Antiviral efficacy of cerium oxide nanoparticles (preprint)

Nanomaterials have been proposed as good candidates for the elimination of viruses due to their multimodal mechanisms of action. Here, we tested the antiviral potential of cerium dioxide (CeO 2 ) nanoparticles which is largely unexplored.Two types of nano-CeO 2 particles with opposing surface charge, nano-CeO 2 (+) and nano-CeO 2 (-), were assessed for their capability to decrease the plaque forming units (PFU) of four enveloped and two non-enveloped viruses during 1 h exposure. Statistically significant antiviral activity of nano-CeO 2 towards enveloped coronavirus SARS-CoV-2 and influenza virus A/WSN/1933 was registered already at 20 mg Ce/l. Significant decrease in PFU of other two enveloped viruses, transmissible gastroenteritis virus TGEV and bacteriophage φ6 was evidenced at 200 mg Ce/l. For all the enveloped viruses the maximum reduction of PFU after 1 h exposure to nano-CeO 2 exceeded 2 logs, that has been considered as the lowest biologically meaningful activity in antiviral applications. For most of the enveloped viruses, 1 h exposure to nano-CeO 2 resulted in ≥ 4 log reduction in PFU. As expected, the sensitivity of non-enveloped viruses towards nano-CeO 2 was significantly lower than that of enveloped viruses. Until the highest tested concentration, 2000 mg Ce/l neither of the nano-CeO 2 showed an effect on picornavirus EMCV and only nano-CeO 2 (-) caused a slight non-monotonic response in MS2 bacteriophage.Parallel testing of antiviral activity of Ce 3+ ions and SiO 2 nanoparticles allows to conclude that nano-CeO 2 activity was due to neither released Ce 3+ ions nor nonspecific effects of any nanosized particulate. Interestingly, we did not evidence any significant antiviral activity of Ag nanoparticles at 1 h exposure. This result referred to notably higher antiviral activity of nano-CeO 2 compared with nanosilver that has been generally considered as active against viruses. Although exhibiting antiviral effects, the antibacterial activity of nano-CeO 2 was very low. These results along with non-existent cytotoxicity nano-CeO 2 allow us to propose nano-CeO 2 for specific antiviral applications.

Bovine Colostrum Derived Antibodies Against SARS-CoV-2 Show Great Potential to Serve as a Prophylactic Agent (preprint)

Severe acute respiratory syndrome Coronavirus 2 (SARS-CoV-2) until now imposes a serious burden to health systems globally. Despite worldwide vaccination, social distancing and wearing masks, the spread of the virus is still ongoing. One of the mechanisms how neutralizing antibodies (NAbs) block virus entry into cells encompasses interaction inhibition between the cell surface receptor angiotensin-converting enzyme 2 (ACE2) and the spike (S) protein of SARS-CoV-2. SARS-CoV-2 specific NAb development can be induced in the blood of cattle. Pregnant cows produce NAbs upon immunization, and antibodies move into the colostrum just before calving. Here we immunized cows with SARS-CoV-2 S1 receptor binding domain (RBD) protein in proper adjuvant solutions, followed by one boost with SARS-CoV-2 trimeric S protein, and purified immunoglobulins from colostrum. We demonstrate that this preparation indeed blocks interaction between the trimeric S protein and ACE2 in different in vitro assays. Moreover, we describe the formulation of purified immunoglobulin preparation into a nasal spray. When administered to human subjects, the formulation persists on the nasal mucosa for at least 4 hours as determined by a clinical study. Therefore, we are presenting a solution that shows great potential to serve as a prophylactic agent against SARS-CoV-2 infection as an additional measure to vaccination and wearing masks. Moreover, our technology allows for a rapid and versatile adaption for preparing prophylactic treatments against other diseases by using the defined characteristics of antibody movement into the colostrum.

Synergistic Interferon Alpha-based Drug Combinations Inhibit SARS-CoV-2 and other viral Infections in Vitro (preprint)

There is an urgent need for new antivirals with powerful therapeutic potential and tolerable side effects. In the present study, we found that recombinant human interferon-alpha (IFNa) triggered cell intrinsic and extrinsic antiviral responses and reduced replication of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) in human lung epithelial Calu-3 cells. However, IFNa alone was insufficient to completely abolish SARS-CoV-2 replication. Combinations of IFNa with camostat, remdesivir, EIDD-2801, cycloheximide or convalescent serum showed strong synergy and effectively inhibited SARS-CoV-2 infection. Additionally, we demonstrated synergistic antiviral activity of IFNa2a with pimodivir against influenza A virus (FluAV) infection in human lung epithelial A549 cells, as well as of IFNa2a with lamivudine against human immunodeficiency virus 1 (HIV-1) infection in human TZM-bl cells. Our results indicate that IFNa2a-based combinational therapies help to reduce drug dose and improve efficacy in comparison with monotherapies, making them attractive targets for further pre-clinical and clinical development.

