Broad-spectrum Antiviral Compounds Research Articles

In vitro broad-spectrum antiviral activity of MIT-001, a mitochondria-targeted reactive oxygen species scavenger, against severe acute respiratory syndrome coronavirus 2 and multiple zoonotic viruses

The COVID-19 pandemic caused by SARS-CoV-2 becomes a serious threat to global health and requires the development of effective antiviral therapies. Current therapies that target viral proteins have limited efficacy with side effects. In this study, we investigated the antiviral activity of MIT-001, a small molecule reactive oxygen species (ROS) scavenger targeting mitochondria, against SARS-CoV-2 and other zoonotic viruses in vitro. The antiviral activity of MIT-001 was quantified by RT-qPCR and plaque assay. We also evaluated the functional analysis of MIT-001 by JC-1 staining to measure mitochondrial depolarization, total RNA sequencing to investigate gene expression changes, and immunoblot to quantify protein expression levels. The results showed that MIT-001 effectively inhibited the replication of B.1.617.2 and BA.1 strains, Zika virus, Seoul virus, and Vaccinia virus. Treatment with MIT-001 restored the expression of heme oxygenase-1 (HMOX1) and NAD(P)H: quinone oxidoreductase 1 (NqO1) genes, anti-oxidant enzymes reduced by SARS-CoV-2, to normal levels. The presence of MIT-001 also alleviated mitochondrial depolarization caused by SARS-CoV-2 infection. These findings highlight the potential of MIT-001 as a broad-spectrum antiviral compound that targets for zoonotic RNA and DNA viruses, providing a promising therapeutic approach to combat viral infection.

Identification of tanshinone I as cap-dependent endonuclease inhibitor with broad-spectrum antiviral effect.

The spread of avian-borne, tick-borne, and rodent-borne pathogens has the potential to pose a serious threat to human health, and candidate vaccines as well as therapeutics for these pathogens are urgently needed. Tanshinones, especially tanshinone I, were identified as a cap-dependent endonuclease inhibitor with broad-spectrum antiviral effects on negative-stranded, segmented RNA viruses including bandavirus, orthomyxovirus, and arenavirus from natural products, implying an important resource of candidate antivirals from the traditional Chinese medicines. This study supplies novel candidate antivirals for the negative-stranded, segmented RNA virus and highlights the endonuclease involved in the cap-snatching process as a reliable broad-spectrum antiviral target.

Inhibitory Potential of Chitosan Derivatives against Severe Acute Respiratory Syndrome Coronavirus 2: An In Silico Prospective

The present work was designed to investigate the antiviral potential of novel monomeric and oligomeric chitosan derivatives through in silico approaches. The goal was to identify potent broad-spectrum antiviral compounds as promising drug candidates against severe acute respiratory syndrome coronavirus 2 and understand their mode of action. Chitosan biopolymer and its derivatives were virtually screened against the spike glycoprotein and human angiotensin-converting enzyme 2 (ACE2) receptor of novel coronavirus-19. Hydroxypropyl trimethyl ammonium chloride chitosan (HTCC), a polymeric chitosan, has been reported to interact with the corona viral spike (S) protein and blocks its interaction with the ACE2 receptor. The enhancement of antiviral activity relies on better biocompatibility, structural correlations, variation in the degree of deacetylation, and molecular weight of modified chitosan derivatives. The chitosan derivatives constructively interact with viral S protein. Among the chitosan derivatives, N-carboxymethyl chitosan (NCMC) displayed efficient binding affinity. Comparing NCMC to mHTCC, monomeric chitosan, for their interaction with the S protein, receptor binding domain site, and ACE2 receptor, NCMC displayed better binding affinity of −7.9, −6.3, and −7.4 with binding energies of −6.2, −4.8, and −5.5 kcal/mol, respectively. Furthermore, through flexible docking, the interactions of the S protein with ACE2 receptor and ligand mHTCC-S protein complex and NCMC-S protein complex with ACE2 receptor were calculated, showing an efficient reduction of binding energy from −901.2 kJ/mol to −765.06 kJ/mol and −814.72 kJ/mol, respectively. This points to the decrease binding affinity of the viral S protein for the ACE2 receptor in the presence of NCMC/mHTCC. For the first time, the computational study envisages the antiviral efficiency of NCMC, mHTCC, and biocompatible chitosan derivatives as a preventive intervention against COVID-19.

