Combination of Azidothymidine (AZT) and (E)-5-(2-Bromovinyl)-2′-deoxyuridine (BVDU) Inhibits the Replication of Herpes Simplex Virus Type 1 (HSV-1) and Type 2 (HSV-2) and Varicella Zoster Virus (VZV) Strains That Are Deficient in the Expression of the Viral Thymidine Kinase (tk)

Abstract

Combinations of high concentrations of AZT with BVDU, acyclovir (ACV) or ganciclovir (GCV) show antagonism against TK⁺ HSV-1, but not TK⁺ VZV strains, in cell cultures. When BVDU and AZT were used in combination against TK^? HSV-1, TK^? HSV-2 and TK^? VZV strains, a pronounced inhibition of viral replication was observed. This potentiating effect was not seen if AZT was combined with ACV or GCV.     

3′非翻译区靶向人工微小RNA对PRRSV在猪肺巨噬细胞中复制的抑制作用（英文）北大核心CSCD

【目的】研究重组腺病毒(rAd)传送的3′非翻译区(UTR)靶向amiR3UTR对猪繁殖与呼吸综合征病毒(PRRSV)在猪肺巨噬细胞(PAM)中复制的抑制作用。【方法】用表达amiR3UTR或对照amiRcon的腺病毒载体转染AAV-293细胞,获得rAd-amiR3UTR-GFP和rAd-amiRcon-GFP,用定量RT-PCR检测amiR3UTR在rAd转导细胞中的表达,用定量RT-PCR、Western blotting和病毒滴定检测amiR3UTR对PRRSV复制的抑制作用。【结果】原代PAM及其细胞系3D4/163均能被rAd-amiR3UTR-GFP转导,但前者转导效率很低;rAd-amiR3UTR-GFP转导细胞能有效表达amiR3UTR,且表达具有剂量和时间依赖性;rAd表达的amiR3UTR能显著抑制不同毒株PRRSV在PAM细胞中的复制,且抑制作用具有剂量依赖性。【结论】amiR3UTR能抑制不同毒株PRRSV在PAM中的复制,其rAd有望作为抗PRRSV新策略进行深入研究。     

Broad antiviral and anti-inflammatory activity of Qingwenjiere mixture against SARS-CoV-2 and other human coronavirus infections

BackgroundQingwenjiere Mixture (QJM) is a traditional Chinese medicine (TCM) that has been shown to have remarkable clinical efficacy against COVID-19. However, little is known about the antiviral and anti-inflammatory activities of QJM against a wider range of human coronavirus (HCoV) strains.PurposeThe study aims to investigate the antiviral and anti-inflammatory activities of QJM, as well as the underlying mechanisms against HCoV infections.MethodsThe chemical compositions from QJM were analyzed by LC-MS. The inhibitory effect of QJM on infections of HCoV-OC43, HCoV-229E, HCoV-NL63, and SARS-CoV-2 was evaluated in HRT-18 cells, Huh7 cells, LLC-MK2 cells, and Vero-E6 cells, respectively, by using cytopathic effect (CPE) inhibition assay or RT-qPCR detection of viral n, s, or RdRp/Hel genes. The expression of pro-inflammatory cytokines induced by HCoV-OC43, HCoV-229E, and SARS-CoV-2, as well as the host ace2 gene was also determined by RT-qPCR assay. Furthermore, the expression of key molecules in the NF-κB/MAPKs signaling pathways was determined by western blot.ResultsIn alcohol-extraction groups of QJM and reference decoction pieces, 53 similar ion peaks were identified, the majority of which were phenylpropanoids, iridoids, and flavonoids. In addition, QJM reduced CPE caused by HCoVs and the expression of viral n genes or N protein. Pretreatment with QJM also exerted inhibitory effect on viral n gene expression. QJM also inhibited the expression of RdRp/Hel and s genes of SARS-CoV-2, as well as the host ace2 gene. Besides, QJM markedly reduced virus-induced mRNA expression of a panel of pro-inflammatory cytokines, such as IL-6, CXCL-8/IL-8, CXCL-10/IP-10, CCL-5/RANTES, TNF-α, IFN-α, CCL-2/MCP-1, CXCL-9/MIG, and IL1-α. We further showed that QJM inhibited the phosphorylation of NF-κB p65, and JNK, ERK 1/2, and p38 MAPKs in HCoV-OC43-infected HRT-18 cells.ConclusionsQJM has broad antiviral and anti-inflammatory activity against both common and newly emerged HCoVs possibly by inhibiting the activation of key components in NF-κB/MAPKs signaling pathway. QJM also has a prevention effect against HCoV infections and inhibits the host receptor required for virus entry. These results indicate that QJM may have the therapeutic potential in the treatment of diseases caused by a broad range of HCoVs.     

