RESUMEN
Immunocompromised patients with coronavirus disease 2019 were prospectively enrolled from March to November 2022 to understand the association between antibody responses and severe acute respiratory syndrome coronavirus 2 shedding. A total of 62 patients were analyzed, and the results indicated a faster decline in genomic and subgenomic viral RNA in patients with higher neutralizing and S1-specific immunoglobulin G (IgG) antibodies (both P < .001). Notably, high neutralizing antibody levels were associated with a significantly faster decrease in viable virus cultures (P = .04). Our observations suggest the role of neutralizing antibodies in prolonged virus shedding in immunocompromised patients, highlighting the potential benefits of enhancing their humoral immune response through vaccination or monoclonal antibody treatments.
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Anticuerpos Neutralizantes , Anticuerpos Antivirales , COVID-19 , Huésped Inmunocomprometido , Inmunoglobulina G , SARS-CoV-2 , Esparcimiento de Virus , Humanos , COVID-19/inmunología , COVID-19/virología , SARS-CoV-2/inmunología , Anticuerpos Antivirales/sangre , Anticuerpos Antivirales/inmunología , Masculino , Estudios Prospectivos , Femenino , Persona de Mediana Edad , Anticuerpos Neutralizantes/sangre , Anticuerpos Neutralizantes/inmunología , Inmunoglobulina G/sangre , Inmunoglobulina G/inmunología , Anciano , ARN Viral , Adulto , Formación de Anticuerpos/inmunologíaRESUMEN
We assessed susceptibility of dogs to SARS-COV-2 Delta and Omicron variants by experimentally inoculating beagle dogs. Moreover, we investigated transmissibility of the variants from infected to naive dogs. The dogs were susceptible to infection without clinical signs and transmitted both strains to other dogs through direct contact.
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COVID-19 , Animales , Perros , COVID-19/veterinaria , SARS-CoV-2RESUMEN
There are limited data supporting current Centers for Disease Control and Prevention guidelines for the isolation period in moderate to severely immunocompromised patients with coronavirus disease 2019 (COVID-19). Adult COVID-19 patients who underwent solid organ transplantation (SOT) or received active chemotherapy against hematologic malignancy were enrolled and weekly respiratory samples were collected. Samples with positive genomic real-time polymerase chain reaction results underwent virus culture and rapid antigen testing (RAT). A total of 65 patients (40 with hematologic malignancy and 25 SOT) were enrolled. The median duration of viable virus shedding was 4 weeks (interquartile range: 3-7). Multivariable analysis revealed that B-cell depletion (hazard ratio [HR]: 4.76) was associated with prolonged viral shedding, and COVID-19 vaccination (≥3 doses) was negatively associated with prolonged viral shedding (HR: 0.22). The sensitivity, specificity, positive predictive value, and negative predictive value of RAT for viable virus shedding were 79%, 76%, 74%, and 81%, respectively. The negative predictive value of RAT was only 48% (95% confidence interval [CI]: 33-65) in the samples from those with symptom onset ≤20 days, but it was as high as 92% (95% CI: 85-96) in the samples from those with symptom onset >20 days. About half of immunocompromised COVID-19 patients shed viable virus for ≥4 weeks from the diagnosis, and virus shedding was prolonged especially in unvaccinated patients with B-cell-depleting therapy treatment. RAT beyond 20 days in immunocompromised patients had a relatively high negative predictive value for viable virus shedding.
