Coronavirus vaccine

Abstract

The present disclosure relates to the field of RNA for preventing or treating coronavirus infections. In particular, the present disclosure relates to methods and agents for vaccination against coronavirus infections and for inducing effective coronavirus antigen-specific immune responses, such as antibody and / or T cell responses. Specifically, in one embodiment, the present disclosure relates to a method for inducing an immune response against coronavirus S proteins, particularly the S protein of SARS-CoV-2, in a subject, comprising administering to the subject RNA encoding a peptide or protein comprising an epitope of SARS-CoV-2 spike protein (S protein), i.e., a vaccine RNA encoding a vaccine antigen.

Description

[Technical Field]

[0001] The present disclosure relates to the field of RNA for preventing or treating coronavirus infections. In particular, the present disclosure relates to methods and agents for vaccination against coronavirus infections and for inducing effective coronavirus antigen-specific immune responses, such as antibody and / or T cell responses. These methods and agents are particularly useful for preventing or treating coronavirus infections. Administration of the RNA disclosed herein to a subject can protect the subject from coronavirus infection. Specifically, in one embodiment, the present disclosure relates to a method for inducing an immune response against coronavirus S proteins, particularly the S protein of SARS-CoV-2, in the subject, comprising administering to the subject RNA encoding a peptide or protein comprising an epitope of the SARS-CoV-2 spike protein (S protein), i.e., a vaccine RNA encoding a vaccine antigen. Administering the RNA encoding the vaccine antigen to the subject can provide the vaccine antigen (after expression of the RNA by appropriate target cells) for inducing an immune response against the vaccine antigen (and disease-associated antigens) in the subject.

[0002] Coronaviruses are single-stranded, positive-stranded RNA ((+)ssRNA) enveloped viruses that encode four structural proteins: the spike protein (S), the envelope protein (E), the membrane protein (M), and the nucleocapsid protein (N). The spike protein (S protein) is responsible for receptor recognition, cell binding, infection via the endosomal pathway, and genome release driven by fusion of the viral membrane with the endosomal membrane. Although sequences differ among different family members, conserved regions and motifs exist within the S protein, dividing it into two subdomains: S1 and S2. The S2 domain, which contains the transmembrane domain, is responsible for membrane fusion, while the S1 domain recognizes virus-specific receptors and binds to target host cells. Receptor-binding domains (RBDs) have been identified in several coronavirus isolates, and the general structure of the S protein has been defined (Figure 1).

[0003] In December 2019, an outbreak of pneumonia of unknown cause occurred in Wuhan, China, and a novel coronavirus (Severe Acute Respiratory Syndrome Coronavirus 2, SARS-CoV-2) was identified as the underlying cause. The genetic sequence of SARS-CoV-2 was made available to the WHO and the public (MN908947.3), and the virus was classified as a member of the Betacoronavirus subfamily. Sequence analysis revealed a phylogenetic tree that revealed a closer relationship to isolates of Severe Acute Respiratory Syndrome (SARS) virus than to another coronavirus that infects humans, namely, Middle East Respiratory Syndrome (MERS) virus.

[0004] SARS-CoV-2 infection and the resulting disease COVID-19 are spreading globally, affecting an increasing number of countries. On March 11, 2020, the WHO characterized the COVID-19 outbreak as a pandemic. As of December 1, 2020, there have been more than 63 million confirmed cases of COVID-19 worldwide, with over 1.4 million deaths, affecting 191 countries / territories. The ongoing pandemic remains a significant challenge to global public health and economic stability.

[0005] Due to the lack of pre-existing immunity to SARS-CoV-2, all individuals are at risk of infection. Following infection, some, but not all, individuals develop protective immunity in terms of neutralizing antibody responses and cell-mediated immunity. However, the extent and duration of this protection are currently unknown. According to the WHO, 80% of infected individuals recover without needing hospital care, 15% develop more severe disease, and 5% require intensive care. Increasing age and underlying health conditions are considered risk factors for developing severe disease.

