Repurposing of poly-gamma-glutamic acid, immunotherapeutic drug for cervical intraepithelial neoplasia 1, as covid-19 treatment

Covid-19 virus variants identified so far are due to viral genetic diversity, genetic evolution, and variable infectivity, suggesting that high infection rates and high mortality rates may be contributed by these mutations. And it has been reported that the targeting strategies for innate immunity should be less vulnerable to viral evolution, variant emergence and resistance. Therefore, the most effective solution to Covid-19 infection has been proposed to prevent and treat severe exacerbation of patients with moderate disease by enhancing human immune responses such as NK cell and T cell. In previous studies, we demonstrated for the first time that gamma-PGA induced significant antitumor activity and antiviral activity by modulating NK cell-mediated cytotoxicity. Especially intranasal administration of gamma-PGA was found to effectively induce protective innate and CTL immune responses against viruses and we found out that gamma-PGA can be an effective treatment for cervical intraepithelial neoplasia 1 through phase 2b clinical trial. In this study, the possibility of gamma-PGA as a Covid-19 immune modulating agent was confirmed by animal experiments infected with Covid-19 viruses. After oral administration of gamma-PGA 300mug/mouse once a day for 5 days in a K18-hACE2 TG mouse model infected with SARS-CoV-2 (NCCP 43326;original strain) and SARS-CoV-2 (NCCP 43390;Delta variant), virus titer and clinical symptom improvement were confirmed. In the RjHan:AURA Syrian hamster model infected with SARS-CoV-2 (NCCP 49930;Delta variant), 350 or 550 mug/head of gamma-PGA was administered orally for 10 days once a day. The virus for infection was administered at 5 x 104 TCID50, and the titer of virus and the improvement of pneumonia lesions were measured to confirm the effectiveness in terms of prevention or treatment. In the mouse model infected with original Covid-19 virus stain, the weight loss was significantly reduced and the survival rate was also improved by the administration of gamma-PGA. And gamma-PGA alleviated the pneumonic lesions and reduced the virus titer of lung tissue in mice infected with delta variant. In the deltavariant virus infected hamster model, gamma-PGA showed statistically significant improvement of weight loss and lung inflammation during administration after infection. This is a promising result for possibility of Covid-19 therapeutics along with the efficacy results of mouse model, suggesting gammaPGA can be therapeutic candidate to modulate an innate immune response for Covid-19.

"Actionable" host genetic markers for repurposing "add-on therapy" in COVID-19

Background/Objectives: COVID-19 still represents a lifethreatening disease in individuals with a specific genetic background. We successfully applied a new Machine Learning method on WES data to extract a set of coding variants relevant for COVID- 19 severity. We aim to identify personalized add-on therapy. Method(s): A subset of identified variants, "actionable" by repurposed drugs, were functionally tested by in vitro and in vivo experiments. Result(s): Males with either rare loss of function variants in the TLR7 gene or L412F polymorphism in the TLR3 gene benefit from IFN-gamma, which is specifically defective in activated PBMCs, restoring innate immunity. Females heterozygous for rare variants in the ADAMTS13 gene and males with D603N homozygous polymorphism in the SELP gene benefit from Caplacizumab, which reduces vWF aggregation and thrombus formation. Males with either the low-frequency gain of function variant T201M in CYP19A1 gene or with poly-Q repeats >=23 in the AR gene benefit from Letrozole, an aromatase inhibitor, which restores normal testosterone levels, reducing inflammation and which rescues male golden hamsters from severe COVID-19. Conclusion(s): By adding these commonly used drugs to standard of care of selected patients, the rate of intubation is expected to decrease consistently, especially in patients with high penetrance rare genetic markers, mitigating the effect of the pandemic with a significant impact on the healthcare system.

COVID-associated pernio: Mechanistic insights to an abortive, seronegative SARS-CoV-2 infection