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COVID-19 , Neoplasias Hematológicas , Adulto , Humanos , COVID-19/diagnóstico , SARS-CoV-2 , Estudios Prospectivos , Vacunas contra la COVID-19 , Neoplasias Hematológicas/complicaciones , Esparcimiento de Virus , ARN Viral/análisisRESUMEN
Human immunodeficiency virus-1 (HIV-1) transactivator (Tat)-mediated transcription is essential for HIV-1 replication. It is determined by the interaction between Tat and transactivation response (TAR) RNA, a highly conserved process representing a prominent therapeutic target against HIV-1 replication. However, owing to the limitations of current high-throughput screening (HTS) assays, no drug that disrupts the Tat-TAR RNA interaction has been uncovered yet. We designed a homogenous (mix-and-read) time-resolved fluorescence resonance energy transfer (TR-FRET) assay using europium cryptate as a fluorescence donor. It was optimized by evaluating different probing systems for Tat-derived peptides or TAR RNA. The specificity of the optimal assay was validated by mutants of the Tat-derived peptides and TAR RNA fragment, individually and by competitive inhibition with known TAR RNA-binding peptides. The assay generated a constant Tat-TAR RNA interaction signal, discriminating the compounds that disrupted the interaction. Combined with a functional assay, the TR-FRET assay identified two small molecules (460-G06 and 463-H08) capable of inhibiting Tat activity and HIV-1 infection from a large-scale compound library. The simplicity, ease of operation, and rapidity of our assay render it suitable for HTS to identify Tat-TAR RNA interaction inhibitors. The identified compounds may also act as potent molecular scaffolds for developing a new HIV-1 drug class.
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VIH-1 , Productos del Gen tat del Virus de la Inmunodeficiencia Humana , Humanos , Productos del Gen tat del Virus de la Inmunodeficiencia Humana/genética , Productos del Gen tat del Virus de la Inmunodeficiencia Humana/química , VIH-1/fisiología , Transferencia Resonante de Energía de Fluorescencia , Transactivadores , ARN Viral/genéticaRESUMEN
Increasing evidence suggests incomplete recovery of COVID-19 patients, who continue to suffer from cardiovascular diseases, including cerebral vascular disorders (CVD) and neurological symptoms. Recent findings indicate that some of the damaging effects of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection, especially in the brain, may be induced by the spike protein, leading to the disruption of the initial blood-brain barrier (BBB). SARS-CoV-2-infected cells and animals exhibit age-dependent pathogenesis. In this study, we identified endothelial BACE1 as a critical mediator of BBB disruption and cellular senescence induced by the SARS-CoV-2 spike S1 subunit protein. Increased BACE1 in human brain microvascular endothelial cells (HBMVEC) decreases the levels of tight junction proteins, including ZO-1, occludin, and claudins. Moreover, BACE1 overexpression leads to the accumulation of p16 and p21, typical hallmarks of cellular senescence. Our findings show that the SARS-CoV-2 spike S1 subunit protein upregulated BACE1 expression in HBMVECs, causing endothelial leakage. In addition, the SARS-CoV-2 spike S1 subunit protein induced p16 and p21 expression, indicating BACE1-mediated cellular senescence, confirmed by ß-Gal staining in HBMVECs. In conclusion, this study demonstrated that BACE1-mediated endothelial cell damage and senescence may be linked to CVD after COVID-19 infection.
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COVID-19 , Enfermedades Cardiovasculares , Secretasas de la Proteína Precursora del Amiloide/metabolismo , Animales , Ácido Aspártico Endopeptidasas/metabolismo , Encéfalo/metabolismo , Enfermedades Cardiovasculares/metabolismo , Células Endoteliales/metabolismo , Humanos , SARS-CoV-2 , Glicoproteína de la Espiga del Coronavirus/metabolismoRESUMEN
The heterocyclic indole structure has been shown to be one of the most promising scaffolds, offering various medicinal advantages from its wide range of biological activity. Nonetheless, the significance of 3-oxindole has been less known. In this study, a series of novel 3-oxindole-2-carboxylates were synthesized and their antiviral activity against human immunodeficiency virus-1 (HIV-1) infection was evaluated. Among these, methyl (E)-2-(3-chloroallyl)-4,6-dimethyl-one (6f) exhibited the most potent inhibitory effect on HIV-1 infection, with a half-maximal inhibitory concentration (IC50) of 0.4578 µM but without severe cytotoxicity (selectivity index (SI) = 111.37). The inhibitory effect of these compounds on HIV-1 infection was concordant with their inhibitory effect on the viral replication cycle. Mode-of-action studies have shown that these prominent derivatives specifically inhibited the Tat-mediated viral transcription on the HIV-1 LTR promoter instead of reverse transcription or integration. Overall, our findings indicate that 3-oxindole derivatives could be useful as a potent scaffold for the development of a new class of anti-HIV-1 agents.