6-Azathymidine-4′-thionucleosides: synthesis and antiviral evaluation

The synthesis of dideoxy-6-azathymidine 4′-thionucleoside 1-(2,3-dideoxy-4-thio-β-D-erythro-pentofuranosyl)-(6-azathymidine) (2), and the L-nucleoside, 1-(4-thio-β-L-erythro-pentofuranosyl)-(6-azathymidine) (3) and their evaluation against a wide panel of antiviral assays are described. The L-thionucleoside (3) was devoid of antiviral activity. The dideoxy-thionucleoside (2) was moderately active against vaccinia virus (VV) and the herpes simplex virus strains HSV-1 (strain KOS) and HSV-2 (strain G) (MIC 12 μM) and retained inhibitory activity vs a thymidine kinase-deficient strain HSV-1/TK^–, suggesting that (2) is not dependent on viral TK-catalysed phosphorylation for antiviral activity and/or may use an alternative metabolic activation pathway.     

Design and synthesis of pyridone inhibitors of non-nucleoside reverse transcriptase

Next generation NNRTIs are sought which possess both broad spectrum antiviral activity against key mutant strains and a high genetic barrier to the selection of new mutant viral strains. Pyridones were evaluated as an acyclic conformational constraint to replace the aryl ether core of MK-4965 (1) and the more rigid indazole constraint of MK-6186 (2). The resulting pyridone compounds are potent inhibitors of HIV RT and have antiviral activity in cell culture that is superior to other next generation NNRTI’s.     

SYNTHESIS AND IN VITRO ACTIVITY OF D- AND L-ENANTIOMERS OF 5-(TRIFLUOROMETHYL)URACIL NUCLEOSIDE DERIVATIVES

Recently, β-L-nucleoside analogues have emerged as a new class of sugar modified nucleosides with potential antiviral and/or antitumoral activity. As a part of our ongoing research on this topic, we decided to synthesize 5-CF₃-β-L-dUrd (7), the hitherto unknown L-enantiomer of Trifluridine, an antiherpetic drug approved by FDA but only used in topical applications due to concomitant cytotoxicity. 5-CF₃-β-L-dUrd (7) as well as some other related L-nucleoside derivatives were stereospecifically prepared and tested in vitro against viral (HSV-1 and HSV-2) and human thymidine kinases (TK).     

Durable Suppression of HIV-1 after Virologic Monitoring-Based Antiretroviral Adherence Counseling in Rakai,Uganda

ObjectivesHIV viral load is recommended for monitoring antiretroviral treatment and identifying treatment failure. We assessed the durability of viral suppression after viral load-triggered adherence counseling among patients with HIV viremia 6 months after ART initiation.DesignObservational cohort enrolled in an antiretroviral treatment program in rural Uganda.MethodsParticipants who underwent routine viral load determination every 24 weeks and had at least 48 weeks of follow-up were included in this analysis. Patients with viral loads >400 copies/ml at 24 weeks of treatment were given additional adherence counseling, and all patients were followed to assess the duration of viral suppression and development of virologic failure.Results1,841 participants initiating antiretroviral therapy were enrolled in the Rakai Health Sciences Program between June 2005 and June 2011 and were followed with viral load monitoring every 24 weeks. 148 (8%) of patients did not achieve viral suppression at 24 weeks and were given additional adherence counseling. 85 (60%) of these patients had undetectable viral loads at 48 weeks, with a median duration of viral suppression of 240 weeks (IQR 193-288 weeks). Failure to achieve an undetectable viral load at 48 weeks was associated with age <30 years and 24 week viral load >2,000 copies/ml in multivariate logistic regression analysis.ConclusionsThe majority of patients with persistent viremia who were provided adherence counseling achieved robust viral suppression for a median 4.6 years. Access to virologic monitoring and adherence counseling is a priority in resource-limited settings.     

