Approved Antiviral Drugs Research Articles

A Rapid Screening Assay Identifies Monotherapy with Interferon-ß and Combination Therapies with Nucleoside Analogs as Effective Inhibitors of Ebola Virus.

To date there are no approved antiviral drugs for the treatment of Ebola virus disease (EVD). While a number of candidate drugs have shown limited efficacy in vitro and/or in non-human primate studies, differences in experimental methodologies make it difficult to compare their therapeutic effectiveness. Using an in vitro model of Ebola Zaire replication with transcription-competent virus like particles (trVLPs), requiring only level 2 biosafety containment, we compared the activities of the type I interferons (IFNs) IFN-α and IFN-ß, a panel of viral polymerase inhibitors (lamivudine (3TC), zidovudine (AZT) tenofovir (TFV), favipiravir (FPV), the active metabolite of brincidofovir, cidofovir (CDF)), and the estrogen receptor modulator, toremifene (TOR), in inhibiting viral replication in dose-response and time course studies. We also tested 28 two- and 56 three-drug combinations against Ebola replication. IFN-α and IFN-ß inhibited viral replication 24 hours post-infection (IC50 0.038μM and 0.016μM, respectively). 3TC, AZT and TFV inhibited Ebola replication when used alone (50–62%) or in combination (87%). They exhibited lower IC50 (0.98–6.2μM) compared with FPV (36.8μM), when administered 24 hours post-infection. Unexpectedly, CDF had a narrow therapeutic window (6.25–25μM). When dosed >50μM, CDF treatment enhanced viral infection. IFN-ß exhibited strong synergy with 3TC (97.3% inhibition) or in triple combination with 3TC and AZT (95.8% inhibition). This study demonstrates that IFNs and viral polymerase inhibitors may have utility in EVD. We identified several 2 and 3 drug combinations with strong anti-Ebola activity, confirmed in studies using fully infectious ZEBOV, providing a rationale for testing combination therapies in animal models of lethal Ebola challenge. These studies open up new possibilities for novel therapeutic options, in particular combination therapies, which could prevent and treat Ebola infection and potentially reduce drug resistance.

Verapamil has Antiviral Activities that Target Different Steps of the Influenza Virus Replication Cycle

Safe and efficacious therapeutic options for infections caused by highly pathogenic influenza viruses (IV) remain limited. Viruses resistant to approved antiviral drugs are increasingly emerging and, in some cases, existing vaccines are insufficient to prevent seasonal epidemics or pandemics caused by these viruses. As a consequence, alternative antiviral strategies that, for example, target cellular factors or mechanisms essential for IV propagation have attracted increasing interest over the past years. Here, we characterized antiviral effects of the calcium channel blocker Verapamil on different steps of the IV replication cycle. Our data suggest that Verapamil (i) efficiently reduces influenza A and B virus titres in cell culture at non-toxic concentrations, (ii) does not influence virus entry, (iii) impairs the replication and transcription activity of the viral polymerase, (iv) decreases viral protein production, (v) reduces virus-induced NF-κB activation and (vi) does not induce the emergence of resistant IV variants, resulting in strongly reduced infectious particle formation.

In silico prediction of ebolavirus RNA polymerase inhibition by specific combinations of approved nucleotide analogues

Background and objectiveThe urgency of ebolavirus drug development is obvious in light of the current local epidemic in Western Africa with high morbidity and a risk of wider spread. We present an in silico study as a first step to identify inhibitors of ebolavirus polymerase activity based on approved antiviral nucleotide analogues. Study designSince a structure model of the ebolavirus polymerase is lacking, we performed combined homology and ab initio modeling and report a similarity to known polymerases of human enterovirus, bovine diarrhea virus and foot-and-mouth disease virus. This facilitated the localization of a nucleotide binding domain in the ebolavirus polymerase. We next performed molecular docking studies with nucleotides (ATP, CTP, GTP and UTP) and nucleotide analogues, including a variety of approved antiviral drugs. Results and conclusionsSpecific combinations of nucleotide analogues significantly reduce the ligand-protein interaction energies of the ebolavirus polymerase for natural nucleotides. Any nucleotide analogue on its own did not reduce ligand-protein interaction energies. This prediction encourages specific drug testing efforts and guides future strategies to inhibit ebolavirus replication.

