From Immunogen to COVID-19 vaccines: Prospects for the post-pandemic era

Abstract

The COVID-19 pandemic has affected millions of people and posed an unprecedented burden on healthcare systems and economies worldwide since the outbreak of the COVID-19. A considerable number of nations have investigated COVID-19 and proposed a series of prevention and treatment strategies thus far. The pandemic prevention strategies implemented in China have suggested that the spread of COVID-19 can be effectively reduced by restricting large-scale gathering, developing community-scale nucleic acid testing, and conducting epidemiological investigations, whereas sporadic cases have always been identified in numerous places. Currently, there is still no decisive therapy for COVID-19 or related complications. The development of COVID-19 vaccines has raised the hope for mitigating this pandemic based on the intercross immunity induced by COVID-19. Thus far, several types of COVID-19 vaccines have been developed and released to into financial markets. From the perspective of vaccine use in globe, COVID-19 vaccines are beneficial to mitigate the pandemic, whereas the relative adverse events have been reported progressively. This is a review about the development, challenges and prospects of COVID-19 vaccines, and it can provide more insights into all aspects of the vaccines.

Graphical Abstract

1. Introduction

The current coronavirus outbreak occurred in Wuhan, China, was first reported to the World Health Organization (WHO) on December 31, 2019. The WHO first named this novel coronavirus “2019-nCoV” on January 12, 2020 and formally named the disease “coronavirus disease 2019” or “COVID-19” on February 11, 2020[1]. It was also termed SARS-CoV-2 due to the similarity with severe acute respiratory syndrome coronavirus (SARS-CoV) [2]. The COVID-19 caused by SARS-CoV-2 has become a pandemic in almost all nations. During the COVID-19 pandemic, nations worldwide have taken the corresponding prophylactic measures. The initial measures comprise reducing social gathering, nucleic acid screening, and epidemiological investigations [3], [4], followed by the introduction of antiviral drugs and anti-inflammatory drugs (e.g., inhibitors of SARS spike proteins, neuraminidase inhibitors, ribavirin, α-interferon, remdesivir and glucocorticoids) [5], [6]. Subsequently, the USA, the UK, China, and several nations announced the successful development of COVID-19 vaccine candidates and have used the vaccines in practice. The existing large-scale clinical trials have suggested that the development of COVID-19 vaccines is beneficial to control and prevent the pandemic [7], [8].

2. COVID-19 and immune system

SARS-CoV-2 is a novel type of β-coronavirus with a viral envelope and a positive-sense, single-stranded RNA. The RNA comprises nearly 30,000 base pairs and 14 open reading frames (ORF) encoding multiple proteins [9], [10]. ORF1a/b took up 2/3 of 5’ end genome encodes 16 non-structural proteins (NSPs), whereas the structural proteins (e.g., nucleocapsid, membrane, envelope, and spike proteins) were encoded by ORF2–10, and eight accessory proteins were encoded by ORF3a/b, ORF6, ORF7a/b, ORF8, ORF9, and ORF10 [11], [12]. Angiotensin-converting enzyme 2 (ACE2) was confirmed as a receptor of SARS-CoV-2 and expressed abundantly in lung epithelium of human being [13]. SARS-CoV-2, specifically bound to the surface receptors, releases the internal genome into the host cytoplasm after membrane fusion, which requires the S protein-mediated [12], [13]. The S protein contains two functional subunits, including S1 and S2, of which S1 is responsible for the specific recognition and binding of the ACE2 receptor, and S2 subunit induces the fusion of the viral envelope and the host cell membrane [13], [14]. Furthermore, existing research has suggested that the type II transmembrane serine protein (TMPRSS2) of host cells, when subjected to SARS-CoV-2 infection, plays a certain role in activating virus S protein and promoting virus invasion into host cells [15].

SARS-CoV-2 exhibits stronger and faster ability of viral replication than SARS-CoV. SARS-CoV-2 proliferates in immune organs (e.g., lymph nodes and spleen), which destroys lymphocytes and monocytes, thus resulting in a decreased level of lymphocytes. Pathological anatomy has suggested that COVID-19 primarily causes deep airway and alveolar damage. Although considerable lymphocytes are infiltrated in the alveolar space and pulmonary septum, there are also excessive activation of CD4 + and CD8 + T cells, which are manifested as increased pro-inflammatory Th1, Th17 transformed from CD4 +T cells, as well as high cytotoxicity of CD8 +T cells. Likewise, more neutrophil and macrophage infiltration, severe "cytokine storm", stronger complement activation, and lower levels of lymphocyte counts were indicated in the outcomes of clinical and autopsy of critically ill patients with COVID-19 [16], [17], [18]. SARS CoV-2 can invade a wide variety of immune cells in the host and mediate the abnormal immune response by inducing disorder and death of the immune cells, thus disrupting the immune system [19].

Innate immunity serves as the first line of defense against microbes, including SARS-CoV-2. Dendritic cells (DCs), monocytes, granulocytes, natural killer cells (NK) and macrophages are effector cells. Macrophages and natural killer cells are capable of recognizing viruses through pattern recognition receptors, recruiting neutrophils and monocytes to gather at the site of infection, secreting active mediators and inflammatory factors, killing viruses and tissue cells non-specifically, removing necrotic cells and activating coagulation system and fibroblasts [19], [20]. Moreover, pattern recognition receptors (PRRs) and associated signaling pathways take on a critical significance to the innate immune immunity, thus stimulating the production of Type I/III interferons (IFNs), chemokines, and pro-inflammatory cytokines and activating inflammatory responses [21]. When the body suffered from the invasion of COVID-19, the PRRs of host cell specifically recognize the pathogen-associated molecular patterns (PAMPs) of SARS-CoV-2 to initiate innate immune response. PAMPs comprise unique viral genome nucleic acid structures and viral replication intermediates, which are primarily recognized by Toll-like Receptors (TLRs) and/or Retinoic acid-inducible gene i (RIG-i) (e.g., receptors (RLRs)), which belong to PRRs [21], [22].

Existing research has suggested that TLRs and RLRs are critical to initiating the innate immune response [21], [23]. TLRs are transmembrane protein receptors in which the extracellular domain covers the leucine-rich repeats, and the intracellular domain contains specific TIR domain to bind TLRs to the interleukin 1 (IL-1) receptor. The intracellular of TLRs (TLR7/8 and TLR3) are primarily responsible for recognizing nucleic acids, whereas extracellular TLRs (TLR1/2 and TLR4) largely recognize viral proteins and lipids [24]. Myeloid differentiation factor 88 (MyD88) is recruited to the TIR domain of TLR7/8, and a random TLR-MyD88 complex is formed, thus up-regulating the expression of cytokines and IFN by activating the downstream effectors [25], [26], [27]. TLR3 induces the expression of IFN through the TIR domain that contains adaptor inducing interferonβ (TRIF) pathway [21], [28]. Besides, RLRs are sensors specifically recognizing a wide variety of dsRNAs in cytoplasm [21], [23], [26]. The C-terminal domain of RIG-I binds to double-stranded RNA (dsRNA), and the dsRNA-CTD interacts with the helicase 2i (Hel-2i) of RID-I to release the free caspase activation and recruitment domain (CARDs). As a result, to a cascade response is generated to stimulate IRF3/IRF7 phosphorylation and translocation to the nucleus, thus up-regulating the expression of IFN and cytokines [29], [30], [31].

A study enrolled 55 patients with COVID-19, and the immune characteristics were summarized. The number of DCs was down-regulated, which is the main source of IFN. Besides, IFN-β was not identified, while a low level of IFN-α2 was reported in the plasma of severe COVID-19 patients [32]. A study contained 15 moderate-severe COVID-19 patients and four healthy individuals. Antigen-presenting cells in the blood collected on the 1st day and the 4th day after admission were established through single-cell RNA sequencing. The result indicated that the antigen presenting cells (APCs) of severe patients had defects in antiviral immune response (e.g., increased proapoptotic pathways in plasmacytoid DC (pDC), decreased innate immune receptors TLR9 and DHX36 and CLEC9a+DC cell subtypes, and reduced MHC-II trans-activator activities in cDC1c+DCs). In addition, antiviral interferon stimulated genes in monocyte subsets were down-regulated [33]. Moreover, the number of NK cells in some patients with COVID-19 was down-regulated and accompanied with the reduced ability of secreting IFN-γ, IL-2, and TNF-α. Another existing study has suggested that COVID-19 patients showed higher proportions of T cells and NK cells at the early recovery stage and at the late recovery stage. Compared with early recovery stage patients, patients at the late recovery stage have more lymphocytes, thus indicating the signs of gradual recovery [34], [35]. The above phenomena suggest that COVID-19 may have impaired innate immune system at an early stage.

