. Inactivation of this pathway by either genetic or pharmacological inhibition phenocopied UL138 expression and reduced VPA-responsive IE transcript and protein accumulation. This work reveals a link between cytoplasmic pathogen sensing and epigenetic control of viral lytic phase transcription and suggests that manipulation of pattern recognition receptor signaling pathways could aid in the refinement of MIEP regulatory strategies to target latent viral reservoirs.
Asunto(s)
Citomegalovirus , Proteínas de la Membrana , Células Mieloides , Nucleotidiltransferasas , Proteínas Serina-Treonina Quinasas , Transducción de Señal , Ácido Valproico , Humanos , Ácido Valproico/farmacología , Células Mieloides/virología , Células Mieloides/metabolismo , Células Mieloides/efectos de los fármacos , Transducción de Señal/efectos de los fármacos , Proteínas de la Membrana/metabolismo , Proteínas de la Membrana/genética , Citomegalovirus/fisiología , Citomegalovirus/efectos de los fármacos , Citomegalovirus/genética , Nucleotidiltransferasas/metabolismo , Nucleotidiltransferasas/genética , Proteínas Serina-Treonina Quinasas/metabolismo , Proteínas Serina-Treonina Quinasas/genética , Infecciones por Citomegalovirus/virología , Infecciones por Citomegalovirus/metabolismo , Infecciones por Citomegalovirus/genética , Latencia del Virus/efectos de los fármacos , Transcripción Genética/efectos de los fármacos , Diferenciación Celular/efectos de los fármacos , Regulación Viral de la Expresión Génica/efectos de los fármacos , Genes Inmediatos-Precoces , Interferón beta/metabolismo , Interferón beta/genéticaRESUMEN
The unique virus-cell interaction in Epstein-Barr virus (EBV)-associated malignancies implies targeting the viral latent-lytic switch is a promising therapeutic strategy. However, the lack of specific and efficient therapeutic agents to induce lytic cycle in these cancers is a major challenge facing clinical implementation. We develop a synthetic transcriptional activator that specifically activates endogenous BZLF1 and efficiently induces lytic reactivation in EBV-positive cancer cells. A lipid nanoparticle encapsulating nucleoside-modified mRNA which encodes a BZLF1-specific transcriptional activator (mTZ3-LNP) is synthesized for EBV-targeted therapy. Compared with conventional chemical inducers, mTZ3-LNP more efficiently activates EBV lytic gene expression in EBV-associated epithelial cancers. Here we show the potency and safety of treatment with mTZ3-LNP to suppress tumor growth in EBV-positive cancer models. The combination of mTZ3-LNP and ganciclovir yields highly selective cytotoxic effects of mRNA-based lytic induction therapy against EBV-positive tumor cells, indicating the potential of mRNA nanomedicine in the treatment of EBV-associated epithelial cancers.
Asunto(s)
Infecciones por Virus de Epstein-Barr , Herpesvirus Humano 4 , Liposomas , Nanopartículas , Transactivadores , Humanos , Herpesvirus Humano 4/genética , Transactivadores/metabolismo , Transactivadores/genética , Infecciones por Virus de Epstein-Barr/virología , Infecciones por Virus de Epstein-Barr/tratamiento farmacológico , Animales , Nanopartículas/química , Línea Celular Tumoral , Ratones , ARN Mensajero/genética , ARN Mensajero/metabolismo , Activación Viral/efectos de los fármacos , Ensayos Antitumor por Modelo de Xenoinjerto , Regulación Viral de la Expresión Génica/efectos de los fármacos , Ratones Desnudos , FemeninoRESUMEN
Bovine alphaherpesvirus 1 (BoHV-1) infection causes respiratory tract disorders and immune suppression and may induce bacterial pneumonia. BoHV-1 establishes lifelong latency in sensory neurons after acute infection. Reactivation from latency consistently occurs following stress or intravenous injection of the synthetic corticosteroid dexamethasone (DEX), which mimics stress. The immediate early transcription unit 1 (IEtu1) promoter drives expression of infected cell protein 0 (bICP0) and bICP4, two viral transcriptional regulators necessary for productive infection and reactivation from latency. The IEtu1 promoter contains two glucocorticoid receptor (GR) responsive elements (GREs) that are transactivated by activated GR. GC-rich motifs, including consensus binding sites for specificity protein 1 (Sp1), are in the IEtu1 promoter sequences. E2F family members bind a consensus sequence (TTTCCCGC) and certain specificity protein 1 (Sp1) sites. Consequently, we hypothesized that certain E2F family members activate IEtu1 promoter activity. DEX treatment of latently infected calves increased the number of E2F2+ TG neurons. GR and E2F2, but not E2F1, E2F3a, or E2F3b, cooperatively transactivate a 436-bp cis-regulatory module in the IEtu1 promoter that contains both GREs. A luciferase reporter construct containing a 222-bp fragment downstream of the GREs was transactivated by E2F2 unless two adjacent Sp1 binding