Junin Virus Research Articles

Cellular N-myristoyl transferases Are Required for Mammarenavirus Multiplication.

The mammarenavirus matrix Z protein plays critical roles in virus assembly and cell egress, whereas heterotrimer complexes of a stable signal peptide (SSP) together with glycoprotein subunits GP1 and GP2, generated via co-and post-translational processing of the surface glycoprotein precursor GPC, form the spikes that decorate the virion surface and mediate virus cell entry via receptor-mediated endocytosis. The Z protein and SSP undergo N-terminal myristoylation by host cell N-myristoyltransferases (NMT1 and NMT2), and G2A mutations that prevent myristoylation of Z or SSP have been shown to affect Z mediated virus budding and GP2 mediated fusion activity required to complete the virus cell entry process. In the present work, we present evidence that the validated on-target specific pan NMT inhibitor DDD85464 exerts a potent antiviral activity against the prototypic mammarenavirus lymphocytic choriomeningitis virus (LCMV) that correlated with reduced Z budding activity and GP2 mediated fusion activity, as well as proteasome mediated degradation of the Z protein. The potent anti-mammarenaviral activity of DDD85646 was also observed with the hemorrhagic fever causing mammarenaviruses Junin (JUNV) and Lassa (LASV) viruses. Our results support exploration of NMT inhibition as a broad-spectrum antiviral against human pathogenic mammarenaviruses.

Coadministration of LHF-535 and favipiravir protects against experimental Junín virus infection and disease

Argentine hemorrhagic fever, caused by Junín virus (JUNV), is the most common of the South American arenaviral hemorrhagic fevers. The disease has a case fatality rate of 15–30% in untreated patients. Although early intervention with immune plasma is effective, diminishing stocks and limited availability outside of Argentina underscores the need for new therapeutics. Ideally, these would be broadly active agents effective against all the pathogenic arenaviruses. The fusion inhibitor LHF-535 and the nucleoside analog favipiravir have shown promise in animal models of Lassa fever, a disease endemic in parts of Africa and the most prominent of the arenaviral hemorrhagic fevers. Against JUNV, a high dose of favipiravir is required to achieve protection in the gold-standard guinea pig infection model. Here, we demonstrate a synergistic effect by the coadministration of LHF-535 with a sub-optimal dose of favipiravir in guinea pigs challenged with JUNV. Administered individually, LHF-535 and sub-optimal favipiravir only delayed the onset of severe disease. However, combined dosing of the drugs afforded complete protection against lethal JUNV infection in guinea pigs. The benefits of the drug combination were also evident by the absence of viremia and infectious virus in tissues compared to guinea pigs treated with only the placebos. Thus, combined targeting of JUNV-endosomal membrane fusion and the viral polymerase with pan-arenaviral LHF-535 and favipiravir may expand their indication beyond Lassa fever, providing a significant barrier to drug resistance.

Glycoprotein-Specific Polyclonal Antibodies Targeting Machupo Virus Protect Guinea Pigs against Lethal Infection.

Convalescent plasma has been shown to be effective at protecting humans against severe diseases caused by New World (NW) arenaviruses, including Junin virus (JUNV) and Machupo virus (MACV). This plasma contains antibodies against the full complement of structural proteins including the nucleocapsid and envelope glycoproteins (GPcs) consisting of GP1 and GP2. To gain insights into the protective and cross-protective properties of anti-GPc-specific polyclonal antibodies, we evaluated the ability of a DNA vaccine-produced anti-GPc rabbit antisera targeting MACV strain Carvallo to provide heterologous protection against another MACV strain termed Chicava in the Hartley guinea pig model. The neutralizing activity of the rabbit antisera against the heterologous MACV strains Chicava and Mallale was found to be 54-fold and 23-fold lower, respectively, compared to the titer against the homologous MACV strain Carvallo in the PRNT50 assay. Despite lower neutralizing activity against the strain Chicava, the rabbit antisera protected 100% of the guinea pigs from this strain when administered up to four days post-infection, whereas all the control animals succumbed to the disease. Using vesicular stomatitis virus (VSV) particles pseudotyped with MACV GPc, we identified a single amino acid difference at position 122 between the strains Chicava and Carvallo GPc that significantly influenced the neutralization activity of the rabbit antisera. These findings indicate that polyclonal antibodies targeting the MACV glycoproteins can protect against lethal infection in a post-challenge setting. These data will help guide future antibody-based therapeutics development against NW arenaviruses.