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Fármacos Anti-VIH , Infecciones por VIH , VIH-1 , Fármacos Anti-VIH/química , Fármacos Anti-VIH/farmacología , Infecciones por VIH/tratamiento farmacológico , Humanos , Oxindoles/farmacología , Transcripción Viral , Replicación Viral , Productos del Gen tat del Virus de la Inmunodeficiencia Humana/metabolismoRESUMEN
BACKGROUND: Integrase (IN) is an essential protein for HIV replication that catalyzes insertion of the reverse-transcribed viral genome into the host chromosome during the early steps of viral infection. Highly active anti-retroviral therapy is a HIV/AIDS treatment method that combines three or more antiviral drugs often formulated from compounds that inhibit the activities of viral reverse transcriptase and protease enzymes. Early IN inhibitors (INIs) mainly serve as integrase strand transfer inhibitors (INSTI) that disrupt strand transfer by binding the catalytic core domain of IN. However, mutations of IN can confer resistance to INSTI. Therefore, non-catalytic integrase inhibitors (NCINI) have been developed as next-generation INIs. METHODS: In this study, we evaluated and compared the activity of INSTI and NCINI according to the analysis method. Antiviral activity was compared using p24 ELISA with MT2 cell and TZM-bl luciferase system with TZM-bl cell. Each drug was serially diluted and treated to MT2 and TZM-b1 cells, infected with HIV-1 AD8 strain and incubated for 5 and 2 days, respectively. Additionally, to analyze properties of INSTI and NCINI, transfer inhibition assay and 3'-processing inhibition assay were performed. RESULTS: During screening of INIs using the p24 ELISA and TZM-bl luciferase systems, we found an inconsistent result with INSTI and NCINI drugs. Following infection of MT2 and TZM-bl cells with T-tropic HIV-1 strain, both INSTI and NCINI treatments induced significant p24 reduction in MT2 cells. However, NCINI showed no antiviral activity in the TZM-bl luciferase system, indicating that this widely used and convenient antiretroviral assay is not suitable for screening of NCINI compounds that target the second round of HIV-1 replication. CONCLUSION: Accordingly, we recommend application of other assay procedures, such as p24 ELISA or reverse transcription activity, in lieu of the TZM-bl luciferase system for preliminary NCINI drug screening. Utilization of appropriate analytical methods based on underlying mechanisms is necessary for accurate assessment of drug efficacy.
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Fármacos Anti-VIH/farmacología , Inhibidores de Integrasa VIH/análisis , Inhibidores de Integrasa VIH/farmacología , Integrasa de VIH/metabolismo , VIH-1/efectos de los fármacos , Línea Celular , Infecciones por VIH/tratamiento farmacológico , Inhibidores de Integrasa VIH/clasificación , Secuenciación de Nucleótidos de Alto Rendimiento , Humanos , Replicación Viral/efectos de los fármacosRESUMEN
We isolated Japanese encephalitis virus genotype 5 from human specimens in South Korea. Whole-genome analysis showed 90.4% identity with other genotype 5 viruses from humans. This virus had a unique insertion in the NS4A gene. However, the envelope protein contained Lys 84, which was specific to strains of genotype 5 viruses from South Korea.