sites were mutated. Chromatin immunoprecipitation studies revealed that E2F2 occupied IEtu1 promoter sequences when the BoHV-1 genome was transfected into mouse neuroblastoma (Neuro-2A) or monkey kidney (CV-1) cells. In summary, these findings revealed that GR and E2F2 cooperatively transactivate IEtu1 promoter activity, which is predicted to influence the early stages of BoHV-1 reactivation from latency. IMPORTANCE: Bovine alpha-herpesvirus 1 (BoHV-1) acute infection in cattle leads to establishment of latency in sensory neurons in the trigeminal ganglia (TG). A synthetic corticosteroid dexamethasone consistently initiates BoHV-1 reactivation in latently infected calves. The BoHV-1 immediate early transcription unit 1 (IEtu1) promoter regulates expression of infected cell protein 0 (bICP0) and bICP4, two viral transcriptional regulators. Hence, the IEtu1 promoter must be activated for the reactivation to occur. The number of TG neurons expressing E2F2, a transcription factor and cell cycle regulator, increased during early stages of reactivation from latency. The glucocorticoid receptor (GR) and E2F2, but not E2F1, E2F3a, or E2F3b, cooperatively transactivated a 436-bp cis-regulatory module (CRM) in the IEtu1 promoter that contains two GR responsive elements. Chromatin immunoprecipitation studies revealed that E2F2 occupies IEtu1 promoter sequences in cultured cells. GR and E2F2 mediate cooperative transactivation of IEtu1 promoter activity, which is predicted to stimulate viral replication following stressful stimuli.
Asunto(s)
Ciclo Celular , Factor de Transcripción E2F2 , Regulación Viral de la Expresión Génica , Herpesvirus Bovino 1 , Proteínas Inmediatas-Precoces , Regiones Promotoras Genéticas , Receptores de Glucocorticoides , Activación Transcripcional , Animales , Bovinos , Ratones , Sitios de Unión , Línea Celular , Dexametasona/farmacología , Factor de Transcripción E2F2/metabolismo , Regulación Viral de la Expresión Génica/efectos de los fármacos , Regulación Viral de la Expresión Génica/genética , Infecciones por Herpesviridae/virología , Infecciones por Herpesviridae/metabolismo , Infecciones por Herpesviridae/veterinaria , Infecciones por Herpesviridae/genética , Herpesvirus Bovino 1/genética , Herpesvirus Bovino 1/fisiología , Proteínas Inmediatas-Precoces/genética , Neuronas/virología , Receptores de Glucocorticoides/metabolismo , Elementos de Respuesta/genética , Factor de Transcripción Sp1/metabolismo , Transactivadores/metabolismo , Ganglio del Trigémino/citología , Ganglio del Trigémino/virología , Activación Viral , Latencia del VirusRESUMEN
Within the first 15 minutes of infection, herpes simplex virus 1 immediate early proteins repurpose cellular RNA polymerase (Pol II) for viral transcription. An important role of the viral-infected cell protein 27 (ICP27) is to facilitate viral pre-mRNA processing and export viral mRNA to the cytoplasm. Here, we use precision nuclear run-on followed by deep sequencing (PRO-seq) to characterize transcription of a viral ICP27 null mutant. At 1.5 and 3 hours post infection (hpi), we observed increased total levels of Pol II on the mutant viral genome and accumulation of Pol II downstream of poly A sites indicating increased levels of initiation and processivity. By 6 hpi, Pol II accumulation on specific mutant viral genes was higher than that on wild-type virus either at or upstream of poly A signals, depending on the gene. The PRO-seq profile of the ICP27 mutant on late genes at 6 hpi was similar but not identical to that caused by treatment with flavopiridol, a known inhibitor of RNA processivity. This pattern was different from PRO-seq profiles of other α gene mutants and upon inhibition of viral DNA replication with PAA. Together, these results indicate that ICP27 contributes to the repression of aberrant viral transcription at 1.5 and 3 hpi by inhibiting initiation and decreasing RNA processivity. However, ICP27 is needed to enhance processivity on most late genes by 6 hpi in a mechanism distinguishable from its role in viral DNA replication.IMPORTANCEWe developed and validated the use of a processivity index for precision nuclear run-on followed by deep sequencing data. The processivity index calculations confirm infected cell protein 27 (ICP27) induces downstream of transcription termination on certain host genes. The processivity indices and whole gene probe data implicate ICP27 in transient immediate early gene-mediated repression, a process that also requires ICP4, ICP22, and ICP0. The data indicate that ICP27 directly or indirectly regulates RNA polymerase (Pol II) initiation and processivity on specific genes at specific times post infection. These observations support specific and varied roles for ICP27 in regulating Pol II activity on viral genes in addition to its known roles in post transcriptional mRNA processing and export.
