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EIDD-2801 resists to infection and co-infection of SARS-CoV-2 and influenza virus

Virology Journal volume 22, Article number: 126 (2025) Cite this article

Abstract

Background

The coronavirus disease 2019 (COVID-19) pandemic has exerted a catastrophic impact on public health. Meanwhile, the seasonal influenza outbreak overlaps with the current pandemic wave. There is still an urgent need to develop effective therapeutic agents for the treatment of co-infection of multiple respiratory viruses. This study aimed to investigate antiviral effects of EIDD-2801, an orally bioavailable ribonucleoside analog, and its potent therapeutic effects in co-infection of multiple respiratory viruses.

Methods

BALB/c mice and hamsters were infected with IFV or SARS-CoV-2, then were dosed orally with EIDD-2801 to measure the antiviral effects of EIDD-2801. Viral replication and mRNA transcription were evaluated by quantitative polymerase chain reaction (qPCR) and protein expression by Western Blot. Influenza viral titer was assessed using EID50 assay.

Results

EIDD-2801 was found to be significantly effective against influenza A virus and influenza B virus. The antiviral activity against SARS-CoV-2 and further co-infection with influenza virus was also distinct. EIDD-2801 had potent antiviral effects against multiple respiratory viruses both in vitro and in vivo.

Conclusion

This study demonstrated that the small-molecule compound EIDD-2801, an orally available broad-spectrum antiviral agent, significantly inhibited the infection of influenza virus and SARS-CoV-2 and effectively protected animals from lethal influenza virus co-infection.

Introduction

The coronavirus disease 2019 (COVID-19) pandemic caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has posed significant threats to public health and economic development worldwide [1]. Most infected patients show pneumonia symptoms ranging from respiratory complications to acute respiratory distress syndrome and multiple organ dysfunction [2]. These symptoms are associated with the respiratory system, resulting in the impairment of lung function [3,4,5]. Vaccines are widely available in disease prevention. However, the emergence of variants of concern (VOCs) including B.1.1.7 (alpha) [6], B.1.351 (beta) [7], P.1 (gamma) [8], B.1.617.2 (delta) [9] and B1.1529 (omicron) [10] lineages, characterized by enhanced transmissibility and immune evasion capabilities, poses significant risks of exacerbating pandemic trajectories [11,12,13]. More importantly, the co-circulation of multiple viruses [14, 15] not only exacerbates the clinical severity of co-infections but also heightens concerns regarding potential cross-transmission risks [16, 17].

Influenza virus, a highly contagious respiratory virus, also can cause respiratory complications and co-infection with other pathogens [18,19,20]. Both influenza virus and SARS-CoV-2 are airborne transmitted viruses following similar transmission routes [21,22,23]. They infect the upper and lower respiratory tracts and have the same target cells despite the different cellular receptors [24, 25]. Previous clinical studies suggested that co-infection of Influenza A virus and other respiratory viruses, including SARS-CoV-2 and other circulating influenza virus strains resulted in more severe disease and higher fatality [26,27,28]. Thus, recurring influenza seasons during the current pandemic potentially make the threats to public health even more critical. In the context of influenza and COVID-19 outbreaks overlap in many regions, difficulties in disease management call for the development of broad-spectrum antiviral agents for the control of co-infection of emerging viruses.

EIDD-2801 (MK-4482), an orally available pro-drug of N4-hydroxycytidine (NHC), exhibits antiviral activities against influenza virus [29, 30] and SARS-CoV-2 [31,32,33,34,35] in vitro and in vivo. As a cytidine analogue, EIDD-2801 inhibits viral RNA-dependent RNA polymerase (RdRp) activity by inducing random low-frequency C-U and G-A transitions, resulting in deleterious mutations and inhibition of viral RNA replication [36,37,38].

In this study, we evaluated the effects of EIDD-2801 against a panel of mouse-adapted viral strains, including influenza A virus (IAV), influenza B virus (IBV) and SARS-CoV-2. The results indicated that therapeutic administration of EIDD-2801 dramatically reduced viral load of divergent influenza virus and SARS-CoV-2 in upper respiratory tract and lung. Notably, oral administration of EIDD-2801 exhibits significant antiviral potency for the treatment of influenza virus and SARS-CoV-2 mono-infection and co-infection.

Materials and methods

Cells, viruses and compounds

MDCK, Vero-E6, and Huh7 cell lines used in the experiments were cultured in Dulbecco’s Modified Eagle’s Medium (DMEM) (Gibco, US) supplemented with 10% fetal bovine serum (FBS) (Gibco, US) and 100 U/mL penicillin-streptomycin at 37 ℃ in 5% CO2. Influenza A virus H1N1-UI182 strain and H3N2 strain, Influenza B virus IBV/S9-E2 strain, and IBV/S9-MD strains were saved as previously described [39]. All SARS-CoV-2 strains used in this study were isolated and stored by Changchun Veterinary Research Institute according to biosafety norms. Oseltamivir phosphate, zanamivir, and baloxavir were purchased from Med Chem Express (Shanghai, CN) and EIDD-2801 was a gift from Prof. Wu Zhong [40]. All compounds were dissolved in phosphate-buffered saline (PBS) and stored at -20 ℃. All working standards were diluted in DMEM or PBS from stock solution within 1 day before the experiment and stored at 4 °C for a maximum of 6 days.

Immunofluorescence assay

MDCK cells were seeded in 48-well plates and infected with H1N1-UI182 (MOI = 0.05), IAV/H3N2 (MOI = 0.01) or IBV/S9-E2 (MOI = 0.05) respectively at 37 °C for 1 h when cell confluence reached 70–90%, then the cells were further incubated with DMEM with 2% FBS and 20 µM EIDD-2801. At 48 h post-infection (hpi), the cells were washed three times with PBS and fixed with 4% paraformaldehyde (PFA) for 20 min. After that, the cells were infiltrated with 0.2% Triton-X100 and blocked in 2% Bovine Serum Albumin (BSA) dissolved in Phosphate-buffered saline with tween (PBST) for 1 h. Anti- IAV NP antibody (Abcam, ab128193, Shanghai, China, 1:200) or Anti-IBV NP antibody (GeneTex, GTX636194, CA, USA, 1:1000) was added respectively and incubated with cells overnight at 4 ℃. After the primary antibodies, cells were washed three times using PBS and incubated with Alexa Fluor 488-labeled goat anti-Rabbit IgG antibody(Abcam, ab150077, Shanghai, China) for 2 h without light. After nuclei staining with Hoechst (Thermo Fisher Scientific, H3569, 1 µg/mL) for 10 min, fluorescent images were taken with a fluorescence microscope (Carl Zeiss, Germany) and the cell number was quantified using Image-J software.