Design,synthesis, antimicrobial activity and anti-HIV activity evaluation of novel hybrid quinazoline–triazine derivatives

Abstract

A series of novel hybrid quinazoline–triazine derivatives was designed and synthesized from cyanuric chloride and anthranilic acid through sequential reactions, which contain different pharmacophores like quinazoline and substituted diaryl triazine (DATA) linked with ethylene diamine. All the newly synthesized compounds were characterized by infrared, ¹H-NMR, ¹³C-NMR, MS and elemental analysis. Further, we evaluated the in vitro anti-HIV activity of the newly synthesized compounds against HIV-1 (III_B) and HIV-2 (ROD) viral strains and as well as in vitro antimicrobial activity against four bacteria (Staphylococcus aureus, Bacillus cereus, Pseudomonas aeruginosa, Klebsiella pneumoniae) and two fungi (Aspergillus clavatus, Candida albicans) using the paper agar streak dilution method. The bioassay results indicate that four compounds namely 7d, 7n, 7r and 7s could be considered as possible potential agents.     

3-Trifluoromethylpyrazolones derived nucleosides: Synthesis and antiviral evaluation

Dengue (DENV) viral infection is a global public health problem that infrequently develops life threatening diseases such as dengue hemorrhagic fever (DFS) and dengue shock syndrome (DSS). Middle East respiratory syndrome coronavirus (MERS-CoV) is a highly pathogenic human corona virus with 38% fatality rate of infected patients. A series of 4-arylhydrazono-5-trifluoromethyl-pyrazolones, their ribofuranosyl, and 5′-deoxyribofuranosyl nucleosides were synthesized, geometry optimized using Density functional theory (DFT), and evaluated for their antiviral activity. 2-Nitrophenylhydrazonopyra-zolone derivative 5 showed significant activity against MERS-CoV (EC₅₀ = 4.6?μM). The nucleoside analog 8 showed moderate activity against DENV-2 (EC₅₀ = 10?μM), while the activity was abolished with the corresponding 5′-deoxyribonucleoside analogs. The identified hits in this study set this category of compounds for further future optimizations.     

Biological and antipathogenic activities of ribosome-inactivating proteins from Phytolacca dioica L.

BackgroundThe species from the genus Phytolacca constitute one of the best sources of ribosome-inactivating proteins (RIPs) that have been used both in the therapy against virus and tumors and in the construction of transgenic plants resistant to virus, bacteria, fungi and insects. Here we investigate new activities of three representative RIPs from Phytolacca dioica (dioicin 2, PD-S2 and PD-L4).ResultsThe three RIPs displayed, in addition to already reported activities, rRNA N-glycosylase activities against plant, bacterial and fungal ribosomes. Additionally dioicin 2 and PD-L4 displayed endonuclease activity on a supercoiled plasmid DNA, and dioicin 2 and PD-S2 arrested the growth of the fungus Penicillium digitatum. Furthermore, dioicin 2 induced caspase activation and apoptosis in cell cultures.ConclusionsThe different activities of the RIPs from Phytolacca dioica may explain the antipathogenic properties attributed to these RIPs in plants and their antiviral and antitumoral effects. In spite of the similarity in their rRNA N-glycosylase and DNA polynucleotide:adenosine glycosylase activities, they differed in their activities against viral RNA, plasmid DNA, fungi and animal cultured cells. This suggests that the presence of isoforms might optimize the response of the plant against several types of pathogens.General significanceRIPs from Phytolacca can induce plant resistance or tumor cell death not only by means of ribosome inactivation but also by the activities found in this report. Furthermore, the induction of cell death by different mechanisms turns these RIPs into more useful tools for cancer treatment rendering the selection of RIP-resistant mutants impossible.     