Design and characterization of BSA-mycophenolic acid nanocomplexes: Antiviral activity exploration

The interactions between bovine serum albumin (BSA) and mycophenolic acid (MPA) were investigated in silico through molecular docking and in vitro, using fluorescence spectroscopy. Dynamic light scattering and scanning electron microscopy were used to figure out the structure of MPA-Complex (MPA-C). The binding affinity between MPA and BSA was determined, yielding a Kd value of (12.0 ± 0.7) μM, and establishing a distance of 17 Å between the BSA and MPA molecules. The presence of MPA prompted protein aggregation, leading to the formation of MPA-C. The cytotoxicity of MPA-C and its ability to fight Junín virus (JUNV) were tested in A549 and Vero cell lines. It was found that treating infected cells with MPA-C decreased the JUNV yield and was more effective than free MPA in both cell line models for prolonged time treatments. Our results represent the first report of the antiviral activity of this type of BSA-MPA complex against JUNV, as assessed in cell culture model systems. MPA-C shows promise as a candidate for drug formulation against human pathogenic arenaviruses.

Alternative translation contributes to the generation of a cytoplasmic subpopulation of the Junín virus nucleoprotein that inhibits caspase activation and innate immunity.

The highly pathogenic arenavirus, Junín virus (JUNV), expresses three truncated alternative isoforms of its nucleoprotein (NP), i.e., NP53kD, NP47kD, and NP40kD. While both NP47kD and NP40kD have been previously shown to be products of caspase cleavage, here, we show that expression of the third isoform NP53kD is due to alternative in-frame translation from M80. Based on this information, we were able to generate recombinant JUNVs lacking each of these isoforms. Infection with these mutants revealed that, while all three isoforms contribute to the efficient control of caspase activation, NP40kD plays the predominant role. In contrast to full-length NP (i.e., NP65kD), which is localized to inclusion bodies, where viral RNA synthesis takes place, the loss of portions of the N-terminal coiled-coil region in these isoforms leads to a diffuse cytoplasmic distribution and a loss of function in viral RNA synthesis. Nonetheless, NP53kD, NP47kD, and NP40kD all retain robust interferon antagonistic and 3'-5' exonuclease activities. We suggest that the altered localization of these NP isoforms allows them to be more efficiently targeted by activated caspases for cleavage as decoy substrates, and to be better positioned to degrade viral double-stranded (ds)RNA species that accumulate in the cytoplasm during virus infection and/or interact with cytosolic RNA sensors, thereby limiting dsRNA-mediated innate immune responses. Taken together, this work provides insight into the mechanism by which JUNV leverages apoptosis during infection to generate biologically distinct pools of NP and contributes to our understanding of the expression and biological relevance of alternative protein isoforms during virus infection.IMPORTANCEA limited coding capacity means that RNA viruses need strategies to diversify their proteome. The nucleoprotein (NP) of the highly pathogenic arenavirus Junín virus (JUNV) produces three N-terminally truncated isoforms: two (NP47kD and NP40kD) are known to be produced by caspase cleavage, while, here, we show that NP53kD is produced by alternative translation initiation. Recombinant JUNVs lacking individual NP isoforms revealed that all three isoforms contribute to inhibiting caspase activation during infection, but cleavage to generate NP40kD makes the biggest contribution. Importantly, all three isoforms retain their ability to digest double-stranded (ds)RNA and inhibit interferon promoter activation but have a diffuse cytoplasmic distribution. Given the cytoplasmic localization of both aberrant viral dsRNAs, as well as dsRNA sensors and many other cellular components of innate immune activation pathways, we suggest that the generation of NP isoforms not only contributes to evasion of apoptosis but also robust control of the antiviral response.

