1 Introduction
Severe acute respiratory syndrome
coronavirus 2 (SARS-CoV-2) appeared in Wuhan, China, in late of 2019,
causing coronavirus disease 2019 (COVID-19) and infecting approximately
560 million individuals worldwide based on July 2022 data, and death of
approximately 6.5 million individuals.1 The
seriousness of the global situation called for urgent development of
COVID-19 vaccine and several vaccines were approved at unprecedented
speed and used clinically throughout the world.2-4
The approved vaccines are delivered by intramuscular (IM) injection and
induce SARS-CoV-2 specific IgG in serum and showed effectiveness in
prevention of severe infection and death.5 However,
vaccines administrated intramuscularly are, in general, known to be
minimally effective in inducing mucosal secretory IgA (S-IgA), which
plays an important role in boosting mucosal immunity, and preventing
respiratory and gastrointestinal tract infections and viral
transmission.6 Recent reports have described
intramuscular administration of COVID-19 mRNA vaccine (e.g.,
Comirnaty)-induced SARS-CoV-2 specific IgA at mucosal
sites.7, 8 However, the effect of mucosal IgA induced
by mRNA vaccine is presumably limited in providing protection against
virus infection through mucosal sites.9 Thus, although
the first-generation COVID-19 vaccines produced limited benefits, such
as reduction of severe infection and death in pandemic emergency
situation, it is necessary to develop the next generation COVID-19
vaccines that can induce effective mucosal S-IgA, in addition to
systemic IgG and IgA, to achieve protection against infection and
prevention of transmission of the virus.
Mucosal vaccines that are directly administered onto mucosal surfaces,
such as the nasal cavity and gastrointestinal tract, can induce mucosal
S-IgA.10 Since the mucosa is equipped with exclusion
systems, namely the cilia, mucus and various proteases, against foreign
substances including vaccine antigens,11 the use of
vaccines that contain antigen alone is unlikely to produce protective
immunity. Thus, for successful mucosal vaccination, live virus vaccines
and mucosal adjuvants that enhance the vaccine effects, have been
developed worldwide.12 However, the live attenuated
vaccines, such as Flumist®, have problems related to
their efficacy and safety and thus their worldwide use is
limited.13
To overcome these problems, we reported previously the use of an antigen
delivery type adjuvant,14 pulmonary surfactant
(PS)-based compound, surfacten®, as an effective and
safe intranasal mucosal adjuvant for influenza ether-split hemagglutinin
vaccine (HAv) and that it enhanced the production of both HAv-specific
IgG in serum and S-IgA in the respiratory mucosae of mice and
swine.15, 16 Based on these results, we then
successfully developed synthetic surfactant (SSF), which mimicked human
PS, consisting of three major lipids
[1,2-dipalmitoyl-phosphatidylcholine (DPPC), phosphatidylglycerol (PG)
and palmitic acid (PA)] plus the K6L16 peptide [which resembles
surfactant protein C (SP-C)], as a potent and safe synthetic mucosal
adjuvant.17, 18 In addition, to increase the
effectiveness of the vaccine, we prepared the antigen and SSF complex by
lyophilization, thus allowing the lyophilized powder to be suspended in
carboxy vinyl polymer (CVP) to increase the viscosity of the antigen and
SSF complex solution in order to prolong antigen uptake on the mucosal
surface.18 The final vaccine solution was termed
antigen-SF-10. In a series of studies, we reported that intranasal
application of antigen-SF-10 enhanced the absorption of antigens to
nasal antigen presenting cells and induced systemic as well as
respiratory humoral immunity in mice and cynomolgus
monkeys18-20 and cell-mediated immunity in
mice.21
The present study is an extension to the above work and was designed to
determine the outcome of mucosal application of recombinant S1 spike
protein of SARS-CoV-2, as an antigen combined with SF-10 (S1-SF-10) on
respiratory and systemic immunity against SARS-CoV-2 infection. For this
purpose, S1-SF-10 vaccine was inoculated intratracheally (IT) through
nasal cavity to the lower respiratory tract (S1-SF-10-IT) of mice, the
site of viral infection and proliferation. To test the effectiveness of
a new administration method of intratracheal vaccination, we analyzed
the levels of induced S1-specific antibodies, the antibody secreting
cells and S1-responsive Th cytokine secreting cells in lung lymphocytes
and splenocytes. We also compared the level of protective immunity
induced by S1-SF-10-IT and intramuscular injection of S1 with an already
reported potent adjuvant AS03 (S1-AS03-IM) against SARS-CoV-2 infection
using the S1/angiotensin converting enzyme 2 (ACE2) binding inhibition
assay.