Introduction: SARS-CoV-2 replicates primarily in the airways but generates a systemic immune response mediated by Type I interferons (IFN-I). Pernio is a rare skin manifestation of disorders characterized by excessive IFN-I signalling. Although pernio increased in incidence during the pandemic, the relationship to SARS-CoV-2 remains controversial. Because of the pivotal nature of interferons in COVID-19 outcomes, pernio offers a window to investigate the biology underlying host resiliency to SARS-CoV-2 infection. Method(s): To further assess COVID-associated pernio, we characterized clinical samples from affected patients across 4 waves of the pandemic and investigated mechanistic feasibility in a rodent model. Patients were followed longitudinally with banking of blood and tissue. Golden hamsters were mock-treated or intra-nasally infected with SARS-CoV-2 and harvested at 3-and 30-days post-infection. Result(s): In affected tissue, immunophenotyping utilizing multiplex immunohistochemistry profiled a robust IFN-1 signature characterized by plasmacytoid dendritic cell activation. Viral RNA was detectable in a subset of cases using in situ hybridization for the SARS-CoV-2 S gene transcript. Profiling of the systemic immune response did not reveal a durable type 1 interferon signature. Consistent with previous literature, antibody and T-cell specific responses to SARS-CoV-2 were not detected. Nasopharyngeal SARS-CoV-2 inoculation in hamsters resulted in rapid dissemination of viral RNA and the generation of an IFN-I response that were both detectable in the paws of infected animals. Conclusion(s): Our data support a durable local IFN signature, with direct evidence of viral SARS-CoV-2 RNA in acral skin and suggest that COVID-associated pernio results from an abortive, seronegative SARS-CoV-2 infection.

The Omicron Sub-Variant BA.4 Displays a Remarkable Lack of Clinical Signs in a Golden Syrian Hamster Model of SARS-CoV-2 Infection.

The ongoing emergence of SARS-CoV-2 virus variants remains a source of concern because it is accompanied by the potential for increased virulence as well as evasion of immunity. Here we show that, although having an almost identical spike gene sequence as another Omicron variant (BA.5.2.1), a BA.4 isolate lacked all the typical disease characteristics of other isolates seen in the Golden Syrian hamster model despite replicating almost as effectively. Animals infected with BA.4 had similar viral shedding profiles to those seen with BA.5.2.1 (up to day 6 post-infection), but they all failed to lose weight or present with any other significant clinical signs. We hypothesize that this lack of detectable signs of disease during infection with BA.4 was due to a small (nine nucleotide) deletion (∆686-694) in the viral genome (ORF1ab) responsible for the production of non-structural protein 1, which resulted in the loss of three amino acids (aa 141-143).

The Highly Conserved Stem-Loop II Motif Is Dispensable for SARS-CoV-2.

The stem-loop II motif (s2m) is an RNA structural element that is found in the 3' untranslated region (UTR) of many RNA viruses, including severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). Though the motif was discovered over 25 years ago, its functional significance is unknown. In order to understand the importance of s2m, we created viruses with deletions or mutations of the s2m by reverse genetics and also evaluated a clinical isolate harboring a unique s2m deletion. Deletion or mutation of the s2m had no effect on growth in vitro or on growth and viral fitness in Syrian hamsters in vivo. We also compared the secondary structure of the 3' UTR of wild-type and s2m deletion viruses using selective 2'-hydroxyl acylation analyzed by primer extension and mutational profiling (SHAPE-MaP) and dimethyl sulfate mutational profiling and sequencing (DMS-MaPseq). These experiments demonstrate that the s2m forms an independent structure and that its deletion does not alter the overall remaining 3'-UTR RNA structure. Together, these findings suggest that s2m is dispensable for SARS-CoV-2. IMPORTANCE RNA viruses, including severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), contain functional structures to support virus replication, translation, and evasion of the host antiviral immune response. The 3' untranslated region of early isolates of SARS-CoV-2 contained a stem-loop II motif (s2m), which is an RNA structural element that is found in many RNA viruses. This motif was discovered over 25 years ago, but its functional significance is unknown. We created SARS-CoV-2 with deletions or mutations of the s2m and determined the effect of these changes on viral growth in tissue culture and in rodent models of infection. Deletion or mutation of the s2m element had no effect on growth in vitro or on growth and viral fitness in Syrian hamsters in vivo. We also observed no impact of the deletion on other known RNA structures in the same region of the genome. These experiments demonstrate that s2m is dispensable for SARS-CoV-2.

Evaluation of Efficacy of the Amino Acid-Peptide Complex Administered Intragastrically to Golden Hamsters Experimentally Infected with Sars-Cov-2