COMUNIDADES DE ROEDORES DE CULTIVOS DE SORGO EN LA PROVINCIA DE CÓRDOBA Y SU POSIBLE VINCULACIÓN CON LA FIEBRE HEMORRÁGICA ARGENTINA

The composition of rodent communities in fields dedicated to sorghum culture was studied in seven sites situated along two transects about 500 km long, in the SE of Córdoba Province (Argentina). Four of the sites are in the endemic area of ArgentineHemorrhagic Fever and the other three in the marginal area, free of Junin virus. The animáis were trapped during a short period in May and June, 1977. Eight Crieetidae (Calomys musculinus, Akodon azarae, Calomys laucha, Oligoryzomys flavescens, Cabreramys benefactus, Akodon dolores, Graomys griseoflavus y Calomys callosus) and one Muridae species (Mus musculus) were obtained. Comparative analysis of the species composition of the communities has been performed by using the Mountford Índex.Total number of specimens from Calomys plus Akodon represented, in all the samples, more than 80%. The most abundant species in six out of seven samples was C. musculinus, the most important reservoir of Junin virus. The structure of this species populationswas very similar for most of the endemic and marginal areas.

The Arenaviridae Family: Knowledge Gaps, Animal Models, Countermeasures, and Prototype Pathogens.

Lassa virus (LASV), Junin virus (JUNV), and several other members of the Arenaviridae family are capable of zoonotic transfer to humans and induction of severe viral hemorrhagic fevers. Despite the importance of arenaviruses as potential pandemic pathogens, numerous gaps exist in scientific knowledge pertaining to this diverse family, including gaps in understanding replication, immunosuppression, receptor usage, and elicitation of neutralizing antibody responses, that in turn complicates development of medical countermeasures. A further challenge to the development of medical countermeasures for arenaviruses is the requirement for use of animal models at high levels of biocontainment, where each model has distinct advantages and limitations depending on, availability of space, animals species-specific reagents, and most importantly the ability of the model to faithfully recapitulate human disease. Designation of LASV and JUNV as prototype pathogens can facilitate progress in addressing the public health challenges posed by members of this important virus family.

Application of multiplex realtime PCR detection for hemorrhagic fever syndrome viruses

BackgroundMultiplex real-time PCR is a quick and cost effective method for detection of various gene simultaneously. HFSV (Hemorrhagic Fever Syndrome Virus) is a newly emerging infectious disease because of globalization and climate change. We tried to develop a molecular diagnostic technique for various causative viruses and evaluate its usefulness for improving public health. MethodsMolecular diagnostic test method that qualitatively detects viruses causing viral hemorrhagic fevers hired Taq-Man Real-time RT-PCR technique. The Ct value was experimentally observed three or more times at the RNA concentration before and after the detection limit. After designing a multiplex real-time RT-PCR test for target gene of selected 17 viruses, the detection limit for each target and the presence or absence of cross-reaction and interference reaction were evaluated to determine its availability. ResultsSix kinds of viruses, including Crimean-Congo hemorrhagic fever virus, Omsk hemorrhagic fever virus, Sabia virus, Chapare virus, Yellow fever virus, and Variola virus (A4L gene, B12R gene), were able to confirm the detection limit of 0.5 copies/μl, and other Ebola virus, Marburg virus, Rift Valley fever virus, Kyasanur Forest disease virus, Junin virus, Guanarito virus, Machupo virus, Chikungunya virus, Hantavirus, Dengue virus types 1–4, and Lassa virus (L gene, GPC gene), and 11 kinds of viruses, the detection limit was confirmed at 5 copies/μl. No cross-reaction or interference between detected genes was observed. ConclusionThe virus test method developed through this study using multiplex is expected to be used for public health and quarantine as a test method that can be used when a hemorrhagic fever virus of unknown cause is introduced.