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Virus de la Encefalitis Japonesa (Especie) , Encefalitis Japonesa , Secuencia de Aminoácidos , Virus de la Encefalitis Japonesa (Especie)/genética , Encefalitis Japonesa/epidemiología , Genotipo , Humanos , Filogenia , República de CoreaRESUMEN
BACKGROUND AND AIM: Interferon-stimulated gene 20 (ISG20) is an interferon-inducible exonuclease that inhibits the replication of several RNA viruses. In patients with chronic hepatitis B, ISG20 expression is related to the interferon-α treatment response. However, the molecular mechanism of ISG20-mediated anti-hepatitis B virus (HBV) activity is unclear. METHODS: We have investigated the effect of ISG20 on antiviral activity to address that. The life cycle of HBV was analyzed by the ectopic expression of ISG20 in HepG2 and HepG2-NTCP cells. Finally, to provide physiological relevance of our study, the expression of ISG20 from chronic hepatitis B patients was examined. RESULTS: Interferon-stimulated gene 20 was mainly induced by interferon-ß and dramatically inhibited HBV replication. In addition, ISG20 decreased HBV gene expression and transcription. Although ISG20 inhibited HBV replication by reducing viral enhancer activity, the expression of transcription factors that bind the HBV enhancer was not affected. Particularly, ISG20 suppressed HBV enhancer activity by binding to the enhancer II and core promoter (EnhII/Cp) region. CONCLUSION: Our findings suggest that ISG20 exerts the anti-HBV activity by acting as a putative repressor binding to the HBV EnhII/Cp region.
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Exorribonucleasas/genética , Exorribonucleasas/metabolismo , Expresión Génica , Virus de la Hepatitis B/genética , Virus de la Hepatitis B/fisiología , Hepatitis B Crónica/tratamiento farmacológico , Hepatitis B Crónica/genética , Interferón-alfa/uso terapéutico , Interferón beta/uso terapéutico , Activación Viral/genética , Exorribonucleasas/fisiología , Células Hep G2 , Humanos , Interferón-alfa/farmacología , Interferón beta/farmacología , Regiones Promotoras Genéticas , Factores de Transcripción/genética , Factores de Transcripción/metabolismo , Replicación Viral/genéticaRESUMEN
A major obstacle in the treatment of human immunodeficiency virus type 1 (HIV-1) is its ability to establish latent infection. To find novel biomarkers associated with the mechanism of HIV-1 latent infection, we identified 70 candidate genes in HIV-1 latently infected cells through the integrated analysis in a previous study. It is important to select more effective biomarkers among 70 candidates and to verify the possibility of selected biomarkers for HIV-1 latency. We identified the 24 and 25 genes from 70 candidate genes in significantly enriched categories selected by Database for Annotation, Visualization and Integrated Discovery (DAVID) software and Gene Set Enrichment Analysis (GSEA) software, respectively. Also, we investigated genes regulated in both HIV-1 latently infected cell lines and PBMCs from HIV-1 infected patients and found the genes with a common pattern of expression levels in both cell lines and PBMCs. Consequently, we identified nine genes, APBB2, GMPR, IGF2BP3, LRP1, MAD2L2, MX1, OXR1, PTK2B, and TNFSF13B, via integrated analysis. Especially, APBB2 and MAD2L2 were identified in both DAVID and GSEA software. Our findings suggest that nine genes were identified via integrated analysis as potential biomarkers and in particular, APBB2 and MAD2L2 may be considered as more significant biomarkers for HIV-1 latency.
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Infecciones por VIH/metabolismo , VIH-1/fisiología , Latencia del Virus , Proteínas Adaptadoras Transductoras de Señales/genética , Proteínas Adaptadoras Transductoras de Señales/metabolismo , Biomarcadores/metabolismo , Línea Celular , Células Cultivadas , Infecciones por VIH/genética , Infecciones por VIH/patología , Infecciones por VIH/virología , Humanos , Proteínas Mad2/genética , Proteínas Mad2/metabolismoRESUMEN