In vitro infection experiment of SARS-CoV-2

Vero-E6 and Huh7 cells were pre-seeded in 96-well plates at the density of 8000 cells/well. When cell confluence reached 70–90%, cells were infected with diluted SARS-CoV-2 variants (BJ01, Delta, BA.2, BF.7 and XBB.1.9.1) at a multiplicity of infection (MOI) of 0.01 for 1 h. Following viral adsorption, a concentration gradient of EIDD-2801 (0.003, 0.03, 0.3, 3, 30 and 60 µM) was administered and maintained under standard culture conditions (37 °C, 5% CO2) for 36 h. Then 200 µL of supernatant in each group was collected to detect viral RNA copy numbers using qRT-PCR.

Quantitative real-time PCR (qRT-PCR)

200 µL cell supernatants or supernatants from the lysed tissues were harvested for RNA extraction (TIANGEN, YDP804-T1) and RNA copies were quantified by qRT-PCR according to the manufacturer’s instructions. The primers designed to amplify the SARS-CoV-2 ORFlab gene were as follows:

Forward sequence: 5′-CCCTGTGGGTTTTACACTTAA-3′.

Reverse sequence: 5′-ACGATTGTGCATCAGCTGA-3′.

Probe sequence: 5′-FAM-CCGTCTGCGGTATGTGGAAAGGTTATGG-BHQ1-3′.

The primers designed to amplify the H1N1 M gene were as follows:

Forward sequence: 5′-GTCTTCTAACCGAGGTCGAAA-3′.

Reverse sequence: 5′-AAGATCTGTGTTCTTTCCTGCAAA-3′.

Probe sequence: 5′-FAM-CCCTCAAAGCCGAGATCGC-TAMRA-3′.

In vivo infection experiment of influenza viruses in mouse model

Six to eight-week-old female BALB/c mice were purchased from Vital River Laboratory Animal Technology Co., Ltd (Beijing, China) and were quarantine-housed in individually ventilated cages. Then the mice were randomly assigned into 3 groups, including blank control (Control), virus control (Virus), and drug-treated group (Virus + Drug), with 12 mice per group. All mice were infected with influenza viruses by nasal drip (except the blank group), and were treated with PBS or oseltamivir phosphate (25 mg/kg/day, orally), zanamivir (5 mg/kg/day, intraperitoneally), baloxavir acid (5 mg/kg/day, subcutaneously) and EIDD-2801 (500 mg/kg/day, orally) for 5 days. For mono-infection of influenza virus, mice were intranasally infected with 50 µL viral dilutions (10×MLD50 of H1N1-UI182 or 1×MLD50 of IBV/S9-MD) separately. For co-infection, mice were intranasally infected with 50 µL viral dilutions of 1×MLD50 H1N1-UI182 and 0.1×MLD50 of IBV/S9-MD. Body weight changes and survival status were monitored daily for 14 days. Mice with more than a 25% decrease in body weight were euthanized humanely. At 5 days post-infection (dpi), 5 mice were randomly selected from each group and euthanized by exsanguination under deep anesthetization, and a part of the organs was collected and fixed with 4% paraformaldehyde (PFA) for pathological analysis. The other part collected lungs was immersed in Radio Immunoprecipitation Assay Lysis (RIPA) buffer (Beyotime, China) with protease inhibitor (MCE, HY-K0011, Shanghai, China) for Western Blot (WB) or in DMEM for quantification of the viral titer.

Co-infection experiment of SARS-CoV-2 and influenza A virus in golden hamster

4-week-old female golden hamsters were purchased from Vital River Laboratory Animal Technology Co., Ltd. (Beijing, China) and were intranasally inoculated with 100 µL virus dilution of SARS-CoV-2/Omicron BF.7 (2000 PFU) and H1N1-UI182 (1×MLD50). The co-infected animals were treated with vehicle (PBS) or EIDD-2801 (350 mg/kg/day, orally) for 5 days. Body weight changes were monitored for 14 days. Nasal lavage fluid was collected at 2 and 4 dpi separately. At 3 days post-infection, 4 hamsters were randomly selected from each group and euthanized by exsanguination under deep anesthetization. Tissues were harvested following euthanasia to measure lung weight and quantify viral loads.

Western blot

Lung tissues harvested from experimental animals were lysed with RIPA buffer. After protein quantification using Enhanced BCA Protein Assay Kit (Beyotime, China), lysates were mixed with 5× loading buffer (Beyotime, China) and boiled for 5 min. Lung tissue homogenates were prepared from n = 3–5 animals per group, and equal amounts of protein (10 µg) were loaded per lane. The proteins were transferred to a polyvinylidene difluoride (PVDF) membrane after being subjected to SDS-PAGE. The PVDF membranes was blocked in 5% BSA solution for 1 h. After blocking, the membranes were incubated with either Anti-IAV NP antibody (GeneTex, GTX125989, CA, USA, 1:5000) and Anti-IAV M antibody (GeneTex, GTX636677, CA, USA, 1:5000) or Anti-IBV NP (GeneTex, GTX636194, CA, USA, 1;5000) and Anti-IBV M antibody (GeneTex, GTX128537, CA, USA, 1:5000) overnight at 4 ℃. Anti-β-actin (Abcam, ab6276, Shanghai, China, 1:5000) or anti-GAPDH (Abcam, ab8245, Shanghai, China, 1:1000) was used as a loading control. Following three washes with Tris-buffered saline containing 0.1% Tween-20 (TBST; Beyotime, China), the membranes were incubated with species-specific secondary antibodies: HRP-conjugated goat anti-rabbit IgG (Abcam, ab205718; 1:1000) or goat anti-mouse IgG (Abcam, ab205719; 1:1000) in TBST for 1 h at room temperature (RT). Finally, the membranes were incubated by enhanced chemiluminescence (ECL) Western Blot substrate after washed another three times with TBST for the analysis using a fluorescence image analysis system (Tanon, China). Band intensities were quantified using Image-J, and results were normalized to loading controls.

Statistical analysis

All the calculated results were presented as mean ± standard deviation (SD). Statistical analyses were performed using one-way ANOVA or student’s t-test with GraphPad Prism 9.0 software. Compared with the Virus-infected group, the statistical significances were defined as P < 0.05 (*), and the higher significance was denoted by P < 0.01 (**) and P < 0.001 (***).