[0006] Symptoms of COVID-19 typically include cough and fever, and chest radiographs show ground-glass opacities or patchy shadows. However, many patients present without fever or radiographic changes, and the infection may be asymptomatic, which is relevant for transmission control. In symptomatic subjects, disease progression can lead to acute respiratory distress syndrome requiring mechanical ventilation and subsequent multiple organ failure and death. Common symptoms in hospitalized patients (ordered from most to least frequent) include fever, dry cough, shortness of breath, fatigue, muscle pain, nausea / vomiting or diarrhea, headache, weakness, and nosebleeds. In approximately 3% of individuals with COVID-19, anosmia (loss of smell) or anageusia (loss of taste) may be the only presenting symptom.

[0007] Although people of all ages can present with the disease, the case fatality rate (CFR) is particularly elevated in those over 60 years of age. Comorbidities are also associated with increased CFR, including cardiovascular disease, diabetes, hypertension, and chronic respiratory disease. Healthcare workers are overrepresented among COVID-19 patients due to occupational exposure to infected patients.

[0008] In most cases, molecular tests are used to detect SARS-CoV-2 and confirm infection. Reverse transcription-polymerase chain reaction (RT-PCR) testing, which targets SARS-CoV-2 viral RNA, is the gold standard in vitro method for diagnosing suspected cases of COVID-19. Samples to be tested are collected via nasal and / or throat swabs.

[0009] Among other things, the present disclosure provides insight into the immune responses elicited by exposure (e.g., by vaccination and / or infection) to different SARS-CoV-2 variants or immunogenic polypeptides (e.g., S proteins), or immunogenic fragments thereof. For example, in some embodiments, administering RNA encoding the S protein of the BA.2 and / or BA.4 / 5 omicron SARS-CoV-2 variants, or an immunogenic fragment thereof, can result in an improved immune response, including, for example, improved neutralization of the omicron BA.4 and / or omicron BA.5 SARS-CoV-2 variants and / or broader cross-neutralization of variants of concern (e.g., omicron variants) (e.g., increased neutralization titers against more variants of concern (e.g., omicron variants)). In some embodiments, the present disclosure provides insights that a bivalent coronavirus vaccine (e.g., a bivalent BA.4 / 5 vaccine comprising a first RNA encoding a SARS-CoV-2 S protein or immunogenic fragment thereof of the Wuhan strain and a second RNA encoding a SARS-CoV-2 S protein or immunogenic fragment thereof comprising one or more mutations characteristic of the BA.4 / 5 omicron variant) can provide broader cross-neutralization against the SARS-CoV-2 Wuhan strain and certain variants thereof (e.g., in some embodiments, variants that are prevalent and / or spreading rapidly in a relevant jurisdiction, e.g., certain omicron variants) in certain subjects compared to a monovalent coronavirus vaccine (e.g., a vaccine comprising an RNA encoding a SARS-CoV-2 S protein of a coronavirus strain or variant thereof). In some embodiments, such broader cross-neutralization can be observed in unvaccinated subjects. In some embodiments, such broader cross-neutralization can be observed in subjects without a coronavirus infection (e.g., a SARS-CoV-2 infection).In some embodiments, such broader cross-neutralization may be observed in subjects who previously received a SARS-CoV-2 vaccine (e.g., in some embodiments, an RNA vaccine encoding the SARS-CoV-2 S protein, e.g., in some embodiments, the Wuhan strain). In some embodiments, such broader cross-neutralization may be observed in young pediatric subjects (e.g., subjects aged 6 months to less than 2 years, and / or 2 years to less than 5 years).