The development of coronavirus infection outbreak into a pandemic, coupled with the lack of effective COVID-19 therapies, is a challenge for the entire pharmaceutical industry. This study aimed to assess the treatment and preventive efficacy of the amino acid-peptide complex (APC) in male Syrian hamsters infected with SARSCoV-2 (intranasal administration of 26 mul of the virus culture, titer of 4 x 104 TCD50/ml). In a modeled COVID-19 case, APC administered for treatment and preventive purposes reduced lung damage. Compared to the positive control group, test group had the lung weight factor 15.2% smaller (trend), which indicates a less pronounced edema. Microscopic examination revealed no alveolar edema, atypical hypertrophied forms of type II alveolocytes, pulmonary parenchyma fibrinization. The macrophage reaction intensified, which is probably a result of the APC-induced activation of regenerative processes in the lung tissues. Spleens of the animals that received APC for therapeutic and preventive purposes were less engorged and had fewer hemorrhages. The decrease of body weight of the test animals that received APC for treatment and prevention was insignificant (p < 0.05), which indicates a less severe course of COVID-19. Administered following a purely therapeutic protocol, APC proved ineffective against SARS-CoV-2 post-infection. Thus, APC-based drug used as a therapeutic and preventive agent reduces pulmonary edema and makes morphological signs of lung tissue damage less pronounced in male Syrian hamsters infected with SARS-CoV-2.Copyright © Extreme Medicine.All right reserved.

Experimental and Clinical Evaluation of Mefloquine Effectiveness against the Infection Caused by Sars-Cov-2

The efficacy of mefloquine has not been studied in the in vivo experiments and clinical trials involving COVID-19 patients. The study was aimed to assess the effects of mefloquine on the SARS-CoV-2 accumulation in the lungs of infected animals and to study the efficacy and safety of mefloquine compared to hydroxychloroquine in patients with COVID-19. During the experiment, a total of 96 Syrian hamsters were infected with SARS-CoV-2. Accumulation of the virus in lungs was compared in the groups of animals treated with mefloquine and ribavirin and in the control group. During the clinical trial, the mefloquine and hydroxychloroquine safety and efficacy in patients with mild and moderate COVID-19 (172 individuals) was assessed based on the symptom changes over time and the computed tomography results. The experiment showed that the SARS-CoV-2 accumulation in the lungs of Syrian hamsters 6 days after infection and mefloquine treatment was 2.2 +/- 0.18 lg PFU/g, which was lower (p < 0.05) than in the control group (3.5 +/- 0.21 lg PFU/g) and ribavirin group (5.2 +/- 0.05 lg PFU/g). During the clinical trial, it was found that 50.0% of patients in the mefloquine group and 32.4% in the hydroxychloroquine group (p < 0.05) developed a mild disease, and the completely resolved respiratory failure was registered in 76.5% and 44.6%, respectively (p < 0.001). Adverse events were observed in 86.7 % and 77% of patients in the mefloquine and hydroxychloroquine groups, respectively (p > 0.05). Thus, during the experiment, mefloquine contributed to the faster virus titer reduction in the lungs. During the clinical trial, the mefloquine efficacy was non-inferiority or, based on a number of indicators, higher compared to hydroxychloroquine, with comparable safety.Copyright © Extreme Medicine.All right reserved.

Nirmatrelvir and Molnupiravir Coadministration Modifies Preclinical Viral Exposure

Background: Implementation of vaccination programmes has had a transformational impact on control of the SARS-CoV-2 pandemic, but the need for effective antiviral drugs remains. Molnupiravir (MPV) targets viral RNA polymerase inhibiting replication via lethal mutagenesis and nirmatrelvir (NTV) is a protease inhibitor boosted with ritonavir when given clinically. This study aimed to assess the virological efficacy of NTV and MPV individually and in combination against the SARS-CoV-2 BA.1 Omicron variant in a K18-hACE2 mouse model. Method(s): K18-hACE2 mice were inoculated intranasally with 103 PFU of SARSCoV-2 BA.1 Omicron (B.1.1.529). After 24 hours, mice were orally dosed q12H, as outlined in Figure 1. At 2, 3, and 4-days post infection mice were sacrificed, and lung samples harvested. Animals were weighed and monitored daily throughout. Subsequently, viral replication in the lung was quantified using qRT-PCR to measure total (N-gene) and sub-genomic (E-gene) viral RNA. Data were normalized to 18S for quantitation. Viral exposures expressed as Areas Under viral load Curves (AUCs) were calculated by the trapezoidal method using mean values at each timepoint. Separate studies in Syrian golden hamsters using individual drugs were also conducted, and total serum IgG was measured by ELISA at 4-days post infection. Result(s): Mice gained weight in all groups post-treatment, with no significant difference between groups. A reduction in lung viral exposure was evident in all treatment groups compared to the vehicle control dosed mice (Figure 1). Coadministration of NTV with MPV displayed a trend towards lower lung viral exposure compared to the vehicle control with ~40-and ~45-fold reduction in AUC for N-and SgE-gene assays, respectively. Dosed individually, NTV and MPV reduced viral exposure 5.7-and 7.7-fold for the N-gene assay, respectively. Differences in total serum IgG concentrations were evident between vehicle and NTV-(34-fold reduction, P=0.018), and MPV-(4.2-fold reduction, P=0.053) treated hamsters. Conclusion(s): These data show virological efficacy of NTV and MPV against the SARS-CoV-2 BA.1 Omicron variant. The combination of NTV and MPV demonstrated a lower viral RNA exposure in the lung than either drug alone, albeit not statistically significant. Initial data indicate potential immune alterations in NTV and MPV dosed hamsters. Studies to clarify the utility of NTV/ MPV combinations and further characterize the impact of antiviral therapy on IgG are warranted.