Results

Potent inhibitory activity and therapeutic effect of EIDD-2801 against influenza viruses in vitro and in vivo

Consistent with prior studies [29, 30, 41], EIDD-2801 demonstrated potent antiviral activity against influenza viruses in this study. In MDCK cells infected with H1N1-UI182, EIDD-2801 treatment reduced IAV NP protein expression by approximately 45% (Fig. S1A, B) and viral titers by 2.5 log10 compared to the control group (Fig. S1C). Similarly, for H3N2, IAV NP protein levels decreased by 96% (Fig. S1D, E) with a 2 log10 reduction in viral titers (Fig. S1F). Notably, EIDD-2801 also exhibited efficacy against IBV/S9-E2, achieving a 63% inhibition of NP protein expression (Fig. S1G, H) and a 1.4 log10 decrease in viral loads (Fig. S1I). To evaluate the in vivo antiviral efficacy of EIDD-2801 against both IAV and IBV, BALB/c mice infected with influenza virus were treated with EIDD-2801 and compared to multiple standard anti-influenza drugs. Briefly, BALB/c mice were intranasally inoculated with a lethal dose of H1N1-UI182 or a non-lethal dose of IBV/S9-MD and subsequently treated with EIDD-2801 (500 mg/kg/day) via oral gavage starting 12 h post-infection (Fig. 1A). After 5 consecutive days of treatment, EIDD-2801 administration significantly protected infected mice from weight loss in both IAV and IBV mouse models, demonstrating superior efficacy compared to oseltamivir phosphate and zanamivir (Fig. 1B, D). Unexpectedly, EIDD-2801 treatment exhibited higher protection efficacy (62.5%) than oseltamivir phosphate (50%) in H1N1-UI182-infected mice (Fig. 1C), highlighting its superior antiviral potential. Necropsy of lung tissues revealed that both IAV and IBV infections caused dark red discoloration with patchy hemorrhage on the lung surface (indicated by black arrows), accompanied by severe pathological damage and a significant increase in lung index (Fig. 1E, F). These effects were markedly alleviated by EIDD-2801 treatment. Meanwhile, Western Blot results revealed that EIDD-2801 treatment significantly inhibited viral protein expression in IAV and IBV. Compared to the viral control group, EIDD-2801 treatment significantly reduced the expression levels of IAV NP and IAV M to below the limit of detection (LOD) in mouse lungs (Fig. 1G). Similarly, for IBV-infected mice (Fig. 1H), EIDD-2801 treatment resulted in a 95% reduction in the expression levels of IBV NP and IBV M compared to the untreated control group (Fig. 1H). These data indicate that oseltamivir can still protect mice from lethal H1N1-UI182 infection; however, its effects on reducing lung damage and viral load are less pronounced. Furthermore, zanamivir failed to ameliorate IBV-induced lung damage. Collectively, the antiviral efficacy of EIDD-2801 was superior to that of both oseltamivir and zanamivir.

Fig. 1
figure 1

Administration of EIDD-2801 effectively protects mice against lethal and non-lethal influenza virus challenge. A Schematic of the mouse pathogenesis study. BALB/C mice were orally administered EIDD-2801 at a dose of 500 mg/kg for up to 5 days following intranasal infection with H1N1-UI182 (10×MLD50) or IBV/S9-MD (1×MLD50). B Body weight change of mice infected with H1N1-UI182. C Survival curves of mice infected with H1N1-UI182. D Body weight change of mice infected with IBV/S9-MD. E-F Representative images of lung tissue are presented, with photomicrographs showing histopathological changes (scale bars = 200 μm). Hematoxylin and eosin staining (H&E) and lung index (%) in groups infected with IAV E or IBV/S9-MD F. G Western Blot analysis of influenza A virus nucleoprotein (IAV NP) and matrix protein (IAV M) expression in lung tissue homogenates from H1N1-UI182-infected mice (n = 3). H Western Blot analysis of influenza B virus nucleoprotein (IBV NP) and matrix protein (IBV M) expression in lung tissue homogenates from IBV/S9-MD-infected mice (n = 3). Band intensities were measured and the expression of β-actin or GAPDH was used as loading control. Statistically significant differences between groups were determined by the Student’s t test (*) 0.01 < P < 0.05, (***) P < 0.001, “ns” denotes no significance

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In vivo efficacy of EIDD-2801 against co-infection of influenza A virus and influenza B virus

To evaluate the antiviral efficacy of EIDD-2801 against co-infected with both IAV and IBV, BALB/c mice were simultaneously infected with H1N1-UI182 and IBV/S9-MD. Compared to the Virus group, mice treated with EIDD-2801 exhibited significantly reduced weight loss, surpassing even the effects observed in the oseltamivir-treated group (Fig. 2A). Furthermore, oral administration of EIDD-2801 ensured complete survival of the mice, comparable to the outcomes achieved with oseltamivir or baloxavir treatment (Fig. 2B). EIDD-2801 treatment led to a significant reduction in the expression of IAV M and IBV M in co-infected mouse lung tissues, with both proteins barely detectable on Western Blot (Fig. 2C). In addition, EIDD-2801 significantly alleviated extensive hemorrhage, inflammation, and edema in mouse lung tissues caused by co-infection with IAV and IBV (Fig. 2D). We further observed that EIDD-2801 treatment significantly reduced the lung index and histopathological scores (Fig. 2E, F). Consistent with Western Blot analysis, the viral load in mouse nasal turbinates was significantly decreased by 1.5 to 2 Log10 in both the EIDD-2801 and baloxavir treatment groups (Fig. 2G). These findings demonstrate that EIDD-2801 effectively protects mice against lethal mixed infections of IAV and IBV.