[0010] In some embodiments, the present disclosure provides the insight that exposure to at least two certain SARS-CoV-2 variants or immunogenic polypeptides (e.g., S proteins), or immunogenic fragments thereof, can result in a synergistically improved immune response (e.g., higher neutralization titers, broader cross-neutralization, and / or an immune response that is less susceptible to immune escape) compared to exposure to other combinations of single SARS-CoV-2 strains and / or SARS-CoV-2 variants. In some embodiments, the present disclosure provides the insight that exposure to the S protein or immunogenic fragment thereof from the Wuhan strain (e.g., by vaccination and / or infection) and exposure to the S protein or immunogenic fragment thereof of the Omicron BA.1 variant (e.g., by vaccination and / or infection) can result in a synergistically improved immune response (e.g., higher neutralization titers, broader cross-neutralization, and / or an immune response that is less susceptible to immune escape) compared to other combinations of single SARS-CoV-2 strains and / or SARS-CoV-2 variants. In some embodiments, the present disclosure provides the insight that exposure to the S protein or immunogenic fragment thereof from the Wuhan strain (e.g., by vaccination and / or infection) and exposure to the S protein or immunogenic fragment thereof of the Omicron BA.4 or BA.5 variant (e.g., by vaccination and / or infection) can result in a synergistic improvement in the immune response (e.g., higher neutralization titers, broader cross-neutralization, and / or an immune response that is less susceptible to immune escape) compared to exposure to other combinations of one SARS-CoV-2 strain and / or SARS-CoV-2 variant.In some embodiments, the present disclosure provides the insight that (i) exposure (e.g., by vaccination and / or infection) to an S protein from a strain / variant selected from the Wuhan strain, alpha variant, beta variant, delta variant, Omicron BA.1, and a sublineage derived from any of the foregoing strains / variants, or an immunogenic fragment thereof, in combination with (ii) exposure (e.g., by vaccination and / or infection) to an S protein from a strain / variant selected from the group consisting of Omicron BA.2, Omicron BA.4, Omicron BA.5, and a sublineage derived from any of the foregoing strains / variants, can result in a synergistic improvement in the immune response (e.g., higher neutralization titers, broader cross-neutralization, and / or an immune response that is less susceptible to immune escape) compared to exposure to other combinations of one SARS-CoV-2 strain and / or SARS-CoV-2 variant.

[0011] The present disclosure also provides important insights into how immune responses develop in subjects following exposure (e.g., vaccination and / or infection) to multiple different SARS-CoV-2 strains. Among other things, disclosed herein is the finding that different combinations of SARS-CoV-2 variants induce different immune responses. Specifically, the present disclosure provides the insight that exposure to certain combinations of SARS-CoV-2 variants can induce improved immune responses (e.g., higher neutralizing titers, broader cross-neutralization, and / or immune responses that are less susceptible to immune escape). In some embodiments, improved immune responses can be produced when a subject is delivered two or more antigens (e.g., as polypeptides or RNA encoding such polypeptides) that each share few epitopes. In some embodiments, an improved immune response may be produced when a subject is delivered (e.g., as a polypeptide or RNA encoding such a polypeptide) a combination of SARS-CoV-2 S proteins that share 50% or less (e.g., 40% or less, 30% or less, 20% or more) epitopes (e.g., including amino acid mutations) that can be bound by neutralizing antibodies. In some embodiments, an improved immune response may be produced by delivering (a) a SARS-CoV-2 S protein or immunogenic fragment thereof from the Wuhan strain of SARS-CoV-2, an alpha variant, a beta variant, or a delta variant, and (b) an S protein or immunogenic fragment thereof from a SARS-CoV-2 omicron variant, as a polypeptide or RNA encoding such a polypeptide. In some embodiments, an improved immune response may be produced by delivering, as a polypeptide or RNA encoding such a polypeptide, (a) the SARS-CoV-2 S protein or an immunogenic fragment thereof from the Wuhan strain of SARS-CoV-2, an alpha variant, a beta variant, or a delta variant, and (b) the S protein or an immunogenic fragment thereof of a SARS-CoV-2 omicron variant that is not the BA.1 omicron variant.In some embodiments, an improved immune response may be produced by delivering, as a polypeptide or RNA encoding such polypeptide, (a) the S protein or immunogenic fragment thereof from the Wuhan strain, an alpha variant, a beta variant, a delta SARS-CoV-2 variant, or a BA.1 omicron variant, and (b) the S protein or immunogenic fragment thereof of a SARS-CoV-2 omicron variant that is not a BA.1 omicron variant. In some embodiments, an improved immune response may be produced by delivering, as a polypeptide or RNA encoding such polypeptide, (a) the SARS-CoV-2 S protein or immunogenic fragment thereof from the Wuhan strain, an alpha variant, a beta variant, or a delta variant, and (b) the S protein or immunogenic fragment thereof of a BA.2 or BA.4 or BA.5 SARS-CoV-2 omicron variant.

