Infectious Disease
Establishment of a nonhuman primate model for development of vaccines and anti-viral drugs against COVID-19
2021 年 3 巻 3 号 p. 109-111
詳細
2021 年 3 巻 3 号 p. 109-111
COVID-19 is caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) and has continued to spread worldwide since late 2019 [1]. It was speculated that SARS-CoV-2 would not infect regular laboratory mice since SARS-CoV-2 is similar to SARS-CoV, which caused severe acute respiratory syndrome in 2003 and did not propagate in laboratory mice [2]. Therefore, hamsters, human ACE2 transgenic mice, rhesus monkeys, and cynomolgus monkeys have been used since the early stages of the COVID-19 pandemic to analyze its pathogenicity and evaluate the efficacy of vaccinations and other therapeutics [3,4,5,6]. We established a nonhuman primate model in our animal biosafety level 3 facility to ascertain the pathogenicity of SARS-CoV-2 and to further develop vaccines and drugs.
Rhesus and cynomolgus monkeys are laboratory animals that are susceptible to SARS-CoV-2 infection and have immunity and metabolism similar to those of humans. Therefore, macaque monkeys are better suited as experimental animals than hamsters and human ACE2 transgenic mice when evaluating the efficacy of vaccines and drugs.
We first examined the pathogenicity of SARS-CoV-2 in cynomolgus monkeys as per our previous studies on influenza virus infection [7,8,9]. We inoculated the SARS-CoV-2 WK-521 strain, which was isolated by the National Institute of Infectious Diseases in Japan, into the conjunctivas, nasal cavities, oral cavities, and tracheas of 3 cynomolgus monkeys [10]. The body temperature of each infected macaque rose markedly for 3 to 7 days after virus inoculation. The infectious virus was detected in swab samples of the conjunctiva, nasal cavity, oral cavity, and trachea of all the macaques on the day after virus inoculation and in nasal and oral swab samples on the seventh day. Infectious viruses were not detected in swab samples after day 10. Using quantitative reverse transcription PCR (RT-PCR), a large amount of viral RNA was detected in swab samples on day 1. The amount gradually decreased; however viral RNA was detected even on day 28.
Viral pneumonia in cynomolgus macaques infected with SARS-CoV-2 was observed on chest X-ray radiographs. A ground-glass appearance in the lungs, especially in the peripheral pulmonary areas, was confirmed between days 3 and 7, but not after day 10. Histological pneumonia was observed in all macaques infected with WK-521. Congestion and neutrophilic infiltration were observed in the lungs of infected macaques 3 days after infection (manuscript in preparation) (Fig. 1). Thickened alveolar walls with severe lymphoid infiltration were observed on day 7. Thickened alveolar walls with infiltration of macrophages were observed in the lungs on day 28, indicating regenerative activity. Thus, cynomolgus monkeys exhibited clinical signs of disease similar to those of humans and viral propagation occurred in respiratory organs.
Histological pneumonia in cynomolgus macaques infected with severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). Hematoxylin and eosin sections of lung tissues of macaques autopsied on day 3 (A), day 7 (B), and day 28 (C), respectively, after being infected with SARS-CoV-2. Bars=100 μm.
Immune responses against SARS-CoV-2 were examined in cynomolgus macaques. One of the macaques exhibited acute increases in plasma cytokine levels after infection, particularly the inflammatory cytokines interleukin-6 (IL-6), monocyte chemotactic protein-1 (MCP-1), and IL-10 on day 1. A neutralizing antibody against SARS-CoV-2 was detected in this monkey on day 10 after virus inoculation; however, neutralizing antibodies were not detected in the other two macaques until day 28. T lymphocyte responses specific to the SARS-CoV-2 antigen, including interferon (IFN)-γ and IL-2 production, were detected in all macaques on day 14. In the two monkeys in which the neutralizing antibody was not detected, the increase in the number of IFN-γ-producing cells preceded that of IL-2-producing cells. Thus, it was revealed that not only neutralizing antibodies but also T-lymphocyte responses specific to SARS-CoV-2 contributed to the elimination of the virus and recovery from infection.
We have evaluated vaccines and drugs based on these findings. Using cynomolgus macaques, we have already examined the efficacy of two vaccines, an attenuated vaccinia virus carrying the SARS-CoV-2 spike (S) gene [11], and an mRNA vaccine containing the SARS-CoV-2 receptor-binding domain (RBD) gene of the S gene. These vaccines effectively induced neutralizing antibodies and prevented an increase in body temperature, propagation of SARS-CoV-2, and viral pneumonia in macaques (manuscript in preparation) [12] (preprint).
We are planning to expand the ABSL3 room at the Research Center for Animal Life Science, Shiga University of Medical Science, using support from the Japan Agency for Medical Research and Development (AMED). This project is expected to accelerate the development of prophylaxis and therapeutics against COVID-19 in cooperation with other laboratories and companies.
These studies were conducted in strict accordance with the Guidelines for the Husbandry and Management of Laboratory Animals of the Research Center for Animal Life Science at Shiga University of Medical Science and the Standards Relating to the Care and Fundamental Guidelines for Proper Conduct of Animal Experiments and Related Activities in Academic Research Institutions under the jurisdiction of the Ministry of Education, Culture, Sports, Science and Technology, Japan.
The authors have nothing to disclose.
This work was supported by grants from AMED under grant numbers 19fk0108172, 20nk0101615, 20fk0108410, and 20fk0108538. I thank Drs. Hirohito Ishigaki, Misako Nakayama, Yoshinori Kitagawa, and Cong Thanh Nguyen for conducting studies on COVID-19, my laboratory staff for supporting the experiments, and Drs. Fumihiko Yasui, Michinori Kohara, Ken J. Ishii, and Yoshihiro Kawaoka for their collaboration.
