scholarly journals Repurposing of drugs as novel influenza inhibitors from clinical gene expression infection signatures

2018 ◽  
Author(s):  
Andrés Pizzorno ◽  
Olivier Terrier ◽  
Claire Nicolas de Lamballerie ◽  
Thomas Julien ◽  
Blandine Padey ◽  
...  
Keyword(s):  
Influenza Virus ◽  
Phase Ii ◽  
Cholesterol Metabolism ◽  
Ex Vivo ◽  
Drug Repurposing ◽  
Influenza Viruses ◽  
Bioactive Molecules ◽  
Computational Screening

AbstractBackground:Influenza virus infections remain a major and recurrent public health burden. The intrinsic ever-evolving nature of this virus, the suboptimal efficacy of current influenza inactivated vaccines, as well as the emergence of resistance against a limited antiviral arsenal, highlight the critical need for novel therapeutic approaches. In this context, the aim of this study was to develop and validate an innovative strategy for drug repurposing as host-targeted inhibitors of influenza viruses and the rapid evaluation of the most promising candidates in Phase II clinical trials.Methods:We exploited in vivo global transcriptomic signatures of infection directly obtained from a patient cohort to determine a shortlist of already marketed drugs with newly identified, host-targeted inhibitory properties against influenza virus. The antiviral potential of selected repurposing candidates was further evaluated in vitro, in vivo and ex vivo.Results:Our strategy allowed the selection of a shortlist of 35 high potential candidates out of a rationalized computational screening of 1,309 FDA-approved bioactive molecules, 31 of which were validated for their significant in vitro antiviral activity. Our in vivo and ex vivo results highlight diltiazem, a calcium channel blocker currently used in the treatment of hypertension, as a promising option for the treatment of influenza infections. Additionally, transcriptomic signature analysis further revealed the so far undescribed capacity of diltiazem to modulate the expression of specific genes related to the host antiviral response and cholesterol metabolism. Finally, combination treatment with diltiazem and virus-targeted oseltamivir neuraminidase inhibitor further increased antiviral efficacy, prompting rapid authorization for the initiation of a Phase II clinical trial.Conclusions:This original, host-targeted, drug repurposing strategy constitutes an effective and highly reactive process for the rapid identification of novel anti-infectious drugs, with potential major implications for the management of antimicrobial resistance and the rapid response to future epidemic or pandemic (re)emerging diseases for which we are still disarmed.

Abstract 14532: Computational Drug Repurposing Identifies a Piperlongumine Analog That Controls a Gstp1-iscu Pathogenic Axis in Pulmonary Hypertension

Circulation ◽  
2020 ◽  
Vol 142 (Suppl_3) ◽  
Author(s):  
Jimin Yang
Keyword(s):  
Drug Repurposing ◽  
Gene Clusters ◽  
Loss Of Function ◽  
Sequencing Data ◽  
Glutathione S Transferase ◽  
Computational Screening ◽  
Therapeutic Development ◽  
Pulmonary Arterial