[0012] In some embodiments, the present disclosure also provides insight that administration of multiple doses (e.g., at least two, at least three, at least four, or more doses) of a coronavirus vaccine described herein (e.g., a bivalent vaccine described herein, such as a bivalent BA.4 / 5 vaccine) can provide certain beneficial effect(s) on antibody affinity for one or more SARS-CoV-2 strains or variants thereof. In some embodiments, such beneficial effect(s) on antibody affinity may be observed for antibodies to certain omicron variants. By way of example only, in some embodiments, such beneficial effect(s) on antibody affinity may be observed for antibodies to certain omicron variants that share at least one common epitope with, for example, the Wuhan strain.

[0013] Also disclosed herein are compositions that can result in improved immune responses (e.g., immune responses with broader cross-neutralizing activity, more potent neutralization, and / or immune responses that are less susceptible to immune escape). In some embodiments, the compositions described herein comprise two or more antigens or nucleic acids (e.g., RNA) encoding such antigens that have few shared epitopes. In some embodiments, the compositions described herein deliver, as polypeptides or nucleic acids encoding such polypeptides, a combination of SARS-CoV-2 S proteins or immunogenic fragments thereof that share 50% or less (e.g., 40% or less, 30% or less, 20% or less, or more) of epitopes (e.g., including amino acid mutations) that can be bound by neutralizing antibodies. In some embodiments, the compositions described herein comprise (a) RNA encoding a SARS-CoV-2 S protein or an immunogenic fragment thereof from the Wuhan strain, an alpha variant, a beta variant, or a delta variant, and (b) RNA encoding an S protein from an omicron variant of SARS-CoV-2 (e.g., in some embodiments, an S protein from a BA.1, BA.2, or BA.4 / 5 omicron variant) or an immunogenic fragment thereof. In some embodiments, the compositions described herein comprise (a) RNA encoding a SARS-CoV-2 S protein or an immunogenic fragment thereof from the Wuhan strain, an alpha variant, a beta variant, or a delta variant, and (b) RNA encoding an S protein or an immunogenic fragment thereof of an omicron variant of SARS-CoV-2 that is not the BA.1 omicron variant. In some embodiments, the compositions described herein comprise (a) RNA encoding the SARS-CoV-2 S protein of the Wuhan strain, alpha variant, beta variant, or delta variant, or the BA.1 omicron variant, or an immunogenic fragment thereof, and (b) RNA encoding the S protein of an omicron variant that is not the BA.1 omicron variant, or an immunogenic fragment thereof.In some embodiments, the compositions described herein comprise (a) RNA encoding a SARS-CoV-2 S protein of the Wuhan strain of SARS-CoV-2, an alpha variant, a beta variant, or a delta variant, and (b) RNA encoding an S protein or an immunogenic fragment thereof of the BA.2 or BA.4 or BA.5 omicron variant of SARS-CoV-2. In some embodiments, the compositions described herein comprise RNA encoding an S protein or an immunogenic fragment thereof from the BA.2 omicron variant of SARS-CoV-2. In some embodiments, the compositions comprise RNA encoding an S protein or an immunogenic fragment thereof from the BA.4 or BA.5 omicron variant of SARS-CoV-2.

[0014] SARS-CoV-2 is an RNA virus with four structural proteins. One of these, the spike protein, is a surface protein that binds to angiotensin-converting enzyme 2 (ACE-2) present on host cells. Therefore, the spike protein is considered to be a relevant antigen for vaccine development.

[0015] BNT162b2 (SEQ ID NO: 20) is an mRNA vaccine for COVID-19 prevention that has demonstrated greater than 95% efficacy in preventing COVID-19. The vaccine is made with 5'-capped mRNA encoding the full-length SARS-CoV-2 spike glycoprotein (S) encapsulated in lipid nanoparticles (LNPs). The final product is presented as an injectable dispersion concentrate containing BNT162b2 as the active substance. Other ingredients are ALC-0315 (4-hydroxybutyl)azanediyl)bis(hexane-6,1-diyl)bis(2-hexyldecanoate), ALC-0159 (2-[(polyethylene glycol)-2000]-N,N-ditetradecylacetamide), 1,2-distearoyl-sn-glycero-3-phosphocholine (DSPC), cholesterol, potassium chloride, potassium dihydrogen phosphate, sodium chloride, disodium phosphate hydrate, sucrose, and water for injection.