Fig. 2
figure 2

EIDD-2801 protects mice from lethal co-infection of influenza A virus and influenza B virus. Mice co-infected with both H1N1-UI182 and IBV/S9-MD were monitored daily for clinical signs. A Body weight of infected mice. B Survival curves of infected mice. C Western Blot analysis of IAV M and IBV M protein expression in lung tissue homogenates (n = 5). The expression of β-actin was used as loading control D Representative images and photomicrographs of lung tissue (H&E staining) from mice co-infected with H1N1-UI182 and IBV/S9-MD. E Lung index of infected mice. F Histopathological scores of the lung tissues. G Viral titers in turbinates collected from euthanized mice at 5 dpi. Differences were considered significant at (**) 0.001 < P < 0.01, (***) P < 0.001, “ns” denotes no significance

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In vivo efficacy of EIDD-2801 against co-infection of influenza A virus and SARS-CoV-2

EIDD-2801 has been approved for clinical trials targeting SARS-CoV-2 and demonstrated strong antiviral activity against multiple variants [42,43,44,45]. Given its potent antiviral effects against influenza viruses, assessing its therapeutic potential in combating co-infections of influenza A virus and SARS-CoV-2 is of great importance. Firstly, in vitro experiments demonstrated that EIDD-2801 significantly reduced the viral load of multiple variants in a dose-dependent manner in the supernatants of Vero-E6 (Fig. S2A) and Huh7 (Fig. S2B) cells, These variants included isolated BJ01 strain, Delta variant, and Omicron variants (BA.2, BF.7, and XBB.1.9.1). Next, H1N1-UI182 and Omicron BF.7 were selected to establish a co-infection model in golden hamsters. Compared with the blank control group, infected hamsters exhibited substantial body weight loss within the first 4 days post-infection, whereas EIDD-2801 treatment moderately alleviated this weight loss (Fig. 3A). Macroscopic analysis of extracted lungs revealed that extensive hemorrhage (marked by the black circles), pulmonary swelling and pathological damage caused by mixed viral infection were enormously alleviated following oral administration of EIDD-2801 (Fig. 3B), Quantitative analysis further demonstrated a marked reduction in lung index and histopathological severity score compared to virus control (Fig. 3C, D). Then we quantified viral RNA copy numbers in tracheas, lungs, turbinates and nasal lavage. qRT-PCR analysis revealed reductions in viral loads across all tissues. For SARS-CoV-2, EIDD-2801 treatment decreased ORF1ab RNA levels by 1 Log10 in tracheas, 1.5 Log10 in lungs, and 1 Log10 in turbinates compared to virus control (Fig. 3E). Notably, ORF1ab RNA copies in nasal lavage exhibited a reduction, with 1.3 Log10 decrease at 2 dpi and 1.2 Log10 decrease at 4 dpi (Fig. 3F). Similarly, EIDD-2801 treatment reduced H1N1 M gene RNA levels by 2 log10 in tracheas, 1.5 log10 in lungs, and 1 log10 in turbinates (Fig. 3G). Furthermore, H1N1 M gene RNA copies in nasal lavage exhibited a decline, with reductions of 1.6 Log10 at 2 dpi and 1.3 Log10 at 4 dpi (Fig. 3H). These results suggested that orally administered EIDD-2801 could not only inhibit viral replication of influenza A virus and SARS-CoV-2 but also markedly improve prognosis level of mixed infection of these two viruses.

Fig. 3
figure 3

EIDD-2801 improves prognosis of mice co-infected with influenza virus and SARS-CoV-2. Mice co-infected with both H1N1-UI182 and Omicron BF.7 were monitored daily for clinical signs. A Body weight change of co-infected golden hamsters. B Representative images of lung tissue are presented, with photomicrographs showing histopathological changes (scale bars = 200 μm). C Lung index of hamsters 3 days after infection. D Histopathological scores of hamster lungs. E Viral RNA copy numbers of SARS-CoV-2 ORF1ab RNA in tracheas, lungs and turbinates. F Viral RNA copy numbers of SARS-CoV-2 ORF1ab RNA in nasal lavages. G Viral RNA copy numbers of H1N1 M gene in tracheas, lungs and turbinates. G Viral RNA copy numbers of H1N1 M gene in nasal lavages. Differences were considered significant at (**) 0.001 < P < 0.01, (***) P < 0.001

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Discussion

Multiple respiratory viruses, including influenza virus and coronavirus, are primarily transmitted through respiratory droplets and close contact and their co-circulation can exacerbate the severity of respiratory infections. Among these, influenza A and B viruses, which have posed significant public health challenges during the COVID-19 pandemic are highly prone to co-infect with SARS-CoV-2. Such co-infections may amplify pathogenic effects due to synergistic interactions between viral mechanisms, leading to increased disease severity [46,47,48]. Positive antiviral strategies including mask-wearing, social distancing, and vaccines were proven to be effective in prevention and reducing morbidity and mortality of patients [49, 50]. However, the effectiveness of vaccines may wane over time because of the high variability and immune evasion capacity of influenza viruses and SARS-CoV-2 Omicron subvariants [51, 52]. Therefore, it is essential to develop effective and broad-spectrum antiviral agents against mono-infection and co-infection of influenza viruses and SARS-CoV-2 strains. Currently, massive drug development against these viruses has been undertaken, no fully effective agents have been approved by FDA for the treatment of co-infection of IAV and SARS-CoV-2. Take S protein-specific neutralizing antibodies (nAbs) for instance, are ineffective against influenza virus infection and were restricted in the therapeutics against newly emerging Omicron variants and subvariants [53,54,55].

EIDD-2801 initially emerged as a possible antiviral for alphaviruses and influenza viruses due to its conspicuous effect in cell cultures. Earlier studies showed that EIDD-2801 is efficacious against influenza A and B viruses in human airway epithelial cultures. In addition, the drug demonstrated reduced shed virus titers and inflammatory infiltrates in nasal lavages and revealed a high genetic barrier to resistance in the ferret model of influenza when administered in therapeutic dosing [29, 56]. In addition, EIDD-2801 displayed an inhibitory effect on the replication and transmission of influenza viruses in guinea pig and mouse models [30]. This ribonucleoside analog has also shown in vitro and in vivo activity against a broad range of viruses including flaviviruses, filoviruses, pneumoviruses, orthomyxoviruses, and coronaviruses by increasing the rate of mutation in the viral genome of virus to induce random low-frequency nucleoside transitions and viral error catastrophe and was in preclinical development for the treatment of influenza [30, 57, 58]. The antiviral activity against SARS-CoV-2 was quickly confirmed in precious SARS-CoV-2 clinical trials during the COVID-19 pandemic, and EIDD-2801 was authorized by EUA for treatment of COVID-19 as an oral antiviral [32, 35, 57]. EIDD-2801 retained activity against all major SARS-CoV-2 variants and can delivered orally due to its great pharmacokinetic properties compared with other approved antiviral treatments such as remdesivir and reconvalescent serum which are poorly suitable for transmission control [32, 59, 60].

