Study: A SCID mouse model to evaluate the efficacy of antivirals against SARS-CoV-2 infection. Image Credit: Kateryna Kon / Shutterstock

Evaluation of the efficacy of antiviral agents against SARS-CoV-2 in a mouse model

In a recent study posted to the bioRxiv* pre-print server, researchers from Belgium evaluated the efficacy of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) antiviral drugs in a severe combined immunodeficient (SCID) mouse model.

Study: A SCID mouse model to evaluate the efficacy of antivirals against SARS-CoV-2 infection† Image Credit: Kateryna Kon/Shutterstock

Background

Due to the inability of ancestral SARS-CoV-2 strains to bind to the mouse angiotensin-converting enzyme 2 (ACE2) receptor, mouse modified (MA) viruses or genetically modified (GM) mice were used to produce SARS-CoV 2 to perform. -related preclinical studies.

However, some of the evolved SARS-CoV-2 variants of concern (VOCs), such as the beta (B.1.351) VOC, bind to the mouse ACE2 receptor and replicate in some wild-type (wt) mouse species, including C57BL/6 and BALB/c mice. The B.1.351 VOC carries spike (S) protein mutations, primarily the N501Y, allowing for efficient mouse receptor binding in wt mice. Similarly, a study showed that the K417N mutation was common in an MA SARS-CoV-2 variant.

Together, these findings indicated that both the K417N and E484K mutations in the receptor binding domain (RBD) of the beta variant facilitate efficient binding to murine ACE2 receptors than SARS-CoV-2 alpha variants.

About the study

In the present study, researchers used the SCID mouse infection model to determine the in vivo efficacy of antiviral agents against SARS-CoV-2, molnupiravir and nirmatrelvir. By using a small animal model, researchers were able to test small amounts of antiviral drugs in appropriate housing conditions and avoid using the MA SARS-CoV-2 strains and GM mice.

The team intranasally infected nine mice of each species with 105 median tissue culture infectious doses (TCID50) of the Beta B.1.351 variant. They euthanized all mice on day 3 post-infection (pi) to harvest their lungs and quantify the infectious viral titers.

Furthermore, the researchers examined the replication kinetics of Beta (B.1.351) VOC in SCID mice. To this end, they infected seven to nine week old male SCID mice with 105 TCID50 of the B.1.351 VOC. Between three and seven days pi, the team euthanized 10 lab animals and harvested their lungs to quantify the infectious viral titers.

Finally, to assess the potential antiviral efficacy of molnupiravir and nirmatrelvir against the beta variant (B.1.351), SCID male mice were treated twice daily with oral doses of a drug or vehicle. In addition, they treated test animals for three days from the day of infection with 200 mg/kg molnupiravir and 300 mg/kg nirmatrelvir. The team euthanized antiviral treated mice on day 3 p.i. before collecting lungs.

Replication of beta (B.1.351) SARS-CoV-2 in different mice.  Infectious viral titers in the lungs of male SCID, male BALB/c and male C57BL/6 mice infected with 105 TCID50 of beta-SARS-CoV-2 variants (pi) 3 days post-infection are expressed as log10 TCID50 per mg of lung tissue.  Individual data and median values ​​are displayed.  Data were analyzed using the Mann™ Whitney U test.  *P < 0.05, **P<0,01).  Gegevens zijn afkomstig van twee onafhankelijke experimenten met:

Replication of beta (B.1.351) SARS-CoV-2 in different mice. Infectious viral titers in the lungs of male SCID, male BALB/c and male C57BL/6 mice infected with 105 TCID50 of beta SARS-CoV-2 variants 3 days post-infection (pi) are expressed as log10 TCID50 per mg of lung tissue. Individual data and median values ​​are displayed. Data were analyzed using the Mann™ Whitney U test. *P<0.05, **P<0.01). Data are from two independent experiments with n=9 per group.

Study findings

Beta B.1.351 infected male SCID mice had high infectious viral titers and aberrant lung pathology on day 3pi. The authors noted significantly higher infectious viral titers in the lungs of infected SCID mice than in infected BALB/c and C57BL/6 mice.