[0016] In some embodiments, a different buffer may be used instead of PBS. In some embodiments, the buffer is formulated in a Tris buffer. In some embodiments, the formulation includes ALC-0315 (4-hydroxybutyl)azanediyl)bis(hexane-6,1-diyl)bis(2-hexyldecanoate), ALC-0159 (2-[(polyethylene glycol)-2000]-N,N-ditetradecylacetamide), DSPC (1,2-distearoyl-sn-glycero-3-phosphocholine), cholesterol, sucrose, trometamol (Tris), trometamol hydrochloride, and water.

[0017] In some embodiments, the concentration of RNA in the pharmaceutical RNA preparation is about 0.1 mg / ml. In some embodiments, about 30 μg of RNA is administered by administering about 200 μL of the RNA preparation. In some embodiments, the RNA in the pharmaceutical RNA preparation is diluted prior to administration (e.g., diluted to a concentration of about 0.05 mg / ml). In some embodiments, the administration volume is about 200 μl to about 300 μl. In some embodiments, the RNA in the pharmaceutical RNA preparation is formulated in about 10 mM Tris buffer and about 10% sucrose.

[0018] In some embodiments, the concentration of RNA in the pharmaceutical RNA preparation is about 0.1 mg / ml and is formulated in about 10 mM Tris buffer, about 10% sucrose, and a dose of about 10 μg, or the RNA is administered by diluting the pharmaceutical RNA preparation about 1:1 and administering about 200 μl of the diluted pharmaceutical RNA preparation. In some embodiments, the concentration of RNA in the pharmaceutical RNA preparation is about 0.1 mg / ml and is formulated in about 10 mM Tris buffer, about 10% sucrose, and a dose of about 10 μg of RNA, and is administered by diluting the pharmaceutical RNA preparation about 1:5.75 and administering about 200 μl of the diluted pharmaceutical RNA preparation.

[0019] The sequence of the SARS-CoV-2 S protein of the Wuhan strain disclosed herein was selected based on the sequences of "SARS-CoV-2 isolate Wuhan-Hu-1": GenBank: MN908947.3 (complete genome) and GenBank: QHD43416.1 (spike surface glycoprotein).

[0020] In some embodiments, the active substance consists of a single-stranded 5' cap codon-optimized mRNA that is translated into a spike antigen of SARS-CoV-2. In some embodiments, the encoded spike antigen protein sequence contains two proline mutations that stabilize an antigenically optimal pre-fusion confirmation (P2S). In some embodiments, the RNA does not contain any uridines; for example, modified N1-methylpseudouridine can be used in place of uridine in RNA synthesis. The mRNA disclosed herein can be translated into the SARS-CoV-2 S protein in a host cell. The S protein is then expressed on the cell surface, where it can induce an adaptive immune response. The S protein has been identified as a target for neutralizing antibodies against the virus and can be considered a relevant vaccine component.

[0021] BNT162b2 can be administered intramuscularly (IM) in two 30 μg doses of diluted vaccine solution given approximately 21 days apart (e.g., to adult unvaccinated subjects (i.e., subjects 12 years of age or older who have not previously received a SARS-CoV-2 vaccine)).

[0022] The recent emergence of novel circulating variants of SARS-CoV-2 raises serious concerns regarding the geographic and temporal effectiveness of vaccine interventions. One of the earliest variants to emerge and rapidly become dominant globally was D614G.

[0023] The alpha variant (B.1.1.7, VOC202012 / 01, also known as 501Y.V1 or GRY) was initially detected in the United Kingdom. The alpha variant has numerous mutations, including several in the S gene. It has been shown to be inherently more transmissible, with growth rates estimated to be 40–70% higher than other SARS-CoV-2 strains in multiple countries (Volz et al., 2021, Nature, https: / / doi.org / 10.1038 / s41586-021-03470-x; Washington et al., 2021, Cell https: / / doi.org / 10.1016 / j.cell.2021.03.052).