Background and Hypothesis: Pulmonary arterial hypertension (PAH) is an incurable vascular disease for which chemotherapies are being considered for therapeutic development. There is no method reported to date for effective computational screening of these drugs for this disease. Big data analyses that leverage the molecular parallels between cancer and PH may define novel pathogenic mechanisms and facilitate repurposing of chemotherapies for PAH. More specifically, while functional deficiency of the iron-sulfur (Fe-S) biogenesis gene ISCU and oxidative metabolism in human pulmonary arterial endothelial cells (PAECs) is known to drive PAH, the pathogenic regulation of ISCU is not fully defined, and no tailored drugs have been identified to bolster ISCU activity. Methods and Results: We applied a computational algorithm EDDY (Evaluating Differential DependencY), which analyzes RNA sequencing data from 810 cancer cell lines exposed to 368 small molecules, in order to identify chemotherapeutics that depended upon rewired PH-related gene clusters. The top ranked drug was a piperlongumine (PL) analog (BRD2889) that was predicted to extensively rewire dependencies across PH gene clusters, mediated by ISCU. In vitro, coupling gain- and loss-of-function analyses of GSTP1 with BRD2889 exposure in PAECs, we found that BRD2889 inhibits glutathione S-transferase P1 (GSTP1), an enzyme which in turn catalyzes ISCU glutathionylation and increases its stability in hypoxia. Consequently, BRD2889 and GSTP1 knockdown phenocopy one another by increasing Fe-S-dependent Complex I activity and mitochondrial oxygen consumption while ameliorating pathogenic apoptosis. Consistent with these computational and in vitro results, in a mouse model of PAH (IL-6 transgenic mice in hypoxia), BRD2889 improved hemodynamic and molecular disease manifestations in vivo. Conclusions: Using a novel computational platform, we identified a coordinated connection between BRD342289 and GSTP1-ISCU axis, crucial to PAEC metabolism. This study offers insight to fundamental PH pathobiology and sets the stage for accelerated repurposing of chemotherapies such as BRD342289 in PH.

scholarly journals N-linked glycosylation plays an important role in budding of NA protein and virulence of influenza viruses

2020 ◽  
Author(s):  
Danqi Bao ◽  
Ruixue Xue ◽  
Min Zhang ◽  
Chenyang Lu ◽  
Tianxin Ma ◽  
...  
Keyword(s):  
Influenza Virus ◽  
Virus Replication ◽  
Immune Escape ◽  
H1n1 Influenza ◽  
Influenza Viruses ◽  
H1n1 Influenza Virus ◽  
Protein Loss ◽  
Knock Out

Neuraminidase (NA) has multiple functions in the life cycle of influenza virus, especially in the late stage of virus replication. Both of Hemagglutinin (HA) and NA are highly glycosylated proteins. N-linked glycosylation (NLG) of HA has been reported to contribute to immune escape and virulence of influenza viruses. However, the function of NLG of NA remains largely unclear. In this study, we found that NLG is critical for budding ability of NA. Tunicamycin treatment or NLG knock-out significantly inhibited the budding of NA. Further studies showed that the NLG knock-out caused attenuation of virus in vitro and in vivo. Notably the NLG at 219 position plays an important role in budding, replication, and virulence of H1N1 influenza virus. To explore the underlying mechanism, unfolded protein response (UPR) was determined in NLG knock-out NA overexpressed cells, which showed that the mutant NA was mainly located in ER, and the UPR markers BIP and p-eIF2α were upregulated, and XBP1 was downregulated. All the results indicated that NLG knock-out NA was stacked in ER and triggered UPR, which might shut down the budding process of NA. Overall, the study shed light on the function of NLG of NA in virus replication and budding. IMPORTANCE NA is a highly glycosylated protein. Nevertheless, how the NLG affects the function of NA protein remains largely unclear. In this study, we found that NLG plays important roles in budding and Neuraminidase activity of NA protein. Loss of NLG attenuated viral budding and replication. Especially the 219 NLG site mutation significantly attenuated the replication and virulence of H1N1 influenza virus in vitro and in vivo, which suggested that NLG of NA protein is a novel virulence marker for influenza viruses.

scholarly journals High-Yield Expression and Purification of Recombinant Influenza Virus Proteins from Stably-Transfected Mammalian Cell Lines

Vaccines ◽  
2020 ◽  
Vol 8 (3) ◽  
pp. 462
Author(s):  
Jeffrey W. Ecker ◽  
Greg A. Kirchenbaum ◽  
Spencer R. Pierce ◽  
Amanda L. Skarlupka ◽  
Rodrigo B. Abreu ◽  
...  
Keyword(s):  
Influenza Virus ◽  
Cell Lines ◽  
Mammalian Cell ◽  
Recombinant Proteins ◽  
Influenza Viruses ◽  
Scale Production ◽  
Vaccine Platform ◽  
Mammalian Cell Lines