Direct inhibitory effect on viral entry of influenza A and SARS‐CoV‐2 viruses by azithromycin

ObjectivesUsing strategy of drug repurposing, antiviral agents against influenza A virus (IAV) and newly emerging SARS‐coronavirus 2 (SARS‐CoV‐2, also as 2019‐nCoV) could be quickly screened out.Materials and MethodsA previously reported engineered replication‐competent PR8 strain carrying luciferase reporter gene (IAV‐luc) and multiple pseudotyped IAV and SARS‐CoV‐2 virus was used. To specifically evaluate the pH change of vesicles containing IAV, we constructed an A549 cell line with endosomal and lysosomal expression of pHluorin2.ResultsHere, we identified azithromycin (AZ) as an effective inhibitor against multiple IAV and SARS‐CoV‐2 strains. We found that AZ treatment could potently inhibit IAV infection in vitro. Moreover, using pseudotyped virus model, AZ could also markedly block the entry of SARS‐CoV‐2 in HEK293T‐ACE2 and Caco2 cells. Mechanistic studies further revealed that such effect was independent of interferon signalling. AZ treatment neither impaired the binding and internalization of IAV virions, nor the viral replication, but rather inhibited the fusion between viral and vacuolar membranes. Using a NPC1‐pHluorin2 reporter cell line, we confirmed that AZ treatment could alkalize the vesicles containing IAV virions, thereby preventing pH‐dependent membrane fusion.ConclusionsOverall, our findings demonstrate that AZ can exert broad‐spectrum antiviral effects against IAV and SARS‐CoV‐2, and could be served as a potential clinical anti‐SARS‐CoV‐2 drug in emergency as well as a promising lead compound for the development of next‐generation anti‐IAV drugs.     

动态脑电图与常规脑电图应用于病毒性脑炎诊断的效果分析

目的:探讨动态脑电图与常规脑电图应用于病毒性脑炎的应用价值。方法:选取150例病毒性脑炎患者,随机分为两组,每组各75例,常规脑电图(REEG)组采用常规脑电图检查,动态脑电图(AEEG)组采用动态脑电图检查;观察并记录脑电图异常率,不同程度病情脑电图异常率的例数,评价动态脑电图与常规脑电图对病毒性脑炎的检测灵敏度和准确度。结果:AEEG组检出的脑电图异常率明显高于REEG组(P0.05)。不同程度病情脑电图检出的患者比例,两组相比,差异没有统计学意义(F=-0.085,P0.05)。REEG组中,轻度与中度病毒性脑炎检出率相比,差异没有统计学意义(P0.05),中度与重度病毒性脑炎检出率相比,差异没有统计学意义(P0.05),重度病毒性脑炎检出率明显高于轻度(P0.05)。AEEG组中,轻度与中度病毒性脑炎检出率相比,差异没有统计学意义(P0.05),重度病毒性脑炎检出率明显高于中度和轻度(P0.05),AEEG组重度病毒性脑炎检出率明显高于REEG组(P0.05)。结论:动态脑电图作为一种无创性检查,对于病毒性脑炎具有极好的检出率,灵敏度高,适用于病毒性脑膜炎的早期辅助诊断。     

Characterization of GPI2A,a Potent Inhibitor of HIV-1 Gene Expression and Viral Replication

Abstract

Fully thioated antisense molecules are often cytotoxic and non-specitic in action. GPI2A is thioated at 7 base positions. GPI2A posses sequence-specific activity against HIV-1 gene expression and viral replication without sigdcant cytotoxicity. Partial thioation did not compromise its uptake, cellular distribution and nuclease resistance.     

5′-Nor Carbocyclic Ribavirin

Abstract

An efficient synthesis of 5′-nor carbocyclic ribavirin (4) is described in 13 steps from conveniently available (+)-(1R,4S)-4-hydroxy-2-cyclopenten-1-yl acetate (6). Compound 4 was evaluated against the following viruses: herpes simplex type 1 and 2, vaccinia, cowpox, smallpox, Ebola, hepatitis B, hepatitis C, adenovirus type 1, influenza A (H1N1 and H3N2), influenza B, parainfluenza type 3, Pichinde, Punta Toro A, respiratory syncytial, rhinovirus type 2, Venezuelan equine encephalitis, yellow fever, and West Nile. No activity was found nor was there any cytotoxicity to the viral host cells.     