[0024] The beta variant (also known as B.1.351 or GH / 501Y.V2) was first detected in South Africa. The beta variant has several mutations in the S gene. Three of these mutations are at sites within the RBD associated with immune evasion: N501Y (shared with alpha), E484K, and K417N.

[0025] The gamma variant (also known as P.1 or GR / 501Y.V3), first detected in Brazil, has several mutations affecting the spike protein, including two mutations shared with the beta (N501Y and E484K) and a different mutation at position 417 (K417T).

[0026] The delta variant (also known as B.1.617.2 or G / 478K.V1) was first documented in India. The delta variant has several point mutations affecting the spike protein, including P681R (a mutation position shared with alpha and adjacent to the furin cleavage site) and L452R, which is within the RBD and has been linked to increased binding to ACE2 and resistance to neutralizing antibodies. There is also a deletion in the spike protein at positions 156 / 157.

[0027] These four VOCs have become prevalent worldwide and have become the predominant variants in the geographic regions in which they were first identified.

[0028] On November 24, 2021, the Omicron (B.1.1.529) variant was first reported to WHO from South Africa.

[0029] Since its first appearance in November 2021, SARS-CoV-2 Omicron and its sublineages have significantly impacted the epidemiological situation of the COVID-19 pandemic (WHO Technical Advisory Group on SARS-CoV-2 Virus Evolution (TAG-VE): Classification of Omicron (B.1.1.259): SARS-CoV-2 Variant of Concern (2021), WHO Headquarters (HQ), WHO Health Emergencies Programme, Enhancing Response to Omicron SARS-CoV-2 variant: Technical brief and priority actions for Member States (2022)). Significant modifications in the spike (S) glycoprotein of the first Omicron variant, BA.1, resulting in the loss of many neutralizing antibody epitopes (M. Hoffmann et al., "The Omicron variant is highly resistant against antibody-mediated neutralization: Implications for control of the COVID-19 pandemic", Cell 185, 447-456.e11 (2022)), enabled BA.1 to partially escape immunity based on the previously established SARS-CoV-2 wild-type strain (Wuhan-Hu-1) (V. Servellita, et al., "Neutralizing 30 immunity in vaccine breakthrough infections from the SARS-CoV-2 Omicron and Delta variants", Cell 185, 1539-1548.e5 (2022); Y. Cao et al., "Omicron escapes the majority of existing SARS-CoV-2 neutralizing antibodies", Nature 602,657-663(2022)).Therefore, breakthrough infection with Omicron is more common among vaccinated individuals than with previous variants of concern (VOCs). While Omicron BA.1 has been replaced by the BA.2 variant in many countries around the world, other variants, such as BA.1.1 and BA.3, have temporarily and / or locally gained traction but have not become globally dominant (S. Xia et al., "Origin, virological features, immune evasion, and intervention of SARS-CoV-2 Omicron sublineages. Signal Transduct. Target. Ther. 7, 241 (2022)"; H. Gruell et al., "SARS-CoV-2 Omicron sublineages exhibit distinct antibody escape patterns," Cell Host Microbe 7, 241 (2022)). Subsequently, in the United States, Omicron BA.2.12.1 replaced BA.2 as dominant, while in Europe, parts of Africa, and Asia / Pacific, BA.4 and BA.5 replaced BA.2 as dominant (H. Gruell et al., “SARS-CoV-2 Omicron sublineages exhibit distinct antibody escape patterns,” Cell Host Microbe 7, 241 (2022); European Centre for Disease Prevention and Control, Weekly COVID-19 country overview - Country overview report: Week 31 2022 (2022); J. Hadfield et al., “Nextstrain: Real-time tracking of pathogen evolution,” Bioinformatics 34, 4121-4123 (2018)).Currently, Omicron XBB.1.5 is dominant worldwide, including in the United States (Centers for Disease Control and Prevention. COVID Data Tracker. Atlanta, GA: US Department of Health and Human Services, CDC; 2023, January 22. https: / / covid.cdc.gov / coviddata-tracker (2022)). Omicron has acquired many modifications (amino acid exchanges, insertions, or deletions) in the S glycoprotein, some of which are shared among all Omicron VOCs and others that are specific to one or more Omicron sublineages. Antigenically, BA.2.12.1 shows high similarity to BA.2 but not to BA.1, whereas BA.4 and BA.5, along their lineage, diverge significantly from their ancestor BA.2 and further from BA.1 (AZMykytyn et al., “Antigenic cartography of SARS-CoV-2 reveals that Omicron BA.1 and BA.2 are antigenically distinct,” Sci. Immunol. 7, eabq4450 (2022)). Key differences between BA.1 and the remaining Omicron VOCs include Δ143-145, L212I, or ins214EPE in the S glycoprotein N-terminal domain and G446S or G496S in the receptor-binding domain (RBD). The amino acid changes T376A, D405N, and R408S in the RBD are, respectively, shared with BA.2 and its descendants but not found in BA.1. In addition, some modifications are specific to individual BA.2 progeny VOCs, such as L452Q in BA.2.12.1 or L452R and F486V in BA.4 and BA.5 (which encode the same S sequence). Most of these shared VOC-specific modifications have been shown to play important roles in immune escape from monoclonal antibodies and polyclonal sera against the wild-type S glycoprotein.In particular, BA.4 / BA.5-specific modifications are strongly involved in the immune evasion of these VOCs (P. Wang et al., "Antibody resistance of SARS-CoV-2 variants B.1.351 and B.1.1.7. Nature 593, 130-135 (2021); Q. Wang et al., "Antibody evasion by SARS-CoV-2 Omicron subvariants BA.2.12.1, BA.4, & BA.5. Nature 608, 5 603-608 (2022)). Summary of the Invention