Influenza viruses infect millions of people each year, resulting in significant morbidity and mortality in the human population. Therefore, generation of a universal influenza virus vaccine is an urgent need and would greatly benefit public health. Recombinant protein technology is an established vaccine platform and has resulted in several commercially available vaccines. Herein, we describe the approach for developing stable transfected human cell lines for the expression of recombinant influenza virus hemagglutinin (HA) and recombinant influenza virus neuraminidase (NA) proteins for the purpose of in vitro and in vivo vaccine development. HA and NA are the main surface glycoproteins on influenza virions and the major antibody targets. The benefits for using recombinant proteins for in vitro and in vivo assays include the ease of use, high level of purity and the ability to scale-up production. This work provides guidelines on how to produce and purify recombinant proteins produced in mammalian cell lines through either transient transfection or generation of stable cell lines from plasmid creation through the isolation step via Immobilized Metal Affinity Chromatography (IMAC). Collectively, the establishment of this pipeline has facilitated large-scale production of recombinant HA and NA proteins to high purity and with consistent yields, including glycosylation patterns that are very similar to proteins produced in a human host.

scholarly journals LACK OF IDENTITY IN NEUTRALIZING AND HEMAGGLUTINATION-INHIBITING ANTIBODIES AGAINST INFLUENZA VIRUSES

1950 ◽  
Vol 91 (1) ◽  
pp. 65-86 ◽  
Author(s):  
Duard L. Walker ◽  
Frank L. Horsfall
Keyword(s):  
Influenza Virus ◽  
Hemagglutination Inhibition ◽  
Neutralizing Antibodies ◽  
Neutralizing Antibody ◽  
Immune Serum ◽  
Influenza Viruses ◽  
In Ovo ◽  
Neutralization Line

There is an exponential linear relationship between the quantity of influenza virus neutralized and the quantity of immune serum employed in in ovo neutralization. The slope of the neutralization line is extremely steep. The concentration of neutralizing antibody can be measured with considerable precision in ovo if the constant virus-varying serum technique is utilized. The amounts of hemagglutination-inhibiting and neutralizing antibodies which are absorbed by a given quantity of influenza virus (PR8) were found to be predictable and the degree of reactivity of these two antibodies was shown to be directly related to the extent of immunization. It was demonstrated that there are marked discrepancies in correlation between antibody titers obtained by in vitro hemagglutination-inhibition and in vivo neutralization techniques and that neutralizing antibody is preferentially absorbed by a given quantity of virus. Inasmuch as the results were found not to be attributable to peculiarities of the techniques employed, it appears that the antibodies measured by hemagglutination-inhibition in vitro and by neutralization in vivo are not identical.

scholarly journals Ethanol Extract of Caesalpinia decapetala Inhibits Influenza Virus Infection In Vitro and In Vivo

Viruses ◽  
2020 ◽  
Vol 12 (5) ◽  
pp. 557 ◽  
Author(s):  
Li Zhang ◽  
Jungang Chen ◽  
Chang Ke ◽  
Haiwei Zhang ◽  
Shoujun Zhang ◽  
...  
Keyword(s):  
Influenza Virus ◽  
Influenza A ◽  
Antiviral Agents ◽  
Influenza Virus Infection ◽  
Virus Titer ◽  
Ethanol Extract ◽  
Influenza Viruses ◽  
Virus Infections

Influenza virus infections can lead to viral pneumonia and acute respiratory distress syndrome in severe cases, causing significant morbidity and mortality and posing a great threat to human health. Because of the diversity of influenza virus strains and drug resistance to the current direct antiviral agents, there have been no effective drugs as yet to cure all patients infected by influenza viruses. Natural products from plants contain compounds with diverse structures that have the potential to interact with multiple host and virus factors. In this study, we identified the ethanol extract of Caesalpinia decapetala (Roth) Alston (EEC) as an inhibitor against the replication of a panel of influenza A and B viruses both on human pulmonary epithelial A549 and human monocytic U937 cells. The animal study revealed that EEC administration reduces the weight loss and improves the survival rate of mice infected with lethal influenza virus. Also, EEC treatment attenuated lung injury and reduced virus titer significantly. In conclusion, we showed that EEC has antiviral activity both in vitro and in vivo, suggesting that the plant C. decapetala has the potential to be further developed as a resource of new anti-influenza drugs.