B cells and neutralizing antibodies significantly make up humoral immune activation, thus effectively clearing pathogens and preventing reinfection [36]. Activated B cell matured from Naive B cells through the activation of T follicular helper cells will progress to memory B cells and IgG‐producing plasmablasts [36], [37]. Currently, SARS-CoV-2-specific IgM and IgG serve as one of the etiological diagnostic criteria [37], [38]. As indicated by a study involving 175 convalescent patients with mild COVID-19 disease, the neutralizing antibodies in COVID-19 patients were low in the first 10 days after onset and remained stable after reaching their maximum peak in 10–15 days [39]. Likewise, the plasma of 14 convalescents with mild SARS-CoV-2 infection was collected and IgM and IgG antibodies against NP and S-RBD were detected, and IgG remained at a high level 2 weeks after discharge [40]. The positive rate of COVID-19-specific antibodies in patients tended to increase over time, and the seroconversion of antibodies occurred in 19 days. Besides, all patients produced IgG, and the positive rate of IgM reached up to 94.1% at 20–22 days after the onset of symptoms [41]. The result indicated that the seroconversion of IgG and IgM in critically ill patients was higher, thus suggesting that a higher titer of antibody is independently correlated with a worse clinical classification[42], [43].

Cellular immunity is considered a vital part of adaptive immunity, playing a certain role in the anti-virus process. T cells are the main effector cells involved in cellular immunity and provide guarantee for innate immunity and humoral immunity [44], [45]. CD4 +T cells will differentiate into different effector T cells under the action of different cytokines, while CD8 +T cells will mature into cytotoxic T cells (CTL) [46]. The result of the pathological examination of COVID-19 patients suggested that the number of CD4 +T and CD8 +T cells in peripheral blood was reduced, and that in the spleen and lymph nodes was also down-regulated, accompanied by lymphocyte degeneration and necrosis [17], [44], [47]. Existing research has revealed that the function and number of T cells is an independent predictor with the prognosis of COVID-19 infection. Most COVID-19 patients are characterized by the down-regulated number of CD4 +T and CD8 +T cells and an imbalance ratio of T cell subsets [44], [45], [48]. Besides the change of the number of lymphocytes, an overactivated state of T cells were identified [17], [49]. Especially in severe ill patients the number of CD4 +T cells and CD8 +T cells are significantly reduced, accompanied with the reduced ability on secreting TNF-α, IFN-γ of CD4 +T cells, and CD8 +T cells exhaustion [50], [51].

The endogenous protein synthesized by SARS-CoV-2 in the host cells activates specific CD8 +T cells through the major histocompatibility complex I (MHC-Ⅰ), thus facilitating their proliferation and generating cytotoxic effects on target cells [52]. CD4 +T cells can differentiate into different T helper cells through the binding of MHC-II antigen complex of antigen presenting cells (APCs) [53]. CD4 +T cells are primarily differentiated to Th1 (helper T cell), Th2, Th17, and Treg [54]. Th1 can facilitate the proinflammatory response, while Th2 is capable of regulating the anti-inflammatory response [55], [56].

It keeps a relative balance between Th1 and Th2 cells under normal conditions [55], [56], [57]. Th1 and Th2 cells secrete cytokines to promote their own proliferation and negatively cross-regulate each other, maintaining the balance of Th1/Th2 is conducive to humoral and cellular immune [57], [58], [59]. The massive replication of COVID-19 in the host cells activates CD4 +T cells to transform into Th1 cells, In addition, B cells can be activated by Th1 and produce IL-6 and antibodies (e.g., IgG), which promotes the transformation of CD4 +T cells into Th1 cells as well [60], [61]. Th1 cells can secrete considerable proinflammatory factors, such as IL-6, IFN-γ, TNF-α and granulocyte macrophage colony-stimulating factor (GM-CSF) [57], [58], [60], [62]. GM-CSF induces monocytes to transform into M1 macrophages, which can produce of IL-6, IL-18, iNOS and other cytokines [62], [63]. The large-scale cytokines can increase glycolysis, and the released lactic acid is capable of further facilitating the transformation of Th1, thus secreting more cytokines and impairing the cytolytic ability of CD8 +T cells [63], [64], [65]. Besides, CD4 +T cell subsets will be differentiated into Th2, secrete IL-4, IL-10, TGF-β and other anti-inflammatory cytokines, thus inhibiting the proliferation of T cells and inducing monocytes to transform into M2 macrophages [61], [66], [67]. In addition, the imbalance between Th17 and Treg is the focus of COVID-19 infection. Th17 and Treg maintain dynamic balance in normal circumstances [68], [69]. Th17 belongs to proinflammatory cells, while Treg can inhibit effector T cells [70]. During the infection, TGF-β and IL-6 can induce the expression of retinoid related orphan receiver γt（RORγT), thus facilitating the differentiation of Th17 and secretion of proinflammatory cytokines. The cytokines (e.g., IL-17, IL-21, and IL-22) further promote the differentiation and proliferation of Th17 while enhancing the inflammatory response and the autoimmune response [69], [71], [72]. Furthermore, TGF-β can induce the expression of forehead box P3 (FoxP3) and promote differentiation into Treg, which secretes TGF-β and IL-10, resulting in inhibiting the inflammation response [71], [73]. Once the balance of T cell subset is broken during COVID-19 infection, it can also lead to the immunosuppressive state of cytokine accumulation and lymphocytes reduction [68], [69], [71].

With the similarity to the Tregs, Myeloid-derived suppressor cells (MDSCs) have the immunosuppressive ability [74]. There are PMN-MDSCs derived from myeloid granulocytes/multinuclear lineages and M-MDSCs derived from bone marrow mononuclear lineages, which are correlated with the development of COVID-19 [75]. Studies have shown that the accumulation of M-MDSCs and PMN-MDSCs with strong immunosuppression in severe ill patients with COVID-19. Analyzing from the peripheral blood cells of COVID-19 patients showed that HLA-DR low monocytes with M-MDSCs characteristics accumulated in the peripheral blood and immature neutrophils with immunosuppressive characteristics (PMN-MDSCs) accumulated in the blood and lungs [76], [77].

Clinical data showed that the cytokine storm caused by COVID-19 is correlated with severity ill patients. Cytokine storm ( Fig. 1 ), i.e., hypercytokinemia, refers to the excessive activation of the immune system after the organism is infected with microbes, such that a wide variety of cytokines are rapidly produced on a large scale in the body fluid. The immune response will be excessively activated and amplified in a chain if the pathogen cannot be eliminated in time [78]. Besides, the aggregation of considerable immune cells and cytokines can trigger the formation of thrombosis. The reasons for the above result are presented as follows: [1] IL-6 and IL-1β facilitate the formation of tissue factors (TFs) of monocytes, macrophages, and vascular epithelial cells in the blood; [2] TNF-α and IL-1β promote the formation of plasma fibrinogen; [3] the adhesion, activation, and aggregation of platelet, an inflammatory component, promote the formation of thrombus [79], [80], [81].

Fig. 1.

This figure illustrates the cascade immune response, the endogenous protein synthesized by COVID-19 in the host cells activates and promotes the proliferation of specific CD8 +T cells through MHC-Ⅰ, which produce cytotoxic effects on the target cells. CD4 +T cells can differentiate into different Th cells（include Th1, Th2, Th17 and Treg） through the binding of MHC-II antigen complex of antigen presenting cells(APC). The massive replication of COVID-19 activates CD4 +T cells to differentiated into Th1 cells, which can secrete a large number of proinflammatory factors, such as IL-6, IFN-γ and granulocyte macrophage colony-stimulating factor (GM-CSF). B cells can be activated by Th1 and produce IL-6 and IgG and IgM. GM-CSF induces monocytes to differentiated into M1 macrophages, which produce IL-6, IL-18, iNOS and a large number of inflammatory chemokines and increase glycolysis in the tissues, while the released lactic acid can further damage the cytolytic ability of CD8 +T cells. CD4 +T cell can also differentiate into Th2, which secrete IL-4, IL-10, TGF-β and other anti-inflammatory factors, inhibit the proliferation of T cells and induce monocytes to transform into M2 macrophages. On the other hand, TGF-β and IL-6 can induce the expression of expression of RORγT and promote CD4 +T cells to differentiate into Th17, which can secrete proinflammatory cytokines, such as IL-17, IL-21 and IL-22. The cytokines thereby further promote the differentiation and proliferation of Th17 and enhance the inflammatory response and autoimmune response. TGF-β can also induce the expression of forkhead box P3 (FoxP3) and promote the differentiation of CD4 +T cells into Treg, which secretes TGF-β and IL-10.

3. The current research status of the COVID-19 vaccines

3.1. Antigen selection for COVID-19 vaccines

COVID-19 is highly infectious and destructive to the immune system (e.g., innate and adaptive immunity). Immunotherapy has been confirmed to be effective in preventing the infectious diseases [82], [83]. Vaccination is a crucial part of immunotherapy, and the vaccine regulates the function of the immune system by simulating the immunogenic characteristics of a part or the whole of virus [7], [83]. Besides immunogenic proteins, RNA or DNA of COVID-19 also have the antigenic properties, which are manifested by inducing heterologous expression of immune proteins or peptides in the host. However, the antigenic components can be directly or indirectly recognized by APCs and play a certain role in immune responses [84], [85].