Temporal changes in pathology and viral RNA distribution in guinea pigs following separate infection with two New World Arenaviruses.

Numerous arenaviruses have been identified throughout the Americas and a subset of these viruses cause viral hemorrhagic fever in humans. This study compared the pathology and viral RNA distribution in Hartley guinea pigs challenged with two human-disease causing New World arenaviruses, Junin virus (JUNV) or Guanarito virus (GTOV). Histopathologic analysis and RNA in situ hybridization revealed similar pathology and viral RNA distribution for both groups of animals challenged with either JUNV or GTOV on days 3, 7, 10 and 12 post exposure (PE). Gross lesions were first observed on day 7 and primarily involved the lungs and liver. The most severe histologic lesions occurred in the lymph nodes, spleen, and thymus and included lymphoid depletion and necrosis which increased in severity over time. Extensive necrosis was also observed in the bone marrow on day 12. Minimal to mild inflammation with and without necrosis was observed in the choroid plexus of the brain, choroid of the eye, intestinal tract, lung and adrenal gland. Significant liver lesions were rare, consisting predominantly of hepatocyte vacuolation. Viral RNA labeling was identified in nearly all organs examined, was often extensive in certain organs and generally increased over time starting on day 7. Our data demonstrate the guinea pig may serve as a useful model to study New World arenavirus infection in humans and for the evaluation and development of medical countermeasures.

Effects of the Natural Flavonoid Quercetin on Arenavirus Junín Infection.

There is no specific chemotherapy approved for the treatment of pathogenic arenaviruses that cause severe hemorrhagic fever (HF) in the population of endemic regions in America and Africa. The present study reports the effects of the natural flavonoid quercetin (QUER) on the infection of A549 and Vero cells with Junín virus (JUNV), agent of the Argentine HF. By infectivity assays, a very effective dose-dependent reduction of JUNV multiplication was shown by cell pretreatment at 2-6 h prior to the infection at non-cytotoxic concentrations, with 50% effective concentration values in the range of 6.1-7.5 µg/mL. QUER was also active by post-infection treatment but with minor efficacy. Mechanistic studies indicated that QUER mainly affected the early steps of virus adsorption and internalization in the multiplication cycle of JUNV. Treatment with QUER blocked the phosphorylation of Akt without changes in the total protein expression, detected by Western blot, and the consequent perturbation of the PI3K/Akt pathway was also associated with the fluorescence redistribution from membrane to cytoplasm of TfR1, the cell receptor recognized by JUNV. Then, it appears that the cellular antiviral state, induced by QUER treatment, leads to the prevention of JUNV entry into the cell.

N6-methyladenosine is required for efficient RNA synthesis of Ebola virus and other haemorrhagic fever viruses

ABSTRACT N6-methyladenosine (m6A) is one of the most abundant modifications of cellular RNA, where it serves various functions. m6A methylation of many viral RNA species has also been described; however, little is known about the m6A epitranscriptome of haemorrhagic fever-causing viruses like Ebola virus (EBOV). Here, we analysed the importance of the methyltransferase METTL3 for the life cycle of this virus. We found that METTL3 interacts with the EBOV nucleoprotein and the transcriptional activator VP30 to support viral RNA synthesis, and that METTL3 is recruited into EBOV inclusions bodies, where viral RNA synthesis occurs. Analysis of the m6A methylation pattern of EBOV mRNAs showed that they are methylated by METTL3. Further studies revealed that METTL3 interaction with the viral nucleoprotein, as well as its importance for RNA synthesis and protein expression, is also observed for other haemorrhagic fever viruses such as Junín virus (JUNV) and Crimean-Congo haemorrhagic fever virus (CCHFV). The negative effects on viral RNA synthesis due to loss of m6A methylation are independent of innate immune sensing, as METTL3 knockout did not affect type I interferon induction in response to viral RNA synthesis or infection. Our results suggest a novel function for m6A that is conserved among diverse haemorrhagic fever-causing viruses (i.e. EBOV, JUNV and CCHFV), making METTL3 a promising target for broadly-acting antivirals.