BACKGROUND: A reservoir of HIV-1 is a major obstacle in eliminating HIV-1 in patients because it can reactivate in stopping antiretroviral therapy (ART). Histone modifications, such as acetylation and methylation, play a critical role in the organization of chromatin domains and the up- or downregulation of gene expression. Although many studies have reported that an epigenetic mechanism is strongly involved in the maintenance of HIV-1 transcriptional latency, neither the epigenetic control of viral replication nor how HIV-1 latency is maintained is not fully understood. RESULTS: We re-analyzed a high throughput parallel DNA sequencing (ChIP-seq) data from previous work to investigate the effect of histone modifications, H3K4me3 and H3K9ac, on HIV-1 latency in terms of chromosome distribution. The outputs of ChIP-seq from uninfected CD4+ T cell lines and HIV-1 latently infected cells were aligned to hg18 using bowtie and then analyzed using various software packages. Certain chromosomes (16, 17, 19, and 22) were significantly enriched for histone modifications in both decreased and increased islands. In the same chromosomes in HIV-1 latently infected cells, 38 decreased and 41 increased islands from common islands of H3K4me3 and H3K9ac were selected for functional annotation. In Gene Ontology analysis, the 38 genes associated with decreased islands were involved in the regulation of biological process, regulation of cellular process, biological regulation, and purinergic receptor signaling pathway, while the 41 genes associated with increased islands were involved in nucleic acid binding, calcium-activated cation channel activity, DNA binding, and zinc ion binding. In Pathway Commons analysis, the 38 genes were strongly involved in the p63 transcription factor network, while the 41 genes were involved in the RNA polymerase III transcription termination pathway. Several genes such as Nuclear factor I X (NFIX) and TNF receptor association factor 4 (TRAF4) were selected as candidate genes for HIV latency. Especially, NFIX was highly expressed in HIV-1 latently infected cell lines and showed a dramatic reduction in expression after phorbol-13-myristate-12-acetate (PMA) treatment. CONCLUSIONS: These results show that the unique enrichment of histone modifications and its linked genes in specific chromosomes might play a critical role in the establishment and maintenance of HIV-1 latency.
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VIH-1/genética , VIH-1/fisiología , Código de Histonas , Latencia del Virus , Línea Celular , Infecciones por VIH/virología , Secuenciación de Nucleótidos de Alto Rendimiento , Factores de Transcripción NFI/metabolismo , Análisis de Secuencia de ADN , Factor 4 Asociado a Receptor de TNF/metabolismoRESUMEN
BACKGROUND: Despite the successful inhibition of human immunodeficiency virus type 1 (HIV-1) replication by combination antiretroviral therapy, cells latently infected with HIV-1 remaining in patients are a major obstacle for eradication of HIV-1 infection. The tumor suppressor factor p53 is activated by HIV-1 infection, and restricts HIV-1 replication. However, a therapeutic strategy based on p53 activity has not been considered for elimination of latently infected cells. METHODS: Apoptotic cells were analyzed using flow cytometry with anti-annexin A5-FITC Ab and PI staining upon treatment of anticancer drugs. The expression and activation of p53 and apoptotic molecules in latently HIV-1-infected T cells were compared using Western blot analysis. The role of p53 in the anticancer drug treatment-induced apoptosis of cells latently infected with HIV-1 was determined by knock-down experiment using siRNA against p53. RESULTS: Upon treatment with 5-fluorouracil (5-FU), apoptosis was increased in latently infected ACH2 cells encoding competent p53 compared with uninfected parent A3.01 cells, while the apoptosis of latently infected p53 null J1.1 cells was less than that of uninfected cells. Treatment with 5-FU increased the levels of cleaved caspase-3 and PARP in ACH2 cells compared with uninfected and latently infected p53 null J1.1 cells. The levels of expression and activation of p53 were higher in both latently infected ACH2 and NCHA2 cells than in uninfected cells. Furthermore, the activation levels of p53 in both cells were further increased upon 5-FU treatment. Consistent with p53 status, apoptosis was markedly increased in ACH2 and NCHA2 cells compared with uninfected and latently infected J1.1 cells upon treatment with other anticancer drugs such as doxorubicin and etoposide. Inhibition of p53 in cells with latent HIV-1 infection diminished apoptosis upon 5-FU treatment. CONCLUSION: Evidence described here indicate that when treated with anticancer drugs, apoptosis of cells with latent HIV-1 infection was increased via the p53 activation pathway and may provide information for application of anticancer drugs to selectively eliminate HIV-1 reservoirs.