NPP-669, a Novel Broad-Spectrum Antiviral Therapeutic with Excellent Cellular Uptake, Antiviral Potency, Oral Bioavailability, Preclinical Efficacy, and a Promising Safety Margin.

DNA viruses are responsible for many diseases in humans. Current treatments are often limited by toxicity, as in the case of cidofovir (CDV, Vistide), a compound used against cytomegalovirus (CMV) and adenovirus (AdV) infections. CDV is a polar molecule with poor bioavailability, and its overall clinical utility is limited by the high occurrence of acute nephrotoxicity. To circumvent these disadvantages, we designed nine CDV prodrug analogues. The prodrugs modulate the polarity of CDV with a long sulfonyl alkyl chain attached to one of the phosphono oxygens. We added capping groups to the end of the alkyl chain to minimize β-oxidation and focus the metabolism on the phosphoester hydrolysis, thereby tuning the rate of this reaction by altering the alkyl chain length. With these modifications, the prodrugs have excellent aqueous solubility, optimized metabolic stability, increased cellular permeability, and rapid intracellular conversion to the pharmacologically active diphosphate form (CDV-PP). The prodrugs exhibited significantly enhanced antiviral potency against a wide range of DNA viruses in infected human foreskin fibroblasts. Single-dose intravenous and oral pharmacokinetic experiments showed that the compounds maintained plasma and target tissue levels of CDV well above the EC50 for 24 h. These experiments identified a novel lead candidate, NPP-669. NPP-669 demonstrated efficacy against CMV infections in mice and AdV infections in hamsters following oral (p.o.) dosing at a dose of 1 mg/kg BID and 0.1 mg/kg QD, respectively. We further showed that NPP-669 at 30 mg/kg QD did not exhibit histological signs of toxicity in mice or hamsters. These data suggest that NPP-669 is a promising lead candidate for a broad-spectrum antiviral compound.

Rapid discovery and classification of inhibitors of coronavirus infection by pseudovirus screen and amplified luminescence proximity homogeneous assay

To identify potent antiviral compounds, we introduced a high-throughput screen platform that can rapidly classify hit compounds according to their target. In our platform, we performed a compound screen using a lentivirus-based pseudovirus presenting a spike protein of coronavirus, and we evaluated the hit compounds using an amplified luminescence proximity homogeneous assay (alpha) test with purified host receptor protein and the receptor binding domain of the viral spike. With our screen platform, we were able to identify both spike-specific compounds (class I) and broad-spectrum antiviral compounds (class II). Among the hit compounds, thiosemicarbazide was identified to be selective to the interaction between the viral spike and its host cell receptor, and we further optimized the binding potency of thiosemicarbazide through modification of the pyridine group. Among the class II compounds, we found raloxifene and amiodarone to be highly potent against human coronaviruses including Middle East respiratory syndrome coronavirus (MERS-CoV), severe acute respiratory syndrome coronavirus (SARS-CoV), and SARS-CoV-2. In particular, using analogs of the benzothiophene moiety, which is also present in raloxifene, we have identified benzothiophene as a novel structural scaffold for broad-spectrum antivirals. This work highlights the strong utility of our screen platform using a pseudovirus assay and an alpha test for rapid identification of potential antiviral compounds and their mechanism of action, which can lead to the accelerated development of therapeutics against newly emerging viral infections.