Effective antigens are capable of stimulating the organisms to produce innate and adaptive immunity, thus promoting the loss of pathogenicity and resistance to viral invasion. The antigenic targets of coronavirus vaccines have become clearer through preclinical research on SARS-CoV and MERS-CoV vaccines [86], [87]. Antibodies bound to the spike protein, especially to its receptor-binding domain (RBD), can prevent their attachment to the host cell and neutralize the virus [88]. Since the outbreak of the COVID-19 pandemic we have learned much about the immune response to SARS-CoV-2 after natural infection. Existing research has suggested the antibodies directed to the spike protein neutralize the virus whether they target the RBD or other regions of the S protein [12], [13], [89], [90].

3.2. The Evolution of COVID-19 vaccines

Based on development and toxicological data of the existing vaccines, COVID-19 vaccines can quickly pass through the exploration stage of SARS CoV-2 vaccines, and the vaccine production process can be adjusted in accordance with existing vaccines or candidate vaccines [91]. In addition, the phase of clinical trial can be partially overlapped, the phase III trial can be rapidly performed during the mid-term analysis of the phase I/II data [92]. According to the WHO report dated to 30 September, 2022 [93], there were 172 vaccines at the stage of clinical development, as well as 199 at the stage of pre-clinical development. A total of 172 vaccine candidates for clinical application comprised 55 (32%) protein subunit vaccines, 23 (13%) virtual vector vaccines (non-replicating), 16 (9%) DNA vaccines, 22 (13%) inactive virus vaccines, 40 (23%) RNA vaccines, 23 (13%) virtual vector vaccines (replicating), 6 (4%) virus like particle vaccines, 2 (1%) VVr+antigen presenting cells (APCs), 2 (1%) live attenuated virus vaccines, 1 (1%) VVnr+antigen presenting cells (APCs), as well as 1 (1%) bacillus antigen spot excess vector vaccine ( Table 1)( Fig. 2). Different vaccine candidates have different doses. To be specific, the numbers of 1 dose vaccines, 2 dose vaccines, 1 dose vaccines, and TBD/ND have reached 40 (23%), 97 (56%), 2 (1%), and 33 (19%), respectively. Moreover, there are 14 vaccines of China in phase III clinical trials overseas and 7 vaccines have gained approval for commercial marketing or emergency use. Furthermore, two vaccines candidates qualify for the World Health Organization's list of emergency uses [93].

Table 1.

The number and percentage of the candidates of the clinical and pre-clinical vaccines.

Platform	Vaccines in clinical development（No. and %）		Vaccines in pre-clinical development（No. and %）
protein subunit	55	31.98%	77	38.69%
RNA	40	23.26%	25	12.56%
DNA	16	9.30%	16	8.04%
Inactivated Virus	22	12.79%	11	5.53%
Viral Vector (non-replicating)	23	13.37%	24	12.06%
Viral Vector (replicating)	4	2.33%	20	10.05%
Virus Like Particle	6	3.49%	19	9.55%
Live Attenuated Virus	2	1.16%	3	1.51%
VVr + Antigen Presenting Cell	2	1.16%	0	0
VVnr + Antigen Presenting Cell	1	0.58%	0	0
Bacterial antigen-spore expression vector	1	0.58%	0	0
bacteria vector（replicating）	0	0	1	0.50%
Live attenuated bacterial vector	0	0	2	1.01%
Cellular based vaccine	0	0	1	0.50%
total	172	100.00%	199	100.00%

(according to the data of World Health Organization, until September 30, 2022, https://www.who.int/publications/m/item/draft-landscape-of-covid-19-candidate-vaccines.)

Fig. 2.

The World Health Organization (WHO) maintains a working document that includes most of the vaccines (until September 30, 2022) in clinical and pre-clinical development. Available from: https://www.who.int/publications/m/item/draft-landscape-of-covid-19-candidate-vaccines.

3.3. Classification of vaccines

The most used vaccine types worldwide comprise nucleic acid vaccine, protein subunit vaccine, live attenuated vaccine, inactivated vaccine, virally vector vaccines, as well as virus like particles vaccine. Vaccines have unique design techniques and characteristics, as listed in Table 2. The induced immunogens are different due to different vaccine design methods, whereas the immune responses remain similar. Nucleic acid vaccines are relatively complex. DNA vaccines should enter the nucleus of host cells to further synthesize corresponding immunogenic proteins. mRNA is capable of synthesizing corresponding immunogenic proteins after entering the cytoplasm of host cells, and the protein coded by nucleic acid vaccines can induce immune response after being secreted by host cells. Moreover, virally vector vaccines cover virtual vector vaccines (non-replicating) and virtual vector vaccines (replicating). Both live attenuated vaccine and vector vaccines (replicating) exhibit the ability of duplication. In general, the protein subunit vaccine, the protein coded by nucleic acid vaccines and virtual vector vaccines, live attenuated vaccine, inactivated vaccine, virally vector vaccines and virus like particles vaccine can be recognized by (APCs), and these vaccines can further activate CD8 +T cells through MHC-II by presenting antigens to CD4 +T cells. The memory B cells and memory CTLs will be produced by the cascade immune response attributed to the vaccines. ( Fig. 3).

Table 2.

Summary and comparison of the different kinds of vaccines.

Classification	Typical vaccine (s)	Design technique	Advantage	Disadvantage	Reference
Nucleic vaccine	mRNA vaccine: BNT162b2 (Pfizer/BioNTech); mRNA-1273 (Moderna) DNA vaccine: INO-4800 (Inovio Pharmaceuticals)	It needs to synthesize corresponding the RNA/DNA of viral proteins.	It can be modified directly on the original sequence to facilitate a timely update and improve production speed.	It is difficult to store and transport mRNA vaccines. DNA vaccine has a risk of integration into the genome sequence of the host.	Ref[94], Ref[95], Ref[96], Ref[97], Ref[98], Ref[99], Ref[100], Ref[101], Ref[102].
Protein subunit vaccine	NVX-CoV2373 (Novavax)	Purified immunogenic protein or peptide of virus were extracted to stimulate the body to produce immune response.	The research and development ideas of protein subunit are relatively clear. It is characterized by high safety and stability	It requires adjuvant and booster injections. It takes time to determining optimal antigen combination.	Ref[103], Ref[104], Ref[105].
Live attenuated vaccine	COVI-VAC（Codagenix）	The weakened viruses retain the ability to replicate in the body and do not cause disease or cause limited disease	It is characterized by high yield and lasting immunity	The production and formulation of live attenuated COVID-19 vaccines is labor intensive and requires strict quality control, which will slow down large-scale production	Ref[106], Ref[107], Ref[108], Ref[109], Ref[110], Ref[111].
Inactivated vaccine	BBIBP⁃CorV (Sinopharm (Beijing)); CoronaVac (Sinovac CoronaVac); Covaxin (Bharat Biotech);	Inactivate viruses can act as immunogens to maintain the integrity of viral particles, which can not replicate.	It can induce a wider range of antibodies against more epitopes and it is cheaper, and easier to handle	It may lead to changes in immunogenicity, hypersensitivity and possible Th2 bias. It requires lots of antigen to elicit an antibody response.	Ref[112], Ref[113], Ref[114], Ref[115], Ref[116].
virally vector vaccines	AZD1222 (AstraZeneca); Ad5-nCoV (CanSino Biologics); Ad26. COV2. S (Johnson & Johnson); Sputnik V (Gamaleya Research Institute).	By using adenovirus as the vector, it can achieve eterologous expression of target genes of SARS-CoV-2	It can induce higher titers of neutralizing antibodies.	The people will have weakened response or failed to respond to the vaccines when inoculated or tolerated to the vector virus.	Ref[117], Ref[118], Ref[119], Ref[120], Ref[121].
virus like particles vaccine	CoVLP (Medicago&GSK)	It can trigger immune responses by simulating the structure of the SARS-CoV-2 and it is suitable for antigen presenting cells	Similar to the protein structures of SARS- CoV-2, it can induce immune responses effectively.	It is difficult to synthesize the proteins of SARS-CoV-2 and assemble virus particles.	Ref[11], Ref[12], Ref[122], Ref[123], Ref[124], Ref[125].

Fig. 3.

This figure illustrates the principles of different vaccines. nucleic acid vaccines are relatively complex. DNA vaccines need to enter the nucleus of host cells to further synthesize corresponding immunogenic proteins. mRNA can synthesize corresponding immunogenic proteins after entering the cytoplasm of host cells, the protein coded by nucleic acid vaccines can induce immune response after being secreted by host cells; virally vector vaccines contains virtual vector vaccines (non replicating) and virtual vector vaccines (replicating). Both live attenuated vaccine and vector vaccines (replicating) have the ability of duplication. Totally, the protein subunit vaccine, the protein coded by nucleic acid vaccines and virtual vector vaccines, live attenuated vaccine, inactivated vaccine, virally vector vaccines and virus like particles vaccine can be recognized by antigen presenting cells (APCs), which can further activate CD4 +T cells through MHC-II antigen complex. The memory B cells and memory CD8 +T cells will be produced by the cascade immune response caused by vaccines.