The roles of XJ13 and XJ44-specific mutations within the Candid #1 GPC in Junin virus attenuation.

Junin virus (JUNV) is a member of the Arenaviridae family of viruses and is the pathogen responsible for causing Argentine hemorrhagic fever, a potentially lethal disease endemic to Argentina. A live attenuated vaccine for human use, called Candid#1, is approved only in Argentina. Candid#1 vaccine strain of Junin virus was obtained through serial passage in mouse brain tissues followed by passage in Fetal Rhesus macaque lung fibroblast (FRhL) cells. Previously, the mutations responsible for attenuation of this virus in Guinea pigs were mapped in the gene encoding for glycoprotein precursor (GPC) protein. The resulting Candid#1 glycoprotein complex has been shown to cause endoplasmic reticulum (ER) stress in vitro resulting in the degradation of the GPC. To evaluate the attenuating properties of specific mutations within GPC, we created recombinant viruses expressing GPC mutations specific to key Candid#1 passages and evaluated their pathogenicity in our outbred Hartley guinea pig model of Argentine hemorrhagic fever. Here, we provide evidence that early mutations in GPC obtained through serial passaging attenuate the visceral disease and increase immunogenicity in guinea pigs. Specific mutations acquired prior to the 13th mouse brain passage (XJ13) are responsible for attenuation of the visceral disease while having no impact on the neurovirulence of Junin virus. Additionally, our findings demonstrate that the mutation within an N-linked glycosylation motif, acquired prior to the 44th mouse brain passage (XJ44), is unstable but necessary for complete attenuation and enhanced immunogenicity of Candid#1 vaccine strain. The highly conserved N-linked glycosylation profiles of arenavirus glycoproteins could therefore be viable targets for designing attenuating viruses for vaccine development against other arenavirus-associated illnesses.

Arenaviruses: Old viruses present new solutions for cancer therapy.

Viral-based cancer therapies have tremendous potential, especially in the context of treating poorly infiltrated cold tumors. However, in tumors with intact anti-viral interferon (IFN) pathways, while some oncolytic viruses induce strong innate and adaptive immune responses, they are neutralized before exerting their therapeutic effect. Arenaviruses, particularly the lymphocytic choriomeningitis virus (LCMV) is a noncytopathic virus with preferential cancer tropism and evolutionary mechanisms to escape the immune system for longer and to block early clearance. These escape mechanisms include inhibition of the MAVS dependent IFN pathway and spike protein antigen masking. Regarding its potential for cancer treatment, LCMV is therefore able to elicit long-term responses within the tumor microenvironment (TME), boost anti-tumor immune responses and polarize poorly infiltrating tumors towards a hot phenotype. Other arenaviruses including the attenuated Junin virus vaccine also have anti-tumor effects. Furthermore, the LCMV and Pichinde arenaviruses are currently being used to create vector-based vaccines with attenuated but replicating virus. This review focuses on highlighting the potential of arenaviruses as anti-cancer therapies. This includes providing a molecular understanding of its tropism as well as highlighting past and present preclinical and clinical applications of noncytophatic arenavirus therapies and their potential in bridging the gap in the treatment of cancers weakly responsive or unresponsive to oncolytic viruses. In summary, arenaviruses represent promising new therapies to broaden the arsenal of anti-tumor therapies for generating an immunogenic tumor microenvironment.

Vaccine Candidates against Arenavirus Infections.

The viral family Arenaviridae contains several members that cause severe, and often lethal, diseases in humans. Several highly pathogenic arenaviruses are classified as Risk Group 4 agents and must be handled in the highest biological containment facility, biosafety level-4 (BSL-4). Vaccines and treatments are very limited for these pathogens. The development of vaccines is crucial for the establishment of countermeasures against highly pathogenic arenavirus infections. While several vaccine candidates have been investigated, there are currently no approved vaccines for arenavirus infection except for Candid#1, a live-attenuated Junin virus vaccine only licensed in Argentina. Current platforms under investigation for use include live-attenuated vaccines, recombinant virus-based vaccines, and recombinant proteins. We summarize here the recent updates of vaccine candidates against arenavirus infections.