Interferon alpha-based combinations suppress SARS-CoV-2 infection in vitro and in vivo (preprint)

There is an urgent need for new antivirals with powerful therapeutic potential and tolerable side effects. In the present study, we found that recombinant human interferon-alpha (IFNa) triggers intrinsic and extrinsic cellular antiviral responses, as well as reduces replication of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) in vitro. Although IFNa alone was insufficient to completely abolish SARS-CoV-2 replication, combinations of IFNa with remdesivir or other antiviral agents (EIDD-2801, camostat, cycloheximide, or convalescent serum) showed strong synergy and effectively inhibited SARS-CoV-2 infection in human lung epithelial Calu-3 cells. Furthermore, we showed that the IFNa-remdesivir combination suppressed virus replication in human lung organoids, and that its single prophylactic dose attenuated SARS-CoV-2 infection in lungs of Syrian hamsters. Transcriptome and metabolomic analyses showed that the combination of IFNa-remdesivir suppressed virus-mediated changes in infected cells, although it affected the homeostasis of uninfected cells. We also demonstrated synergistic antiviral activity of IFNa2a-based combinations against other virus infections in vitro. Altogether, our results indicate that IFNa2a-based combination therapies can achieve higher efficacy while requiring lower dosage compared to monotherapies, making them attractive targets for further pre-clinical and clinical development.

Seroprevalence of SARS-CoV-2 IgG antibodies in two regions of Estonia (KoroSero-EST-1) (preprint)

Background: In Estonia, during the first wave of COVID-19 total number of cases confirmed by PCR was 13.3/10,000, similar in most regions, including capital Tallinn, but in the hotspot of Estonian epidemic, an island Saaremaa, the cumulative incidence was 166.1/10,000. Aim: We aimed to determine the prevalence of SARS-CoV-2 IgG antibodies in these two regions, symptoms associated with infection and factors associated with antibody concentrations. Methods: Participants were selected using stratified (formed by age decades) random sampling and recruited by general practitioners. IgG were determined from sera by four assays. Symptoms of acute respiratory illness associated with seropositivity were analyzed by multiple correspondence analysis, antibody concentrations by multiple linear regression. Results: Total of 3608 individual were invited and 1960 recruited From May 8 to July 31, 2020. Seroprevalence was 1.5% (95% confidence interval (CI) 0.9-2.5) and 6.3% (95% CI 5.0-7.9), infection fatality rate 0.1% (95% CI 0.0-0.2) and 1.3% (95% CI 0.4-2.1) in Tallinn and Saaremaa, respectively. Of seropositive subjects 19.2% (14/73) had acute respiratory illness. Fever, diarrhea and the absence of cough and runny nose were associated with seropositivity in individuals aged 50 or more years. IgG concentrations were higher if fever, difficulty breathing, shortness of breath, chest pain or diarrhea was present, or hospitalization required. Conclusion: Similarly to other European countries the seroprevalence of SARS-CoV-2 in Estonia was low even in the hotspot region Saaremaa suggesting that majority of population is still susceptible to SARS-CoV-2. Focusing only on respiratory symptoms may delay accurate diagnosis of SARS-CoV-2 infection.

Identification of novel antiviral drug combinations in vitro and tracking their development (preprint)

Combination therapies have become a standard for the treatment for HIV and HCV infections. They are advantageous over monotherapies due to better efficacy and reduced toxicity, as well as the ability to prevent the development of resistant viral strains and to treat viral co-infections. Here, we identify several new synergistic combinations against emerging and re-emerging viral infections in vitro. We observed synergistic activity of nelfinavir with investigational drug EIDD-2801 and convalescent serum against SARS-CoV-2 infection in human lung epithelial Calu-3 cells. We also demonstrated synergistic activity of vemurafenib combination with emetine, homoharringtonine, gemcitabine, or obatoclax against echovirus 1 infection in human lung epithelial A549 cells. We also found that combinations of sofosbuvir with brequinar and niclosamide were synergistic against HCV infection in hepatocyte derived Huh-7.5 cells, whereas combinations of monensin with lamivudine and tenofovir were synergistic against HIV-1 infection in human cervical TZM-bl cells. Finally, we present an online resource that summarizes novel and known antiviral drug combinations and their developmental status. Overall, the development of combinational therapies could have a global impact improving the preparedness and protection of the general population from emerging and re-emerging viral threats.

Antiviral options against SARS-CoV-2 infection (preprint)

As of June 2020, the number of people infected with severe acute respiratory coronavirus 2 (SARS-CoV-2) continues to skyrocket, with more than 6,5 million cases worldwide. Both the World Health Organization (WHO) and United Nations (UN) has highlighted the need for better control of SARS-CoV-2 infections. However, developing novel virus-specific vaccines, monoclonal antibodies and antiviral drugs against SARS-CoV-2 can be time-consuming and costly. Convalescent sera and safe-in-man broad-spectrum antivirals (BSAAs) are readily available treatment options. Here we developed a neutralization assay using SARS-CoV-2 strain and Vero-E6 cells. We identified most potent sera from recovered patients for treatment of SARS-CoV-2-infected patients. We also screened 136 safe-in-man broad-spectrum antivirals against SARS-CoV-2 infection in Vero-E6 cells and identified nelfinavir, salinomycin, amodiaquine, obatoclax, emetine and homoharringtonine. We found that combinations of virus-directed nelfinavir along with host-directed amodiaquine exhibited the highest synergy. Finally, we developed a website to disseminate the knowledge on available and emerging treatments of COVID-19.