3.4. Nucleic acid vaccines

In general, nucleic acid vaccines of COVID-19 comprise a segment of DNA or mRNA sequence, capable of encoding spike(S) protein or RBD [126]. BNT162b2 (Pfizer and BioNTech) and mRNA-1273 (Moderna) are mRNA vaccines based on mRNA molecules with synthetic stable S protein of COVID-19 [127], [128]. The advantage of mRNA vaccines is that only the corresponding RNA/DNA of the viral protein should be synthesized, which increases the production speed. It can be modified directly on the original sequence to facilitate a timely update and metabolized and eliminated by the natural mechanism of the host [129], [130]. mRNA-1273 vaccine is the first mRNA vaccine of COVID-19. It showed that neutralizing antibodies were detected in all patients after two-dose inoculation in Phase I clinical trial. The results of phase III trial showed that mRNA-1273 is safe and effective in preventing COVID-19 during the pandemic [91]. Clinical trials also show that the BNT162b2 was safe for immunocompromised patients and highlight the need for additional vaccine doses in certain immunocompromised patient groups to improve immunity [94], [95].

The conditions for storage and transportation of mRNA vaccine are relatively strict though this vaccine is highly efficient and relatively easy to manufacture on a large scale. Multiple dose vials of BNT162b2 vaccine are stored at − 80–60ºC, while thaw and store undiluted vials are stored at 2–8ºC for up to 5 days, the thaw undiluted vials at ambient temperature (up to 25 ℃) maintain for 30 min. Meanwhile, Multiple-dose vials of mRNA-1273 vaccine are stored at − 25 − 15ºC, the mRNA-1273 vaccine remains stable at 2–8℃for 30 days and remains only 12 h at ambient temperature [96], [97], [131]. It is more difficult to store mRNA vaccines for some underdeveloped nations or regions. Furthermore, current clinical data suggests that cellular immunogenicity of mRNA vaccines may need to be further improved [94]. RNA/DNA vaccines are relatively safe and well tolerated. Compared with mRNA vaccines, DNA vaccines exhibit higher stability. INO-4800 (Inovio) is a SARS-CoV-2 Spike DNA-based vaccine, transporting DNA of SARS-CoV-2 to host cells through plasmids. Unlike mRNA vaccines, DNA vaccines should enter the nucleus and display its function, such that there is a risk of integration into the host genome sequence. In the phase Ⅰ of clinical trial, INO-4800 vaccine induced humoral and cellular immunity in the vaccinated subjects, which is characterized by functional antibody and T cell reaction [132], [133]. Clinical study data indicated that DNA vaccines (39.5%, 29.3%) achieved significantly lower local and systemic reaction rates than RNA vaccines (89.4%, 83.3%) [98]. DNA vaccines are expected to be extensively employed through continuous improvement and development. In brief, the prospect of nucleic acid vaccines should be very bright.

3.5. Protein subunit vaccines

Protein subunit vaccines comprise purified immunogenic protein or peptide of virus, full-length spike or the corresponding ectodomain, and they have served as the antigen. The research and development ideas regarding protein subunit vaccines have been relatively clear [99]. NVX-CoV2373 produced by Novavax is one of the typical protein subunit vaccines, which has been proved to induce higher titers of neutralizing antibodies. It was composed of trimeric full-length S protein and saponin-based Matrix-M1 adjuvant [100]. Heath PT, et al. [100] conducted a phase Ⅲ clinical trial of NVX-CoV2373 at 33 sites in the UK, this research indicated an efficacy of 86.3% against the B.1.1.7 (or alpha) variant and 96.4% against non-B.1.1.7 variants. Reactogenicity of NVX-CoV2373 was generally mild and transient. The incidence of serious adverse events was low and similar in both groups of the clinical trial. Clinical trials conducted by Dunkle LM, et al. [101] in the US and Mexico in 2021 also concluded that NVX-CoV2373 is safe and effective for the prevention of Covid-19. At present, a protein subunit vaccine of China has entered the phase III clinical trial. Instead of using the full-length S protein, the RBD of S protein has been selected as the antigen of this vaccine. The phase III clinical trial involved 28,873 adult volunteers (aged 18 years and older) and indicated the protective effects of COVID-19, alpha mutant and delta mutant were 81.76%, 92.93% and 77.54%, respectively [102]. Since protein subunit vaccine exhibits low immunogenicity, adjuvant and booster injections are required, and it takes time to determining optimal antigen combination to potentiate the immune responses [134].

3.6. Live attenuated vaccines

The live attenuated vaccines (LAV) are based on the live microorganisms that have been weakened, which retain the ability to replicate in the body and cause limited disease, they are highly effective at inducing strong immune response and long-lasting immune memory. Thousands of people were protected from disabling and deadly diseases using LAV in the past [103], [104]. LAVs were attenuated mainly through adaptation to cold culture conditions or through non-human animals [135], COVI-VAC (a intranasal delivery vaccine) produced by Codagenix/Serum has entered Phase Ⅰ clinical trials[136]. The LAV candidates are achieved by isolating and culturing SARS-CoV-2 in vitro. The above low pathogenic mutants will modulate neutrophil influx, alleviate lung injury and enhance the immune response in animal trials [105]. LAV vaccines with the strongest immunogenicity, which can also effectively stimulate B lymphocytes and T lymphocytes and enhance humoral and cellular immunity. Besides, LAV candidates are subjected to several risks (e.g., viral recombination (especially homologous recombination) and phenotypic/genotypic reversion to virulent strains) [103], [106].

Advantages of LAV candidates of COVID-19 include targeting and stimulating robust mucosal and cellular immunity, which are critical for protection without adjuvants. However, there are concerns that LAV may be excreted in feces and lead to the transmission to unvaccinated individuals. LAV inoculation may also increase the risk of recombination between vaccine strains and circulating wild-type viruses, such that new mutations are caused [104], [107], [137]. Furthermore, the production and formulation of LAV is labor-intensive and requires strict quality control, hindering large-scale production [108].

3.7. Inactivated vaccines

Inactivated vaccines are produced using chemical agents to inactivate viruses grown in vitro. Vaccines can be the integrity of viral particles to maintain immunogens. Inactivated vaccines are injected to induce immune response. BBIBP⁃CorV (Sinopharm), CoronaVac (Sinovac Biotech), and Covaxin (Bharat) belong to typical inactivated vaccines [108], [138]. Vero cells in the production process were infected with SARS-CoV-2 from COVID-19 patients, and the most replicable HB02 strain was selected and then purified. The P1 stock was obtained through adaptive culture, subculture, and expansion in Vero cells. The P7 stock BJ-P-0207 was employed as the original strain, and the virus strain was inactivated using β-propiolactone [139]. The phase III clinical trial of BBIBP-CorV exhibited the safety and effectiveness. High titers of antibodies were produced after 2 doses when inoculation was completed, the positive rate of neutralizing antibodies reached 99.52%, and the protective effect against COVID-19 was obtained as 79.34% [109].

Conventional inactivated vaccines show their advantages. Inactivated vaccines are capable of inducing a wider range of antibodies against more epitopes of COVID-19 [110]. Although it may cause changes in immunogenicity, hypersensitivity and Th1/Th2/Th17 imbalance (Th2 bias), most of the above drawbacks have been ameliorated by adding appropriate adjuvants and adjusting vaccine doses, immunization schedules and the routes of administration [111]. Moreover, the incidence of adverse reactions of was low, and no death was reported in the clinical trials, thus suggesting that the vaccines are safe. However, the production of inactivated vaccines is limited and mainly correlated with the short duration of immune memory, thus requiring vaccination with larger amounts or the association of inactivated microorganisms with adjuvants [112]. Inactive vaccines are cheaper, safer, and easier to inject though the immune responses induced by inactive viruses are generally weaker than those elicited by attenuated viruses [103].

3.8. Virally vector vaccines

In general, virus vector vaccine, i.e., recombinant virus vector vaccine, employs adenovirus as the vector to replace some genes in adenovirus genes with target genes to achieve heterologous expression of target genes. There are two major classes of virally vectored vaccines, including virtual vector vaccines (non-replicating), 2) virtual vector vaccines (replicating) [113]. AZD1222 (AstraZeneca), Ad5-nCoV (Cansino Biologics), Sputnik V (Gamaleya) and Ad26. COV2. S (Janssen) are typical virus vector vaccines. AZD1222 adopts weakened chimpanzee adenovirus (ChAdOx1) to encode the information of wild type SARS CoV 2 spike protein, while Ad5 nCoV adopts human adenovirus serotype 5 vector (Ad5). Sputnik V uses Ad26 vector for priming and Ad5 vector for boost) [113], [140], [141]. In a phase Ⅲ trial of AZD-1222 experiment, 8895 adult participants were enrolled and two doses of vaccines were administered at an interval of 4 weeks. The results indicated that the efficacy was obtained as 62.1% [140]. However, the safety of recombinant adenovirus vector vaccine has aroused extensive attention due to the occurrence of venous thromboembolism in several individuals vaccinated with AZD1222 vaccine [141]. Moreover, a Phase Ⅲ trial of Sputnik V achieved 91.6% efficacy against COVID-19, and the vaccine was effectively tolerated [114], [115]. A phase Ⅲ trial of Ad5 nCoV involving Pakistan, Mexico, Russia, Chile, and Argentina confirmed that the efficacy reached 57.5% (95% CI 39.7–70.0, p = 0.0026) of one dose of Ad5 nCoV at 28 days or more post-vaccination [116].