Cap-snatching inhibitors of influenza virus are inhibitory to the in vitro transcription of rice stripe virus

Rice stripe virus (RSV) is one of the most important viral pathogens of rice in East Asia. The transcription of RSV is initiated by cap-snatching, a mechanism shared by influenza virus (IFV). This lends the possibility that antiviral compounds targeting the cap-snatching of IFV, many of which have been commercially available, may inhibit RSV transcription. A convenient and inexpensive system allowing researchers to test this idea, however, has been unavailable to date. Here, we show that purified RSV performs transcription in vitro and the transcription was readily detectable by nested reverse transcription-polymerase chain reaction (RT-PCR). With this system, we tested the effects of 2,4-dioxo-4-phenylbutanoic acid (DPBA) and pimodivir, two well-known IFV cap-snatching inhibitors, as well as ribavirin, a broad-spectrum antiviral compound whose targets remain elusive. In reaction mixtures containing 2.5 ng/µL of purified RSV, DPBA and pimodivir abolished RSV transcription at a concentration of 10 and 100 µM, respectively. In contrast, no inhibitory effect was detected from ribavirin, even at a concentration as high as 400 µM. These results suggest that at least some cap-snatching inhibitors of IFV are inhibitory to RSV transcription. These compounds, which can be identified with the experimental system described here, may serve as starting points in developing antivirals against RSV or related plant viruses.

The synergy between nucleotide biosynthesis inhibitors and antiviral nucleosides: New opportunities against viral infections?

5'-Phosphorylated nucleoside derivatives are molecules that can be found in all living organisms and viruses. Over the last century, the development of structural analogs that could disrupt the transcription and translation of genetic information culminated in the development of clinically relevant anticancer and antiviral drugs. However, clinically effective broad-spectrum antiviral compounds or treatments are lacking. This viewpoint proposes that molecules that inhibit nucleotide biosynthesis may sensitize virus-infected cells toward direct-acting antiviral nucleosides. Such potentially synergistic combinations might allow the repurposing of drugs, leading to the development of new combination therapies.

A hnRNPA2B1 agonist effectively inhibits HBV and SARS-CoV-2 omicron in vivo.

The twenty-first century has already recorded more than ten major epidemics or pandemics of viral disease, including the devastating COVID-19. Novel effective antivirals with broad-spectrum coverage are urgently needed. Herein, we reported a novel broad-spectrum antiviral compound PAC5. Oral administration of PAC5 eliminated HBV cccDNA and reduced the large antigen load in distinct mouse models of HBV infection. Strikingly, oral administration of PAC5 in a hamster model of SARS-CoV-2 omicron (BA.1) infection significantly decreases viral loads and attenuates lung inflammation. Mechanistically, PAC5 binds to a pocket near Asp49 in the RNA recognition motif of hnRNPA2B1. PAC5-bound hnRNPA2B1 is extensively activated and translocated to the cytoplasm where it initiates the TBK1-IRF3 pathway, leading to the production of type I IFNs with antiviral activity. Our results indicate that PAC5 is a novel small-molecule agonist of hnRNPA2B1, which may have a role in dealing with emerging infectious diseases now and in the future.

Rocaglates as Antivirals: Comparing the Effects on Viral Resistance, Anti-Coronaviral Activity, RNA-Clamping on eIF4A and Immune Cell Toxicity.

Rocaglates are potent broad-spectrum antiviral compounds with a promising safety profile. They inhibit viral protein synthesis for different RNA viruses by clamping the 5′-UTRs of mRNAs onto the surface of the RNA helicase eIF4A. Apart from the natural rocaglate silvestrol, synthetic rocaglates like zotatifin or CR-1-31-B have been developed. Here, we compared the effects of rocaglates on viral 5′-UTR-mediated reporter gene expression and binding to an eIF4A-polypurine complex. Furthermore, we analyzed the cytotoxicity of rocaglates on several human immune cells and compared their antiviral activities in coronavirus-infected cells. Finally, the potential for developing viral resistance was evaluated by passaging human coronavirus 229E (HCoV-229E) in the presence of increasing concentrations of rocaglates in MRC-5 cells. Importantly, no decrease in rocaglate-sensitivity was observed, suggesting that virus escape mutants are unlikely to emerge if the host factor eIF4A is targeted. In summary, all three rocaglates are promising antivirals with differences in cytotoxicity against human immune cells, RNA-clamping efficiency, and antiviral activity. In detail, zotatifin showed reduced RNA-clamping efficiency and antiviral activity compared to silvestrol and CR-1-31-B, but was less cytotoxic for immune cells. Our results underline the potential of rocaglates as broad-spectrum antivirals with no indications for the emergence of escape mutations in HCoV-229E.