NAFAMOSTAT AS NASAL CHEMOPROPHYLAXIS FOR SARS-CoV-2 IN HAMSTERS

Background: Chemoprophylaxis is a critical tool for many infectious diseases, and in COVID-19 may have particular benefit for vulnerable patients that do not maximally benefit from vaccination. Nafamostat inhibits TMPRSS2, which catalyses a critical cell entry pathway for SARS-CoV-2. This study sought to assess efficacy of intranasal nafamostat against airborne transmission of SARSCoV-2 in Syrian Golden hamsters. Method(s): Male hamsters were intranasally administered water or 5 mg/kg nafamostat in water twice daily for 5 days (sentinels). One day after treatment initiation, sentinels were co-housed with an untreated hamster that was intranasally inoculated with 1 x 104 PFU of Wuhan SARS-CoV-2 (donor). Sentinels were separated from the donor by a perforated divider, allowing airflow between zones but not contact. Hamsters were weighed and throat-swabbed throughout. At day 4, all animals were culled, and lung and nasal turbinates were harvested. N-RNA was quantified relative to 18S-RNA by qPCR. A 2-way ANOVA with Bonferroni correction was applied to compare weight changes in the nafamostat group to those in controls. An unpaired t-test was used to compare viral RNA in lung and nasal turbinate between groups. Result(s): SARS-CoV-2 viral RNA was significantly lower in the nasal turbinates of nafamostat-treated hamsters compared to water-treated controls (P = 0.012;Figure 1). Within the lung, SARS-CoV-2 RNA was undetectable in the nafamostat-treated hamsters, but was detectable in the water-treated controls. Viral RNA was undetectable in the swabs of the nafamostat-treated hamsters at all timepoints, but was quantifiable in the water-treated control group from day 3. Body weight of the nafamostat-treated hamsters was significantly lower (P = < 0.001) than in the water-treated animals throughout. SARS-CoV-2 viral RNA was detectable in the donor hamsters lung, nasal turbinate and swab samples confirming validity of the experiment. Conclusion(s): This study demonstrated a protective effect of intranasal nafamostat against airborne SARS-CoV-2 transmission in Syrian golden hamsters. A phase IIa study of intravenously administered nafamostat yielded no evidence of clinical efficacy in hospitalised patients, but further investigation of intranasally administered nafamostat in a prophylactic setting may be warranted.

Characterization of the Immune Reaction to Covid-19 Treatment with Dexamethasone: A Narrative Review

Purpose of Study: The spread of SARS-CoV-2 and the resulting Coronavirus Disease 2019 (COVID-19) continues to manifest in individuals in varying severity with limited treatment options available. Despite research efforts put forth in developing therapeutic options for treatment of COVID-19 disease, effective and well understood mechanisms remain limited. Corticosteroid treatment with dexamethasone was shown to be beneficial for those with severe illness early in the pandemic with little understanding of its beneficial mechanism. This narrative review describes the current findings regarding the mechanism of action of dexamethasone treatment in the setting of SARS-CoV-2 infection. Methods Used: A comprehensive search of Embase and PubMed was conducted in consultation with a health sciences librarian. Search terms included (1) COVID-19 (2) dexamethasone (3) animal model and (4) immune response. No limits were used on the search and other reviews were excluded. Search results were screened based on titles and s before being selected for full text review. Outcomes recorded included characterization of the microenvironment of lung tissue following SARS-CoV-2 through cytokine measurement, histopathological staining and analysis of lung tissue, and clinical outcomes such as survival time. Summary of Results: The search resulted in 100 articles. Of these, 8 articles were identified that met the inclusion criteria. Three conducted experiments with Syrian hamsters, two with mice, two with alveolar macrophages, and one study was conducted with human subjects. Dexamethasone treatment was found to diminish inflammatory cytokine levels and preserve the integrity of lung tissue in several animal models and in vitro experiments in the setting of SARS-CoV-2 infection. Dexamethasone treatment was also found to reduce inflammatory cell infiltration of lung tissue infected with SARS-CoV-2. In humans, combination therapy of low dose dexamethasone with spironolactone proved more effective at lowering inflammatory markers than high dose dexamethasone alone. Conclusion(s): Collectively, the articles included in this review support the use of dexamethasone treatment in SARS-CoV-2 infection. Protective effects exhibited with dexamethasone treatment suggest that its action may be linked to the inflammatory nature of COVID-19 disease. Macrophage regulation and diminished inflammatory cytokine levels were hypothesized as possible mechanistic features of dexamethasone action but lacked exact characterization. Further exploration of combination treatment with dexamethasone and its mechanism of action is needed to identify specific and effective therapeutic strategies in the future.