In this study, we confirmed the antiviral activity of EIDD-2801 against divergent authentic influenza virus strains in vitro, as well as therapeutic in vivo effects against mono-infection and co-infection of mouse-adapted influenza A virus and B virus in BALB/c mice. EIDD-2801 is potent in inhibiting infection of SARS-CoV-2 variants and subvariants such as BA.2, BF.7, and XBB which are resistant to most nAbs and antibodies in the serum of SARS-CoV-2 patients or vaccinated people. Furthermore, the results of co-infection of IAV and SARS-CoV-2 in golden hamsters showed that EIDD-2801 can dramatically reduce viral titers in tracheas, lungs, and turbinates and alleviate lung lesions. Collectively, we have demonstrated that EIDD-2801 exhibits significant antiviral activity against divergent influenza virus and SARS-CoV-2 strains, and strong protection for lethal mono-infection or co-infection of IAV and IBV, as well as non-lethal co-infection of IAV and SARS-CoV-2. Therefore, EIDD-2801 is a promising candidate as a broad-spectrum drug for the therapeutics and prevention of single infection and co-infection of IAV and SARS-CoV-2.

Conclusions

Overall, this study demonstrates that EIDD-2801 exhibits prominent antiviral activity against influenza virus and SARS-CoV-2. These results indicate that oral administration of EIDD-2801 provides significant protection and reduces viral burden during lethal challenges involving both mono-infection and co-infection with influenza virus and SARS-CoV-2, tthereby offering valuable support for the development of broad-spectrum antiviral therapies. Currently, research on EIDD-2801’s activity against other common respiratory viruses, such as respiratory syncytial virus (RSV), human metapneumovirus (hMPV), and others, remains limited. To fully explore its potential as a broad-spectrum antiviral agent, further studies are needed to systematically evaluate its antiviral spectrum against a wider range of respiratory pathogens.

Data availability

The datasets used and/or analyzed during the current study are available from the corresponding author on reasonable request.

References

  1. Li Q, Guan X, Wu P, Wang X, Zhou L, Tong Y, et al. Early transmission dynamics in Wuhan, China, of novel Coronavirus–Infected pneumonia. N Engl J Med. 2020;382:1199–207. https://doiorg.publicaciones.saludcastillayleon.es/10.1056/NEJMoa2001316.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  2. Huang C, Wang Y, Li X, Ren L, Zhao J, Hu Y, et al. Clinical features of patients infected with 2019 novel coronavirus in Wuhan, China. Lancet. 2020;395:497–506. https://doiorg.publicaciones.saludcastillayleon.es/10.1016/S0140-6736(20)30183-5.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  3. Harrison AG, Lin T, Wang P. Mechanisms of SARS-CoV-2 transmission and pathogenesis. Trends Immunol. 2020;41:1100–15. https://doiorg.publicaciones.saludcastillayleon.es/10.1016/j.it.2020.10.004.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  4. Sia SF, Yan L-M, Chin AWH, Fung K, Choy K-T, Wong AYL, et al. Pathogenesis and transmission of SARS-CoV-2 in golden hamsters. Nature. 2020;583:834–8. https://doiorg.publicaciones.saludcastillayleon.es/10.1038/s41586-020-2342-5.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  5. Chan JF-W, Zhang AJ, Yuan S, Poon VK-M, Chan CC-S, Lee AC-Y, et al. Simulation of the clinical and pathological manifestations of coronavirus disease 2019 (COVID-19) in a golden Syrian Hamster model: implications for disease pathogenesis and transmissibility. Clin Infect Dis. 2020;ciaa325. https://doiorg.publicaciones.saludcastillayleon.es/10.1093/cid/ciaa325.

  6. Mlcochova P, Kemp SA, Dhar MS, Papa G, Meng B, Ferreira IATM, et al. SARS-CoV-2 B.1.617.2 Delta variant replication and immune evasion. Nature. 2021;599:114–9. https://doiorg.publicaciones.saludcastillayleon.es/10.1038/s41586-021-03944-y.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  7. Garcia-Beltran WF, Lam EC, St. Denis K, Nitido AD, Garcia ZH, Hauser BM, et al. Multiple SARS-CoV-2 variants escape neutralization by vaccine-induced humoral immunity. Cell. 2021;184:2372–e23839. https://doiorg.publicaciones.saludcastillayleon.es/10.1016/j.cell.2021.03.013.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  8. Zhou D, Dejnirattisai W, Supasa P, Liu C, Mentzer AJ, Ginn HM, et al. Evidence of escape of SARS-CoV-2 variant B.1.351 from natural and vaccine-induced Sera. Cell. 2021;184:2348–e23616. https://doiorg.publicaciones.saludcastillayleon.es/10.1016/j.cell.2021.02.037.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  9. Ong SWX, Chiew CJ, Ang LW, Mak TM, Cui L, Toh MPHS, et al. Clinical and virological features of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) variants of concern: A retrospective cohort study comparing B.1.1.7 (Alpha), B.1.351 (Beta), and B.1.617.2 (Delta). Clin Infect Dis. 2022;75:e1128–36. https://doiorg.publicaciones.saludcastillayleon.es/10.1093/cid/ciab721.

    Article  PubMed  Google Scholar 

  10. Andrews N, Stowe J, Kirsebom F, Toffa S, Rickeard T, Gallagher E, et al. Covid-19 vaccine effectiveness against the Omicron (B.1.1.529) variant. N Engl J Med. 2022;386:1532–46. https://doiorg.publicaciones.saludcastillayleon.es/10.1056/NEJMoa2119451.

    Article  CAS  PubMed  Google Scholar 

  11. Planas D, Veyer D, Baidaliuk A, Staropoli I, Guivel-Benhassine F, Rajah MM, et al. Reduced sensitivity of SARS-CoV-2 variant Delta to antibody neutralization. Nature. 2021;596:276–80. https://doiorg.publicaciones.saludcastillayleon.es/10.1038/s41586-021-03777-9.