Alteration of thiol-disulphide homeostasis in acute tonsillopharyngitis

Objective: Thiol-disulphide homeostasis (TDH) has a critical role in various clinical disorders. We aimed to assess the association of TDH with acute tonsillopharyngitis (AT) in children.

Methods: This study included 94 (73 viral and 21 bacterial) tonsillopharyngitis patients and 88 control children. Their native thiol, total thiol, and disulphide levels were measured.

Results: Viral and bacterial tonsillopharyngitis patients had lower native thiol levels compared with healthy children (P?P?=?0.008, respectively). Both groups had lower total thiol levels compared with control children (P?=?0.002 for viral, P?=?0.011 for bacterial). The disulphide levels were lower in bacterial than in viral tonsillopharyngitis patients (P?=?0.04), and there was a significant difference between viral tonsillopharyngitis patients and the control group (P?P?P?=?0.017 for bacterial). The disulphide/native thiol and disulphide/total thiol ratios were significantly higher in viral (P?P?=?0.017 for both) than in healthy children. In all patients, a correlation was found between the levels of C-reactive protein (CRP) and native thiol (r?=??0.211, P?=?0.04), CRP and total thiol (r?=??0.217, P?=?0.036), white blood cell (WBC) and native thiol (r?=??0.228, P?=?0.002), WBC and total thiol (r?=??0.191, P?=?0.01), and WBC and disulphide (r?=?0.160, P?=?0.03).

Discussion: TDH is altered in AT in children. The alteration is more prominent in viral than in bacterial tonsillopharyngitis.     

IFNL4 rs12979860 polymorphism influences HBV DNA viral loads but not the outcome of HBV infection in Moroccan patients

ObjectivesThe interferon (IFN) is known to bridge innate and adaptive immune responses, and to play a critical role particularly against hepatitis B virus (HBV) infection. Defects in IFN signals may result, therefore, in attenuated responses against HBV. Accordingly, polymorphisms in genes coding for immune response effectors may affect the clinical outcome of HBV infection. We analyzed the putative association between IFNL4 rs12979860 polymorphism and the outcome of HBV infection in Moroccan patients.MethodsIn this study, 237 chronic HBV (CHB) patients and 129 spontaneously resolved HBV (SRB) individuals were enrolled and genotyped using a predesigned Taqman allelic discrimination assay.ResultsOur data show a significant increase of HBV DNA loads in patients with IFNL4 rs12979860 CC genotype compared to patients with CT and TT genotypes (p = 0.0008). However, there was no consistent association between IFNL4 rs12979860 polymorphism and the outcome of HBV infection.ConclusionsAlthough IFNL4 rs12979860 polymorphism seems to modulate circulating HBV DNA levels, it is disconnected from chronic disease progression. This observation suggests that the role of rs12979860 in liver disease is restricted to viral control and inactive in the deleterious immune pathology that affects liver tissue. Taken together, our data suggest that rs12979860 CC genotypes could be useful as a predictor of success or failure of IFN-based therapy in chronic HBV-infected patients.     

Studies Designed to Increase the Stability and Antiviral Activity (HCMV) of the Active Benzimidazole Nucleoside,TCRB

Abstract

The potent activity of 2,5,6-trichloro-1-(ß-D-ribofuranosyl)benzimidazole (TCRB) against Human Cytomegalovirus with the concomitant low cellular toxicity at concentrations that inhibit viral growth prompted considerable interest in this research area. This interest was moderated by the pharmacokinetic studies of TCRB in rats and monkeys that revealed the instability of TCRB in vivo. These studies suggested that the instability was due to a cleavage of the glycosidic bond in vivo which released the heterocycle (2,5,6-trichlorobenzimidazole) into the bloodstream. This prompted us to initiate synthetic studies designed to increase the stability of the glycosidic bond of TCRB and BDCRB. Several synthetic approaches to address this and other problems are presented.     

Modified Hammerhead Ribozymes as Potential Therapeutics

Abstract

Hammerhead ribozymes were modified in the 2′-position by flouro-, amino-, deoxy-, O-methyl- and methoxyethoxy-groups to stabilize against degradations. They were tested for their ability to cut specifically oncogenic N-ras RNA in vitro. Exogenous delivery by lipofection as well as viral vector mediated transfection showed comparable results in reducing N-ras mRNA.