2. Requirements of a SARS-CoV-2 animal model
A perfect animal model should be permissive to infection and be able to mimic the clinical pathology of disease as observed in humans (Sutton & Subbarao, 2015). These models play a crucial role in studying host-virus interactions that contribute to disease pathogenesis and transmission. A protocol for the development of animal models is mentioned by the ‘FDA animal efficacy rule’(Product Development Under the Animal Rule , 2015). Accordingly, these models should have the same receptors as those present in humans that help viruses in the attachment and entry process, and the outcome of the infection should be as severe as in humans. In many emerging disease studies, in vitro studies cannot completely simulate human pathophysiology. Also, the immunological components are very complex in humans, which cannot be proven in the in-vitro experiments. However, despite the differences in animal model translation to humans, a lot of critical information related to the pathogenesis, prevention and treatment of newly emerging infectious diseases can still be discovered.
Animal models can be categorized as small or large. Small animal models include the use of mice, rats, ferrets, hamsters and rabbits, which are relatively smaller in size and require limited space. Small animal models offer several advantages because the animals and reagents specific to the animals are easily available, the animals can be handled with less effort and cost, and allows the use of a large number of animals to provide data for strong statistical analyses (Sutton & Subbarao, 2015). The small animal model is limited by the significant intrinsic biological differences between humans and rodents or small mammals, which have led to the requirement that viruses must be adapted to the animals, or the animals, must be genetically manipulated to recapitulate the human system (Subbarao et al., 2004). In addition, the animals’ short life span hampers the ability to monitor the long-term prognosis of the disease (Gretebeck & Subbarao, 2015). Large animal models such as non-human primates (NHP) are more reliable models to replicate human disease pathogenesis as they are physiologically, immunologically and genetically more closely related to humans (Fujiyama et al., 2002; Lu et al., 2008). The major limitation of using NHP models are the high cost and resources involved in the study, which limits the number of animals that may be included in a study and thus adversely affecting the statistical power of the outcome. In addition, most NHPs are outbred animals and have a wide variability in genetic backgrounds, which sometimes make it difficult to interpret the outcome of a study due to variability in results among individual animals (Lu et al., 2008).
For the ongoing COVID-19 pandemic, direct human clinical trials are complicated by the mild to severe forms of the disease due to genetic diversities, age of the host, comorbidities, multiple infections along with other preexisting diseases (P. K. Chan & Chan, 2013; Gold et al., 2020; ”What explains Covid-19’s lethality for the elderly? Scientists look to ‘twilight’ of the immune system,” 2020). The virus uses the ACE-2 receptor that is expressed in cells found in the heart, lungs, gastrointestinal tract, and renal tract (Hamming et al., 2004; Hoffmann et al., 2020). After entry, viral replication kinetics in most of the target cells remains unknown. Air droplets and aerosols are critical aspects of the global spread of SARS-CoV-2 (Hamid, Mir, & Rohela, 2020; Meselson, 2020). Regardless, there is an urgent need to understand the risks factors of transmission for SARS-CoV-2 (Figure 2 ). The symptoms of COVID-19 with other specific and non-specific upper respiratory tract symptoms are similar in nature, which make it difficult to distinguish from other diseases (Bhatraju et al., 2020). A case report from Eiju General Hospital Tokyo, Japan, showed the existence of the Influenza virus along with SARS-CoV-2 showing similar types of symptoms that posed a difficulty in the differential diagnosis of the two diseases (Azekawa, Namkoong, Mitamura, Kawaoka, & Saito, 2020). Another case in Rhode Island showed the co-infection of SARS-CoV-2 with Human metapneumovirus, where a patient tested positive for human metapneumovirus but failed to test for SARS-CoV-2. The symptoms did not subside even after treatment for metapneumovirus. Nevertheless, when the patient was tested for SARS-CoV-2 by PCR and the results came back as positive (Touzard-Romo, Tape, & Lonks, 2020). A travel-related case of an 80-year old male from Japan, with a history of diabetes, showed coinfection of SARS-CoV-2 and Legionella with respiratory distress and gastrointestinal symptoms. The patient passed away after 13 days but was later confirmed to be infected with COVID-19 (Arashiro et al., 2020). These individual case reports compel the need for suitable animal studies of SARS-CoV-2 co-infection with other pathogens with similar disease manifestations for better clinical outcomes.