Efficacy of Plant-Made Human Recombinant ACE2 against COVID-19 in a Golden Syrian Hamster Model.

Coronavirus disease 2019 (COVID-19) is a novel infectious respiratory disease caused by SARS-CoV-2. We evaluated the efficacy of a plant-based human recombinant angiotensin-converting enzyme 2 (hrACE2) and hrACE2-foldon (hrACE2-Fd) protein against COVID-19. In addition, we analyzed the antiviral activity of hrACE2 and hrACE2-Fd against SARS-CoV-2 using real-time reverse-transcription PCR and plaque assays. The therapeutic efficacy was detected using the Golden Syrian hamster model infected with SARS-CoV-2. Both hrACE2 and hrACE2-Fd inhibited SARS-CoV-2 by 50% at concentrations below the maximum plasma concentration, with EC50 of 5.8 µg/mL and 6.2 µg/mL, respectively. The hrACE2 and hrACE2-Fd injection groups showed a tendency for decreased viral titers in nasal turbinate tissues on day 3 after virus inoculation; however, this decrease was not detectable in lung tissues. Histopathological examination on day 9 after virus inoculation showed continued inflammation in the SARS-CoV-2 infection group, whereas decreased inflammation was observed in both the hrACE2 and hrACE2-Fd injection groups. No significant changes were observed at other time points. In conclusion, the potential therapeutic efficacy of plant-based proteins, hrACE2 and hrACE2-Fd, against COVID-19 was confirmed in a SARS-CoV-2-inoculated Golden Syrian hamster model. Further preclinical studies on primates and humans are necessary to obtain additional evidence and determine the effectiveness of these therapies.

SARS-CoV-2 Variant Pathogenesis Following Primary Infection and Reinfection in Syrian Hamsters.

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), the causative agent of COVID-19, has evolved into multiple variants. Animal models are important to understand variant pathogenesis, particularly for variants with mutations that have significant phenotypic or epidemiological effects. Here, cohorts of naive or previously infected Syrian hamsters (Mesocricetus auratus) were infected with variants to investigate viral pathogenesis and disease protection. Naive hamsters infected with SARS-CoV-2 variants had consistent clinical outcomes, tissue viral titers, and pathology, while hamsters that recovered from initial infection and were reinfected demonstrated less severe clinical disease and lung pathology than their naive counterparts. Males had more frequent clinical signs than females in most variant groups, but few sex variations in tissue viral titers and lung pathology were observed. These findings support the use of Syrian hamsters as a SARS-CoV-2 model and highlight the importance of considering sex differences when using this species. IMPORTANCE With the continued circulation and emergence of new severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) variants, understanding differences in the effects between the initial infection and a subsequent reinfection on disease pathogenesis is critical and highly relevant. This study characterizes Syrian hamsters as an animal model to study reinfection with SARS-CoV-2. Previous infection reduced the disease severity of reinfection with different SARS-CoV-2 variants.

EXPERIMENTAL MODEL OF SARS COV-2 IN YOUNG SYRIAN HAMSTERS FOR PRECLINICAL STUDIES

SARS-CoV-2 researchers faced the problem of specificity for ACE-2 receptors. In mice and rats, ACE-2 has a low affinity for this virus. In contrast, the Syrian hamster (Mesocricetus auratus) when infected with SARS-CoV-2 show clinical signs of infection with this virus. Therefore, it is necessary to assess the severity of pathological lesions and immune response for relevance to preclinical new drugs against SARS-CoV-2. In this study, we studied blood counts, serum levels of Il-6, TNF-α, IFN-α and IFN-γ and the histological state of the lungs of Syrian hamsters. In our study Syrian hamsters showed clinical signs of COVID-19. In infected animals, a decrease in the level of leukocytes and granulocytes was noted. There was some suppression of the immune response, without an increase in the main biomarkers of ARDS. The exception was high levels of TNF-α. Histological examination showed an early exudative phase of moderate ARDS. There were no significant gender differences in the production of pro-inflammatory cytokines. In this study, it was shown that infection with COVID-19 in young Syrian hamsters is modeled with moderate severity, and can be applied in preclinical studies. © 2022 International Multidisciplinary Scientific Geoconference. All rights reserved.