Virally vector vaccine shows an advantage that the selected antigen is expressed in the background of active heterologous virus infection and exhibits strong immunogenicity [115]. Moreover, it can effectively induce the immune response mediated by B lymphocytes and T lymphocytes [141]. S protein serves as the vital protein of new coronavirus that invades the human body. When COVID-19 spike glycoprotein gene is recombined with adenovirus vector, adenovirus also exhibits the characteristics of coding S protein, thus producing stronger cellular and humoral immune responses and generating antibodies or memory immune cells specifically for the S protein of COVID-19 after inoculation [117].

3.9. Virus like particles vaccines

The virus like particles (VLP) vaccine has been confirmed as a good pattern to simulate pathogenic microorganisms to achieve a good protective effect. COVID-19 comprises three structural proteins, including S, M, and E [11], [12]. Accordingly, VLP can be constructed in vitro to simulate the structures of COVID-19, whereas virus particles are difficult to assemble. The researchers co-transfected three types of mRNA encoding S, M and E proteins into the cultured cells at the molar ratio at 1:2:2. The proteins could be detected by western blotting in the culture medium and purified by sucrose gradient. Lastly, spike protein was interspersed on the surface to form SARS CoV-2 virus like particles [118]. An animal experiment of VLP was performed. The binding antibody was detected in mice 20 days after the first injection, and the antibody titers increased dramatically in mice after a boost and peaked at 3rd week. Furthermore, mice immunized with VLP achieved an altered ratio (0.67:0.29), compared with the CD4⁺:CD8⁺ T cell ratio in the placebo group (0.74:0.20), thus showing a significant increase in CD8⁺ T cell frequency [119].

CoVLP is a recombinant plant-derived VLP vaccine designed by Medicago, Canada. The function of CoVLP was evaluated using homologous and heterologous prime-boost regimen in mice. As revealed by the result, CoVLP is capable of inducing strong and extensive cross-reaction neutralizing antibody (NAb) reaction against several variant virus strains of COVID-19 [142]. During the phase Ⅰ trial of CoVLP, the serum NAb titers were up-regulated significantly, which were ten-fold higher than titers of COVID-19 convalescent. Furthermore, S protein-specific IFN-γ and IL-4 related responses were induced. Currently, the phase II/III trials of CoVLP have been performed [120].

4. Adverse reactions of vaccines

Existing research has suggested that vaccines are highly effective and safe for the COVID-19 pandemic. However, some people experienced adverse effects after being vaccinated with the vaccines [143]. COVID-19 vaccines are more likely to cause adverse reactions than other commercially available vaccines, which should be investigated in depth. Fatigue, muscle pain, headache, chills, joint pain, fever, and skin conditions (e.g., pernio/chilblains, zoster, and herpes simplex flares) have been confirmed as the most common adverse effects [121], [122]. In general, the adverse reaction relatively rare and the symptoms were mild, which usually occurred within the first 3 days after vaccination and relieved within 1–2 days. Furthermore, clinical trials have suggested that some moderate to severe adverse events are rare, whereas they may be correlated with the vaccination (e.g., hypersensitive reaction [123]). Most of adverse events are correlated with the 2nd dose vaccination. Some of the severe complications are summarized in Table 3.

Table 3.

Summary of some different complications of the vaccines.

Complications	Clinicle presentation	Mainly affected areas	Probable mechanisms	Differential diagnosis	Reference
Hypersensitive reaction	hives, throat swelling, shortness of breath, chest tightness, low blood pressure and even loss of consciousness	Skin and respiratory tracts	1. The people with a history of and anaphylaxis or have mast cell activating diseases, 2.women are susceptible to allergic reactions. 3. The excipients of the vaccine may also induce IgE/ non-IgE allergic reactions.	dermatosis	Ref[144], Ref[145], Ref[146], Ref[147], Ref[148], Ref[149], Ref[150], Ref[151].
Immune thrombotic thrombocytopenia	It is characterized by thrombocytopenia, systemic activation of coagulation, and extensive venous thrombosis and platelet factor 4(PF4) antibody. The presentation of cerebral venous thrombosis includes headache, seizures, dyspnea, limbs’weakness, petechiae, and lethargy.	extensive venous(cerebral vessels or dural sinuses, visceral vessels)	1. PF4 antibodies binds to the site on PF4 and activate platelets through the through FcγRIIa receptors. 2. Antibody (Ab2) response is elicted by Vaccine elicited specific antibody (Ab1)	autoimmune heparin-induced thrombocytopenia (HIT)	Ref[152], Ref[153], Ref[154], Ref[155], Ref[156], Ref[157], Ref[158], Ref[159].
Nervous system disease	It characterized mainly as facial paralysis, acute demyelination, seizures and autonomic dysfunction.	Nervous system; Spinal cord	1.a history of underlying neurological diseases; 2. The molecular mimicry is between infectious antigens and self-antigens; 3.adjuvants could aberrantly provoke innate and adaptive immune responses, which promote pro-inflammatory cascades.	Guillain-Barre syndrome (GBS); acute transverse myelitis (ATM)	Ref[159], Ref[160], Ref[161], Ref[162], Ref[163], Ref[160].
Myocarditis	Myocarditis is defined as the presence of inflammatory cellular infiltrate in the myocardium alongside tissue necrosis. The most common clinical symptoms include chest pain, fever, dyspnea, fatigue	Myocardium	1.molecular mimicry between the viral spike protein and self-antigens. 2.preexisting immune dysregulation, leading to polyclonal B-cell expansion, immune complex formation, and inflammation.	coronary heart disease	Ref[164], Ref[165], Ref[166], Ref[167], Ref[168], Ref[169], Ref[170].
Rhabdomyolysis	Symptoms include limb pain, weakness, difficulty walking, and hematuria (tawny urine).	Striae (skeletal) muscle cells	several hypotheses have been proposed, including pathogenic mutations in the RYR1 gene and vaccine-induced complement activation syndrome.	Acute renal failure	Ref[171], Ref[172], Ref[173], Ref[174].

4.1. hypersensitive reaction

Hypersensitive reaction is a severe but rare adverse event, representing hives, throat swelling, shortness of breath, chest tightness, low blood pressure and even loss of consciousness [121], [124]. A study on BNT162b2 and mRNA-1273 showed that 80% of allergic reactions occurred within 15 min, whereas 89% occurred in 30 min after vaccination, The incidence of acute severe allergic reaction of BNT162b2 is 4.7 per million doses, while 2.5 per million doses of mRNA-1273 [125]. Hypersensitive reactions tended to occur in the people with a history of and anaphylaxis or the patients who have mast cell activating diseases, women are susceptible to allergic reactions [175]. In addition, the excipients of the vaccine may induce allergic reactions. Liposome is a vital adjunct of mRNA vaccine, capable of wrapping mRNA and maintaining stability. Polyethylene glycol (PEG) contained in liposome is the potential cause of allergic reaction of mRNA vaccine. Cross reactive anaphylaxis may also occur between the above compounds due to the similar structures to polysorbate. In addition, ammonia methanol in mRNA-1273 vaccine is one of the substances causing allergy [144], [176]. Immunological tests show that non-IgE-mediated immune responses to PEG may be responsible for most individuals, while PEG in its native form cannot be as immunoreactive as PEG-2000/lipid conjugates [177], [145]. Moreover, IgE-mediated allergic reactions have aroused broad attention. However, IgE-mediated hypersensitive reactions to vaccines occur in less than 1 case per million applications, as indicated by the evaluation of European Academy of Allergy and Clinical Immunology [178], [146].

Skin prick test (SPT), basophil activation test (BAT) provide insight into a range of vaccine-related hypersensitive responses [147]. SPT and IDT are correlated with BNT162b2, mRNA-1273, AZD1222, Ad26. COV2. S and also useful tools in a COVID-19 vaccination regimen that enables safe immunization of patients at high allergy risk [148]. Guidance from the Centers for Disease Control and Prevention (USA) recommends that individuals who have an allergic reaction or are allergic to the component of COVID-19 vaccines should avoid the same vaccination[149]. Some research also indicated that most patients allergic to the first vaccine do not respond to the second. Although the probability of allergy after vaccination with COVID-19 vaccines is higher than other vaccines, the above COVID-19 vaccines are effective in curbing the development of COVID-19 [150], [151].

