ABSTRACT
Investigations on the twigs and leaves of Antirhea chinensis have led to the discovery of two undescribed pentacyclic triterpenoids (1 and 2) and nine known analogs. Compounds 1 and 2, each assigned as the ursane and 24-noroleanane-type triterpenoids, featuring similar oxidation pattern of 3ß,6ß,19α-trihydroxy-28-carboxyl. Their structures were elucidated via comprehensive analyses of spectroscopic data. Compound 1 displayed potent anti-HIV activity (EC50 =1.24â µM) and high selectivity index (SI >32.3).
Subject(s)
Rubiaceae , Triterpenes , Triterpenes/chemistry , Plant Leaves/chemistry , Plant Extracts/chemistry , Molecular StructureABSTRACT
An ethanol extract of the powdered twigs of Podocarpus imbricatus afforded 14 new diterpenoids (1-14), which all share an aromatized C ring. These isolates belong to five diterpenoid types that include abietanes (1-3), semperviranes (4-9), totaranes (10-12), a C-17 norabietane (13), and an icetexane (14). Their structures were assigned mainly by analysis of the spectroscopic data, and the absolute configuration of 1 was determined by X-ray crystallography. A biosynthetic pathway for five of the biogenetically related types of diterpenoids was proposed. Compound 7 showed moderate inhibitory activity against Zika virus with an IC50 value of 2.5 µM.
Subject(s)
Diterpenes/chemistry , Tracheophyta/chemistry , Antineoplastic Agents, Phytogenic/chemistry , Drug Screening Assays, Antitumor , Humans , Magnetic Resonance Spectroscopy/methods , Quantum TheoryABSTRACT
An amendment to this paper has been published and can be accessed via a link at the top of the paper.
ABSTRACT
Inactivation of the protein complex 'mechanistic target of rapamycin complex 1' (mTORC1) can increase the nuclear content of transcriptional regulators of metabolism and apoptosis. Previous studies established that nuclear import of signal transducer and activator of transcription-1 (STAT1) requires the mTORC1-associated adaptor karyopherin-α1 (KPNA1) when mTORC1 activity is reduced. However, the role of other mTORC1-interacting proteins in the complex, including 'protein kinase C delta' (PKCδ), have not been well characterized. In this study, we demonstrate that PKCδ, a STAT1 kinase, contains a functional 'target of rapamycin signaling' (TOS) motif that directs its interaction with mTORC1. Depletion of KPNA1 by RNAi prevented the nuclear import of PKCδ in cells exposed to the mTORC1 inhibitor rapamycin or amino acid restriction. Mutation of the TOS motif in PKCδ led to its loss of regulation by mTORC1 or karyopherin-α1, resulting in increased constitutive nuclear content. In cells expressing wild-type PKCδ, STAT1 activity and apoptosis were increased by rapamycin or interferon-ß. Those expressing the PKCδ TOS mutant exhibited increased STAT1 activity and apoptosis; further enhancement by rapamycin or interferon-ß, however, was lost. Therefore, the TOS motif in PKCδ is a novel structural mechanism by which mTORC1 prevents PKCδ and STAT1 nuclear import, and apoptosis.
Subject(s)
Cell Nucleus/metabolism , Mechanistic Target of Rapamycin Complex 1/metabolism , Protein Kinase C-delta/metabolism , Active Transport, Cell Nucleus/drug effects , Amino Acid Motifs , Apoptosis/drug effects , Apoptosis/physiology , Cell Line , Humans , Models, Molecular , Mutation, Missense , Point Mutation , Protein Conformation , Protein Interaction Mapping , Protein Kinase C-delta/chemistry , Protein Kinase C-delta/genetics , RNA Interference , RNA, Small Interfering/genetics , Recombinant Proteins/metabolism , Regulatory-Associated Protein of mTOR/metabolism , STAT1 Transcription Factor/biosynthesis , Sequence Alignment , Sirolimus/pharmacology , alpha Karyopherins/antagonists & inhibitors , alpha Karyopherins/metabolismABSTRACT
Studies aimed at repurposing existing drugs revealed that some antimalarial compounds possess anti-Zika virus (anti-ZIKV) activity. Here, we further tested 14 additional antimalarial drugs and their metabolites or analogs for anti-ZIKV activity using a phenotypic screening approach. We identified four compounds with varying anti-ZIKV activity, including a metabolite of amodiaquine termed desethylamodiaquine (DAQ) and N-desethylchloroquine (DECQ), a metabolite of chloroquine, which both exhibited low micromolar effective concentrations against three different ZIKV strains. Two other compounds termed dihydroartemisinin (DHA) and quinidine (QD) exhibited only partial inhibition of ZIKV replication. Characterization of the inhibitory mechanisms of DAQ and DECQ showed that both drugs target the entry step as well as postentry events of the viral replication cycle. These hits represent attractive starting points for future optimization of new anti-ZIKV drug candidates derived from antimalarial drugs and their analogs.