Activity of Galidesivir in a Hamster Model of SARS-CoV-2.

Coronavirus disease 2019 (COVID-19) has claimed the lives of millions of people worldwide since it first emerged. The impact of the COVID-19 pandemic on public health and the global economy has highlighted the medical need for the development of broadly acting interventions against emerging viral threats. Galidesivir is a broad-spectrum antiviral compound with demonstrated in vitro and in vivo efficacy against several RNA viruses of public health concern, including those causing yellow fever, Ebola, Marburg, and Rift Valley fever. In vitro studies have shown that the antiviral activity of galidesivir also extends to coronaviruses. Herein, we describe the efficacy of galidesivir in the Syrian golden hamster model of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection. Treatment with galidesivir reduced lung pathology in infected animals compared with untreated controls when treatment was initiated 24 h prior to infection. These results add to the evidence of the applicability of galidesivir as a potential medical intervention for a range of acute viral illnesses, including coronaviruses.

Phycobilins as Potent Food Bioactive Broad-Spectrum Inhibitors Against Proteases of SARS-CoV-2 and Other Coronaviruses: A Preliminary Study.

In the 21st century, we have witnessed three coronavirus outbreaks: SARS in 2003, MERS in 2012, and the ongoing pandemic coronavirus disease 2019 (COVID-19). The search for efficient vaccines and development and repurposing of therapeutic drugs are the major approaches in the COVID-19 pandemic research area. There are concerns about the evolution of mutant strains (e.g., VUI – 202012/01, a mutant coronavirus in the United Kingdom), which can potentially reduce the impact of the current vaccine and therapeutic drug development trials. One promising approach to counter the mutant strains is the “development of effective broad-spectrum antiviral drugs” against coronaviruses. This study scientifically investigates potent food bioactive broad-spectrum antiviral compounds by targeting main protease (Mpro) and papain-like protease (PLpro) proteases of coronaviruses (CoVs) using in silico and in vitro approaches. The results reveal that phycocyanobilin (PCB) shows potential inhibitor activity against both proteases. PCB had the best binding affinity to Mpro and PLpro with IC50 values of 71 and 62 μm, respectively. Also, in silico studies with Mpro and PLpro enzymes of other human and animal CoVs indicate broad-spectrum inhibitor activity of the PCB. As with PCB, other phycobilins, such as phycourobilin (PUB), phycoerythrobilin (PEB), and phycoviolobilin (PVB) show similar binding affinity to SARS-CoV-2 Mpro and PLpro.

Beneficial effect of Arbidol in the management of COVID-19 infection.

This study analyzed the effect of Arbidol, a broad-spectrum antiviral compound, on the outcomes of COVID-19 patients. Records of 252 COVID-19 patients were retrospectively analyzed from February 13 to February 29, 2020 in 4 inpatient wards in the Cancer Center, Union Hospital, Tongji Medical College of Huazhong University of Science and Technology, Wuhan, China. The rate of clinical improvement was significantly greater among patients treated with Arbidol than among those who did not receive Arbidol (86.8% vs. 54.2%). In moderately and severely ill patients, the clinical improvement rates in the Arbidol group were 95.6% and 81.7%, respectively, which was significantly higher than in the no-Arbidol group (66.6% and 53.8%). Among critically ill patients, however, there was no significant difference. The levels of hypersensitive C-reactive protein, lactate dehydrogenase, D-dimer, IL-6, and IL-10 were increased in non-improved patients but declined during treatment in the improved patients. This suggests these mediators are associated with the disease severity and could potentially serve as prognostic markers. Moreover, our data demonstrate that Arbidol is effective in the treatment of COVID-19 patients and may serve as a cost-effective antiviral treatment strategy for patients with moderate to severe COVID-19 symptoms.

Berberine and Obatoclax Inhibit SARS-Cov-2 Replication in Primary Human Nasal Epithelial Cells In Vitro.