scholarly journals Cross-Protection of Chicken Immunoglobulin Y Antibodies against H5N1 and H1N1 Viruses Passively Administered in Mice

2011 ◽  
Vol 18 (7) ◽  
pp. 1083-1090 ◽  
Author(s):  
Michael G. Wallach ◽  
Richard J. Webby ◽  
Fakhrul Islam ◽  
Stephen Walkden-Brown ◽  
Eva Emmoth ◽  
...  
Keyword(s):  
Influenza Virus ◽  
Influenza Pandemic ◽  
Influenza Virus Infection ◽  
Influenza Viruses ◽  
Antigenic Drift ◽  
Virus Infectivity ◽  
Lethal Challenge ◽  
H5n1 Influenza

ABSTRACTInfluenza viruses remain a major threat to global health due to their ability to undergo change through antigenic drift and antigenic shift. We postulated that avian IgY antibodies represent a low-cost, effective, and well-tolerated approach that can easily be scaled up to produce enormous quantities of protective antibodies. These IgY antibodies can be administered passively in humans (orally and intranasally) and can be used quickly and safely to help in the fight against an influenza pandemic. In this study, we raised IgY antibodies against H1N1, H3N2, and H5N1 influenza viruses. We demonstrated that, using whole inactivated viruses alone and in combination to immunize hens, we were able to induce a high level of anti-influenza virus IgY in the sera and eggs, which lasted for at least 2 months after two immunizations. Furthermore, we found that by use ofin vitroassays to test for the ability of IgY to inhibit hemagglutination (HI test) and virus infectivity (serum neutralization test), IgYs inhibited the homologous as well as in some cases heterologous clades and strains of viruses. Using anin vivomouse model system, we found that, when administered intranasally 1 h prior to infection, IgY to H5N1 protected 100% of the mice against lethal challenge with H5N1. Of particular interest was the finding that IgY to H5N1 cross-protected against A/Puerto Rico/8/34 (H1N1) bothin vitroandin vivo. Based on our results, we conclude that anti-influenza virus IgY can be used to help prevent influenza virus infection.

scholarly journals 5,6-Epoxycholesterol Isomers Induce Oxiapoptophagy in Myeloma Cells

Cancers ◽  
2021 ◽  
Vol 13 (15) ◽  
pp. 3747
Author(s):  
Oumaima Jaouadi ◽  
Inès Limam ◽  
Mohamed Abdelkarim ◽  
Emna Berred ◽  
Ahlem Chahbi ◽  
...  
Keyword(s):  
Caspase 3 ◽  
Potential Role ◽  
Cholesterol Metabolism ◽  
Ex Vivo ◽  
Myeloma Cell ◽  
Bioactive Molecules ◽  
Myeloma Cells ◽  
Acquired Drug Resistance ◽  
Anti Tumor Activity

Multiple myeloma (MM) is an incurable plasma cell malignancy with frequent patient relapse due to innate or acquired drug resistance. Cholesterol metabolism is reported to be altered in MM; therefore, we investigated the potential anti-myeloma activity of two cholesterol derivatives: the 5,6 α- and 5,6 β-epoxycholesterol (EC) isomers. To this end, viability assays were used, and isomers were shown to exhibit important anti-tumor activity in vitro in JJN3 and U266 human myeloma cell lines (HMCLs) and ex vivo in myeloma patients’ sorted CD138+ malignant cells. Moreover, we confirmed that 5,6 α-EC and 5,6 β-EC induced oxiapoptophagy through concomitant oxidative stress and caspase-3-mediated apoptosis and autophagy. Interestingly, in combination treatment a synergistic interaction was observed between 5,6 α-EC and 5,6 β-EC on myeloma cells. These data highlight a striking anti-tumor activity of 5,6 α-EC and 5,6 β-EC bioactive molecules against human myeloma cells, paving the way for their potential role in future therapeutic strategies in MM.