Subject(s)
Antimalarials/pharmacology , Antiviral Agents/pharmacology , Drug Repositioning , Virus Replication/drug effects , Zika Virus/drug effects , Animals , Antimalarials/metabolism , Antiviral Agents/isolation & purification , Cell Line , Chlorocebus aethiops , Culicidae/cytology , Vero Cells , Zika Virus/physiologyABSTRACT
INTRODUCTION: The Zika virus (ZIKV) infection results in severe neurological complications and has emerged as a threat to public health worldwide. No drugs or vaccines are available for use in the clinic and the need for novel and effective therapeutic agents is urgent. AREAS COVERED: This review describes the latest progress of antiviral development for the treatment of ZIKV infection; it primarily focuses on the literature describing 20 potential anti-ZIKV drugs/agents currently being tested in vivo or in clinical trials. The paper also discusses the need for novel ZIKV inhibitors and the critical issues for successful antiviral drug development. EXPERT OPINION: So far, 20 compounds have been tested in vivo and three in the clinical trials; progressing these compounds to the clinic is a challenge. Novel ZIKV inhibitors that target virus or host factors are urgently needed. Knowledge-driven drug repurposing, structure-based discovery, RNA interference, long noncoding RNAs, miRNAs, and peptide inhibitors may pave the way for the discovery of such novel agents.
Subject(s)
Antiviral Agents/therapeutic use , Drugs, Investigational/therapeutic use , Zika Virus Infection/drug therapy , Animals , Antiviral Agents/pharmacology , Drug Design , Drug Repositioning , Drugs, Investigational/pharmacology , Humans , Viral Vaccines/administration & dosage , Zika Virus/drug effects , Zika Virus/isolation & purification , Zika Virus Infection/virologyABSTRACT
Background: Dolutegravir (DTG) is an integrase strand-transfer inhibitor (INSTI) used for treatment of human immunodeficiency virus (HIV)-infected individuals. Owing to its high genetic barrier to resistance, DTG has been clinically investigated as maintenance monotherapy to maintain viral suppression and to reduce complication and healthcare costs. Our study aims to explain the underlying mechanism related to the emergence of a S230R substitution in patients who experienced virologic failure while using DTG monotherapy. Methods: We evaluated the effect of the S230R substitution in regard to integrase enzyme activity, viral infectivity, replicative capacity, and susceptibility to different INSTIs by biochemical and cell-based assays. Results: The S230R substitution conferred a 63% reduction in enzyme efficiency. S230R virus was 1.29-fold less infectious than wild-type virus but could replicate in PM1 cells without significant delay. Resistance levels against DTG, cabotegravir, raltegravir, and elvitegravir in tissue culture were 3.85-, 3.72-, 1.52-, and 1.21-fold, respectively, in virus with the S230R substitution. Conclusions: Our data indicate that the S230R substitution is comparable to the previously reported R263K substitution in some respects. Virologic failure during DTG monotherapy can occur through the development of the S230R or R263K mutation, without the need for high-level DTG resistance.
Subject(s)
Amino Acid Substitution , Drug Resistance, Viral , HIV Infections/drug therapy , HIV Integrase Inhibitors/therapeutic use , HIV/drug effects , Heterocyclic Compounds, 3-Ring/therapeutic use , Viral Load , HIV/genetics , HIV/growth & development , HIV/isolation & purification , HIV Integrase/genetics , HIV Integrase/metabolism , Humans , Maintenance Chemotherapy/methods , Microbial Sensitivity Tests , Mutation, Missense , Oxazines , Piperazines , Pyridones , Treatment Failure , Virus ReplicationABSTRACT
Zika virus (ZIKV) outbreak has emerged as a global health threat, particularly in tropical areas, over the past few years. No antiviral therapy or vaccine is available at present. For these reasons, repurposing clinically approved drugs against ZIKV infection may provide rapid and cost-effective global health benefits. Here, we explored this strategy and screened eight FDA-approved drugs for antiviral activity against ZIKV using a cell-based assay. Our results show that the antimalarial drug amodiaquine has anti-ZIKV activity with EC50 at low micromolar concentrations in cell culture. We further characterized amodiaquine antiviral activity against ZIKV and found that it targets early events of the viral replication cycle. Altogether, our results suggest that amodiaquine may be efficacious for the treatment of ZIKV infection.