Preclinical studies of antiviral activity of the RPH-137 fusion protein and molnupiravir against COVID-19.

Finding effective and safe medicines to fight SARS-CoV-2 infection is an urgent task. RPH-137 is an original trap fusion protein against SARS-CoV-2 virus. It comprises the angiotensin-converting enzyme type 2 extracellular domain and the human IgG1 Fc fragment. The aim of the study was to carry out a preclinical evaluation of the efficacy of RPH-137 and molnupiravir against SARS-CoV-2 infection. Material(s) and Method(s): the authors analysed RPH-137 expressed in a stable CHO cell line and molnupiravir used as an active pharmaceutical ingredient. Drug-mediated inhibition of virus-induced cytotoxicity was assessed in Vero cell culture. In vivo efficacy assessments were performed in Syrian hamsters. The animals were infected intranasally with SARS-CoV-2 (PIK35 clinical isolate) in the dose of 5 log TCID50. The authors evaluated body weight measurements, lung-body weight ratios, and lung histopathology findings and determined viral RNA levels in oropharyngeal swabs by RT-PCR using the amplification cycle threshold (Ct). The statistical analyses involved one- and two-way ANOVA, Student's t-test, and Mann-Whitney test. Result(s): RPH-137 and molnupiravir inhibited the cytopathic effect of SARS-CoV-2 in Vero cells;the EC50 values of RPH-137 amounted to 4.69 mug/mL (21.3 nM) and 16.24 mug/mL (73.8 nM) for 50 TCID50 and 200 TCID50, respectively, whereas the EC50 values of molnupiravir were 0.63 mug/mL (1900 nM) for both doses. Intramuscular RPH-137 (30 and 80 mg/kg) had no effect on the infection process in Syrian hamsters. The comparison with the challenge control group showed that intraperitoneal RPH-137 (100 mg/kg) had statistically significant effects on a number of parameters, including a 27% reduction in inflammation and a 30% reduction in the total lesion area of the lungs by Day 7. Intragastric molnupiravir (300 mg/kg twice daily) significantly inhibited SARS-CoV-2 infection. Conclusion(s): both RPH-137 and molnupiravir inhibited the cytopathic effect of SARS-CoV-2 in Vero cells. In Syrian hamsters, molnupiravir demonstrated a more pronounced inhibition of SARS-CoV-2 than RPH-137. However, RPH-137 had statistically significant effects on a range of parameters. This offers additional perspectives for further research.Copyright © 2023 Safety and Risk of Pharmacotherapy. All rights reserved.

SARS-CoV-2 Disease Severity in the Golden Syrian Hamster Model of Infection Is Related to the Volume of Intranasal Inoculum.

The golden Syrian hamster (Mesocricetus auratus) is now commonly used in preclinical research for the study of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection and the assessment of vaccines, drugs and therapeutics. Here, we show that hamsters inoculated via the intranasal route with the same infectious virus dose of prototypical SARS-CoV-2 administered in a different volume present with different clinical signs, weight loss and viral shedding, with a reduced volume resulting in reduced severity of disease similar to that obtained by a 500-fold reduction in the challenge dose. The tissue burden of the virus and the severity of pulmonary pathology were also significantly affected by different challenge inoculum volumes. These findings suggest that a direct comparison between the severity of SARS-CoV-2 variants or studies assessing the efficacy of treatments determined by hamster studies cannot be made unless both the challenge dose and inoculation volume are matched when using the intranasal route. Additionally, analysis of sub-genomic and total genomic RNA PCR data demonstrated no link between sub-genomic and live viral titres and that sub-genomic analyses do not provide any information beyond that provided by more sensitive total genomic PCR.

Mucosal immunization with Ad5-based vaccines protects Syrian hamsters from challenge with omicron and delta variants of SARS-CoV-2.