COVID-19 has a variety of clinical outcomes (J. F.-W. Chan et al., 2020; Huang et al., 2020). Most of the patients who are admitted to hospitals with severe clinical manifestations have other comorbid conditions such as diabetes, cardiovascular disease, gastrointestinal disease, or hypertension (Gold et al., 2020). In the case of Influenza, one publication showed that the risk of the Acute Respiratory Distress increases 3.4-fold in the H7N9 infected person with similar comorbidity (H. N. Gao et al., 2013). Age-related co-morbidity has mostly affected the transmission cycle of disease (Sun et al., 2020). The exact mechanism of how these comorbidities deteriorate SARS-CoV-2 patient conditions remains unknown. These comorbid conditions that lead to severe to fatal outcomes are another aspect of COVID-19 pathogenesis that needs a suitable animal model for study.
There is a concerted effort in the research community to develop therapeutics and vaccines against COVID-19. For example, Remdesivir is an antiviral drug that failed the clinical trial for Ebola a decade ago (Mulangu et al., 2019) and now has been proposed for COVID-19 treatment (Beigel et al., 2020) (ClinicalTrials.gov: NCT04257656, NCT04252664, NCT04280705). It was repurposed for COVID-19 treatment because it directly blocked RNA synthesis (Agostini et al., 2018). Preliminary reports for Remdesivir in a clinical trial of 1063 adult patients with lower respiratory tract infection showed a shortening in recovery time in 538 patients given Remdesivir intravenously (median 11 days), compared to the 521 patients given a placebo (median 15 days) (Beigel et al., 2020). However, there were still serious adverse effects reported in COVID-19 patients given Remdesivir, but Remdesivir worked better in comparison to the placebo. To test the efficacy of Remdesivir in disease outcome, a rhesus macaque model of SARS-CoV-2 infection was developed, where infected macaques developed mild to moderate clinical symptoms(Brandi N. Williamson, 2020). The macaques treated with Remdesivir did not show any respiratory distress, and viral titers in bronchoalveolar lavage was reduced after 12 hours of treatment, suggesting that Remdesivir may have some beneficial effect in the treatment of COVID-19 (Brandi N. Williamson, 2020). A study using Lopinavir, a protease inhibitor and Ritonavir, that primes the action of Lopinavir widely used to treat HIV infection, are in clinical trials for COVID-19 (Bhatnagar et al., 2020). The combination of Lopinavir and Ritonavir had shown efficacy against SARS-CoV in mice (Chu et al., 2004) and MERS-CoV in NHPs (J. F. Chan et al., 2015). ACE-2 inhibitors as well as some fusion inhibitors such as Arbidol are also in clinical trials (Chinese Government Clinical Trials: ChiCTR2000029573)(Wang et al., 2020). Clinical trials using convalescent serum as treatment for COVID-19 has already begun at the Icahn School of Medicine at Mount Sinai, New York, USA (Sean T. H. Liu, 2020) . Similarly, polyclonal antibodies have also been employed in clinical trials because polyclonal human immunoglobulin G (IgG) had shown effectiveness against MERS-CoVs.(Luke et al., 2016). Vaccine development is important to the control of disease and protection against the transmission of the virus to immunized individuals. Many groups are working to develop potential vaccines for SARS-CoV-2, out of which some candidates are already in clinical trials (Mukherjee, 2020). Moderna, a vaccine manufacturer, has started phase II clinical trials of their vaccine candidate mRNA- 1273, which passed phase I trial recently by generating desired immune response by natural infection (ClinicalTrials.gov Identifier: NCT04283461) (”Moderna Moderna announces funding award from CEPI to accelerate development of messenger RNA (mRNA) vaccine against novel coronavirus; 2020. [accessed 2020 February15]
,”). A purified inactivated SARS-CoV-2 virus candidate (PiCoVacc) tested in mice, rats and non-human primates have also shown protective immune response suggesting neutralizing activity against SARS-CoV-2 (Q. Gao et al., 2020). Recombinant protein-based vaccines by University of Queensland and viral-vector based vaccines by University of Oxford, England are also being actively tested for vaccine efficacy (”Developing Therapeutics and Vaccines for Coronaviruses,”). Another study has suggested that polio vaccine could be used to prevent SARS-CoV-2 (Yeh et al., 2020).
In order to study various unanswered questions about the disease pathogenesis, suitable animal models are essential. Several studies are in progress to find suitable animal models to study the transmission, disease pathogenesis, and pre-clinical trials of potential therapeutics for the management of COVID-19. However, to successfully end the COVID-19 pandemic, efforts to develop vaccines to prevent the virus spread should synchronize with studies to uncover disease pathogenic mechanisms in comorbid and high-risk co-infection conditions. Although difficult, identifying a model that adequately mimics the human disease of SARS-CoV-2 is bolstered by several studies of SARS-CoV and MERS-CoV, which has provided some preliminary insights on the models. All laboratory animal models, like mice to hamsters then ferret to NHP’s, are equally valuable to dissect out various questions to better understand disease mechanisms.