Asunto(s)
Genoma Viral , Herpesvirus Humano 1 , Proteínas Inmediatas-Precoces , Mutación , ARN Polimerasa II , Transcripción Viral , Animales , Humanos , Línea Celular , Chlorocebus aethiops , Regulación Viral de la Expresión Génica/efectos de los fármacos , Genes Virales/genética , Genoma Viral/genética , Herpes Simple/virología , Herpes Simple/genética , Herpesvirus Humano 1/genética , Herpesvirus Humano 1/fisiología , Proteínas Inmediatas-Precoces/deficiencia , Proteínas Inmediatas-Precoces/genética , Poli A/genética , Poli A/metabolismo , ARN Polimerasa II/metabolismo , ARN Mensajero/genética , ARN Mensajero/metabolismo , ARN Viral/genética , ARN Viral/metabolismo , Células Vero , Transcripción Viral/efectos de los fármacos , Transcripción Viral/genética , Replicación Viral/genéticaRESUMEN
Nucleot(s)ide analogues, the current antiviral treatments against chronic hepatitis B (CHB) infection, are non-curative due to their inability to eliminate covalently closed circular DNA (cccDNA) from the infected hepatocytes. Preclinical studies have shown that coumarin derivatives can effectively reduce the HBV DNA replication. We evaluated the antiviral efficacy of thirty new coumarin derivatives in cell culture models for studying HBV. Furanocoumarins Fc-20 and Fc-31 suppressed the levels of pre-genomic RNA as well as cccDNA, and reduced the secretion of virions, HBsAg and HBeAg. The antiviral efficacies of Fc-20 and Fc31 improved further when used in combination with the hepatitis B antiviral drug Entecavir. There was a marked reduction in the intracellular HBx level in the presence of these furanocoumarins due to proteasomal degradation resulting in the down-regulation of HBx-dependent viral genes. Importantly, both Fc-20 and Fc-31 were non-cytotoxic to cells even at high concentrations. Further, our molecular docking studies confirmed a moderate to high affinity interaction between furanocoumarins and viral HBx via residues Ala3, Arg26 and Lys140. These data suggest that furanocoumarins could be developed as a new therapeutic for CHB infection.
Asunto(s)
Antivirales , ADN Circular , Furocumarinas , Virus de la Hepatitis B , Complejo de la Endopetidasa Proteasomal , Transactivadores , Proteínas Reguladoras y Accesorias Virales , Replicación Viral , Virus de la Hepatitis B/efectos de los fármacos , Virus de la Hepatitis B/genética , Virus de la Hepatitis B/fisiología , Virus de la Hepatitis B/metabolismo , Replicación Viral/efectos de los fármacos , Humanos , Transactivadores/metabolismo , Transactivadores/genética , ADN Circular/metabolismo , ADN Circular/genética , Proteínas Reguladoras y Accesorias Virales/metabolismo , Proteínas Reguladoras y Accesorias Virales/genética , Furocumarinas/farmacología , Antivirales/farmacología , Complejo de la Endopetidasa Proteasomal/metabolismo , ADN Viral/metabolismo , ADN Viral/genética , Regulación hacia Abajo/efectos de los fármacos , Transcripción Genética/efectos de los fármacos , Proteolisis/efectos de los fármacos , Regulación Viral de la Expresión Génica/efectos de los fármacos , Células Hep G2RESUMEN
DNA replication of E1-deleted first-generation adenoviruses (AdV) in cultured cancer cells has been reported repeatedly and it was suggested that certain cellular proteins could functionally compensate for E1A, leading to the expression of the early region 2 (E2)-encoded proteins and subsequently virus replication. Referring to this, the observation was named E1A-like activity. In this study, we investigated different cell cycle inhibitors with respect to their ability to increase viral DNA replication of dl70-3, an E1-deleted adenovirus. Our analyses of this issue revealed that in particular inhibition of cyclin-dependent kinases 4/6 (CDK4/6i) increased E1-independent adenovirus E2-expression and viral DNA replication. Detailed analysis of the E2-expression in dl70-3 infected cells by RT-qPCR showed that the increase in E2-expression originated from the E2-early promoter. Mutations of the two E2F-binding sites in the E2-early promoter (pE2early-LucM) caused a significant reduction in E2-early promoter activity in trans-activation assays. Accordingly, mutations of the E2F-binding sites in the E2-early promoter in a virus named dl70-3/E2Fm completely abolished CDK4/6i induced viral DNA replication. Thus, our data show that E2F-binding sites in the E2-early promoter are crucial for E1A independent adenoviral DNA replication of E1-deleted vectors in cancer cells. IMPORTANCE E1-deleted AdV vectors are considered replication deficient and are important tools for the study of virus biology, gene therapy, and large-scale vaccine development. However, deletion of the E1 genes does not completely abolish viral DNA replication in cancer cells. Here, we report, that the two E2F-binding sites in the adenoviral E2-early promoter contribute substantially to the so-called E1A-like activity in tumor cells. With this finding, on the one hand, the safety profile of viral vaccine vectors can be increased and, on the other hand, the oncolytic property for cancer therapy might be improved through targeted manipulation of the host cell.