SARS-CoV-2 variant clades continue to circumvent antibody responses elicited by vaccination or infection. Current parenteral vaccination strategies reduce illness and hospitalization, yet do not significantly protect against infection by the more recent variants. It is thought that mucosal vaccination strategies may better protect against infection by inducing immunity at the sites of infection, blocking viral transmission more effectively, and significantly inhibiting the evolution of new variants of concern (VOCs). In this study, we evaluated the immunogenicity and efficacy of a mucosally-delivered, non-replicating, adenovirus type 5-vectored vaccine that expresses the spike (S) gene of Wuhan (rAd5-S-Wuhan), delta (rAd5-S-delta), or omicron (rAd5-S-omicron) SARS-CoV-2 VOCs. Hamsters were immunized with these vaccines intranasally prior to challenge with omicron or delta variants. Additionally, one group was vaccinated by oral gavage with rAd5-S-Wuhan prior to challenge with the delta variant. Both intranasal and oral administration of rAd5-S-Wuhan generated cross-reactive serum IgG and mucosal IgA to all variant spike and RBD proteins tested. rAd5-S-omicron and rAd5-S-delta additionally elicited cross-reactive antibodies, though rAd5-S-omicron had significantly lower binding antibody levels except against its matched antigens. Two weeks after the final vaccination, hamsters were challenged with a SARS-CoV-2 variant; omicron or delta. Whether matched to the challenge or with rAd5-S-Wuhan, all vaccines protected hamsters from weight loss and lung pathology caused by challenge and significantly reduced viral shedding compared to placebo. Vaccination with rAd5-S-Wuhan provided significant protection, although there was an improved reduction in shedding and disease pathology in groups protected by the matched VOC vaccines. Nevertheless, Wuhan-based vaccination elicited the most cross-reactive antibody responses generally. Overall, heterologous vaccination via mucosal routes may be advantageous for second-generation vaccines.

Co-administration of Favipiravir and the Remdesivir Metabolite GS-441524 Effectively Reduces SARS-CoV-2 Replication in the Lungs of the Syrian Hamster Model.

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has spread worldwide since December 2019, causing coronavirus disease 2019 (COVID-19). Although vaccines for this virus have been developed rapidly, repurposing drugs approved to treat other diseases remains an invaluable treatment strategy. Here, we evaluated the inhibitory effects of drugs on SARS-CoV-2 replication in a hamster infection model and in in vitro assays. Favipiravir significantly suppressed virus replication in hamster lungs. Remdesivir inhibited virus replication in vitro, but was not effective in the hamster model. However, GS-441524, a metabolite of remdesivir, effectively suppressed virus replication in hamsters. Co-administration of favipiravir and GS-441524 more efficiently reduced virus load in hamster lungs than did single administration of either drug for both the prophylactic and therapeutic regimens; prophylactic co-administration also efficiently inhibited lung inflammation in the infected animals. Furthermore, pretreatment of hamsters with favipiravir and GS-441524 effectively protected them from virus transmission via respiratory droplets upon exposure to infected hamsters. Repurposing and co-administration of antiviral drugs may help combat COVID-19. IMPORTANCE During a pandemic, repurposing drugs that are approved for other diseases is a quick and realistic treatment option. In this study, we found that co-administration of favipiravir and the remdesivir metabolite GS-441524 more effectively blocked SARS-CoV-2 replication in the lungs of Syrian hamsters than either favipiravir or GS-441524 alone as part of a prophylactic or therapeutic regimen. Prophylactic co-administration also reduced the severity of lung inflammation. Moreover, co-administration of these drugs to naive hamsters efficiently protected them from airborne transmission of the virus from infected animals. Since both drugs are nucleotide analogs that interfere with the RNA-dependent RNA polymerases of many RNA viruses, these findings may also help encourage co-administration of antivirals to combat future pandemics.

Oral Administration of Universal Bacterium-Vectored Nucleocapsid-Expressing COVID-19 Vaccine is Efficacious in Hamsters.

Oral delivery of an inexpensive COVID-19 (coronavirus disease 2019) vaccine could dramatically improve immunization rates, especially in low- and middle-income countries. Previously, we described a potential universal COVID-19 vaccine, rLVS ΔcapB/MN, comprising a replicating bacterial vector, LVS (live vaccine strain) ΔcapB, expressing the highly conserved SARS-CoV-2 (severe acute respiratory syndrome coronavirus 2) membrane and nucleocapsid (N) proteins, which, when administered intradermally or intranasally, protects hamsters from severe COVID-19-like disease after high-dose SARS-CoV-2 respiratory challenge. Here, we show that oral administration of the vaccine also protects against high-dose SARS-CoV-2 respiratory challenge; its protection is comparable to that of intradermal, intranasal, or subcutaneous administration. Hamsters were protected against severe weight loss and lung pathology and had reduced oropharyngeal and lung virus titers. Protection against weight loss and histopathology by the vaccine, which in mice induces splenic and lung cell interferon gamma in response to N protein stimulation, was correlated in hamsters with pre-challenge serum anti-N TH1-biased IgG (IgG2/3). Thus, rLVS ΔcapB/MN has potential as an oral universal COVID-19 vaccine. IMPORTANCE The COVID-19 pandemic continues to rage into its fourth year worldwide. To protect the world's population most effectively from severe disease, hospitalization, and death, a vaccine is needed that is resistant to rapidly emerging viral variants of the causative agent SARS-CoV-2, inexpensive to manufacture, store, and transport, and easy to administer. Ideally, such a vaccine would be capable of oral administration, especially in resource-poor countries of the world where there are shortages of needles, syringes and trained personnel to administer injectable vaccines. Here, we show that oral administration of a bacterium-vectored vaccine meeting all these criteria protects naturally susceptible Syrian hamsters from severe COVID-19-like disease, including severe weight loss and lung pathology, after high-dose SARS-CoV-2 respiratory challenge. As the vaccine is based upon inducing immunity to highly conserved SARS-CoV-2 membrane and nucleocapsid proteins, as opposed to the rapidly mutating Spike protein, it should remain resistant to newly emerging SARS-CoV-2 variants.