ID: 68: IFN-β treatment for Ebola virus disease: Bench to bedside

To date there are no approved vaccines or antiviral drugs for the treatment of Ebola virus disease (EVD). While a number of candidate drugs have shown limited efficacy in preclinical studies, differences in experimental methodologies make it difficult to compare their therapeutic effectiveness. Using an in vitro model of Ebola Zaire replication with transcription-competent virus like particles, we compared the antiviral activities of 8 different candidate antivirals from 3 drug classes: IFN-α and IFN-s, viral polymerase inhibitors (lamivudine (3TC), zidovudine (AZT) tenofovir (TFV), favipiravir (FPV), the active metabolite of brincidofovir, cidofovir (CDF), and the estrogen receptor modulator, toremifene (TOR). We also tested 28 two- and 56 three-drug combinations against Ebola replication. IFN-α and IFN-β inhibited viral replication 24 h post-infection (IC50 0.038 μM and 0.016 μM, respectively). 3TC, AZT and TFV inhibited Ebola replication when used alone (50–62%) or in combination (87%). They exhibited lower IC50 (0.98–6.2 μM) compared with FPV (36.8 μM), when administered 24 h post-infection. CDF had a narrow therapeutic window (6.25–25 μM). When dosed >50 μM, CDF treatment enhanced viral infection. IFN- β exhibited strong synergy with 3TC (97.9% inhibition) or in triple combination with 3TC and AZT (98.5% inhibition). Our data indicate that IFN-β is a potent inhibitor of Ebola virus, superior to the antivirals we tested, contributing to the decision to conduct a clinical trial of IFN-β treatment for Ebola virus disease in Coyah, Guinea. Results from the trial, including effects of IFN treatment on viremia, disease symptoms and survival will be presented.

Nucleic acid-based drugs against emerging zoonotic viruses.

Global outbreaks of diseases caused by zoonotic viruses have steadily increased in recent years. Emerging zoonotic viruses are generally phylogenetically diverse, are unpredictable and are known to cause diseases with high case fatality rates in humans and are hard to protect against due to lack of approved antiviral drugs. The aim of this review is to discuss how advances in genomics, rational drug design and innate immune signaling can contribute to the design of nucleic acid-based drugs to combat these emerging threats. Specifically, the antiviral activity of siRNAs, antisense oligonucleotides is mediated by sequence-specific gene silencing, and broad-spectrum innate and antiviral immune responses can be elicited by toll-like receptor agonists. This review will summarize their current state of development, safety and efficacy, and provide perspectives on future development.

Immunogenicity and protective efficacy of an EV71 virus-like particle vaccine against lethal challenge in newborn mice

Enterovirus 71(EV71) has caused severe epidemics of hand, foot and mouth disease (HFMD) in the Asia Pacific in recent years, particularly in infants and pre-school children. It has become a serious public health threat, as currently there are no approved vaccines or antiviral drugs for EV71 infection. Many EV71 vaccines have been under development worldwide, however the main focus is inactivated EV71 vaccines. For example, the inactivated EV71 vaccine has recently finished phase III clinical trial in Mainland China. There have been very few studies on EV71 virus like particles (VLPs). In this study, the immunogenicity and protective potency of the EV71 VLPs produced in insect cells were evaluated in mice with different dosages. Our results showed that EV71 VLPs could elicit high titers of neutralizing antibodies (NTAbs) in a dose-dependent manner and NTAbs were sustained after the second injection with an average GMT (geometric mean titer) level from 19 to 2960 in immunized mice. Survival rates were 100%, 100%, 85%, and 40% after challenge with 15 LD50 (median lethal dose) of EV71 in these newborn mice, respectively. ED50 (50% effective dose) of VLPs was 0.20 μg/dose in newborn mice, while NTAb titer under this dosage was about 50. Passive protection was determined with 2 methods and demonstrated that the survival rates were positively correlated with NTAb titers, which at 24 and 54 induced 50% survival rates in experimental animals. The ED50 of VLP vaccines and the passive NTAb titers were also analyzed. The maternal NTAb titer was similar as the passive NTAb titer in the mouse model challenged with our lethal mouse EV71 strain. Hence, our work has provided preliminary data on the protection potency of VLPs as a vaccine candidate and would facilitate future VLP vaccine development.

Antiviral therapies against Ebola and other emerging viral diseases using existing medicines that block virus entry

Emerging viral diseases pose a threat to the global population as intervention strategies are mainly limited to basic containment due to the lack of efficacious and approved vaccines and antiviral drugs. The former was the only available intervention when the current unprecedented Ebolavirus (EBOV) outbreak in West Africa began. Prior to this, the development of EBOV vaccines and anti-viral therapies required time and resources that were not available. Therefore, focus has turned to re-purposing of existing, licenced medicines that may limit the morbidity and mortality rates of EBOV and could be used immediately. Here we test three such medicines and measure their ability to inhibit pseudotype viruses (PVs) of two EBOV species, Marburg virus (MARV) and avian influenza H5 (FLU-H5). We confirm the ability of chloroquine (CQ) to inhibit viral entry in a pH specific manner. The commonly used proton pump inhibitors, Omeprazole and Esomeprazole were also able to inhibit entry of all PVs tested but at higher drug concentrations than may be achieved in vivo. We propose CQ as a priority candidate to consider for treatment of EBOV.