4.2. Immune thrombotic thrombocytopenia

Vaccine-induced immune thrombotic thrombocytopenia (VITT) is a prothrombotic syndrome identified after the inoculation of adenoviral vector-based vaccines, characterized by thrombocytopenia, systemic activation of coagulation, extensive venous (especially cerebral venous) thrombosis and platelet factor 4(PF4) antibody[179]. Abbattista et al. calculated the incidence of adverse events correlated with VITT of four vaccines candidates (including BNT162b2, mRNA1273, AZD1222 and Ad26. COV2. S) from January 1, 2021 to July 30, 2021. The total reporting rate of thrombosis reached 39.9/million doses, Pfizer vaccine was 15.9/million doses, Moderna was 75.1/million doses, and Johnson&Johnson was 69.4/million doses. Besides, AstraZeneca and was 122.9/million doses, cerebral vein thrombosis accounted for 14.1% of total thrombosis cases, and the total reporting rate of CVT of the four vaccines was 5.6/million doses. The number of thrombotic patients with thrombocytopenia accounted for 10% of total thrombosis cases, and the total reporting rate was 4.1 million doses [180]. Adenoviral vector vaccine elicited antiplatelet auto- and alloantibody was correlated with platelet activation, documented by raised circulating platelet-derived microvesicles, sP-sel and mild blood clotting activation in vivo, especially for AZD1222 and Ad26. COV2. S [179], [181]. PF4 antibody can be bound to specific site on PF4, corresponding to heparin, forming immune complex and activating platelets through Fcγ-Receptor Iia [182]. Cerebral venous sinus thrombosis is a severe pathological disease occurring secondary to blood clots, thus obstructing the cerebral vessels or dural sinuses [183]. Clinical manifestations comprise generalized or localized headache correlated with hemiplegia and aphasia, as well as seizures and encephalopathy occurred in a few of the above cases for intracranial hypertension [184]. Greinacher, et al. have suggested that 11 thrombotic events in German 16 days after vaccination of AZD1222 comprised cerebral vein thrombosis, visceral vein thrombosis, pulmonary embolism and other thrombosis, whereas the CDC(USA) also announced 12 cases of thrombocytopenia attributed to cerebral venous sinus thrombosis after vaccination with Ad26. COV2. S [181], [182]. Although with the similarities to autoimmune heparin-induced thrombocytopenia (HIT) [152], platelet activation in VITT occurs in the presence of PF4 rather than low heparin concentrations. VITT and HIT can be identified through PF4-induced platelet activation test and PF4-induced flow cytometry–based platelet activation test [185], [186]. In general, VITT is correlated with disseminated intravascular coagulation, with significant increased plasma D-dimer and fibrinogen consumption [153], [154]. The mechanism of VITT induce by COVID-19 vaccines remains unclear, whereas viral vector vaccination may increase the transient production of platelet-specific autoantibodies or the cross-reaction between antiviral antibodies and platelet glycoproteins [155]. Furthermore, the vaccine-elicited anti-Spike-specific antibody (Ab1) has an immunogenic amino acid sequence, and it is termed an idiotype, which elicits an anti-idiotype antibody (Ab2) antibody response. Ab2 antibodies can be bound to antibodies that protectively neutralize Ab1 and impair Ab1 efficacy [179].

4.3. Nervous system disease

With the large-scale inoculation of COVID-19 vaccine worldwide, numerous neurological complications have been reported (e.g., Bell’s palsy, Guillain-Barre syndrome (GBS), acute transverse myelitis (ATM), and multiple sclerosis(MS)), primarily characterized as facial paralysis, acute demyelination, seizures, and autonomic dysfunction [156]. However, the correlation between neurological adverse effect and vaccination remains unclear. Several existing research has suggested that post-vaccination demyelination is most likely to trigger clinical disease expression in individuals with an underlying disease process already [157]. Bell’s palsy is an acute unilateral facial nerve palsy, and it is correlated with physical stress, infections, tumor, and vaccination [158]. Sato K, et al. [158] analyzed 303589 reports of vaccination and have indicated that 405 cases of facial paralysis were reported after BNT162b2, and 512 cases were reported after mRNA-1273. As revealed by the results, the incidence of facial nerve palsy was slight but significantly increased after the administration of BNT162b2 (ROR 1.84; 95% CI 1.65–2.06) and RNA-1273 (ROR 1.54; 95% CI 1.39–1.70). Surprisingly, a 61-year-old man developed unilateral facial nerve palsy 5 h after the 1st dose of BNT162b2 vaccination and occurred again 2 days after the 2nd dose [187]. The clinical trial of AZD1222 once paused, followed with a case of transverse myelitis I, i.e., an inflammatory disorder of the spinal cord that occurs due to different etiologies (e.g., infections, autoimmune and demyelinating diseases) [188]. Besides, the Vaccine Adverse Event Reporting System of CDC (USA) announced the occurrence of nine cases of acute TM with BNT162b2, mRNA1273, and AZD1222 [159]. GBS refers to an autoimmune peripheral neuropathy characterized by demyelinating lesions of peripheral nerves and nerve roots and small vessel inflammatory cell infiltration. Waheed S, et al. [189] have suggested that an 82-year-old woman developed Guillain-Barre syndrome after being vaccinated the first dose of Pfizer COVID-19 vaccine. The patient had general malaise in the first week and she developed worsening symptoms and showed difficulty in walking after the vaccination. A reported that a 48-year-old man presented left-sided lower motor neuron facial weakness (House-Brackmann grade III) 10 days after being vaccinated with the first dose of the Vaxzevria vaccine. The patient developed bilateral paralysis(House-Brackmann grade V) and severe back pain on the 13th day after vaccination[160]. In addition, The FDA(USA) has report 100 cases of Guillain Barre syndrome after approximately 12.5 million doses inoculation of Ad26. COV2. S[161]. The temporary correlation between vaccination and neurological complications does not imply causality. However, the pathogenesis remains unclear. The molecular mimicry between infectious antigens and self-antigens has been confirmed as one of the main proposed mechanisms for developing demyelination post-vaccination. Besides, adjuvants (substances increasing the antigen-specific immune responses) is capable of aberrantly provoking innate and adaptive immune responses of mRNA vaccines, which may promote several pro-inflammatory cascades by up-regulating the serum level of inflammatory cytokines [190], [191]. The elevated level of IL-6 in the CSF of patients with myelitis predicts a poor prognosis [162].

4.4. Myocarditis

There have been numerous reports on the onset of myocarditis and pericarditis after inoculation of COVID-19 vaccines; the risk of myocarditis in young people after mRNA vaccination was 5–25 times higher [192]. Myocarditis has been defined as the presence of inflammatory cellular infiltrate in the myocardium alongside tissue necrosis, which is not attributed to coronary heart disease; it is diagnosed in accordance with histological, immunological, and immunohistochemical criteria [193]. Most patients with myocarditis following COVID-19 vaccination demonstrate symptom onset in the first week after the second injection, with the vast majority presenting in the first 4 days, accompanied with an abnormal increase in troponin. The most common clinical symptoms of myocarditis after COVID-19 vaccination comprise chest pain, fever, dyspnea, as well as fatigue [194], [195]. It usually starts within 1–5 days. The research has indicated that only 9.3% of all adverse reactions after BNT162b2 vaccination for the age of 12–17 years were serious adverse events, and myocarditis accounted for 4.3% of the total adverse events [163]. The characteristics of COVID-19 vaccine-associated myocarditis in adolescents were nearly similar to those in adults, and the incidence of myocarditis after mRNA vaccination in males was significantly higher than that in females [196].

CDC(USA) announced on July 07, 2021 that approximately 296 million doses of mRNA vaccines (e.g., BNT162b2 and mRNA-1273) had been inoculated as of June 11, 2021, and over 177 million people have been vaccinated with at least one dose of COVID-19 vaccine, more than 1000 cases of adverse reactions to myocarditis and pericarditis after COVID-19 mRNA vaccine inoculation, For people age 12–29, the rates of myocarditis after the 2nd dose in male and female were nearly 40.6 and 4.2 cases per million, while for people over the age of 30, the rates of myocarditis after the 2nd dose in male and female were nearly 2.4 and 1.0 cases per million[169]. In addition, Mevorach, et al. [164] reported a total of 283 people developed symptoms of myocarditis of approximately 5.1 million people were vaccinated with two doses COVID-19 mRNA vaccines. 142 (50%) patients occurred after the BNT162b2 vaccine, whereas 136 were eventually diagnosed with probable or definitive myocarditis. One hypothesis involves molecular mimicry between the mRNA vaccine-encoded SARS-CoV2 spike glycoprotein and self-antigens, thus causing polyclonal B cell expansion, immune complex formation, and inflammation. The glycoprotein encoded by mRNA vaccine can be bound to the surface of cardiomyocytes via ACE2 receptors or deposited immune complexes, thus serving as an antigenic trigger of inflammation [165]. However, another hypothesis involves the production of antibodies targeting the anti-spike protein antibodies, thus mimicking the spike protein, binding cardiac ACE2 receptors, leading to the formation of immune complexes, and activating the classic complement pathway [166]. As a result, a subset of post-vaccine myocarditis may be caused by an innate inflammatory response to the mRNA-encoded viral spike glycoprotein [167].