Asunto(s)
Adenoviridae , Ciclo Celular , Replicación del ADN , Replicación Viral , Adenoviridae/genética , Adenoviridae/metabolismo , Proteínas E1A de Adenovirus/genética , Proteínas E1A de Adenovirus/metabolismo , Sitios de Unión , Ciclo Celular/efectos de los fármacos , Línea Celular Tumoral , Células/efectos de los fármacos , Células/virología , Replicación del ADN/efectos de los fármacos , ADN Viral/metabolismo , Regulación Viral de la Expresión Génica/efectos de los fármacos , Mutación , Regiones Promotoras Genéticas/genética , Inhibidores de Proteínas Quinasas/farmacología , Replicación Viral/fisiología , HumanosRESUMEN
Rare CD4 T cells that contain HIV under antiretroviral therapy represent an important barrier to HIV cure1-3, but the infeasibility of isolating and characterizing these cells in their natural state has led to uncertainty about whether they possess distinctive attributes that HIV cure-directed therapies might exploit. Here we address this challenge using a microfluidic technology that isolates the transcriptomes of HIV-infected cells based solely on the detection of HIV DNA. HIV-DNA+ memory CD4 T cells in the blood from people receiving antiretroviral therapy showed inhibition of six transcriptomic pathways, including death receptor signalling, necroptosis signalling and antiproliferative Gα12/13 signalling. Moreover, two groups of genes identified by network co-expression analysis were significantly associated with HIV-DNA+ cells. These genes (n = 145) accounted for just 0.81% of the measured transcriptome and included negative regulators of HIV transcription that were higher in HIV-DNA+ cells, positive regulators of HIV transcription that were lower in HIV-DNA+ cells, and other genes involved in RNA processing, negative regulation of mRNA translation, and regulation of cell state and fate. These findings reveal that HIV-infected memory CD4 T cells under antiretroviral therapy are a distinctive population with host gene expression patterns that favour HIV silencing, cell survival and cell proliferation, with important implications for the development of HIV cure strategies.
Asunto(s)
Linfocitos T CD4-Positivos , Regulación Viral de la Expresión Génica , Infecciones por VIH , VIH-1 , Latencia del Virus , Humanos , Linfocitos T CD4-Positivos/citología , Linfocitos T CD4-Positivos/efectos de los fármacos , Linfocitos T CD4-Positivos/metabolismo , Linfocitos T CD4-Positivos/virología , Proliferación Celular/efectos de los fármacos , Supervivencia Celular/efectos de los fármacos , ADN Viral/aislamiento & purificación , Regulación Viral de la Expresión Génica/efectos de los fármacos , Infecciones por VIH/tratamiento farmacológico , Infecciones por VIH/genética , Infecciones por VIH/inmunología , Infecciones por VIH/virología , VIH-1/efectos de los fármacos , VIH-1/genética , VIH-1/aislamiento & purificación , VIH-1/patogenicidad , Memoria Inmunológica , Microfluídica , Necroptosis/efectos de los fármacos , Transducción de Señal/efectos de los fármacos , Transcriptoma/efectos de los fármacos , Latencia del Virus/efectos de los fármacos , Antirretrovirales/farmacología , Antirretrovirales/uso terapéuticoRESUMEN
Across biological scales, gene-regulatory networks employ autorepression (negative feedback) to maintain homeostasis and minimize failure from aberrant expression. Here, we present a proof of concept that disrupting transcriptional negative feedback dysregulates viral gene expression to therapeutically inhibit replication and confers a high evolutionary barrier to resistance. We find that nucleic-acid decoys mimicking cis-regulatory sites act as "feedback disruptors," break homeostasis, and increase viral transcription factors to cytotoxic levels (termed "open-loop lethality"). Feedback disruptors against herpesviruses reduced viral replication >2-logs without activating innate immunity, showed sub-nM IC50, synergized with standard-of-care antivirals, and inhibited virus replication in mice. In contrast to approved antivirals where resistance rapidly emerged, no feedback-disruptor escape mutants evolved in long-term cultures. For SARS-CoV-2, disruption of a putative feedback circuit also generated open-loop lethality, reducing viral titers by >1-log. These results demonstrate that generating open-loop lethality, via negative-feedback disruption, may yield a class of antimicrobials with a high genetic barrier to resistance.