scholarly journals Oseltamivir-Ribavirin Combination Therapy for Highly Pathogenic H5N1 Influenza Virus Infection in Mice

2008 ◽  
Vol 52 (11) ◽  
pp. 3889-3897 ◽  
Author(s):  
Natalia A. Ilyushina ◽  
Alan Hay ◽  
Neziha Yilmaz ◽  
Adrianus C. M. Boon ◽  
Robert G. Webster ◽  
...  
Keyword(s):  
Influenza Virus ◽  
Influenza Virus Infection ◽  
Antiviral Effect ◽  
Influenza Viruses ◽  
Oseltamivir Carboxylate ◽  
Highly Pathogenic ◽  
H5n1 Influenza ◽  
Pathogenic H5n1

ABSTRACT We studied the effects of a neuraminidase inhibitor (oseltamivir) and an inhibitor of influenza virus polymerases (ribavirin) against two highly pathogenic H5N1 influenza viruses. In vitro, A/Vietnam/1203/04 virus (clade 1) was highly susceptible to oseltamivir carboxylate (50% inhibitory concentration [IC50] = 0.3 nM), whereas A/Turkey/15/06 virus (clade 2.2) had reduced susceptibility (IC50 = 5.5 nM). In vivo, BALB/c mice were treated with oseltamivir (1, 10, 50, or 100 mg/kg of body weight/day), ribavirin (37.5, 55, or 75 mg/kg/day), or the combination of both drugs for 8 days, starting 4 h before virus inoculation. Monotherapy produced a dose-dependent antiviral effect against the two H5N1 viruses in vivo. Three-dimensional analysis of the drug-drug interactions revealed that oseltamivir and ribavirin interacted principally in an additive manner, with several exceptions of marginal synergy or marginal antagonism at some concentrations. The combination of ribavirin at 37.5 mg/kg/day and oseltamivir at 1 mg/kg/day and the combination of ribavirin at 37.5 mg/kg/day and oseltamivir at 10 mg/kg/day were synergistic against A/Vietnam/1203/04 and A/Turkey/15/06 viruses, respectively. These optimal oseltamivir-ribavirin combinations significantly inhibited virus replication in mouse organs, prevented the spread of H5N1 viruses beyond the respiratory tract, and abrogated the cytokine response (P < 0.01). Importantly, we observed clear differences between the efficacies of the drug combinations against two H5N1 viruses: higher doses were required for the protection of mice against A/Turkey/15/06 virus than for the protection of mice against A/Vietnam/1203/04 virus. Our preliminary results suggest that oseltamivir-ribavirin combinations can have a greater or lesser antiviral effect than monotherapy, depending on the H5N1 virus and the concentrations used.

scholarly journals Identification of a Type-Specific Promoter Element That Differentiates between Influenza A and B Viruses

2019 ◽  
Vol 93 (23) ◽  
Author(s):  
Shuman Gao ◽  
Wenyu Zhang ◽  
Congyu Lu ◽  
Mengmeng Cao ◽  
Shan Cen ◽  
...  
Keyword(s):  
Influenza Virus ◽  
Polymerase Activity ◽  
Replication Initiation ◽  
Influenza Viruses ◽  
Promoter Element ◽  
Promoter Elements ◽  
Promoter Sequences ◽  
Functional Features