Peptidomimetic ethyl propenoate covalent inhibitors of the enterovirus 71 3C protease: a P2–P4 study

Enterovirus 71 (EV71) is a highly infectious pathogen primarily responsible for Hand, Foot, and Mouth Disease, particularly among children. Currently, no approved antiviral drug has been developed against this disease. The EV71 3C protease is deemed an attractive drug target due to its crucial role in viral polyprotein processing. Rupintrivir, a peptide-based inhibitor originally developed to target the human rhinovirus 3C protease, was found to inhibit the EV71 3C protease. In this communication, we report the inhibitory activities of 30 Rupintrivir analogs against the EV71 3C protease. The most potent inhibitor, containing a P2 ring-constrained phenylalanine analog (compound 9), was found to be two-fold more potent than Rupintrivir (IC50 value 3.4 ± 0.4 versus 7.3 ± 0.8 μM). Our findings suggest that employing geometrically constrained residues in peptide-based protease inhibitors can potentially enhance their inhibitory activities.

The Catalytic Mechanism of Human Parainfluenza Virus Type 3 Haemagglutinin‐Neuraminidase Revealed

AbstractHuman parainfluenza virus type 3 (hPIV‐3) is one of the leading causes for lower respiratory tract disease in children, with neither an approved antiviral drug nor vaccine available to date. Understanding the catalytic mechanism of human parainfluenza virus haemagglutinin‐neuraminidase (HN) protein is key to the design of specific inhibitors against this virus. Herein, we used 1H NMR spectroscopy, X‐ray crystallography, and virological assays to study the catalytic mechanism of the HN enzyme activity and have identified the conserved Tyr530 as a key amino acid involved in catalysis. A novel 2,3‐difluorosialic acid derivative showed prolonged enzyme inhibition and was found to react and form a covalent bond with Tyr530. Furthermore, the novel derivative exhibited enhanced potency in virus blockade assays relative to its Neu2en analogue. These outcomes open the door for a new generation of potent inhibitors against hPIV‐3 HN.

The catalytic mechanism of human parainfluenza virus type 3 haemagglutinin-neuraminidase revealed.

Human parainfluenza virus type 3 (hPIV-3) is one of the leading causes for lower respiratory tract disease in children, with neither an approved antiviral drug nor vaccine available to date. Understanding the catalytic mechanism of human parainfluenza virus haemagglutinin-neuraminidase (HN) protein is key to the design of specific inhibitors against this virus. Herein, we used (1) H NMR spectroscopy, X-ray crystallography, and virological assays to study the catalytic mechanism of the HN enzyme activity and have identified the conserved Tyr530 as a key amino acid involved in catalysis. A novel 2,3-difluorosialic acid derivative showed prolonged enzyme inhibition and was found to react and form a covalent bond with Tyr530. Furthermore, the novel derivative exhibited enhanced potency in virus blockade assays relative to its Neu2en analogue. These outcomes open the door for a new generation of potent inhibitors against hPIV-3 HN.

Antiviral therapies against Ebola and other emerging viral diseases using existing medicines that block virus entry.

Emerging viral diseases pose a threat to the global population as intervention strategies are mainly limited to basic containment due to the lack of efficacious and approved vaccines and antiviral drugs. The former was the only available intervention when the current unprecedented Ebolavirus (EBOV) outbreak in West Africa began. Prior to this, the development of EBOV vaccines and anti-viral therapies required time and resources that were not available. Therefore, focus has turned to re-purposing of existing, licenced medicines that may limit the morbidity and mortality rates of EBOV and could be used immediately. Here we test three such medicines and measure their ability to inhibit pseudotype viruses (PVs) of two EBOV species, Marburg virus (MARV) and avian influenza H5 (FLU-H5). We confirm the ability of chloroquine (CQ) to inhibit viral entry in a pH specific manner. The commonly used proton pump inhibitors, Omeprazole and Esomeprazole were also able to inhibit entry of all PVs tested but at higher drug concentrations than may be achieved in vivo. We propose CQ as a priority candidate to consider for treatment of EBOV.