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Antivirales , Regulación Viral de la Expresión Génica/efectos de los fármacos , Animales , Antivirales/farmacología , Farmacorresistencia Viral , Redes Reguladoras de Genes/efectos de los fármacos , Ratones , SARS-CoV-2/efectos de los fármacos , Replicación ViralRESUMEN
Small interfering RNA (siRNA) constitutes a promising therapeutic modality supporting the potential functional cure of hepatitis B. A novel ionizable lipidoid nanoparticle (RBP131) and a state-of-the-art lyophilization technology were developed in this study, enabling to deliver siRNA targeting apolipoprotein B (APOB) into the hepatocytes with an ED50 of 0.05 mg/kg after intravenous injection. In addition, according to the requirements of Investigational New Drug (IND) application, a potent siRNA targeting hepatitis B virus (HBV) was selected and encapsulated with RBP131 to fabricate a therapeutic formulation termed RB-HBV008. Efficacy investigations in transient and transgenic mouse models revealed that the expressions of viral RNAs and antigens (HBsAg and HBeAg), as well as viral DNA, were repressed, dose-dependently and time-dependently at multilog decreasing amplitude, in both circulation and liver tissue. In contrast, entecavir (ETV), the first-line clinically-employed nucleoside analog drug, barely recused the antigen expression, although it triggered as high as 3.50 log reduction of viral DNA, in line with clinical observations. Moreover, the toxicity profiles suggested satisfactory safety outcomes with ten times the therapeutic window. Therefore, this study provides an effective nucleic acid delivery system and a promising RNAi agent for the treatment of hepatitis B.
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Regulación Viral de la Expresión Génica/efectos de los fármacos , Antígenos de Superficie de la Hepatitis B , Antígenos e de la Hepatitis B , Virus de la Hepatitis B , Hepatitis B , ARN Interferente Pequeño , Células Hep G2 , Hepatitis B/tratamiento farmacológico , Hepatitis B/genética , Hepatitis B/metabolismo , Antígenos de Superficie de la Hepatitis B/biosíntesis , Antígenos de Superficie de la Hepatitis B/genética , Antígenos e de la Hepatitis B/biosíntesis , Antígenos e de la Hepatitis B/genética , Virus de la Hepatitis B/genética , Virus de la Hepatitis B/metabolismo , Humanos , ARN Interferente Pequeño/genética , ARN Interferente Pequeño/farmacologíaRESUMEN
Omicron is an emerging SARS-CoV-2 variant, evolved from the Indian delta variant B.1.617.2, which is currently infecting worldwide. The spike glycoprotein, an important molecule in the pathogenesis and transmissions of SARS-CoV-2 variants, especially omicron B.1.1.529, shows 37 mutations distributed over the trimeric protein domains. Notably, fifteen of these mutations reside in the receptor-binding domain of the spike glycoprotein, which may alter transmissibility and infectivity. Additionally, the omicron spike evades neutralization more efficiently than the delta spike. Most of the therapeutic antibodies are ineffective against the omicron variant, and double immunization with BioNTech-Pfizer (BNT162b2) might not adequately protect against severe disease induced by omicron B.1.1.529. So far, no efficient antiviral drugs are available against omicron. The present study identified the promising inhibitors from seaweed's bioactive compounds to inhibit the omicron variant B.1.1.529. We have also compared the seaweed's compounds with the standard drugs ceftriaxone and cefuroxime, which were suggested as beneficial antiviral drugs in COVID-19 treatment. Our molecular docking analysis revealed that caffeic acid hexoside (-6.4 kcal/mol; RMSD = 2.382 Å) and phloretin (-6.3 kcal/mol; RMSD = 0.061 Å) from Sargassum wightii (S. wightii) showed the inhibitory effect against the crucial residues ASN417, SER496, TYR501, and HIS505, which are supported for the inviolable omicron and angiotensin-converting enzyme II (ACE2) receptor interaction. Cholestan-3-ol, 2-methylene-, (3beta, 5 alpha) (CMBA) (-6.0 kcal/mol; RMSD = 3.074 Å) from Corallina officinalis (C. officinalis) manifested the strong inhibitory effect against the omicron RBD mutated residues LEU452 and ALA484, was magnificently observed as the essential residues in Indian delta variant B.1.617.2 previously. The standard drugs (ceftriaxone and cefuroxime) showed no or less inhibitory effect against RBD of omicron B.1.1.529. The present study also emphasized the pharmacological properties of the considered chemical compounds. The results could be used to develop potent seaweed-based antiviral drugs and/or dietary supplements to treat omicron B.1.1529-infected patients.