ABSTRACT Type A and type B influenza viruses (FluA and FluB viruses) are two major human pathogens that share common structural and functional features. FluA and FluB viruses can reassort within each type but never between the types. Here, we bioinformatically analyzed all promoter sequences of FluA and FluB viruses and confirmed the presence of the type-specific promoter elements. We then studied the promoter elements with cell-based in vivo assays and an in vitro replication initiation assay. Our results identified, for the first time, a type-specific promoter element—the nucleotide at position 5 in the 3′ end of the viral RNA (vRNA)—that plays a key role(s) in modulating polymerase activity in a type-specific manner. Interestingly, swapping the promoter element between FluA and FluB recombinant viruses showed different tolerances: the replacement of FluA virus-specific U5 with FluB virus-specific C5 in influenza virus A/WSN/33 (H1N1) could be reverted to U5 after 2 to 3 passages, while the replacement of FluB virus-specific C5 with FluA virus-specific U5 in influenza virus B/Yamagata/88 could be maintained, but with significantly reduced replication efficiency. Therefore, our findings indicate that the nucleotide variation at position 5 in the 3′ end of the vRNA promoter between FluA and FluB viruses contributes to their RNP incompatibility, which may shed new light on the mechanisms of intertypic exclusion of reassortment between FluA and FluB viruses. IMPORTANCE Genetic reassortment of influenza virus plays a key role in virus evolution and the emergence of pandemic strains. The reassortment occurs extensively within either FluA or FluB viruses but never between them. Here, we bioinformatically compared available promoter sequences of FluA and FluB viruses and confirmed the presence of the type-specific promoter elements. Our in vivo and in vitro mutagenesis studies showed that a type-specific promoter element—the nucleotide at position 5 in the 3′ end of vRNA promoters—plays key roles in modulating polymerase activity. Interestingly, FluA and FluB viruses showed different tolerances upon key promoter element swapping in the context of virus infections. We concluded that the nucleotide at position 5 in the 3′ end of the vRNA promoters of FluA and FluB viruses is a critical type-specific determinant. This work has implications for further elucidating the mechanisms of the intertypic exclusion of reassortment between FluA and FluB viruses.

scholarly journals Promethazine Exhibits Antiparasitic Properties In Vitro and Reduces Worm Burden, Egg Production, Hepatomegaly, and Splenomegaly in a Schistosomiasis Animal Model

2019 ◽  
Vol 63 (12) ◽  
Author(s):  
Daniel B. Roquini ◽  
Ramon M. Cogo ◽  
Ana C. Mengarda ◽  
Susana F. Mazloum ◽  
Cristiane S. Morais ◽  
...  
Keyword(s):  
Animal Model ◽  
Worm Burden ◽  
Egg Production ◽  
Ex Vivo ◽  
Anticholinergic Drug ◽  
Drug Repurposing ◽  
Content Type ◽  
Parasite Motility

ABSTRACT The treatment and control of schistosomiasis, a neglected disease that affects more than 200 million people worldwide, rely on the use of a single drug, praziquantel. A vaccine has yet to be developed, and since new drug design and development is a lengthy and costly process, drug repurposing is a promising strategy. In this study, the efficacy of promethazine, a first-generation antihistamine, was evaluated against Schistosoma mansoni ex vivo and in a murine model of schistosomiasis. In vitro assays demonstrated that promethazine affected parasite motility and viability, and it induced severe tegumental damage in schistosomes. The 50% lethal concentration (LC50) of the drug was 5.84 μM. Similar to promethazine, schistosomes incubated with atropine, a classical anticholinergic drug, displayed reduced motor activity. In an animal model, promethazine treatment was introduced at an oral dose of 100 mg/kg of body weight for five successive days at different intervals from the time of infection for the evaluation of the stage-specific susceptibility (prepatent and patent infections). Various parasitological criteria indicated the following in vivo antischistosomal effects of promethazine: there were significant reductions in worm burden, egg production, hepatomegaly, and splenomegaly. The highest worm burden reduction was achieved with promethazine in patent infections (>90%). Taken together, considering the importance of the repositioning of drugs in infectious diseases, especially those related to poverty, our data revealed the possibility of promethazine repositioning as an antischistosomal agent.

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