    Article  CAS  PubMed  Google Scholar 

  12. Qu P, Xu K, Faraone JN, Goodarzi N, Zheng Y-M, Carlin C, et al. Immune evasion, infectivity, and fusogenicity of SARS-CoV-2 BA.2.86 and flip variants. Cell. 2024;187:585–e5956. https://doiorg.publicaciones.saludcastillayleon.es/10.1016/j.cell.2023.12.026.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  13. Weisblum Y, Schmidt F, Zhang F, DaSilva J, Poston D, Lorenzi JC, et al. Escape from neutralizing antibodies by SARS-CoV-2 Spike protein variants. eLife. 2020;9:e61312. https://doiorg.publicaciones.saludcastillayleon.es/10.7554/eLife.61312.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  14. Kim D, Quinn J, Pinsky B, Shah NH, Brown I. Rates of Co-infection between SARS-CoV-2 and other respiratory pathogens. JAMA 2020;323:2085. https://doiorg.publicaciones.saludcastillayleon.es/10.1001/jama.2020.6266

  15. Kim KW, Deveson IW, Pang CNI, Yeang M, Naing Z, Adikari T, et al. Respiratory viral co-infections among SARS-CoV-2 cases confirmed by Virome capture sequencing. Sci Rep. 2021;11:3934. https://doiorg.publicaciones.saludcastillayleon.es/10.1038/s41598-021-83642-x.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  16. Lai C-C, Wang C-Y, Hsueh P-R. Co-infections among patients with COVID-19: the need for combination therapy with non-anti-SARS-CoV-2 agents? J Microbiol Immunol Infect. 2020;53:505–12. https://doiorg.publicaciones.saludcastillayleon.es/10.1016/j.jmii.2020.05.013.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  17. Lansbury L, Lim B, Baskaran V, Lim WS. Co-infections in people with COVID-19: a systematic review and meta-analysis. J Infect. 2020;81:266–75. https://doiorg.publicaciones.saludcastillayleon.es/10.1016/j.jinf.2020.05.046.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  18. Chan KF, Carolan LA, Korenkov D, Druce J, McCaw J, Reading PC, et al. Investigating viral interference between influenza A virus and human respiratory syncytial virus in a ferret model of infection. J Infect Dis. 2018;218:406–17. https://doiorg.publicaciones.saludcastillayleon.es/10.1093/infdis/jiy184.

    Article  CAS  PubMed  Google Scholar 

  19. Goka E, Vallely P, Mutton K, Klapper P. Influenza A viruses dual and multiple infections with other respiratory viruses and risk of hospitalisation and mortality. Influenza Resp Viruses. 2013;7:1079–87. https://doiorg.publicaciones.saludcastillayleon.es/10.1111/irv.12020.

    Article  Google Scholar 

  20. Zhong P, Zhang H, Chen X, Lv F. Clinical characteristics of the lower respiratory tract infection caused by a single infection or coinfection of the human parainfluenza virus in children. J Med Virol. 2019;91:1625–32. https://doiorg.publicaciones.saludcastillayleon.es/10.1002/jmv.25499.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  21. Murphy K. SARS CoV-2 detection from upper and lower respiratory tract specimens. Chest. 2020;158:1804–5. https://doiorg.publicaciones.saludcastillayleon.es/10.1016/j.chest.2020.07.061.

    Article  CAS  PubMed  Google Scholar 

  22. Van Riel D, Munster VJ, De Wit E, Rimmelzwaan GF, Fouchier RAM, Osterhaus ADME, et al. Human and avian influenza viruses target different cells in the lower respiratory tract of humans and other mammals. Am J Pathol. 2007;171:1215–23. https://doiorg.publicaciones.saludcastillayleon.es/10.2353/ajpath.2007.070248.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  23. Yan R, Zhang Y, Li Y, Xia L, Guo Y, Zhou Q. Structural basis for the recognition of SARS-CoV-2 by full-length human ACE2. Science. 2020;367:1444–8. https://doiorg.publicaciones.saludcastillayleon.es/10.1126/science.abb2762.

    Article  PubMed  PubMed Central  Google Scholar 

  24. Bai L, Zhao Y, Dong J, Liang S, Guo M, Liu X, et al. Coinfection with influenza A virus enhances SARS-CoV-2 infectivity. Cell Res. 2021;31:395–403. https://doiorg.publicaciones.saludcastillayleon.es/10.1038/s41422-021-00473-1.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  25. Huang Y, Skarlupka AL, Jang H, Blas-Machado U, Holladay N, Hogan RJ, et al. SARS-CoV-2 and influenza A virus coinfections in ferrets. J Virol. 2022;96:e01791–21. https://doiorg.publicaciones.saludcastillayleon.es/10.1128/jvi.01791-21.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  26. Zheng X, Wang H, Su Z, Li W, Yang D, Deng F, et al. Co-infection of SARS-CoV-2 and influenza virus in early stage of the COVID-19 epidemic in Wuhan, China. J Infect. 2020;81:e128–9. https://doiorg.publicaciones.saludcastillayleon.es/10.1016/j.jinf.2020.05.041.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  27. Wu X, Cai Y, Huang X, Yu X, Zhao L, Wang F, et al. Co-infection with SARS-CoV-2 and influenza A virus in patient with pneumonia, China. Emerg Infect Dis. 2020;26:1324–6. https://doiorg.publicaciones.saludcastillayleon.es/10.3201/eid2606.200299.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  28. Zheng J, Chen F, Wu K, Wang J, Li F, Huang S, et al. Clinical and virological impact of single and dual infections with influenza A (H1N1) and SARS-CoV-2 in adult inpatients. PLoS Negl Trop Dis. 2021;15:e0009997. https://doiorg.publicaciones.saludcastillayleon.es/10.1371/journal.pntd.0009997.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  29. Toots M, Yoon J-J, Cox RM, Hart M, Sticher ZM, Makhsous N, et al. Characterization of orally efficacious influenza drug with high resistance barrier in ferrets and human airway epithelia. Sci Transl Med. 2019;11:eaax5866. https://doiorg.publicaciones.saludcastillayleon.es/10.1126/scitranslmed.aax5866.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  30. Yoon J-J, Toots M, Lee S, Lee M-E, Ludeke B, Luczo JM, et al. Orally efficacious Broad-Spectrum ribonucleoside analog inhibitor of influenza and respiratory syncytial viruses. Antimicrob Agents Chemother. 2018;62:e00766–18. https://doiorg.publicaciones.saludcastillayleon.es/10.1128/AAC.00766-18.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  31. Fischer WA, Eron JJ, Holman W, Cohen MS, Fang L, Szewczyk LJ, et al. A phase 2a clinical trial of molnupiravir in patients with COVID-19 shows accelerated SARS-CoV-2 RNA clearance and elimination of infectious virus. Sci Transl Med. 2022;14:eabl7430. https://doiorg.publicaciones.saludcastillayleon.es/10.1126/scitranslmed.abl7430.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  32. Sheahan TP, Sims AC, Zhou S, Graham RL, Pruijssers AJ, Agostini ML, et al. An orally bioavailable broad-spectrum antiviral inhibits SARS-CoV-2 in human airway epithelial cell cultures and multiple coronaviruses in mice. Sci Transl Med. 2020;12:eabb5883. https://doiorg.publicaciones.saludcastillayleon.es/10.1126/scitranslmed.abb5883.