4.5. Rhabdomyolysis

Rhabdomyolysis has been confirmed as a complication of COVID-19 infection, whereas recent reports have suggested that rhabdomyolysis is also capable of flaring up in 24 h to a week after COVID-19 vaccination. Symptoms comprise limb pain, weakness, difficulty walking, and hematuria (tawny urine). Rhabdomyolysis refers to the destruction of striae (skeletal) muscle cells and the release of their contents into circulation, especially electrolytes, myoglobin, creatine kinase, lactate dehydrogenase, alanine aminotransferase, as well as aspartate amino transferase [197], [198]. Although there is currently no definite mechanism explaining vaccine-associated rhabdomyolysis, several hypotheses have been proposed (e.g., pathogenic mutations in the RYR1 gene and vaccine-induced complement activation syndrome) [198], [199].

The pathological mechanism of rhabdomyolysis is either direct myometrial injury (trauma: muscle cell damage due to ischemia-reinjection and neutrophil-induced inflammation) or depletion of intramuscular adenosine triphosphate, thus resulting in uncontrolled calcium influx, myogenic fiber network collapse, and subsequent muscle cell death [200]. The severity of rhabdomyolysis ranges from asymptomatic elevations in muscle enzymes to life-threatening complications (e.g., acute kidney injury, disseminated intravascular coagulation, compartment syndrome, as well as electrolyte abnormalities). Resuscitation with intravenous fluids is employed as the primary treatment for the disease. Serum CK levels in patients with rhabdomyolysis tend to be at least 5 times the upper limit of normal (>5000 IU/L) and MRI scans can be used to identify the underlying cause[198].

5. Vaccination strategies

5.1. Mixed vaccinations

Since the outbreak of COVID-19, many nations have conducted vaccine research. So far, there are more than 300 types of vaccines in the world, including vaccines that have not been put into clinical use [93]. However, the effectiveness and safety of vaccines are different, and some vaccines can cause serious adverse events after vaccination. Thus, increasing the efficiency and safety of vaccines is the current trend of vaccine research and development. Currently, vaccination with homologous COVID-19 vaccine has been employed as the main recommended method. Given the uneven distribution of global vaccine resources and the inability of some people to complete homologous vaccination due to adverse reactions, some nations have conducted the heterologous vaccination projects, i.e., two or more different vaccine types are adopted to deliver the same or similar pathogenic antigens, to enhance the immune response and the effectiveness of protection. Several successful research has suggested that mixed vaccination is capable of enhancing immunogenicity and the humoral immune response [168], [201], [170]. The vaccine protection effectiveness is 88%, as indicated by a study on heterologous vaccination (hAdOx1⁃S+mRNA vaccine) based on large samples in Denmark [171]. Besides, there has been research on heterologous vaccination based on inactivated vaccine in China and Brazil. The study of Brazil suggested that vaccine protection effectiveness (92.7%) is significantly improved after inoculation of two doses of Sinovac inactivated vaccine and a booster of BNT162b2 vaccine [172]. The existing data of mixed vaccination have been rare, and the combination of vaccines, the sequence and time of vaccination may affect the effect of vaccination. However, the study of this vaccination method provides a potential mechanism for hybrid COVID-19 vaccines to achieve higher immune responses [199], [173].

5.2. Vaccination of special groups

As vaccines have been leaping forward, the treatment for COVID-19 has shown a bright prospect, whereas there are many people with special constitutions, which comprise pregnant women, chronic disease (e.g., diabetes, chronic kidney disease) and immunosuppressed patients (e.g., malignant tumors, organ transplant patients, autoimmune diseases). Pregnant women were not included in the vaccination program since there has been rare evidence of vaccine safety and efficacy in the crowd at the early stage. Extensive studies have been conducted on COVID-19 vaccination during pregnancy and lactation. The antibody response rate was nearly 100% when pregnant women were vaccinated with mRNA COVID-19 vaccine [174], [202]. compared with 10861 pregnant women who were not vaccinated, the vaccine protection efficiency of 10861 pregnant women who were vaccinated with BNT162b2 was 96% within 7–56 days after inoculation [203]. Women vaccinated with the COVID‐19 vaccine during the first 20 weeks of gestation also had no increased risk of miscarriage compared with the general pregnant population [204]. The safety of COVID-19 vaccine for pregnant women and their offspring cannot be determined since there has been rare large sample research. Accordingly, it is recommended that the above women be postponed from vaccination in China [205]. Pregnant women can accept vaccination if they make an informed choice and lactation is not a contraindication to vaccination, as recommended by the International Federation of Obstetricians and Gynaecologists. Moreover, the American Academy of Breastfeeding Medicine has recommended that lactating women vaccinated with the COVID-19 vaccine can continue to breastfeed [206], [207]. Autoimmune disease is a type of disease caused by the dysfunction of the body's immune system, which breaks the immune tolerance of the immune system to its own components, nearly 100 autoimmune diseases have been confirmed and approximately 3–5% of the population are suffering from different degrees of autoimmune diseases [208]. A multicenter study [209] showed that the seropositivity rate and S1/S2 IgG levels were 86.0% (590/686) and (132.9 ± 91.7) BAU/ml in the patients with autoimmune inflammatory rheumatic diseases after BNT162b vaccination, while the seropositivity rate and S1/S2 IgG level were 100.0% (121/121）and（218.6 ± 82.1）BAU/ml in general population. Mado et al. [210] reported that the antibodies to the S-protein receptor binding domain of a 23-year-old female, who suffered from PPMS and received orezumab treatment, were detected after 4 weeks after the second injection of Pfizer COVID-19 mRNA vaccine. Bitoun et al. [211] illustrated that the patients treated with rituximab showed a lower proportion of neutralizing antibodies to the COVID-19 vaccine.

Tumor patients (e.g., solid tumors and hematologic malignancies) and the patients vaccinated with immunosuppression and immunomodulation therapy were not involved in the COVID-19 vaccine trails in the beginning of COVID-19 pandemic [212]. As vaccine technology has progressively matured and been popularized, several COVID-19 vaccine trails have been performed in tumor patients. Herishanu et al. [213] have suggested that serum antibody reaction was detected in only 39.5% (66/167) of 167 patients with chronic lymphoblastic leukemia who were vaccinated with BNT162b2 COVID-19 vaccine, and 52 patients had significantly lower antibody reaction rate than the health people. As indicated by a study of 95 patients with solid cancer, 56 patients with blood tumor and 54 healthy controls, the proportions of positive anti-S IgG titers in healthy controls, solid cancer patients and blood tumor patients accounted for (32/34, 94%), (21/56, 38%) and (8/44, 18%), respectively, whereas the proportions were (12/12100%), (18/19,95%) and (3/5,60%) 2 weeks after the objects were vaccinated with the 2nd dose of vaccine [214], approximately 21 days after the administration of 1st dose of BNT162b2 vaccine.

This study suggested that organ transplant recipients have weaker responsiveness to COVID-19 vaccines since they have undergone long-term immunosuppressive treatment. The results of the antibody titer of S-protein indicated that the organ transplant patients receiving mRNA had significantly lower antibody response than healthy people, only 6.2% (9/145) showed antibody reaction before the second vaccination of 145 kidney transplant patients vaccinated with the first COVID-19 vaccine (e.g., BNT162b2 and mRNA-1273) [215]. Benotmane et al. [216] have suggested that the positive rate of serum antibody in kidney transplant recipients was increased to 47.8% (98/205) in 28 days after the second dose of mRNA 1273 vaccine was inoculated. The incidence and severity of adverse reactions in organ transplant recipients after 1–2 doses of COVID-19 vaccine were not significantly different from those in the normal population [217]. Vaccinations should be completed in 2 weeks before transplantation, or the first dose should be started in 1 month after transplantation, as recommended in the American Society of Transplantation [218].

6. The VOCs of COVID-19 and the second-generation vaccines

6.1. The VOCs of COVID-19

It is worth mention that the mutation of SARS-CoV-2 gene evolved and caused new outbreaks with the development COVID-19 pandemic in the global. Variants of concern(VOC)/Variant of Interest(VOI) is a monitoring and grading system for COVID-19 variants of World Health Organization (WHO). VOCs include B.1.1.7 (Alpha), B.1.351 (Beta), P1 (Gamma), B.1.617.2 (Delta) and B.1.1.529(Omicron). Alpha and Beta variants were first confirmed on Dec.18, 2020 in United Kingdom and South Africa, respectively. Gamma variant was first confirmed on Jan.11, 2021 in Brizal, Delta variant was first confirmed on May.11, 2021 in India and Omicron variant was first confirmed on Nov.24, 2021 in South Africa [219], [220]. These VOCs will elevate the incidence and mortality of COVID-19 infection in different regions, and prolong the pandemic of COVID-19. Thus far, the Omicron variant, which is easy to spread, has occupied a leading position in the worldwide. Compared with ancestral COVID-19, the mutation of VOCs may lead to the phenotypic changes of spike protein. Whether it will affect the immune protection effect of existing vaccines remains inconclusive because the current research outcomes are still limited.