Antiviral therapies against Ebola and other emerging viral diseases using existing medicines that block virus entry

Emerging viral diseases pose a threat to the global population as intervention strategies are mainly limited to basic containment due to the lack of efficacious and approved vaccines and antiviral drugs. The former was the only available intervention when the current unprecedented Ebolavirus (EBOV) outbreak in West Africa began. Prior to this, the development of EBOV vaccines and anti-viral therapies required time and resources that were not available. Therefore, focus has turned to re-purposing of existing, licenced medicines that may limit the morbidity and mortality rates of EBOV and could be used immediately. Here we test three such medicines and measure their ability to inhibit pseudotype viruses (PVs) of two EBOV species, Marburg virus (MARV) and avian influenza H5 (FLU-H5). We confirm the ability of chloroquine (CQ) to inhibit viral entry in a pH specific manner. The commonly used proton pump inhibitors, Omeprazole and Esomeprazole were also able to inhibit entry of all PVs tested but at higher drug concentrations than may be achievedin vivo. We propose CQ as a priority candidate to consider for treatment of EBOV.

Toward an ounce of HCV prevention

Hepatitis C Virus Chronic hepatitis C virus (HCV) infection causes liver inflammation that can reduce liver function or even cause liver failure. Recent approval of antiviral drugs for HCV provides treatment options; however, these new therapies are expensive, with limited availability, leaving the door open for preventative approaches such as vaccines. Now, Swadling et al. report a preliminary trial of a prime-boost vaccine strategy for HCV in human patients. The strategy induced an immune T cell response similar to one that can control HCV in natural infection. Sci. Transl. Med. 6 , 261ra153 (2014).

Antiviral effects of two Ganoderma lucidum triterpenoids against enterovirus 71 infection

Enterovirus 71 (EV71) is a major causative agent for hand, foot and mouth disease (HFMD), and fatal neurological and systemic complications in children. However, there is currently no clinical approved antiviral drug available for the prevention and treatment of the viral infection. Here, we evaluated the antiviral activities of two Ganoderma lucidum triterpenoids (GLTs), Lanosta-7,9(11),24-trien-3-one,15;26-dihydroxy (GLTA) and Ganoderic acid Y (GLTB), against EV71 infection. The results showed that the two natural compounds display significant anti-EV71 activities without cytotoxicity in human rhabdomyosarcoma (RD) cells as evaluated by 3-(4,5-Dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) cell proliferation assay. The mechanisms by which the two compounds affect EV71 infection were further elucidated by three action modes using Ribavirin, a common antiviral drug, as a positive control. The results suggested that GLTA and GLTB prevent EV71 infection through interacting with the viral particle to block the adsorption of virus to the cells. In addition, the interactions between EV71 virion and the compounds were predicated by computer molecular docking, which illustrated that GLTA and GLTB may bind to the viral capsid protein at a hydrophobic pocket (F site), and thus may block uncoating of EV71. Moreover, we demonstrated that GLTA and GLTB significantly inhibit the replication of the viral RNA (vRNA) of EV71 replication through blocking EV71 uncoating. Thus, GLTA and GLTB may represent two potential therapeutic agents to control and treat EV71 infection.

Dengue virus infection: current concepts in immune mechanisms and lessons from murine models

Dengue viruses (DENV), a group of four serologically distinct but related flaviviruses, are responsible for one of the most important emerging viral diseases. This mosquito-borne disease has a great impact in tropical and subtropical areas of the world in terms of illness, mortality and economic costs, mainly due to the lack of approved vaccine or antiviral drugs. Infections with one of the four serotypes of DENV (DENV-1-4) result in symptoms ranging from an acute, self-limiting febrile illness, dengue fever, to severe dengue haemorrhagic fever or dengue shock syndrome. We reviewed the existing mouse models of infection, including the DENV-2-adapted strain P23085. The role of CC chemokines, interleukin-17 (IL-17), IL-22 and invariant natural killer T cells in mediating the exacerbation of disease in immune-competent mice is highlighted. Investigations in both immune-deficient and immune-competent mouse models of DENV infection may help to identify key host–pathogen factors and devise novel therapies to restrain the systemic and local inflammatory responses associated with severe DENV infection.

Site directed mutagenesis studies on HIV-1 reverse transcriptase (RT) shed light on the mechanism of action of a new Ribonuclease H/ DNA polymerase RT dual inhibitor

Background The viral reverse transcriptase (RT) is a major target in the therapeutic strategy against the human immunodeficiency virus (HIV). This multifunctional enzyme performs the viral genome replication by two associated functions: DNA polymerase and ribonuclease H (RNase H). Both activities are essential for viral replication and validated targets. Currently, none of the approved antiviral drugs acts on both activities, although few classes of compounds have been reported to inhibit both RT functions in vitro [1]. In a previous virtual screening [2] we identified a new isatine-based scaffold for dual functions RT inhibitors that was further derivatized.