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Antineoplásicos/metabolismo , Apoptosis , VIH-1/fisiología , Linfocitos T/efectos de los fármacos , Linfocitos T/virología , Proteína p53 Supresora de Tumor/metabolismo , Latencia del Virus/efectos de los fármacos , Citometría de Flujo , HumanosRESUMEN
OBJECTIVES: Persistent HIV-1 infections are characterized by a long silent infection period in resting CD4+ T cells, which allows them to escape the host immune response. Several HIV-1 latency mechanisms have been reported, but the molecular mechanism underlying polycomb repressor complex (PRC)-mediated HIV-1 latency remains poorly understood. METHODS: Expression of PRC proteins in latent cells was measured by Western blot. Knockdowns of PRC genes were conducted by the specific siRNA and methylations at H3K27 on the proviral LTR were investigated by ChIP assay. RESULTS: PRC proteins (EED, BMI-1, and RNF2) were dramatically downregulated in latent cells after PMA treatment. The downregulation of PRC proteins was followed by a decrease in the methylation of H3K27 and ubiquitination of H2AK119 in the PMA-treated latent cells. siRNA knockdowns of EED and BMI-1 also enhanced HIV-1 reactivation significantly in latently infected cells. By contrast, proteasomal inhibitor MG132 successfully abrogated the PMA-induced downregulation of PRCs. In particular, di-/tri-methylations of histone-3 in the proviral LTR was absent from latent cells after PMA treatment. CONCLUSIONS: This study shows that PRC is strongly related to the control of HIV-1 latency and that PRC-breaking agents may be helpful for purging HIV-1 from latent reservoirs.
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Silenciador del Gen , VIH-1/fisiología , Proteínas del Grupo Polycomb/metabolismo , Provirus/fisiología , Activación Viral , Latencia del Virus , Western Blotting , Inmunoprecipitación de Cromatina , Perfilación de la Expresión Génica , Técnicas de Silenciamiento del Gen , VIH-1/genética , Humanos , Proteínas del Grupo Polycomb/genética , Provirus/genéticaRESUMEN
Mouse models expressing human ACE2 for coronavirus disease 2019 have been frequently used to understand its pathogenesis and develop therapeutic strategies against SARS-CoV-2. Given that human TMPRSS2 supports viral entry, replication, and pathogenesis, we established a double-transgenic mouse model expressing both human ACE2 and TMPRSS2 for SARS-CoV-2 infection. Co-overexpression of both genes increased viral infectivity in vitro and in vivo. Double-transgenic mice showed significant body weight loss, clinical disease symptoms, acute lung injury, lung inflammation, and lethality in response to viral infection, indicating that they were highly susceptible to SARS-CoV-2. Pretreatment with the TMPRSS2 inhibitor, nafamostat, effectively reduced virus-induced weight loss, viral replication, and mortality in the double-transgenic mice. Moreover, the susceptibility and differential pathogenesis of SARS-CoV-2 variants were demonstrated in this animal model. Together, our results demonstrate that double-transgenic mice could provide a highly susceptible mouse model for viral infection to understand SARS-CoV-2 pathogenesis and evaluate antiviral therapeutics against coronavirus disease 2019.
Asunto(s)
Enzima Convertidora de Angiotensina 2 , COVID-19 , Modelos Animales de Enfermedad , Ratones Transgénicos , SARS-CoV-2 , Serina Endopeptidasas , Animales , Enzima Convertidora de Angiotensina 2/genética , Enzima Convertidora de Angiotensina 2/metabolismo , Serina Endopeptidasas/genética , Serina Endopeptidasas/metabolismo , COVID-19/virología , COVID-19/genética , COVID-19/metabolismo , SARS-CoV-2/fisiología , SARS-CoV-2/genética , Humanos , Ratones , Replicación Viral , Benzamidinas , Guanidinas/farmacología , Chlorocebus aethiops , Tratamiento Farmacológico de COVID-19RESUMEN
OBJECTIVES: Recent studies have reported that human immunodeficiency virus type 1 (HIV-1) proviruses are strongly suppressed in the unique epigenetic environments caused by chromatin modifications such as acetylation and methylation. Therefore, optimized therapeutic strategies directed against the virus reservoir using these epigenetic modifying agents (EMAs) should cure HIV infection. METHODS: Cytotoxicity and HIV-1 reactivation were determined using the PrestoBlue™ Cell Viability Reagent and p24 HIV ELISA, respectively. RESULTS: EMAs, including histone deacetylase inhibitors (VPA and SAHA), DNA methyltransferase inhibitor (5'-Aza-CdR), histone methyltransferase inhibitor (ADOX) and 12-O-tetradecanoylphorbol-13-acetate (TPA), were used to reactivate proviruses in HIV-1 latently infected cells. The effect of monotreatment with these EMAs on HIV-1 reactivation was VPA or SAHA > 5'-Aza-CdR > ADOX. Even though cotreatment with these potential HIV-1 reactivating agents did not show any significant reactivation effects in HIV-1 latently infected cells, employing SAHA under TPA treatment demonstrated a dramatic synergistic effect on purging HIV-1 proviruses in all HIV-1 latently infected cells via the ERK and AP-1 pathways. CONCLUSIONS: These results suggest that the combined approaches of EMAs, cotreatment of SAHA and TPA, could provide an effective way to lead a decline of HIV-1 reservoirs in patients.