    Article  CAS  PubMed  Google Scholar 

  33. Johnson MG, Puenpatom A, Moncada PA, Burgess L, Duke ER, Ohmagari N, et al. Effect of molnupiravir on biomarkers, respiratory interventions, and medical services in COVID-19: A randomized, Placebo-Controlled trial. Ann Intern Med. 2022;175:1126–34. https://doiorg.publicaciones.saludcastillayleon.es/10.7326/M22-0729.

    Article  PubMed  Google Scholar 

  34. Strizki JM, Gaspar JM, Howe JA, Hutchins B, Mohri H, Nair MS, et al. Molnupiravir maintains antiviral activity against SARS-CoV-2 variants and exhibits a high barrier to the development of resistance. Antimicrob Agents Chemother. 2024;68:e00953–23. https://doiorg.publicaciones.saludcastillayleon.es/10.1128/aac.00953-23.

    Article  CAS  PubMed  Google Scholar 

  35. Wahl A, Gralinski LE, Johnson CE, Yao W, Kovarova M, Dinnon KH, et al. SARS-CoV-2 infection is effectively treated and prevented by EIDD-2801. Nature. 2021;591:451–7. https://doiorg.publicaciones.saludcastillayleon.es/10.1038/s41586-021-03312-w.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  36. Kabinger F, Stiller C, Schmitzová J, Dienemann C, Kokic G, Hillen HS, et al. Mechanism of molnupiravir-induced SARS-CoV-2 mutagenesis. Nat Struct Mol Biol. 2021;28:740–6. https://doiorg.publicaciones.saludcastillayleon.es/10.1038/s41594-021-00651-0.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  37. Gordon CJ, Tchesnokov EP, Schinazi RF, Götte M. Molnupiravir promotes SARS-CoV-2 mutagenesis via the RNA template. J Biol Chem. 2021;297:100770. https://doiorg.publicaciones.saludcastillayleon.es/10.1016/j.jbc.2021.100770.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  38. Agostini ML, Pruijssers AJ, Chappell JD, Gribble J, Lu X, Andres EL, et al. Small-molecule antiviral _-D-N4-hydroxycytidine inhibits a proofreading-intact coronavirus with a high genetic barrier to resistance. J Virol. 2019;93:e01348-19. https://doiorg.publicaciones.saludcastillayleon.es/10.1128/JVI.01348-19

  39. Luo R, Lv C, Wang T, Deng X, Sima M, Guo J, et al. A potential Chinese medicine monomer against influenza A virus and influenza B virus: isoquercitrin. Chin Med. 2023;18:144. https://doiorg.publicaciones.saludcastillayleon.es/10.1186/s13020-023-00843-4.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  40. Wang Z, Yang S, Dai Q, Guo X, Li Y, Li W, et al. In vitro and in vivo efficacy of molnupiravir against Zika virus infections. Virol Sin. 2023;38:639–42. https://doiorg.publicaciones.saludcastillayleon.es/10.1016/j.virs.2023.05.011.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  41. Toots M, Yoon J-J, Hart M, Natchus MG, Painter GR, Plemper RK. Quantitative efficacy paradigms of the influenza clinical drug candidate EIDD-2801 in the ferret model. Translational Res. 2020;218:16–28. https://doiorg.publicaciones.saludcastillayleon.es/10.1016/j.trsl.2019.12.002.

    Article  Google Scholar 

  42. Imai M, Ito M, Kiso M, Yamayoshi S, Uraki R, Fukushi S, et al. Efficacy of antiviral agents against Omicron subvariants BQ.1.1 and XBB. N Engl J Med. 2023;388:89–91. https://doiorg.publicaciones.saludcastillayleon.es/10.1056/NEJMc2214302.

    Article  PubMed  Google Scholar 

  43. Vangeel L, Chiu W, De Jonghe S, Maes P, Slechten B, Raymenants J, et al. Remdesivir, molnupiravir and nirmatrelvir remain active against SARS-CoV-2 Omicron and other variants of concern. Antiviral Res. 2022;198:105252. https://doiorg.publicaciones.saludcastillayleon.es/10.1016/j.antiviral.2022.105252.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  44. Lieber CM, Cox RM, Sourimant J, Wolf JD, Juergens K, Phung Q, et al. SARS-CoV-2 VOC type and biological sex affect molnupiravir efficacy in severe COVID-19 Dwarf hamster model. Nat Commun. 2022;13:4416. https://doiorg.publicaciones.saludcastillayleon.es/10.1038/s41467-022-32045-1.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  45. Uraki R, Kiso M, Imai M, Yamayoshi S, Ito M, Fujisaki S, et al. Therapeutic efficacy of monoclonal antibodies and antivirals against SARS-CoV-2 Omicron BA.1 in Syrian hamsters. Nat Microbiol. 2022;7:1252–8. https://doiorg.publicaciones.saludcastillayleon.es/10.1038/s41564-022-01170-4.

    Article  CAS  PubMed  Google Scholar 

  46. Kinoshita T, Watanabe K, Sakurai Y, Nishi K, Yoshikawa R, Yasuda J. Co-infection of SARS-CoV-2 and influenza virus causes more severe and prolonged pneumonia in hamsters. Sci Rep. 2021;11:21259. https://doiorg.publicaciones.saludcastillayleon.es/10.1038/s41598-021-00809-2.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  47. Zhang AJ, Lee AC-Y, Chan JF-W, Liu F, Li C, Chen Y, et al. Coinfection by severe acute respiratory syndrome coronavirus 2 and influenza A(H1N1)pdm09 virus enhances the severity of pneumonia in golden Syrian hamsters. Clin Infect Dis. 2021;72:e978–92. https://doiorg.publicaciones.saludcastillayleon.es/10.1093/cid/ciaa1747.

    Article  CAS  PubMed  Google Scholar 

  48. Achdout H, Vitner EB, Politi B, Melamed S, Yahalom-Ronen Y, Tamir H, et al. Increased lethality in influenza and SARS-CoV-2 coinfection is prevented by influenza immunity but not SARS-CoV-2 immunity. Nat Commun. 2021;12:5819. https://doiorg.publicaciones.saludcastillayleon.es/10.1038/s41467-021-26113-1.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  49. Chen Y, Zhao X, Zhou H, Zhu H, Jiang S, Wang P. Broadly neutralizing antibodies to SARS-CoV-2 and other human coronaviruses. Nat Rev Immunol. 2023;23:189–99. https://doiorg.publicaciones.saludcastillayleon.es/10.1038/s41577-022-00784-3.