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Enzima Convertidora de Angiotensina 2/química , Regulación Viral de la Expresión Génica/efectos de los fármacos , Simulación del Acoplamiento Molecular , SARS-CoV-2/metabolismo , Algas Marinas/química , Glicoproteína de la Espiga del Coronavirus/química , Enzima Convertidora de Angiotensina 2/metabolismo , Antivirales/química , Antivirales/farmacología , Humanos , Mutación , Unión Proteica , Conformación Proteica , SARS-CoV-2/genética , Glicoproteína de la Espiga del Coronavirus/genética , Tratamiento Farmacológico de COVID-19RESUMEN
The spread of SARS-CoV-2 and ongoing COVID-19 pandemic underscores the need for new treatments. Here we report that cannabidiol (CBD) inhibits infection of SARS-CoV-2 in cells and mice. CBD and its metabolite 7-OH-CBD, but not THC or other congeneric cannabinoids tested, potently block SARS-CoV-2 replication in lung epithelial cells. CBD acts after viral entry, inhibiting viral gene expression and reversing many effects of SARS-CoV-2 on host gene transcription. CBD inhibits SARS-CoV-2 replication in part by up-regulating the host IRE1α RNase endoplasmic reticulum (ER) stress response and interferon signaling pathways. In matched groups of human patients from the National COVID Cohort Collaborative, CBD (100 mg/ml oral solution per medical records) had a significant negative association with positive SARS-CoV-2 tests. This study highlights CBD as a potential preventative agent for early-stage SARS-CoV-2 infection and merits future clinical trials. We caution against use of non-medical formulations including edibles, inhalants or topicals as a preventative or treatment therapy at the present time.
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Antivirales/farmacología , Cannabidiol/farmacología , Interacciones Huésped-Patógeno/efectos de los fármacos , Inmunidad Innata/efectos de los fármacos , SARS-CoV-2/efectos de los fármacos , Células A549 , Animales , Antivirales/química , COVID-19/virología , Cannabidiol/química , Cannabidiol/metabolismo , Chlorocebus aethiops , Estrés del Retículo Endoplásmico/efectos de los fármacos , Endorribonucleasas/genética , Endorribonucleasas/metabolismo , Células Epiteliales/virología , Femenino , Regulación Viral de la Expresión Génica/efectos de los fármacos , Interacciones Huésped-Patógeno/fisiología , Humanos , Interferones/metabolismo , Ratones , Proteínas Serina-Treonina Quinasas/genética , Proteínas Serina-Treonina Quinasas/metabolismo , SARS-CoV-2/fisiología , Células Vero , Internalización del Virus/efectos de los fármacos , Replicación Viral/efectos de los fármacos , Tratamiento Farmacológico de COVID-19RESUMEN
Non-coding RNAs, particularly lncRNAs and miRNAs, have recently been shown to regulate different steps in viral infections and induction of immune responses against viruses. Expressions of several host and viral lncRNAs have been found to be altered during viral infection. These lncRNAs can exert antiviral function via inhibition of viral infection or stimulation of antiviral immune response. Some other lncRNAs can promote viral replication or suppress antiviral responses. The current review summarizes the interaction between ncRNAs and herpes simplex virus, cytomegalovirus, and Epstein-Barr infections. The data presented in this review helps identify viral-related regulators and proposes novel strategies for the prevention and treatment of viral infection.
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Susceptibilidad a Enfermedades , Interacciones Huésped-Patógeno/genética , ARN no Traducido , Virosis/etiología , Replicación Viral , Animales , Antivirales/farmacología , Antivirales/uso terapéutico , Virus ADN/fisiología , Modelos Animales de Enfermedad , Susceptibilidad a Enfermedades/inmunología , Regulación de la Expresión Génica/efectos de los fármacos , Regulación Viral de la Expresión Génica/efectos de los fármacos , Interacciones Huésped-Patógeno/inmunología , Humanos , Técnicas de Diagnóstico Molecular , Unión Proteica , Especificidad de la Especie , Virosis/diagnóstico , Virosis/tratamiento farmacológico , Virosis/metabolismo , Replicación Viral/efectos de los fármacosRESUMEN
Raddeanin A (RA) has indicated suppressive effects on various human tumor cells, and insufficient vitamin D was associated with human papillomavirus (HPV) persistence and gynecological tumors. However, combined effects of RA and vitamin D on HPV-positive cells remain elusive. Herein, we aimed to investigate the combined effects of RA and 1É,25(OH)2D3 (VD3) on cellular viability and modulation of HPV18E6/E7, programmed cell death 1 ligand (PD-L1) and vitamin D receptor (VDR) expression in HeLa cells in vitro. HeLa cells were treated with RA alone or VD3 combined with RA. Cell viability was measured using 3-(4,5-dimethyl-2-thiazolyl)-2,5-diphenyl-2-H-tetrazolium bromide (MTT), and apoptosis was detected by flow cytometry. Real-time PCR (qRT-PCR) and Western blot were used to determine the gene/protein expression levels. The autophagosomes were observed by Transmission electron microscopy (TEM). The result showed that cell viability was inhibited by RA, and apoptosis in HeLa cells treated with RA was elevated accordingly. The expression of Bax, Cleaved-caspase-3, Cleaved-caspase-9 and Cleaved-PARP increased, and Bcl-2 decreased. The autophagy was induced by RA, as evidenced by elevated autophagosomes and the increased LC3-II/I ratio and Beclin-1. The expression of HPV18E6/E7, PD-L1 and VDR was reduced by RA. Moreover, RA combined with VD3 had a stronger effect on HeLa cells than RA alone. In conclusion, RA inhibits HeLa proliferation and induces apoptosis and autophagy via suppressing HPV18E6/E7, PD-L1 and VDR, and VD3 showed reinforced effects of RA on HeLa cells. Therefore, combined usage of VD3 with RA might be a potential novel immunotherapy strategy for HPV-related diseases.