Subject(s)
Amodiaquine/pharmacology , Antiviral Agents/pharmacology , Virus Replication/drug effects , Zika Virus/drug effects , Animals , Antimalarials/pharmacology , Cell Line , Drug Evaluation, Preclinical , Drug Repositioning , Humans , Microbial Sensitivity Tests , Zika Virus/physiologyABSTRACT
INTRODUCTION: To date, three HIV integrase strand transfer inhibitors (INSTIs), i.e. raltegravir, elvitegravir and dolutegravir, have been approved for clinical use. Recent research has focused on new integrase inhibitors including those targeting non-catalytic sites of HIV integrase. Areas covered: This paper reviews two investigational INSTIs in phase I and II clinical trials, bictegravir (BIC) and cabotegravir (CAB), as well as an investigational noncatalytic integrase inhibitor (NCINI) termed BI 224436. Expert opinion: Data from phase I and II clinical trials demonstrate that CAB has good efficacy and is well-tolerated. CAB is promising because it can be formulated both orally and as a long-acting (LA) injectable for treatment and prevention of HIV infection. Since LA-CAB formulation offers the possibility of favourable dosing, it may help individuals who struggle with adherence issues. BIC also represents a promising safe, effective and well-tolerated drug that can be administered as a single once-daily regimen in coformulation with emtricitabine and tenofovir alafenamide (FTC/TAF). Ongoing phase III trials should clarify optimal doses and reveal the potential clinical advantages of these new drugs and formulations over other current regimens. Exploration of novel HIV integrase inhibitors acting through mechanisms different from those of INSTIs is still needed.
Subject(s)
Drugs, Investigational/administration & dosage , HIV Infections/drug therapy , HIV Integrase Inhibitors/administration & dosage , Amides , Animals , Clinical Trials, Phase I as Topic , Clinical Trials, Phase II as Topic , Drugs, Investigational/adverse effects , Drugs, Investigational/pharmacology , HIV Infections/enzymology , HIV Integrase Inhibitors/adverse effects , HIV Integrase Inhibitors/pharmacology , Heterocyclic Compounds, 3-Ring , Heterocyclic Compounds, 4 or More Rings/administration & dosage , Heterocyclic Compounds, 4 or More Rings/adverse effects , Heterocyclic Compounds, 4 or More Rings/pharmacology , Humans , Medication Adherence , Piperazines , Pyridones/administration & dosage , Pyridones/adverse effects , Pyridones/pharmacologyABSTRACT
We evaluated Sofosbuvir (SOF), the anti-hepatitis C virus prodrug of ß-d-2'-deoxy-2'-α-fluoro-2'-ß-C-methyluridine-5'-monophosphate, for potential inhibitory activity against DENV replication. Both cell-based and biochemical assays, based on use of purified DENV full-length NS5 enzyme, were studied. Cytopathic effect protection and virus yield reduction assays confirmed that SOF possessed anti-DENV activity in cell culture with a 50% effective concentration (EC50) of 4.9 µM and 1.4 µM respectively. Real-time RT-PCR verified that SOF inhibits generation of viral RNA with an EC50 of 9.9 µM. Purified DENV NS5 incorporated the active triphosphate form (SOF-TP) into nascent RNA, causing chain-termination. Relative to the natural UTP, the incorporation efficiency of SOF-TP was low (discrimination value = 327.5). In a primer extension assay, SOF-TP was active against DENV NS5 wild-type polymerase activity with an IC50 of 14.7 ± 2.5 µM. The S600T substitution in the B Motif of DENV polymerase conferred 4.3-fold resistance to SOF-TP; this was due to decreased incorporation efficiency rather than enhanced excision of the incorporated SOF nucleotide. SOF has antiviral activity against DENV replication. The high discrimination value in favor of UTP in enzyme assays may not necessarily preclude antiviral activity in cells. SOF may be worthy of evaluation against severe DENV infections in humans.