A clinical trail of Ad26. COV2. S vaccination conducted in South Africa showed that the vaccine efficacy was 52% 14 days after the first innoculation and 64% 28 days after the first innoculation. However, it has no effect on Beta variant. Another clinical trials of ChAdOx1-nCoV-19 illustrated that the overall vaccine efficacy of ChAdOx1-nCoV-19 was only 22%, and only 10% for the Beta variant. Meanwhile, the vaccine efficacy of NVX-CoV2373 for SARS-CoV-2, Alpha variant and Beta variant were 95.6%, 85.6% and 49.4%, respectively [219].

Wang L, et al. analyzed the ability of 14 COVID-19 mutants (belonging to VOCs and VOIs) to escape from the neutralizing antibody induced by mRNA vaccines. This research showed that the serum neutralization and replication were reduced in varying degrees after vaccination. Although the Omicron variant showed the highest escape from neutralization, the sera maintained moderate neutralization against this variant after a third dose of COVID-19 mRNA vaccin [221]. A study conducted in Israel showed that the vaccine efficacy of fully vaccination with BNT162b2 against the confirmed or possible Beta variant infection was 72% and against symptomatic confirmed or possible Beta varrient infection was 100% [222]. Similarly, the sera from the non-human primates or human subjects that received mRNA-1273 vaccination could effectively neutralize the experimental pseudovirus with spike glycoprotein containing B.1.1.7 lineage (Alpha), although the neutralization titer against B.1.351 lineage (Beta) had been reduced by 6.4-fold. However, the sera could still completely neutralize B.1.351 at a titer of 1:290, which was sufficient to prevent severe COVID-19 infection [223]. In addition, a study found that the convalescent sera of the unvaccinated patients who infected with ancestral COVID-19 showed a reduced neutralizing capacity on Beta and Omicron variants. Compared with ancestral COVID-19, the sera from individuals who received 3 doses of mRNA-1273 or BNT162b2 demonstrated lower neutralizing capacity on the Omicron variant. However, sera from individuals who infected with ancestral COVID-19 and subsequently vaccinated with 2 doses of BNT162b2 induced significantly higher neutralizing antibody titer against ancestral COVID-19 and all VOCs [224].

In addition to humoral immunity, cellular immunity may also contribute to the protection of COVID-19 and VOCs. COH04S1 is a synthetic multiantigen COVID-19 vaccine, which expresses potent spike(S) and nucleocapsid(N) antigen. A study showed that individuals maintained cross-reactive cellular immunity for more than 6 months after vaccination of COH04S1 or BNT162b2. Although reduced activity of neutralizing antibodies in sera induced by COH04S1 or BNT162b2 against Delta and Omicron variants in relative to ancestral SARS-CoV-2, S-specific T cells elicited by COH04S1 and BNT162b2 and N-specific T cells elicited by COH04S1 demonstrated potent cross-reactivity against ancestral SARS-CoV-2 and the major VOCs [225] (The Third dose of CoronVac vaccination induces broad and potent adaptive immune responses that recognize SARS-CoV-2 Delta and Omicron variants）.

6.2. The second-generation vaccines

Although many countries or regions have carried out large-scale vaccination, COVID-19 is still prevalent all over the world, which is closely related to the persistent mutation of COVID-19 and VOCs/VOIs. VOCs reduce the neutralizing ability of vaccine induced antibodies and enhance virus transmission. Studies have shown that most VOCs, especially the dominant Omicron variant, can evade the immune response and reduce the effectiveness of the currently approved mRNA vaccines, which pose a huge risks to human health around the world [226], [227], [228]. Vaccines against volatile organic compounds have also been studied to find the second-generation vaccine with stronger protection and longer protection time for COVID-19 variants, as well as vaccines that can be vaccinated without injection, so as to help establish protective immunity against COVID-19 and its new variants. So far, the second-generation vaccines mainly include: 1. Recombinant protein vaccine for new variants; 2. mRNA vaccine against variant strains; 3. Multi-antigen modified vaccine based on viral vector.

Recently, the RBD-conjugated nanoparticle vaccines targeting ancestral COVID-19, Delta, Beta and Gamma variants demonstrated the effect on eliciting rich neutralizing antibodies and protecting hACE2 mice from the infection of the ancestral COVID-19 and the VOCs. In addition, it was also found that the re-booster of the trivalent vaccine elicited more extensive cross-protective neutralizing antibodies, which target almost all COVID-19 variants (including the Omicron variant) in rhesus macaques [229].

Compared to the mRNA vaccine candidate CVnCov, CV2CoV, a second-generation mRNA vaccine, has optimized non-coding regions and enhances antigen expression, which elicited comparable immunogenicity with BNT162b2 in macaques. CV2CoV can induce higher titers of neutralizing antibodies, memory B cell and T cell responses against COVID-19 variants (including Delta variant) in cynomolgus macaques [230]. CV2CoV elicited as well higher levels of neutralising antibodies and induced cross-neutralisation of the Alpha, Beta and the variant (B1.1.298) in rats [231].

The development of virus vector based vaccines provides promising strategies for second-generation COVID-19 vaccines, especially the vaccines with stable antigen expression or multi-antigen modification. A noval vaccine AVX/COVID-12-HEXAPRO（Patria）, which can be administed intranasally or intramuscularly, contains a live Newcastle disease virus vector and expresses the spike protein stabilized. The application of AVX/COVID-12-HEXAPRO(Patria) elicited potent serum neutralizing antibody response in the pig models. In addition, substantial reactivity to B.1.1.7, B.1.351 and P.1 spike mutants was detected [232]. COH04S1 is a synthetic multi-antigen modified COVID-19 vaccine based on Ankara virus vector, which can simultaneously express spike and nucleocapsid antigens. Intramuscular or intranasal inoculation of COH04S1 can induce effective Th1 biased antigen specific humoral immunity and cross neutralization antibodies in Syrian hamsters, and can prevent lower respiratory tract infection and lung injury caused by intranasal attack of COVID-19. The similar protection can be detected in non-human primates prime or booster vaccination of COH04S1 [233].

7. Prospects and challenge

7.1. Nanotechnology in COVID-19 vaccine

The application of nanotechnology in COVID-19 vaccine has been expected as a future trend of the vaccine development. Nanoparticles, as antigen presentation tools, show the advantages as follows: 1. Nanoparticles can be mixed with antigens as adjuvants. 2. They are capable of encapsulating antigen components and maintaining antigen stability. 3. They serve as an antigen display platform to add a variety of antigen components [234]. Lipid nanoparticles (LNPs) present a novel colloidal drug delivery system and differ from liposomes as micellar structures in the core that can be modified by formulation and synthesis parameters [235]. Coupling of nanoparticles maintains the stability of antigen, delays the degradation of antigen, and significantly enhances the protective immunity of protein vaccine. The nanoparticle vaccine can be designed to 20–200 nm, close to the effective recognition size of antigen presenting cells and other immune cells [236]. Homotype SARS-CoV-2 nanoparticles can modulate the immune response to produce IgG, which are bound to the zoonotic RBD and neutralizes heterologous coronaviruses after booster immunization [237]. Ferritin refers to a type of non-viral self-assembled protein, which can be used to construct ferritin nanoparticles (SpFN). At present, the COVID-19 vaccine developed by the Walter Reed Army Research Institute (WRAIR) can cause effective immune response while providing extensive protection [238]. A nanoparticle vaccine covalently conjugates the self-assembled 24-mer ferritin to the receptor binding domain (RBD) and/or heptad repeat (HR) subunits of the spike (S) protein developed in China [239].

For the future

Since the outbreak of COVID-19, it has been raging around the world for more than three years, thus turning our initial panic into today's calm. In general, since the pathogenesis of COVID-19 was clarified, the global pandemic of COVID-19 has been curbed, and COVID-19 vaccine has also been continuously developed and promoted. In the post pandemic era, recurrent sporadic outbreaks of COVID-19 and viral mutations under immune pressure are still risk factors for population health. Therefore, efficient and safe COVID-19 vaccines (such as bionic nanoparticle vaccines and pan SARS CoV vaccines) should be developed. In addition, a large sample of mixed vaccine can be tested, and the monitoring of special populations can be strengthened, so that the vaccine can benefit more people after the COVID-19 pandemic.

Funding

None.

Author contributions

There are 5 first authors in this manuscript and they have equally contributed to this project. Dr. GGM, ZQC,HSC,WHW and XC were responsible for collecting key information and drafting the manuscript. Dr. HYL,LYZ, and HFZ also collected some information. Furthermore, we have 5 corresponding authors in this manuscript. Dr. HZC,XLL,JFS, YL and TTF have contributed to designed the study and critical revised the manuscript.All authors read and approved the final manuscript.

Consent for publication

Not applicable.

Acknowledgements

None.

Declaration

This manuscript did not receive any funding and there were no conflicts of interest between all authors.

Conflict of interest statement

No conflict of interest exits in the submission of this manuscript, and manuscript is approved by all authors for publication. I would like to declare on behalf of my co-authors that the work described has not been published previously, and not under consideration for publication elsewhere, in whole or in part. All the authors listed have approved the manuscript that is enclosed.