Structure-activity relationship studies of anti-bunyaviral cap-dependent endonuclease inhibitors

Bunyaviruses, including the Lassa virus (LASV), are known to cause hemorrhagic fever and have a high fatality rate among hospitalized patients, as there are few effective treatments. We focused on the fact that bunyaviruses use cap-dependent endonuclease (CEN) for viral replication, which is similar to influenza viruses. This led us to screen carbamoyl pyridone bicycle (CAB) compounds, which compose a series of baloxavir acid (BXA) derivatives, against lymphocytic choriomeningitis virus (LCMV) and Junin virus (JUNV) among the bunyaviruses. This led to the discovery of 1c, which has potent anti-bunyaviral activities. In SAR studies, we found that a large lipophilic side chain is preferred for the 1-position of the CAB scaffold, similar to the influenza CEN inhibitor, and that a small alkyl group for the 3-position shows high activity. Moreover, the 7‑carboxyl group of the scaffold is essential for anti-bunyaviral activities, and the antiviral activity is reduced by conversion to various carboxylic acid bioisosteres. The SAR results are discussed using a binding model of 9d in the active center of the known LCMV CEN crystal structure. These compounds show promise as broad-spectrum anti-bunyavirus therapeutics, given their relatively favorable metabolic stability and PK profiles.

VelcroVax: a "Bolt-On" Vaccine Platform for Glycoprotein Display.

Having varied approaches to the design and manufacture of vaccines is critical in being able to respond to worldwide needs and newly emerging pathogens. Virus-like particles (VLPs) form the basis of two of the most successful licensed vaccines (against hepatitis B virus [HBV] and human papillomavirus). They are produced by recombinant expression of viral structural proteins, which assemble into immunogenic nanoparticles. VLPs can be modified to present unrelated antigens, and here we describe a universal "bolt-on" platform (termed VelcroVax) where the capturing VLP and the target antigen are produced separately. We utilize a modified HBV core (HBcAg) VLP with surface expression of a high-affinity binding sequence (Affimer) directed against a SUMO tag and use this to capture SUMO-tagged gp1 glycoprotein from the arenavirus Junín virus (JUNV). Using this model system, we have solved the first high-resolution structures of VelcroVax VLPs and shown that the VelcroVax-JUNV gp1 complex induces superior humoral immune responses compared to the noncomplexed viral protein. We propose that this system could be modified to present a range of antigens and therefore form the foundation of future rapid-response vaccination strategies. IMPORTANCE The hepatitis B core protein (HBc) forms noninfectious virus-like particles, which can be modified to present a capturing molecule, allowing suitably tagged antigens to be bound on their surface. This system can be adapted and provides the foundation for a universal "bolt-on" vaccine platform (termed VelcroVax) that can be easily and rapidly modified to generate nanoparticle vaccine candidates.

Modulation of the Aryl Hydrocarbon Receptor Signaling Pathway Impacts on Junín Virus Replication.

Junín virus (JUNV), a member of the family Arenaviridae, is the etiological agent of the Argentine hemorrhagic fever, an endemic disease in the rural region of Argentina lacking a specific chemotherapy. Aryl hydrocarbon receptor (AHR) is expressed in several mammalian tissues and has been indicated as a sensor of ligands from variable sources and a modulator of the cell immune response. Interestingly, recent studies have suggested that the activation or depression of the AHR signaling pathway may play a role in the outcome of diverse human viral infections. In the present report, the effect of the pharmacological modulation of AHR on JUNV in vitro infection was analyzed. An initial microarray screening showed that the AHR pathway was overexpressed in JUNV-infected hepatic cells. Concomitantly, the infection of Vero and Huh-7 cells with the JUNV strains IV4454 and Candid#1 was significantly inhibited in a dose-dependent manner by treatment with CH223191, a specific AHR antagonist, as detected by infectivity assays, real-time RT-PCR and immunofluorescence detection of viral proteins. Furthermore, the pro-viral role of AHR in JUNV infection appears to be independent of the IFN-I pathway. Our findings support the promising perspectives of the pharmacological modulation of AHR as a potential target for the control of AHF.