Subject(s)
Antiviral Agents/pharmacology , Dengue Virus/physiology , Sofosbuvir/pharmacology , Virus Replication/drug effects , Cell Line , Dengue/drug therapy , Dengue/virology , Dengue Virus/drug effects , Dengue Virus/enzymology , Drug Evaluation , Humans , Inhibitory Concentration 50 , Molecular Docking Simulation , Uridine Triphosphate/metabolism , Viral Nonstructural Proteins/chemistry , Viral Nonstructural Proteins/genetics , Viral Nonstructural Proteins/metabolismABSTRACT
Objectives: To compare the effectiveness of HIV integrase inhibitor monotherapy between raltegravir and dolutegravir as an approach to simplify therapy. Methods: We evaluated and compared the efficacy of 20 week monotherapy with dolutegravir or raltegravir in humanized mice (HSC-NSG) infected with HIVBaL. Plasma HIV RNA was measured by quantitative RT-PCR (limit of detection of 150 copies/45 µL of plasma) and drug levels by LC-MS/MS. Escape viruses were genotyped and analysed for replication capacity and drug susceptibility in tissue culture. Results: Drug-untreated control mice maintained constant viraemia throughout the study. Virus isolates from these mice were susceptible to both raltegravir (EC50 of <8 nM) and dolutegravir (EC50 of <1 nM). Mice treated with raltegravir or dolutegravir had plasma drug levels comparable to those in humans. Monotherapy with raltegravir initially suppressed HIV viraemia, but failed to maintain suppression in 4/4 mice. Viruses from raltegravir failing mice developed mutations G140G/S and Q148H/K, and were resistant to both raltegravir (EC50 values of >100 nM) and dolutegravir (EC50 values ranging from 8.8 to 13.3 nM). Monotherapy with dolutegravir suppressed viraemia in 5/5 of mice, but viraemia rebounded in one animal. The virus from this mouse had mutations E138K, G140S, Q148H, N155H and S230R, was highly resistant to both raltegravir (EC50 of >1000 nM) and dolutegravir (EC50 of 550 nM), and replicated to levels similar to those of control viruses in PBMCs. Conclusions: Monotherapy with either raltegravir or dolutegravir does not consistently maintain HIV suppression, suggesting that dual therapy may be required in simplification strategies.
Subject(s)
Anti-HIV Agents/therapeutic use , HIV Infections/drug therapy , HIV-1/drug effects , Heterocyclic Compounds, 3-Ring/therapeutic use , Raltegravir Potassium/therapeutic use , Viremia/drug therapy , Animals , Genotype , HIV Infections/virology , HIV Integrase Inhibitors/therapeutic use , HIV-1/genetics , Humans , Mice , Mice, Transgenic , Mutation , Oxazines , Piperazines , Pyridones , RNA, Viral/blood , Viremia/virology , Virus Replication/drug effectsABSTRACT
Human immunodeficiency virus (HIV) infection persists despite decades of active antiretroviral therapy (ART), effectively preventing viral eradication. Treatment decreases plasma viral RNA, but viral DNA persists, mostly integrated within the genome of nucleated blood cells. Viral DNA blood levels correlate with comorbidities and the rapidity of viral rebound following treatment interruption. To date, no intervention aiming at decreasing HIV DNA levels below those attained through ART has been successful. This includes use of some integrase inhibitors either as part of ART or in treatment intensification studies. We have argued that using the integrase inhibitor dolutegravir (DTG) in similar studies may yield better results, but this remains to be studied. In treatment-experienced individuals, the most frequent substitution associated with failure with dolutegravir is R263K in integrase. R263K decreases integration both in cell-free and tissue culture assays. We investigated here how integrated DNA levels evolve over time during prolonged infections with R263K viruses. To investigate a potential defect in reverse transcription with R263K, the levels of reverse transcripts were measured by quantitative PCR. We measured HIV type 1 (HIV-1) integration in Jurkat cells over the course of 4-week infections using Alu-mediated quantitative PCR. The results show that R263K did not decrease reverse transcription. Prolonged infections with R263K mutant viruses led to less HIV-1 integrated DNA over time compared to wild-type viruses. These tissue culture results help to explain the absence of the R263K substitution in most individuals experiencing failure with DTG and support studies aiming at longitudinally measuring the levels of integrated DNA in individuals treated with this drug.IMPORTANCE Antiretroviral treatment decreases plasma viral RNA, but HIV DNA persists for decades within infected cells. Studies of nonhuman primates have suggested that reducing retroviral DNA levels might represent a path to eradication. The integrase inhibitor dolutegravir is less susceptible than any other anti-HIV drug to the emergence of resistance in treatment-naive individuals. In treatment-experienced individuals, in contrast, rare cases of treatment failure were commonly associated with emergence of an R263K integrase substitution that confers low-level resistance to dolutegravir. It is unclear why this substitution is not more common in individuals experiencing failure with dolutegravir. We report here that R263K progressively diminishes the levels of integrated HIV-1 DNA in tissue culture over multiple cycles of infection. Our results help to explain aspects of the clinical efficacy of dolutegravir and suggest that this drug may be able to reduce HIV DNA levels within infected individuals compared to other drugs.