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) emerged as a new human pathogen in late 2019 and it has infected over 100 million people in less than a year. There is a clear need for effective antiviral drugs to complement current preventive measures, including vaccines. In this study, we demonstrate that berberine and obatoclax, two broad-spectrum antiviral compounds, are effective against multiple isolates of SARS-CoV-2. Berberine, a plant-derived alkaloid, inhibited SARS-CoV-2 at low micromolar concentrations and obatoclax, which was originally developed as an anti-apoptotic protein antagonist, was effective at sub-micromolar concentrations. Time-of-addition studies indicated that berberine acts on the late stage of the viral life cycle. In agreement, berberine mildly affected viral RNA synthesis, but it strongly reduced infectious viral titers, leading to an increase in the particle-to-pfu ratio. In contrast, obatoclax acted at the early stage of the infection, which is in line with its activity to neutralize the acidic environment in endosomes. We assessed infection of primary human nasal epithelial cells that were cultured on an air-liquid interface and found that SARS-CoV-2 infection induced and repressed expression of specific sets of cytokines and chemokines. Moreover, both obatoclax and berberine inhibited SARS-CoV-2 replication in these primary target cells. We propose berberine and obatoclax as potential antiviral drugs against SARS-CoV-2 that could be considered for further efficacy testing.

Different Cases of SARS-CoV-2 Infection and Its Impact on Health and Economy with Special Emphasis on Antiviral Drug Targets

COVID-19 caused by SARS-CoV-2 has not only issued a pandemic situation but also lead to economic disaster and unprecedented health emergency. Being a close relative of Bat corona-virus, SARS and MERS it’s structural and sequence similarity has abled scientists for repurposing of popular drugs like hydroxychloroquine, cloroquine and also scavenging for lead molecules by in-silico and in-vivo or in-vitro approach. The potent drug targets are ACE2; spike protein binding receptor to host cell surface, Mpro; proteo-lytic polyprotein processing enzymes needed for virion maturation and RdRp; RNA dependent RNA polymerase needed for RNA replication. The recent trend focuses on the fact that cocktail of anti-corona virus treatment will be available soon by broad spectrum antiviral compounds. It takes time to develop such drug targets till then social distancing and following of hygiene are the only way to thrive well. This article incorporates the present World scenario related to COVID infection, focuses on its origin and also future possibilities for a COVID free future.

Recent research for COVID-19 drugs and therapies

<p indent="0mm">The sudden outbreak of COVID-19 is causing a severe health crisis to China and the world. The rapid spread coronavirus worldwide highlights the urgent need to find effective drugs and therapies to fight the COVID-19 and related diseases such as acute respiratory disease. Based on the current knowledge on the structure, pathogenic mechanism and pharmacy of coronavirus and previous understanding and experience in treating similar diseases, researchers are testing and screening the approved broad-spectrum antiviral drugs and compounds, and investigating new specific drugs aiming at curing COVID-19. Facing the coronavirus pandemic, it is essential methods and strategies to review existing broad-spectrum antiviral drugs and screen possible effective medicines through some compounds database or repository of structured medical information. Till now, scientists have proposed hundreds of possible drug candidates in a short period of time. This article summarized the potential drugs and treatments proposed by scientists, pharmaceutical industries, and clinicians through literature survey on PubMed—a medical academic database, two clinical trials databases, and internet information published before February 16th, 2020. Then a system review and analysis on the development situations and trends about COVID-19 drugs and therapies were carried out. According to the documents published, scientists and doctors have focused on several possible categories of drugs, including: (1) Chemical compounds, such as nucleoside analogues, protease inhibitors, and other antiviral drugs with multiple mechanisms; (2) biologics, such as monoclonal antibodies, interferon, peptides, plasma of recovered patients and some new kinds of therapies; (3) traditional Chinese medicines, such as some syndrome differentiation therapies. Now, nearly 100 agents or therapies have entered into the clinic research processes and some of them have arrived at the IV phase of clinical trials. Due to the considerable previous experiences and understands that have been accumulated in the researches on SARS-nCoV, Ebola, MERS, and AIDS, with the help of advanced biotechnology and medical technology, scientists can respond faster than ever before, and have investigated a variety of potential treatment options. For example, Remdesivir, which was originally targeted at Ebola virus, Kaletra, which has been used for the treatment of AIDS, Chloroquine, a typical drug used as antimalarial long time ago, glucocorticoids, a common therapies method used to combat SARS, washed microbiota transplantation and Mesenchymal Stem Cell, emerging biological therapies, and combination of multiple drugs or combination of traditional Chinese medicines and modern medicines, are bringing hope to treat COVID-19. Now scientists have proposed several drug candidates (e.g., Remdesivir, Kaletra, Arbidol and Chloroquine) that could be the potential anti-coronavirus agents. All of these drugs are under study in the late phases of clinical trials. While Remdesivir and Kaletra have already shown positive effect on COVID-19 related disease at personalized level. Traditional Chinese Medicines treatment with syndrome differentiation combined with modern medicines also shows a good effect. A considerable number of candidate drugs have shown good results<italic> in vitro</italic> cell tests on COVID-19. Currently, the follow-up animal tests, toxicology tests, and human clinical trials are in progress at various stages. After a period of clinical application research and further validation research on the clinical effect, safety, dosage and indications of these candidate drugs, it is expected that more drugs with significant efficacy will emerge. Finally, we will defeat COVID-19, and the public will live in a healthy and peaceful world.