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Sinergismo Farmacológico , VIH-1/efectos de los fármacos , Inhibidores de Histona Desacetilasas/metabolismo , Acetato de Tetradecanoilforbol/análogos & derivados , Activación Viral/efectos de los fármacos , Latencia del Virus/efectos de los fármacos , Línea Celular , Supervivencia Celular/efectos de los fármacos , Ensayo de Inmunoadsorción Enzimática , Proteína p24 del Núcleo del VIH/análisis , VIH-1/fisiología , Humanos , Acetato de Tetradecanoilforbol/metabolismoRESUMEN
Ongoing emergence of SARS-CoV-2 Omicron subvariants and their rapid worldwide spread pose a threat to public health. From November 2022 to February 2023, newly emerged Omicron subvariants, including BQ.1.1, BF.7, BA.5.2, XBB.1, XBB.1.5, and BN.1.9, became prevalent global strains (>5% global prevalence). These Omicron subvariants are resistant to several therapeutic antibodies. Thus, the antiviral activity of current drugs such as remdesivir, molnupiravir, and nirmatrelvir, which target highly conserved regions of SARS-CoV-2, against newly emerged Omicron subvariants need to be evaluated. We assessed the antiviral efficacy of the drugs using the half-maximal inhibitory concentration (IC50) against human isolates of 23 Omicron subvariants and four former SARS-CoV-2 variants of concern (VOCs) and compared it with the antiviral efficacy of these drugs against the SARS-CoV-2 reference strain (hCoV/Korea/KCDC03/2020). Maximal IC50-fold changes of remdesivir, molnupiravir, and nirmatrelvir were 1.9 (BA.2.75.2), 1.2 (B.1.627.2), and 1.4 (BA.2.3), respectively, compared to median IC50 values of the reference strain. Moreover, median IC50-fold changes of remdesivir, molnupiravir, and nirmatrelvir against the Omicron variants were 0.96, 0.4, and 0.62, respectively, similar to the 1.02, 0.88, and 0.67, respectively, median IC50-fold changes for previous VOCs. Although K90R and P132H in Nsp 5, and P323L, A529V, G671S, V405F, and ins823D in Nsp 12 mutations were identified, these amino acid substitutions did not affect drug antiviral activity. These results indicate that current antivirals retain antiviral efficacy against newly emerged Omicron subvariants. It is important to continue active surveillance and testing of new variants for drug resistance to enable early identification of drug-resistant strains.
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COVID-19 , Humanos , SARS-CoV-2 , Antivirales/farmacología , Lactamas , Leucina , NitrilosRESUMEN
The coronavirus disease 19 (COVID-19) pandemic, caused by the severe acute respiratory syndrome, coronavirus 2 (SARS-CoV-2), has resulted in unprecedented challenges to healthcare worldwide. In particular, the anthroponotic transmission of human coronaviruses has become a common concern among pet owners. Here, we experimentally inoculated beagle dogs with SARS-CoV-2 or Middle East respiratory syndrome (MERS-CoV) to compare their susceptibility to and the pathogenicity of these viruses. The dogs in this study exhibited weight loss and increased body temperatures and shed the viruses in their nasal secretions, feces, and urine. Pathologic changes were observed in the lungs of the dogs inoculated with SARS-CoV-2 or MERS-CoV. Additionally, clinical characteristics of SARS-CoV-2, such as increased lactate dehydrogenase levels, were identified in the current study.