Preclinical studies of antiviral activity of the RPH-137 fusion protein and molnupiravir against COVID-19

Finding effective and safe medicines to fight SARS-CoV-2 infection is an urgent task. RPH-137 is an original trap fusion protein against SARS-CoV-2 virus. It comprises the angiotensin-converting enzyme type 2 extracellular domain and the human IgG1 Fc fragment. The aim of the study was to carry out a preclinical evaluation of the efficacy of RPH-137 and molnupiravir against SARS-CoV-2 infection. Materials and methods: the authors analysed RPH-137 expressed in a stable CHO cell line and molnupiravir used as an active pharmaceutical ingredient. Drug-mediated inhibition of virus-induced cytotoxicity was assessed in Vero cell culture. In vivo efficacy assessments were performed in Syrian hamsters. The animals were infected intranasally with SARS-CoV-2 (PIK35 clinical isolate) in the dose of 5 log TCID50. The authors evaluated body weight measurements, lung-body weight ratios, and lung histopathology findings and determined viral RNA levels in oropharyngeal swabs by RT-PCR using the amplification cycle threshold (Ct). The statistical analyses involved one- and two-way ANOVA, Student's t-test, and Mann–Whitney test. Results: RPH-137 and molnupiravir inhibited the cytopathic effect of SARS-CoV-2 in Vero cells;the EC50 values of RPH-137 amounted to 4.69 μg/mL (21.3 nM) and 16.24 μg/mL (73.8 nM) for 50 TCID50 and 200 TCID50, respectively, whereas the EC50 values of molnupiravir were 0.63 μg/mL (1900 nM) for both doses. Intramuscular RPH-137 (30 and 80 mg/kg) had no effect on the infection process in Syrian hamsters. The comparison with the challenge control group showed that intraperitoneal RPH-137 (100 mg/kg) had statistically significant effects on a number of parameters, including a 27% reduction in inflammation and a 30% reduction in the total lesion area of the lungs by Day 7. Intragastric molnupiravir (300 mg/kg twice daily) significantly inhibited SARS-CoV-2 infection. Conclusions: both RPH-137 and molnupiravir inhibited the cytopathic effect of SARS-CoV-2 in Vero cells. In Syrian hamsters, molnupiravir demonstrated a more pronounced inhibition of SARS-CoV-2 than RPH-137. However, RPH-137 had statistically significant effects on a range of parameters. This offers additional perspectives for further research.

Antiviral Activity of The Preparation Meflokhin® Against COVID-19

The COVID-19 virus has caused a global emergency and has attracted the attention of healthcare professionals and the public around the world. The significant increase in the number of new cases of infection with this virus demonstrates the relevance of the search for drugs that are effective against this pathogen. The aim of this work was to evaluate the antiviral efficacy of Mefloquin® against COVID-19. The antiviral efficacy of Mefloquin® against the new pandemic virus SARS-CoV-2 was studied in in vitro experiments in Vero C1008 cell culture and in vivo on Syrian golden hamsters. The results of the study revealed that the drug Mefloquine® at a concentration of 2.0 µg ml-1, when applied after infection of cells, suppresses the reproduction of the SARS-CoV-2 virus by 1.7-1.9 lg, the inhibition rate is about 99%. When using Mefloquine, pathological changes in the lung tissue were less pronounced than in the control group. 6 days after infection, it was shown that when using Mefloquine, there was a statistically significant decrease in viral load in the lungs of infected Syrian golden hamsters, with an inhibition rate of 95.5%.