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Antígeno B7-H1/metabolismo , Calcitriol/farmacología , Proteínas de Unión al ADN/metabolismo , Proteínas Oncogénicas Virales/metabolismo , Receptores de Calcitriol/metabolismo , Saponinas/farmacología , Autofagosomas/efectos de los fármacos , Autofagosomas/metabolismo , Autofagia , Proliferación Celular/efectos de los fármacos , Supervivencia Celular/efectos de los fármacos , Sinergismo Farmacológico , Regulación Neoplásica de la Expresión Génica/efectos de los fármacos , Regulación Viral de la Expresión Génica/efectos de los fármacos , Células HeLa , Humanos , Microscopía Electrónica de TransmisiónRESUMEN
Therapeutic interventions targeting viral infections remain a significant challenge for both the medical and scientific communities. While specific antiviral agents have shown success as therapeutics, viral resistance inevitably develops, making many of these approaches ineffective. This inescapable obstacle warrants alternative approaches, such as the targeting of host cellular factors. Respiratory syncytial virus (RSV), the major respiratory pathogen of infants and children worldwide, causes respiratory tract infection ranging from mild upper respiratory tract symptoms to severe life-threatening lower respiratory tract disease. Despite the fact that the molecular biology of the virus, which was originally discovered in 1956, is well described, there is no vaccine or effective antiviral treatment against RSV infection. Here, we demonstrate that targeting host factors, specifically, mTOR signaling, reduces RSV protein production and generation of infectious progeny virus. Further, we show that this approach can be generalizable as inhibition of mTOR kinases reduces coronavirus gene expression, mRNA transcription and protein production. Overall, defining virus replication-dependent host functions may be an effective means to combat viral infections, particularly in the absence of antiviral drugs.
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Coronavirus/metabolismo , Virus Sincitial Respiratorio Humano/metabolismo , Serina-Treonina Quinasas TOR/metabolismo , Proteínas Virales/metabolismo , Células A549 , Coronavirus/efectos de los fármacos , Coronavirus/genética , Regulación Viral de la Expresión Génica/efectos de los fármacos , Humanos , Biosíntesis de Proteínas/efectos de los fármacos , Inhibidores de Proteínas Quinasas/farmacología , Inhibidores de Proteínas Quinasas/uso terapéutico , Interferencia de ARN , ARN Interferente Pequeño/metabolismo , Proteína Asociada al mTOR Insensible a la Rapamicina/antagonistas & inhibidores , Proteína Asociada al mTOR Insensible a la Rapamicina/genética , Proteína Asociada al mTOR Insensible a la Rapamicina/metabolismo , Proteína Reguladora Asociada a mTOR/antagonistas & inhibidores , Proteína Reguladora Asociada a mTOR/genética , Proteína Reguladora Asociada a mTOR/metabolismo , Infecciones por Virus Sincitial Respiratorio/tratamiento farmacológico , Infecciones por Virus Sincitial Respiratorio/patología , Infecciones por Virus Sincitial Respiratorio/virología , Virus Sincitial Respiratorio Humano/efectos de los fármacos , Virus Sincitial Respiratorio Humano/aislamiento & purificación , Serina-Treonina Quinasas TOR/antagonistas & inhibidores , Serina-Treonina Quinasas TOR/genética , Proteínas Virales/genéticaRESUMEN
The growing resistance of the influenza virus to widely used competitive neuraminidase inhibitors occupying the active site of the enzyme requires the development of bifunctional compounds that can simultaneously interact with other regulatory sites on the protein surface. When developing such an inhibitor and combining structural fragments that could be located in the sialic acid cavity of the active site and the adjacent 430-cavity, it is necessary to select a suitable linker not only for connecting the fragments, but also to ensure effective interactions with the unique arginine triad Arg118-Arg292-Arg371 of neuraminidase. Using molecular modeling, we have demonstrated the usefulness of the sulfonamide group in the linker design and the potential advantage of this functional group over other isosteric analogues.