Subject(s)
Drug Resistance, Viral , HIV Integrase Inhibitors/pharmacology , HIV Integrase/metabolism , HIV-1/physiology , Heterocyclic Compounds, 3-Ring/pharmacology , Mutation, Missense , Virus Integration , DNA, Viral/analysis , HEK293 Cells , HIV Integrase/genetics , HIV-1/enzymology , Humans , Jurkat Cells , Oxazines , Piperazines , Pyridones , Real-Time Polymerase Chain ReactionABSTRACT
HIV can spread by both cell-free and cell-to-cell transmission. Here, we show that many of the amino acid changes in Env that are close to the CD4 binding pocket can affect HIV replication. We generated a number of mutant viruses that were unable to infect T cells as cell-free viruses but were nevertheless able to infect certain T cell lines as cell-associated viruses, which was followed by reversion to the wild type. However, the activation of JAK-STAT signaling pathways caused the inhibition of such cell-to-cell infection as well as the reversion of multiple HIV Env mutants that displayed differences in their abilities to bind to the CD4 receptor. Specifically, two T cell activators, interleukin-2 (IL-2) and phorbol 12-myristate 13-acetate (PMA), both capable of activation of JAK-STAT pathways, were able to inhibit cell-to-cell viral transmission. In contrast, but consistent with the above result, a number of JAK-STAT and mTOR inhibitors actually promoted HIV-1 transmission and reversion. Hence, JAK-STAT signaling pathways may differentially affect the replication of a variety of HIV Env mutants in ways that differ from the role that these pathways play in the replication of wild-type viruses.IMPORTANCE Specific alterations in HIV Env close to the CD4 binding site can differentially change the ability of HIV to mediate infection for cell-free and cell-associated viruses. However, such differences are dependent to some extent on the types of target cells used. JAK-STAT signaling pathways are able to play major roles in these processes. This work sheds new light on factors that can govern HIV infection of target cells.
Subject(s)
CD4 Antigens/metabolism , HIV Infections/transmission , Janus Kinases/antagonists & inhibitors , STAT Transcription Factors/antagonists & inhibitors , TOR Serine-Threonine Kinases/antagonists & inhibitors , env Gene Products, Human Immunodeficiency Virus/genetics , Binding Sites/genetics , CD4-Positive T-Lymphocytes/immunology , CD4-Positive T-Lymphocytes/virology , HIV Infections/virology , HIV-1/metabolism , Humans , Interleukin-2/metabolism , Janus Kinases/metabolism , Nitriles , Piperidines/pharmacology , Pyrazoles/pharmacology , Pyrimidines/pharmacology , Pyrroles/pharmacology , Pyrrolidines/pharmacology , STAT Transcription Factors/metabolism , Signal Transduction , Sulfonamides/pharmacology , Tetradecanoylphorbol Acetate/pharmacology , Virus Internalization , Virus Replication/geneticsABSTRACT
Background: The viral RNA-dependent RNA polymerase (RdRp) enzymes of the Flaviviridae family are essential for viral replication and are logically important targets for development of antiviral therapeutic agents. Zika virus (ZIKV) is a rapidly re-emerging human pathogen for which no vaccine or antiviral agent is currently available. Methods: To facilitate development of ZIKV RdRp inhibitors, we have established an RdRp assay using purified recombinant ZIKV NS5 polymerase. Results: We have shown that both the hepatitis C virus (HCV) nucleoside inhibitor sofosbuvir triphosphate and a pyridoxine-derived non-nucleoside small-molecule inhibitor, DMB213, can act against ZIKV RdRp activity at IC 50 s of 7.3 and 5.2â µM, respectively, in RNA synthesis reactions catalysed by recombinant ZIKV NS5 polymerase. Cell-based assays confirmed the anti-ZIKV activity of sofosbuvir and DMB213 with 50% effective concentrations (EC 50 s) of 8.3 and 4.6â µM, respectively. Control studies showed that DMB213 did not inhibit recombinant HIV-1 reverse transcriptase and showed only very weak inhibition of HIV-1 integrase strand-transfer activity. The S604T substitution in motif B of the ZIKV RdRp, which corresponds to the S282T substitution in motif B of HCV RdRp, which confers resistance to nucleotide inhibitors, also conferred resistance to sofosbuvir triphosphate, but not to DMB213. Enzyme assays showed that DMB213 appears to be competitive with natural nucleoside triphosphate (NTP) substrates. Conclusions: Recombinant ZIKV RdRp assays can be useful tools for the screening of both nucleos(t)ide compounds and non-nucleotide metal ion-chelating agents that interfere with ZIKV replication.