Virus-like particles for enterovirus 71 produced from Saccharomyces cerevisiae potently elicits protective immune responses in mice

Human Enterovirus 71 (EV71) is recognized as the leading causative agent of hand-foot-and-mouth disease (HFMD) in the Asia-Pacific region in recent years. There are still no approved antiviral drugs or vaccines against EV71 infection yet. In this study, we have developed an advanced platform for production of the virus-like particles (VLPs) for EV71 in Saccharomyces Cerevisiae by co-expressing P1 and 3CD genes of EV71. These VLPs exhibited similar morphology and protein composition as EV71 empty particles produced from EV71-infected cells. Immunization with VLPs in mice elicited robust neutralization antibodies against EV71 and potent cellular immune response. In vivo challenge experiments showed that the immune sera induced by VLP conferred protection in neonate mice against lethal EV71 challenge. Together, our study indicated that VLP from yeast is another potential vaccine candidate against EV71 infection.

The antiviral effect of 7-hydroxyisoflavone against Enterovirus 71 in vitro

Enterovirus 71 (EV71) is the major causative agent of hand foot and mouth disease. And EV71 causes epidemics worldwide, particularly in the Asia–Pacific region. Unfortunately, currently there is no approved vaccine or antiviral drug for EV71-induced disease prevention and therapy. In screening for anti-EV71 candidates, we found that 7-hydroxyisoflavone was active against EV71. 7-Hydroxyisoflavone exhibited strong antiviral activity against three different EV71 strains. The 50% inhibitory concentration range was between 3.25 and 4.92 μM by cytopathic effect assay. 7-Hydroxyisoflavone could reduce EV71 viral RNA and protein synthesis in a dose-dependent manner. Time course study showed that treatment of Vero cells with 7-hydroxyisoflavone at indicated times after EV71 inoculation (0–6 h) resulted in significant antiviral activity. Results showed that 7-hydroxyisoflavone acted at an early step of EV71 replication. 7-Hydroxyisoflavone also exhibited strong antiviral activity against coxsackievirus B2, B3, and B6. In short, 7-hydroxyisoflavone may be used as a lead compound for anti-EV71 drug development.

In vitro evaluation of the antiviral activity of heparan sulfate mimetic compounds against Enterovirus 71

Enterovirus 71 (EV71) is the causative agent of hand foot and mouth disease (HFMD) and can also cause fatal neurological complications for which currently there is no vaccine or approved antiviral drug. Despite suggestions that heparan sulfate (HS)-like compounds are effective antivirals against various viruses, no research has been undertaken to examine their effects upon EV71. Therefore, this study aimed to investigate the in vitro anti-EV71 effect of HS mimetics (heparin, heparan sulfate, and pentosan polysulfate). The results revealed that all of the compounds exhibited significant antiviral actions (p<0.05) against EV71 at concentrations less than 250μg/mL, compared to virus control and positive control, ribavirin. Among the compounds, heparin exhibited the most potent antiviral activity, as 7.81μg/mL of it prevented the infection by more than 90% (p<0.05). Assays to reveal the mode of action revealed that all of the compounds were capable of exerting antiviral activity through hindrance of viral attachment to the cells. In addition, some of the compounds could inhibit viral replication when added to cells 1h before infection, but none significantly reduced viral penetration. Overall, this research revealed that HS mimetic compounds could inhibit EV71 infection, and that HS may be involved in EV71–host cell interactions, as the virus binding to the host cells was significantly hindered by the HS-like compounds but not by ribavirin. Thus, further investigations to discover the molecular mechanisms underlying the anti-EV71 action of HS-like compounds are warranted.

Inhibitors of Dengue virus and West Nile virus proteases based on the aminobenzamide scaffold

Dengue and West Nile viruses (WNV) are mosquito-borne members of flaviviruses that cause significant morbidity and mortality. There is no approved vaccine or antiviral drugs for human use to date. In this study, a series of functionalized meta and para aminobenzamide derivatives were synthesized and subsequently screened in vitro against Dengue virus and West Nile virus proteases. Four active compounds were identified which showed comparable activity toward the two proteases and shared in common a meta or para(phenoxy)phenyl group. The inhibition constants (Ki) for the most potent compound 7n against Dengue and West Nile virus proteases were 8.77 and 5.55μM, respectively. The kinetics data support a competitive mode of inhibition of both proteases by compound 7n. This conclusion is further supported by molecular modeling. This study reveals a new chemical scaffold which is amenable to further optimization to yield potent inhibitors of the viral proteases via the combined utilization of iterative medicinal chemistry/structure–activity relationship studies and in vitro screening.