Recent developments on Junin virus, a causative agent for Argentine haemorrhagic fever.

Junin virus consists of ribonucleic acid as the genome and is responsible for a rapidly changing tendency of the virus. The virus is accountable for ailments in the human body and causes Argentine Haemorrhagic Fever (AHF). The infection is may be transmitted through contact between an infected animal/host and a person, and later between person to person. Prevention of outbreaks of AHF in humans can be a tough practice, as their occurrence is infrequent and unpredictable. In this review, recent information from the past 5years available on the Junin virus including the risk of its emergence, infectious agents, its pathogenesis in humans, available diagnostic and therapeutic approaches, and disease management has been summarised. Altogether, this article would be highly significant in understanding the mechanistic basis behind virus interaction and other processes during the life cycle. Currently, no specific therapeutic options are available to treat the Junin virus infection. The information covered in this review could be important for finding possible treatment options for Junin virus infections.

Topoisomerase II as a Novel Antiviral Target against Panarenaviral Diseases.

Although many arenaviruses cause severe diseases with high fatality rates each year, treatment options are limited to off-label use of ribavirin, and a Food and Drug Administration (FDA)-approved vaccine is not available. To identify novel therapeutic candidates against arenaviral diseases, an RNA polymerase I-driven minigenome (MG) expression system for Lassa virus (LASV) was developed and optimized for high-throughput screening (HTS). Using this system, we screened 2595 FDA-approved compounds for inhibitors of LASV genome replication and identified multiple compounds including pixantrone maleate, a topoisomerase II inhibitor, as hits. Other tested topoisomerase II inhibitors also suppressed LASV MG activity. These topoisomerase II inhibitors also inhibited Junin virus (JUNV) MG activity and effectively limited infection by the JUNV Candid #1 strain, and siRNA knockdown of both topoisomerases (IIα and IIβ) restricted JUNV replication. These results suggest that topoisomerases II regulate arenavirus replication and can serve as molecular targets for panarenaviral replication inhibitors.

Severe mammarenaviral disease in guinea pigs effectively treated by an orally bioavailable fusion inhibitor, alone or in combination with favipiravir

Infections by pathogenic New World mammarenaviruses (NWM)s, including Junín virus (JUNV), can result in a severe life-threatening viral hemorrhagic fever syndrome. In the absence of FDA-licensed vaccines or antivirals, these viruses are considered high priority pathogens. The mammarenavirus envelope glycoprotein complex (GPC) mediates pH-dependent fusion between viral and cellular membranes, which is essential to viral entry and may be vulnerable to small-molecule inhibitors that disrupt this process. ARN-75039 is a potent fusion inhibitor of a broad spectrum of pseudotyped and native mammarenaviruses in cell culture and Tacaribe virus infection in mice. In the present study, we evaluated ARN-75039 against pathogenic JUNV in the rigorous guinea pig infection model. The compound was well-tolerated and had favorable pharmacokinetics supporting once-per-day oral dosing in guinea pigs. Importantly, significant protection against JUNV challenge was observed even when ARN-75039 was withheld until 6 days after the viral challenge when clinical signs of disease are starting to develop. We also show that ARN-75039 combination treatment with favipiravir, a viral polymerase inhibitor, results in synergistic activity in vitro and improves survival outcomes in JUNV-challenged guinea pigs. Our findings support the continued development of ARN-75039 as an attractive therapeutic candidate for treating mammarenaviral hemorrhagic fevers, including those associated with NWM infection.