Subject(s)
Antiviral Agents/pharmacology , Enzyme Inhibitors/pharmacology , RNA-Dependent RNA Polymerase/antagonists & inhibitors , RNA-Dependent RNA Polymerase/metabolism , Zika Virus/enzymology , Drug Discovery/methods , HIV Integrase/metabolism , HIV Reverse Transcriptase/antagonists & inhibitors , RNA, Viral/metabolism , RNA-Dependent RNA Polymerase/chemistry , RNA-Dependent RNA Polymerase/genetics , Recombinant Proteins/metabolism , Sofosbuvir/pharmacology , Virus Replication/drug effects , Zika Virus/drug effects , Zika Virus/physiologyABSTRACT
Dengue virus (DENV) causes a variety of difficult-to-treat diseases that threaten almost half of the world's population. Currently, no effective vaccine or antiviral therapy is available. We have examined a series of synthetic resveratrol analogs to identify potential anti-DENV agents. Here, we demonstrate that two resveratrol analogs, PNR-4-44 and PNR-5-02, possess potent anti-DENV activity with EC50 values in the low nanomolar range. These two resveratrol analogs were shown to mainly target viral RNA translation and viral replication, but PNR-5-02 is also likely to target cellular factors inside host cells. Although the precise molecular mechanism(s) mediating anti-DENV activities have not been elucidated, further structure-guided design might lead to the development of newer improved resveratrol derivatives that might have therapeutic value. J. Med. Virol. 89:397-407, 2017. © 2016 Wiley Periodicals, Inc.
Subject(s)
Antiviral Agents/pharmacology , Dengue Virus/drug effects , Stilbenes/pharmacology , Dengue Virus/physiology , Drug Evaluation, Preclinical , Drug Repositioning , Humans , Microbial Sensitivity Tests , Protein Biosynthesis/drug effects , Resveratrol , Virus Replication/drug effectsABSTRACT
Three integrase strand transfer inhibitors (INSTIs), raltegravir (RAL), elvitegravir (EVG) and dolutegravir (DTG), have been approved by the FDA. Resistance against these three INSTIs have been reported and cross-resistance among them has been documented. Due to extensive and dynamic genetic diversity in different HIV-1 variants, significant differences in susceptibility to the INSTIs have been observed among HIV subtypes. This review summarizes what is known about this topic and discusses possible clinical implications.
Subject(s)
Drug Resistance, Viral/drug effects , Drug Resistance, Viral/genetics , HIV Infections/virology , HIV Integrase Inhibitors/pharmacology , HIV-1/drug effects , HIV-1/genetics , Heterocyclic Compounds, 3-Ring/pharmacology , Humans , Oxazines , Piperazines , Pyridones , Quinolones/pharmacology , Raltegravir Potassium/pharmacologyABSTRACT
Antiretroviral therapy (ART) has greatly decreased HIV-related morbidity and mortality. However, HIV can establish viral reservoirs that evade both the immune system and ART. Dolutegravir (DTG) is a second-generation integrase strand transfer inhibitor (INSTI) related to the first-generation INSTIs raltegravir (RAL) and elvitegravir (EVG). DTG shows a higher genetic barrier to the development of HIV-1 resistance than RAL and EVG. More interestingly, clinical resistance mutations to DTG in treatment-naïve patients have not been observed to date. This review summarizes recent studies on strategies toward a cure for HIV, explores resistance profiles of DTG, and discusses how DTG might help in finding a functional cure for HIV.
