Options for the development of a bivalent vaccine against SARS-CoV-2 and influenza

KEYWORDS:

• Influenza
• SARS-CoV-2
• COVID-19
• coinfection
• bivalent vaccine
• viral vectored vaccine

Unlike most vaccine-controlled viral infections, which require only a single vaccination or a limited number of booster doses to maintain lifelong protective immunity, influenza virus has long been the only pathogen against which annual vaccination campaigns are recommended because the vaccine-induced immunity wanes over time, and the virus mutates very quickly, thereby evading the action of neutralizing antibodies. Since the emergence of the new severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) in 2019, the epidemiological situation has changed dramatically because the new virus has become established in the human population and is subject to constant antigenic drift. As a result, breakthrough infections occur in individuals fully vaccinated with licensed vaccines against the coronavirus disease 2019 (COVID-19) [1]. Moreover, reinfection with antigenically evolved SARS-CoV-2 within 12 months of primary infection is not uncommon [2]. Most probably, the two viruses will become co-circulating worldwide, and regular vaccination against both pathogens is likely to become a necessary measure to reduce the potential burden caused by these infections. Although a clear seasonality for SARS-CoV-2, unlike influenza viruses, has not yet been identified, it is reasonable to assume that vaccination campaigns against both infections could be combined to reduce the overall cost of interventions.

Several randomized controlled trials of co-administration of licensed and/or experimental vaccines against both pathogens have been initiated to explore the possibility of simultaneous protection against SARS-CoV-2 and influenza viruses, and the available data indicate that the reactogenicity profile of such combinations was comparable to that of individual vaccines administered separately (reviewed in [3]). Although the immune responses induced by such combinatory vaccines against both viruses are generally non-inferior to that following vaccination with each vaccine individually, public acceptance of co-administering two different vaccines is expected to be lower than the willingness to receive either vaccine alone [3]. It may also be assumed that a single ‘universal’ vaccine combining influenza virus and SARS-CoV-2 antigens could potentially increase total vaccination coverage against both pathogens, as well as reduce the overall cost of vaccine production.

Such bivalent vaccines against COVID-19 and influenza can be developed using different vaccine platforms, such as nucleic acids (DNA or RNA), virus-like particles (VLPs), or viral vectors that can simultaneously deliver all necessary viral antigens to the target cells. For example, the mRNA-based approach is being studied by Moderna, developer of the licensed mRNA-1273/Spikevax® COVID-19 vaccine; however, preclinical results have not yet been disclosed [3]. A group of researchers from Beijing Institute of Microbiology and Epidemiology (China) developed a combined SARS-CoV-2/influenza vaccine using a lipid nanoparticle (LNP)-encapsulated mRNA vaccine platform. This construct encoded the hemagglutinin (HA) antigen of the H1N1 influenza virus and the receptor-binding domain (RBD) of the SARS-CoV-2 S protein, and immunization of mice with two doses of the vaccine elicited robust humoral and cell-mediated immunity to both pathogens, thereby protecting the animals against influenza and SARS-CoV-2 coinfection [4]. Given that the mRNA COVID-19 vaccines have been successful in inducing protective immunity in humans while also being relatively safe, exploiting this technology to create a bivalent vaccine against SARS-CoV-2 and influenza viruses looks like a promising area of research.

A recent study reported the development of a hybrid vaccine by incorporating the RBD protein fused with granulocyte-macrophage colony-stimulating factor (GM-CSF) into influenza VLPs containing HA and M1 matrix proteins of H1N1 influenza virus [5]. The GM-CSF cytokine, a biological adjuvant for the FDA-approved prostate cancer vaccine, Provenge, enhanced the immunogenicity of the combinatory VLP vaccine, so that the vaccine prototype protected mice against influenza virus and a mouse-adapted SARS-CoV-2.

Other studies explored viral vector platforms to develop bivalent vaccines against COVID-19 and influenza, since some viruses can deliver foreign antigens to the target cells and therefore lead to induction of both humoral and T-cell immunity to the pathogens of interest. One example is a chimpanzee adenovirus serotype 68 vector (AdC68) which encodes the coronavirus RBD fused to the conserved HA stalk domain of influenza virus [6]. Another study discloses a recombinant vesicular stomatitis virus (VSV) vector-based vaccine against two infections. In this case, SARS-CoV-2 RBD protein was expressed along with four copies of influenza M2 ectodomain, a highly conserved influenza virus antigen [7]. Both AdC68-based and the VSV-vectored vaccines induced robust humoral and cellular responses to SARS-CoV-2 and influenza virus antigenic determinants, indicating the feasibility of this approach for designing a hybrid vaccine against both pathogens. It is worth noting that these constructs aim to protect against a wide range of influenza A viruses, but the protective potential of antibodies targeting conserved influenza virus epitopes has yet to be demonstrated in clinical practice.