Antiviral Activity of Benzavir-2 against Emerging Flaviviruses.

Most flaviviruses are arthropod-borne viruses, transmitted by either ticks or mosquitoes, and cause morbidity and mortality worldwide. They are endemic in many countries and have recently emerged in new regions, such as the Zika virus (ZIKV) in South-and Central America, the West Nile virus (WNV) in North America, and the Yellow fever virus (YFV) in Brazil and many African countries, highlighting the need for preparedness. Currently, there are no antiviral drugs available to treat flavivirus infections. We have previously discovered a broad-spectrum antiviral compound, benzavir-2, with potent antiviral activity against both DNA- and RNA-viruses. Our purpose was to investigate the inhibitory activity of benzavir-2 against flaviviruses. We used a ZIKV ZsGreen-expressing vector, two lineages of wild-type ZIKV, and other medically important flaviviruses. Benzavir-2 inhibited ZIKV derived reporter gene expression with an EC50 value of 0.8 ± 0.1 µM. Furthermore, ZIKV plaque formation, progeny virus production, and viral RNA expression were strongly inhibited. In addition, 2.5 µM of benzavir-2 reduced infection in vitro in three to five orders of magnitude for five other flaviviruses: WNV, YFV, the tick-borne encephalitis virus, Japanese encephalitis virus, and dengue virus. In conclusion, benzavir-2 was a potent inhibitor of flavivirus infection, which supported the broad-spectrum antiviral activity of benzavir-2.

Identification of Broad-Spectrum Antiviral Compounds by Targeting Viral Entry.

Viruses are a major threat to human health and economic well-being. In recent years Ebola, Zika, influenza, and chikungunya virus epidemics have raised awareness that infections can spread rapidly before vaccines or specific antagonists can be made available. Broad-spectrum antivirals are drugs with the potential to inhibit infection by viruses from different groups or families, which may be deployed during outbreaks when specific diagnostics, vaccines or directly acting antivirals are not available. While pathogen-directed approaches are generally effective against a few closely related viruses, targeting cellular pathways used by multiple viral agents can have broad-spectrum efficacy. Virus entry, particularly clathrin-mediated endocytosis, constitutes an attractive target as it is used by many viruses. Using a phenotypic screening strategy where the inhibitory activity of small molecules was sequentially tested against different viruses, we identified 12 compounds with broad-spectrum activity, and found a subset blocking viral internalisation and/or fusion. Importantly, we show that compounds identified with this approach can reduce viral replication in a mouse model of Zika infection. This work provides proof of concept that it is possible to identify broad-spectrum inhibitors by iterative phenotypic screenings, and that inhibition of host-pathways critical for viral life cycles can be an effective antiviral strategy.

Novel broad spectrum virucidal molecules against enveloped viruses.