Subject(s)
HIV Infections/drug therapy , HIV Integrase Inhibitors/therapeutic use , Heterocyclic Compounds, 3-Ring/therapeutic use , Drug Resistance, Viral , HIV-1 , Humans , Oxazines , Piperazines , PyridonesABSTRACT
The viral RNA-dependent RNA polymerase (RdRp) activity of the dengue virus (DENV) NS5 protein is an attractive target for drug design. Here, we report the identification of a novel class of inhibitor (i.e., an active-site metal ion chelator) that acts against DENV RdRp activity. DENV RdRp utilizes a two-metal-ion mechanism of catalysis; therefore, we constructed a small library of compounds, through mechanism-based drug design, aimed at chelating divalent metal ions in the catalytic site of DENV RdRp. We now describe a pyridoxine-derived small-molecule inhibitor that targets DENV RdRp and show that 5-benzenesulfonylmethyl-3-hydroxy-4-hydroxymethyl-pyridine-2-carboxylic acid hydroxyamide (termed DMB220) inhibited the RdRp activity of DENV serotypes 1 to 4 at low micromolar 50% inhibitory concentrations (IC50s of 5 to 6.7 µM) in an enzymatic assay. The antiviral activity of DMB220 against DENV infection was also verified in a cell-based assay and showed a 50% effective concentration (EC50) of <3 µM. Enzyme assays proved that DMB220 was competitive with nucleotide incorporation. DMB220 did not inhibit the enzymatic activity of recombinant HIV-1 reverse transcriptase and showed only weak inhibition of HIV-1 integrase strand transfer activity, indicating high specificity for DENV RdRp. S600T substitution in the DENV RdRp, which was previously shown to confer resistance to nucleoside analogue inhibitors (NI), conferred 3-fold hypersusceptibility to DMB220, and enzymatic analyses showed that this hypersusceptibility may arise from the decreased binding/incorporation efficiency of the natural NTP substrate without significantly impacting inhibitor binding. Thus, metal ion chelation at the active site of DENV RdRp represents a viable anti-DENV strategy, and DMB220 is the first of a new class of DENV inhibitor.
Subject(s)
Antiviral Agents/pharmacology , Chelating Agents/pharmacology , Dengue Virus/drug effects , Hydroxamic Acids/pharmacology , Picolines/pharmacology , RNA-Dependent RNA Polymerase/antagonists & inhibitors , Small Molecule Libraries/pharmacology , Sulfones/pharmacology , Viral Nonstructural Proteins/antagonists & inhibitors , Aedes , Amino Acid Substitution , Animals , Antiviral Agents/chemical synthesis , Binding Sites , Catalytic Domain , Cell Line , Chelating Agents/chemical synthesis , Cricetinae , Dengue Virus/enzymology , Dengue Virus/genetics , Dose-Response Relationship, Drug , Drug Design , Epithelial Cells/drug effects , Epithelial Cells/virology , Gene Expression , Histidine/genetics , Histidine/metabolism , Humans , Hydroxamic Acids/chemical synthesis , Kinetics , Molecular Docking Simulation , Oligopeptides/genetics , Oligopeptides/metabolism , Picolines/chemical synthesis , Protein Binding , Protein Structure, Secondary , RNA-Dependent RNA Polymerase/chemistry , RNA-Dependent RNA Polymerase/genetics , RNA-Dependent RNA Polymerase/metabolism , Recombinant Fusion Proteins/chemistry , Recombinant Fusion Proteins/genetics , Recombinant Fusion Proteins/metabolism , Small Molecule Libraries/chemical synthesis , Sulfones/chemical synthesis , Viral Nonstructural Proteins/chemistry , Viral Nonstructural Proteins/genetics , Viral Nonstructural Proteins/metabolismABSTRACT
Two new alkaloids, flueggether A (1) and virosinine A (2), were isolated from a Chinese medicinal plant, Flueggea virosa. Their structures were assigned via spectroscopic methods with the absolute configurations of 1 and 2 being established by X-ray diffraction analysis and calculated electronic circular dichroism data, respectively. Compound 1 represents the first example with an ether bridge of Securinega alkaloid oligomers, and 2 bears a new heterocyclic backbone. Both alkaloids showed mild in vitro anti-HIV activity.