RESUMEN
SARS-CoV-2 variants have continuously emerged globally, including in South Korea. To characterize the molecular evolution of SARS-CoV-2 in South Korea, we performed phylogenetic and genomic recombination analyses using more than 12,000 complete genome sequences collected until October 2022. The variants in South Korea originated from globally identified variants of concern and harbored genetic clade-common and clade-specific amino acid mutations mainly around the N-terminal domain (NTD) or receptor binding domain (RBD) in the spike protein. Several point mutation residues in key antigenic sites were under positive selection persistently with changing genetic clades of SARS-CoV-2. Furthermore, we detected 17 potential genomic recombinants and 76.4% (13/17) retained the mosaic NTD or RBD genome. Our results suggest that point mutations and genomic recombination in the spike contributed to the molecular evolution of SARS-CoV-2 in South Korea, which will form an integral part of global prevention and control measures against SARS-CoV-2.
RESUMEN
To identify potent antiviral compounds, we introduced a high-throughput screen platform that can rapidly classify hit compounds according to their target. In our platform, we performed a compound screen using a lentivirus-based pseudovirus presenting a spike protein of coronavirus, and we evaluated the hit compounds using an amplified luminescence proximity homogeneous assay (alpha) test with purified host receptor protein and the receptor binding domain of the viral spike. With our screen platform, we were able to identify both spike-specific compounds (class I) and broad-spectrum antiviral compounds (class II). Among the hit compounds, thiosemicarbazide was identified to be selective to the interaction between the viral spike and its host cell receptor, and we further optimized the binding potency of thiosemicarbazide through modification of the pyridine group. Among the class II compounds, we found raloxifene and amiodarone to be highly potent against human coronaviruses including Middle East respiratory syndrome coronavirus (MERS-CoV), severe acute respiratory syndrome coronavirus (SARS-CoV), and SARS-CoV-2. In particular, using analogs of the benzothiophene moiety, which is also present in raloxifene, we have identified benzothiophene as a novel structural scaffold for broad-spectrum antivirals. This work highlights the strong utility of our screen platform using a pseudovirus assay and an alpha test for rapid identification of potential antiviral compounds and their mechanism of action, which can lead to the accelerated development of therapeutics against newly emerging viral infections.
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COVID-19 , Coronavirus del Síndrome Respiratorio de Oriente Medio , Humanos , Luminiscencia , Clorhidrato de Raloxifeno , SARS-CoV-2/metabolismo , Antivirales/farmacología , Antivirales/química , Glicoproteína de la Espiga del Coronavirus/metabolismoRESUMEN
The Middle East respiratory syndrome (MERS) is a fatal acute viral respiratory disease caused by MERS-coronavirus (MERS-CoV) infection. To date, no vaccine has been approved for MERS-CoV despite continuing outbreaks. Inactivated vaccines are a viable option when developed using the appropriate inactivation methods and adjuvants. In this study, we evaluated the immunogenicity and protective effects of MERS-CoV vaccine candidates inactivated by three different chemical agents. MERS-CoV was effectively inactivated by formaldehyde, hydrogen peroxide, and binary ethylene imine and induced humoral and cellular immunity in mice. Although inflammatory cell infiltration was observed in the lungs four days after the challenge, the immunized hDPP4-transgenic mouse group showed 100% protection against a challenge with MERS-CoV (100 LD50). In particular, the immune response was highly stimulated by MERS-CoV inactivated with formaldehyde, and all mice survived a challenge with the minimum dose. In the adjuvant comparison test, the group immunized with inactivated MERS-CoV and AddaVax had a higher immune response than the group immunized with aluminum potassium sulfate (alum). In conclusion, our study indicates that the three methods of MERS-CoV inactivation are highly immunogenic and protective in mice and show strong potential as vaccine candidates when used with an appropriate adjuvant.