Viral infections are an important cause of death worldwide. Unfortunately, there is still a lack of antiviral drugs or vaccines for a large number of viruses, and this represents a remarkable challenge particularly for emerging and re-emerging viruses. For this reason, the identification of broad spectrum antiviral compounds provides a valuable opportunity for developing efficient antiviral therapies. Here we report on a class of rhodanine and thiobarbituric derivatives displaying a broad spectrum antiviral activity against seven different enveloped viruses including an HSV-2 acyclovir resistant strain with favorable selectivity indexes. Due to their selective action on enveloped viruses and to their lipid oxidation ability, we hypothesize a mechanism on the viral envelope that affects the fluidity of the lipid bilayer, thus compromising the efficiency of virus-cell fusion and preventing viral entry.

Evaluation of broad-spectrum antiviral compounds against chikungunya infection using a phenotypic screening strategy

Chikungunya fever is an emerging disease and a significant public health problem in tropical countries. Recently reported outbreaks in Brazil in 2015 drew attention to the need to develop prevention and treatment options, as no antiviral chemotherapy or vaccines are currently available for this disease. Two strategies have been proved to accelerate the discovery of new anti-infectives: phenotypic screening and drug repurposing. Phenotypic screening can support the fast interrogation of compounds without the need for a pre-validated drug target, which is not available for the chikungunya virus (CHIKV) and has the additional advantage of facilitating the discovery of antiviral with novel mechanism of action. Drug repurposing can save time and resources in drug development by enabling secondary uses for drugs that are already approved for human treatment, thus precluding the need for several of the mandatory preclinical and clinical studies necessary for drug approval. A phenotypic screening assay was developed by infecting the human hepatoma Huh-7 cells with CHIKV 181/25 and quantifying infection through indirect immunofluorescence. The compound 6-azauridine was used as a positive control drug. The screening assay was validated by testing a commercial library of 1,280 compounds, including FDA-approved drugs, and used to screen a panel of broad-spectrum antiviral compounds for anti-CHIKV activity. A high content assay was set up in Huh-7 cells-infected with CHIKV. The maximum rate of infection peaked at 48 hours post-infection, after which the host cell number was greatly reduced due to a strong cytopathic effect. Assay robustness was confirmed with Z’-factor values &gt;0.8 and high correlation coefficient between independent runs, demonstrating that the assay is reliable, consistent and reproducible. Among tested compounds, sofosbuvir, an anti-hepatitis C virus drug, exhibited good selectivity against CHIKV with an EC50of 11 µM, suggesting it is a promising candidate for repurposing.

Arbidol (Umifenovir): A Broad-Spectrum Antiviral Drug That Inhibits Medically Important Arthropod-Borne Flaviviruses.

Arthropod-borne flaviviruses are human pathogens of global medical importance, against which no effective small molecule-based antiviral therapy has currently been reported. Arbidol (umifenovir) is a broad-spectrum antiviral compound approved in Russia and China for prophylaxis and treatment of influenza. This compound shows activities against numerous DNA and RNA viruses. The mode of action is based predominantly on impairment of critical steps in virus-cell interactions. Here we demonstrate that arbidol possesses micromolar-level anti-viral effects (EC50 values ranging from 10.57 ± 0.74 to 19.16 ± 0.29 µM) in Vero cells infected with Zika virus, West Nile virus, and tick-borne encephalitis virus, three medically important representatives of the arthropod-borne flaviviruses. Interestingly, no antiviral effects of arbidol are observed in virus infected porcine stable kidney cells (PS), human neuroblastoma cells (UKF-NB-4), and human hepatoma cells (Huh-7 cells) indicating that the antiviral effect of arbidol is strongly cell-type dependent. Arbidol shows increasing cytotoxicity when tested in various cell lines, in the order: Huh-7 < HBCA < PS < UKF-NB-4 < Vero with CC50 values ranging from 18.69 ± 0.1 to 89.72 ± 0.19 µM. Antiviral activities and acceptable cytotoxicity profiles suggest that arbidol could be a promising candidate for further investigation as a potential therapeutic agent in selective treatment of flaviviral infections.