From day 4 pi, the infectious virus titers in the lungs started to decrease. After day 3 pi, the test animals also started to gain weight normally. However, when monitored up to day 14 pi, the test animals showed no sign of weight loss or morbidity. Histological examination, however, showed mild peribronchial inflammation. In addition, there was significant perivascular inflammation and intra-alveolar hemorrhage.

Replication kinetics of beta (B.1.351) SARS-CoV-2 in male SCID mice.  (A) Infectious viral loads in the lungs of male SCID mice infected with 105 TCID50 of beta-SARS-CoV-2 variants at different days post-infection (pi) are expressed as log10 TCID50 per mg of lung tissue.  Individual data and median values ​​are displayed.  Data were analyzed using the Mann™ Whitney U test.  ***P = 0.0003, ****P < 0,0001 (B) Gewichtsverandering op verschillende dagen pi in percentage, genormaliseerd naar het lichaamsgewicht op het moment van infectie.  Staven vertegenwoordigen gemiddelden ± SD.  Alle gegevens zijn afkomstig van 2 onafhankelijke experimenten met 10 dieren per groep.  (C) Representatief H & E-beeld van long van SCID-muis geïnfecteerd met de bèta-variant op dag 3 pi met beperkte peri-bronchiale ontsteking (blauwe pijlen), significante peri-vasculaire ontsteking (rode pijlen) en intra-alveolaire bloeding (groene pijl).  Schaal

Replication kinetics of beta (B.1.351) SARS-CoV-2 in male SCID mice. (A) Infectious viral loads in the lungs of male SCID mice infected with 105 TCID50 of beta SARS-CoV-2 variants at different days post-infection (pi) are expressed as log10 TCID50 per mg of lung tissue. Individual data and median values ​​are displayed. Data were analyzed using the Mann™ Whitney U test. ***P = 0.0003, ****P < 0.0001 (B) Weight change on different days pi in percentage, normalized to body weight at the time of infection. Bars represent means ± SD. All data are from 2 independent experiments with 10 animals per group. (C) Representative H&E image of lung from SCID mouse infected with the beta variant at day 3 pi showing limited peri-bronchial inflammation (blue arrows), significant peri-vascular inflammation (red arrows) and intra-alveolar hemorrhage (green arrow). Scale bar = 100 µM

Infected mice treated with SARS-CoV-2 antiviral agents, molnupiravir or nirmatrelvir, showed a significant decrease in infectious virus titres in their lungs. The molnupiravir and nirmatrelvir treated groups showed a decrease of 1.9 and 3.8 log10 TCID50/mg tissue in lung virus titers, respectively. In addition, it significantly improved the lung pathology of the experimental animals. Notably, four and eight of 14 animals in the molnupiravir and nirmatrelvir treated groups, respectively, had no infectious viral titers. As expected, these animals did not lose weight due to the drugs.

conclusions

The findings of the present study demonstrate the benefits of using SCID mice/B.1.351 variant infection models to assess the potential activity of SARS-CoV-2 antivirals in preclinical studies.

The infection model of SCID mice was not suitable for the evaluation of vaccines and therapeutic antibodies against SARS-CoV-2. However, they were found to be beneficial for in vivo efficacy studies of SARS-CoV-2 replication-inhibiting drug molecules.

Both molnupiravir and nirmatrelvir have previously been shown to inhibit the replication of the B.1.351 variant in Syrian hamsters. Likewise, they showed high efficacy in the SCID mouse model. Notably, both drugs reduced viral loads in the lungs of infected mice with similar potency as previously seen in the Syrian hamster model.

These results are encouraging, especially as molnupiravir, marketed as Lagevrio, is being used as an oral drug in several countries to treat coronavirus disease 2019 (COVID-19). Nirmatrelvir and ritonavir tablets, marketed as Paxlovid, are also approved for oral use by the U.S. Food and Drug Administration (FDA) against SARS-CoV-2 and other coronaviruses

*Important announcement

bioRxiv publishes preliminary scientific reports that have not been peer-reviewed and therefore should not be considered conclusive, that should guide clinical practice/health-related behavior or be treated as established information.

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