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Antivirales/farmacología , Inhibidores Enzimáticos/farmacología , Neuraminidasa/metabolismo , Orthomyxoviridae/enzimología , Sulfonamidas/química , Antivirales/síntesis química , Antivirales/química , Dominio Catalítico , Cristalografía por Rayos X , Inhibidores Enzimáticos/síntesis química , Inhibidores Enzimáticos/química , Regulación Viral de la Expresión Génica/efectos de los fármacos , Modelos Moleculares , Simulación del Acoplamiento Molecular , Neuraminidasa/antagonistas & inhibidores , Neuraminidasa/química , Orthomyxoviridae/efectos de los fármacos , Relación Estructura-Actividad , Proteínas Virales/antagonistas & inhibidores , Proteínas Virales/química , Proteínas Virales/metabolismoRESUMEN
Kaposi's sarcoma-associated herpesvirus (KSHV), an oncogenic virus, has two life cycle modes: the latent and lytic phases. KSHV lytic reactivation is important for both viral propagation and KSHV-induced tumorigenesis. The KSHV replication and transcription activator (RTA) protein is essential for lytic reactivation. Hesperetin, a citrus polyphenolic flavonoid, has antioxidant, anti-inflammatory, hypolipidemic, cardiovascular and anti-tumour effects. However, the effects of hesperetin on KSHV replication and KSHV-induced tumorigenesis have not yet been reported. Here, we report that hesperetin induces apoptotic cell death in BCBL-1 cells in a dose-dependent manner. Hesperetin inhibits KSHV reactivation and reduces the production of progeny virus from KSHV-harbouring cells. We also confirmed that HIF1α promotes the RTA transcriptional activities and lytic cycle-refractory state of KSHV-infected cells. Hesperetin suppresses HIF1α expression to inhibit KSHV lytic reactivation. These results suggest that hesperetin may represent a novel strategy for the treatment of KSHV infection and KSHV-associated lymphomas.
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Antivirales/farmacología , Infecciones por Herpesviridae/metabolismo , Herpesvirus Humano 8/efectos de los fármacos , Hesperidina/farmacología , Subunidad alfa del Factor 1 Inducible por Hipoxia/metabolismo , Activación Viral/efectos de los fármacos , Apoptosis/efectos de los fármacos , Regulación Viral de la Expresión Génica/efectos de los fármacos , Infecciones por Herpesviridae/genética , Infecciones por Herpesviridae/fisiopatología , Infecciones por Herpesviridae/virología , Herpesvirus Humano 8/genética , Herpesvirus Humano 8/fisiología , Humanos , Subunidad alfa del Factor 1 Inducible por Hipoxia/genética , Proteínas Virales/genética , Proteínas Virales/metabolismo , Replicación Viral/efectos de los fármacosRESUMEN
Application of differentiation therapy targeting cellular plasticity for the treatment of solid malignancies has been lagging. Nasopharyngeal carcinoma (NPC) is a distinctive cancer with poor differentiation and high prevalence of Epstein-Barr virus (EBV) infection. Here, we show that the expression of EBV latent protein LMP1 induces dedifferentiated and stem-like status with high plasticity through the transcriptional inhibition of CEBPA. Mechanistically, LMP1 upregulates STAT5A and recruits HDAC1/2 to the CEBPA locus to reduce its histone acetylation. HDAC inhibition restored CEBPA expression, reversing cellular dedifferentiation and stem-like status in mouse xenograft models. These findings provide a novel mechanistic epigenetic-based insight into virus-induced cellular plasticity and propose a promising concept of differentiation therapy in solid tumor by using HDAC inhibitors to target cellular plasticity.
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Proteínas Potenciadoras de Unión a CCAAT/genética , Inhibidores de Histona Desacetilasas/farmacología , Carcinoma Nasofaríngeo/tratamiento farmacológico , Factor de Transcripción STAT5/genética , Proteínas de la Matriz Viral/genética , Animales , Desdiferenciación Celular/efectos de los fármacos , Línea Celular Tumoral , Plasticidad de la Célula/efectos de los fármacos , Infecciones por Virus de Epstein-Barr/tratamiento farmacológico , Infecciones por Virus de Epstein-Barr/genética , Infecciones por Virus de Epstein-Barr/patología , Infecciones por Virus de Epstein-Barr/virología , Regulación Neoplásica de la Expresión Génica/efectos de los fármacos , Regulación Viral de la Expresión Génica/efectos de los fármacos , Herpesvirus Humano 4/efectos de los fármacos , Herpesvirus Humano 4/patogenicidad , Xenoinjertos , Humanos , Ratones , Carcinoma Nasofaríngeo/genética , Carcinoma Nasofaríngeo/patología , Carcinoma Nasofaríngeo/virologíaRESUMEN
Hepatitis C virus (HCV) is a positive-sense RNA virus that interacts with a liver-specific microRNA called miR-122. miR-122 binds to two sites in the 5' untranslated region of the viral genome and promotes HCV RNA accumulation. This interaction is important for viral RNA accumulation in cell culture, and miR-122 inhibitors have been shown to be effective at reducing viral titers in chronic HCV-infected patients. Herein, we analyzed resistance-associated variants that were isolated in cell culture or from patients who underwent miR-122 inhibitor-based therapy and discovered three distinct resistance mechanisms all based on changes to the structure of the viral RNA. Specifically, resistance-associated variants promoted riboswitch activity, genome stability, or positive-strand viral RNA synthesis, all in the absence of miR-122. Taken together, these findings provide insight into the mechanism(s) of miR-122-mediated viral RNA accumulation and provide mechanisms of antiviral resistance mediated by changes in RNA structure.