Among a variety of viral vectors that can be used to develop a bivalent vaccine, special attention should be paid to influenza viruses because the vector itself is already capable of providing protection against one of the target pathogens. The establishment of effective reverse genetic systems for influenza A virus several decades ago prompted the development of influenza virus vector-based vaccines that induced protective immunity against various pathogens of interest (reviewed in [8]). The idea of using influenza virus as a vector for designing COVID-19 vaccines has been exploited by several groups of researchers. In some developments, the influenza virus genome was manipulated in such a way that one of the genes was replaced with a genetic construct encoding the coronavirus antigen, rendering the influenza virus incapable of replication. This increases the safety profile of the vaccine, but requires specific cell lines that constitutively express the missing viral protein. One example is the insertion of SARS-CoV-2 RBD fragment in place of the influenza virus HA open reading frame, with subsequent growth of the chimeric virus in MDCK cells stably expressing HA [9]. This bivalent vaccine induced influenza- and RBD-specific humoral and cellular responses in mice, and further studies will clarify whether this prototype is safe and provides protection against both infections. A similar COVID-19 vaccine based on M2-deficient single replication (M2SR) influenza virus vector is under development by the University of Wisconsin, Madison, and the vaccine companies FluGen and Bharat Biotech. Although details of the vaccine design have not yet been disclosed, promising results from recent clinical trials of the M2SR vaccine in healthy adults make this virus a promising platform for vector vaccine design [10].

A slightly different design of influenza virus vector-based vaccines against COVID-19 assumes that the virus retains the ability to replicate after replacing one gene with the gene of interest, most commonly the SARS-CoV-2 RBD fragment. It can be speculated that the active viral replication in vaccinated individual may induce more robust immune responses, compared to viruses with a single replication cycle, although other molecular mechanisms can be involved in the immunogenicity of engineered influenza viruses. In one example, the neuraminidase (NA) coding sequence of influenza virus was replaced by a membrane-anchored form of RBD, while the HA gene was mutated to reduce its affinity for the sialic acid receptor [11]. When administered intranasally to mice, this chimeric vaccine induced neutralizing antibodies to both viruses, although the protective potential of this prototype against SARS-CoV-2 and influenza has not yet been studied in appropriate animal models. A more advanced bivalent vaccine was developed by the Hong Kong University, and in this case, the gene encoding SARS-CoV-2 RBD fragment was inserted instead of the influenza virus NS1 gene. The vaccine has already been fully tested in animal models [12] and passed phase 1–2 clinical trials in adults [13].

It should be noted that the attenuated phenotype of NA- and NS1-modified variants of the influenza virus vector is controlled by a single gene, increasing the risk of reversion to the virulent phenotype in case of reassortment with a wild-type influenza virus. In contrast, attenuated phenotype of cold-adapted influenza viruses, which are the basis of licensed live attenuated influenza vaccines (LAIVs), is controlled by multiple loci, and no reversion to the wild-type phenotype has been reported during decades of the LAIVs’ clinical use [14]. Particular advantages of LAIVs include ease of intranasal administration, established safety profile, the ability to induce mucosal and T-cell responses, and existing production capacities, which make the LAIV platform an attractive alternative to other influenza vector platforms for designing a bivalent vaccine against COVID-19 and influenza [14]. Recently, a prototype T cell-based COVID-19 vaccine was developed by inserting polyepitope cassettes of SARS-CoV-2 into NA or NS1 open reading frames of the LAIV virus. The selected LAIV/SARS-CoV-2 vaccine prototype replicated well in conventional in vitro systems and was attenuated in animal models. Furthermore, this bivalent vaccine proved to be immunogenic and protected Syrian hamsters against both infections, prompting its further preclinical and clinical development [15].

The major limitation of the influenza virus vector-based bivalent vaccines is that they only protect against one subtype of influenza A virus. This would be relevant in the event of influenza pandemic when the bivalent vaccine is prepared using a new pandemic influenza strain; however, current seasonal influenza vaccines contain three or four different influenza viruses, including two type A viruses and one or two type B viruses. Therefore, a seasonal bivalent vaccine effective against both infections must include all WHO-recommended influenza viruses, where one or both influenza A strains are engineered to express SARS-CoV-2 immunogenic fragments. Since the influenza vaccine strains are updated almost annually, the SARS-CoV-2 antigens can also be adjusted to match the dominant strain in circulation.

In summary, many different vaccine platforms are being evaluated for designing bivalent vaccines against SARS-CoV-2 and influenza viruses, but more research is needed to prove the feasibility of these approaches and to find the optimal mode of vaccinating humans, given that they have preexisting immunity to both infections.

Declaration of interest

The authors have no relevant affiliations or financial involvement with any organization or entity with a financial interest in or financial conflict with the subject matter or material discussed in the manuscript. This includes employment, consultancies, honoraria, stock ownership or options, expert testimony, grants or patents received or mending, or royalties.

Reviewer disclosures

Peer reviewers on this manuscript have no relevant financial or other relationships to disclose.

Additional information

Funding

This work was supported by the Russian Science Foundation under Grant [21-75-30003].