    Article  CAS  PubMed  Google Scholar 

  50. McCauley J, Barr IG, Nolan T, Tsai T, Rockman S, Taylor B. The importance of influenza vaccination during the COVID-19 pandemic. Influenza Resp Viruses. 2022;16:3–6. https://doiorg.publicaciones.saludcastillayleon.es/10.1111/irv.12917.

    Article  CAS  Google Scholar 

  51. Wang L, Jiao F, Jiang H, Yang Y, Huang Z, Wang Q, et al. Fusogenicity of SARS-CoV-2 BA.2.86 subvariant and its sensitivity to the prokaryotic Recombinant EK1 peptide. Cell Discov. 2024;10:6. https://doiorg.publicaciones.saludcastillayleon.es/10.1038/s41421-023-00631-2.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  52. Leung C, Konya L, Su L. Postpandemic immunity debt of influenza in the USA and England: an interrupted time series study. Public Health. 2024;227:239–42. https://doiorg.publicaciones.saludcastillayleon.es/10.1016/j.puhe.2023.12.009.

    Article  CAS  PubMed  Google Scholar 

  53. Park S-J, Yu K-M, Kim Y-I, Kim S-M, Kim E-H, Kim S-G et al. Antiviral Efficacies of FDA-Approved Drugs against SARS-CoV-2 Infection in Ferrets. mBio. 2020;11:e01114-20. https://doiorg.publicaciones.saludcastillayleon.es/10.1128/mBio.01114-20

  54. Beigel JH, Tomashek KM, Dodd LE, Mehta AK, Zingman BS, Kalil AC, et al. Remdesivir for the treatment of Covid-19 — Final report. N Engl J Med. 2020;383:1813–26. https://doiorg.publicaciones.saludcastillayleon.es/10.1056/NEJMoa2007764.

    Article  CAS  PubMed  Google Scholar 

  55. Li G, Hilgenfeld R, Whitley R, De Clercq E. Therapeutic strategies for COVID-19: progress and lessons learned. Nat Rev Drug Discov. 2023;22:449–75. https://doiorg.publicaciones.saludcastillayleon.es/10.1038/s41573-023-00672-y.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  56. Toots M, Yoon J-J, Hart M, Natchus MG, Painter GR, Plemper RK. Quantitative efficacy paradigms of the influenza clinical drug candidate EIDD-2801 in the ferret model. Transl Res. 2020;218:16–28. https://doiorg.publicaciones.saludcastillayleon.es/10.1016/j.trsl.2019.12.002.

    Article  PubMed  Google Scholar 

  57. Painter GR, Natchus MG, Cohen O, Holman W, Painter WP. Developing a direct acting, orally available antiviral agent in a pandemic: the evolution of molnupiravir as a potential treatment for COVID-19. Curr Opin Virol. 2021;50:17–22. https://doiorg.publicaciones.saludcastillayleon.es/10.1016/j.coviro.2021.06.003.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  58. Stuyver LJ, Whitaker T, McBrayer TR, Hernandez-Santiago BI, Lostia S, Tharnish PM, et al. Ribonucleoside analogue that blocks replication of bovine viral diarrhea and hepatitis C viruses in culture. Antimicrob Agents Chemother. 2003;47:244–54. https://doiorg.publicaciones.saludcastillayleon.es/10.1128/AAC.47.1.244-254.2003.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  59. Humeniuk R, Mathias A, Cao H, Osinusi A, Shen G, Chng E, et al. Safety, tolerability, and pharmacokinetics of Remdesivir, an antiviral for treatment of COVID-19, in healthy subjects. Clin Translational Sci. 2020;13:896–906. https://doiorg.publicaciones.saludcastillayleon.es/10.1111/cts.12840.

    Article  CAS  Google Scholar 

  60. Painter WP, Holman W, Bush JA, Almazedi F, Malik H, Eraut NCJE, et al. Human safety, tolerability, and pharmacokinetics of Molnupiravir, a novel Broad-Spectrum oral antiviral agent with activity against SARS-CoV-2. Antimicrob Agents Chemother. 2021;65:e02428–20. https://doiorg.publicaciones.saludcastillayleon.es/10.1128/AAC.02428-20.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

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Acknowledgements

The authors would like to thank the research team of Researcher Yuwei Gao (Changchun Veterinary Research Institute, Chinese Academy of Agricultural Sciences, State Key Laboratory of Pathogen and Biosecurity, Key Laboratory of Jilin Province for Zoonosis Prevention and Control, Changchun, China).

Funding

This study was carried out as part of our routine work.

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  1. Bingshuo Qian and Rongbo Luo contributed equally to this work.

Authors and Affiliations

  1. Changchun Veterinary Research Institute, Chinese Academy of Agricultural Sciences, Changchun, 130122, China

    Bingshuo Qian, Rongbo Luo, Beilei Shen, Lingjun Fan, Junkui Zhang, Shijun Zhang, Yan Sun, Xiuwen Deng & Yuwei Gao

  2. School of Pharmacy, Henan University, Kaifeng, 475004, China

    Bingshuo Qian, Junkui Zhang & Xiaobin Pang

  3. National Engineering Research Center for the Emergency Drug, Beijing Institute of Pharmacology and Toxicology, Beijing, 100850, China

    Wu Zhong

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Contributions

Bingshuo Qian: data curation, formal analysis, methodology, software, validation, visualization, writing-original draft. Rongbo Luo: formal analysis, investigation, software, visualization. Beilei Shen: investigation, resources. Lingjun Fan: formal analysis, reviewing and editing. Junkui Zhang: reviewing and editing. Shijun Zhang: reviewing and editing. Yan Sun: reviewing and editing. Xiuwen Deng: reviewing and editing. Xiaobin Pang: project administration, writing-review and editing. Wu Zhong: project administration, writing-review and editing. Yuwei Gao: conceptualization, funding acquisition, project administration, supervision, reviewing and editing.

Corresponding authors

Correspondence to Xiaobin Pang, Wu Zhong or Yuwei Gao.

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Ethics approval and consent to participate

This study and included experimental procedures were conducted in accordance with the Declaration of Helsinki, and approved by the institutional animal care and use committee of Changchun Veterinary Research Institute (IACUC-AMMS-11-2023-001). All animal housing and experiments were performed in strict accordance with ethical guidelines.

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Written informed consent for publication was obtained from all participants.

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Qian, B., Luo, R., Shen, B. et al. EIDD-2801 resists to infection and co-infection of SARS-CoV-2 and influenza virus. Virol J 22, 126 (2025). https://doiorg.publicaciones.saludcastillayleon.es/10.1186/s12985-025-02755-1

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