[0030] The present disclosure generally encompasses immunotherapeutic treatment of a subject, including administration of RNA, i.e., vaccine RNA, encoding an amino acid sequence, i.e., a vaccine antigen, comprising the SARS-CoV-2 S protein, an immunogenic variant thereof, or an immunogenic fragment, i.e., an antigenic peptide or protein, of the SARS-CoV-2 S protein or its immunogenic variant. Thus, the vaccine antigen comprises an epitope of the SARS-CoV-2 S protein for inducing an immune response against the coronavirus S protein, particularly the SARS-CoV-2 S protein, in the subject. The RNA encoding the vaccine antigen is administered (following expression of the polynucleotide by appropriate target cells) to provide an antigen for induction, i.e., stimulation, priming, and / or expansion, of an immune response, e.g., antibodies and / or immune effector cells, targeting the target antigen (coronavirus S protein, particularly the SARS-CoV-2 S protein) or its progressive products. In one embodiment, the immune response induced in accordance with the present disclosure is a B cell-mediated immune response, i.e., an antibody-mediated immune response. Additionally or alternatively, in one embodiment, the immune response induced in accordance with the present disclosure is a T cell-mediated immune response. In one embodiment, the immune response is an anti-coronavirus, particularly an anti-SARS-CoV-2, immune response.

[0031] The vaccines described herein comprise, as an active ingredient, a single-stranded RNA that can be translated into a protein upon entry into the recipient's cells. In addition to a wild-type or codon-optimized sequence encoding an antigen sequence, the RNA may contain one or more structural elements (e.g., a 5' cap, a 5' UTR, a 3' UTR, a poly(A) tail, or a combination thereof) that are optimized for maximum effectiveness of the RNA with respect to stability and translation efficiency. In one embodiment, the RNA contains all of these elements. In one embodiment, the cap1 structure may be utilized as a specific capping structure at the 5' end of the RNA drug substance. In one embodiment, beta-S-ARCA(D1)(m2 7,2’-O GppSpG) or m2 7,3’-O Gppp(m1 2’-O ) ApG can be used as a specific capping structure at the 5' end of an RNA drug substance. The 5'-UTR sequence may optionally be the 5'-UTR sequence of human alpha-globin mRNA (e.g., SEQ ID NO: 12), which has an optimized "Kozak sequence" to enhance translation efficiency. The 3'-UTR sequence may be a combination of two sequence elements (FI elements) derived from the "amino-terminal enhancer of split" (AES) mRNA (designated F) and mitochondrially encoded 12S ribosomal RNA (designated I) (e.g., SEQ ID NO: 13), located between the coding sequence and the poly(A) tail, to ensure higher maximum protein levels and long-term mRNA persistence. These were identified through an ex vivo selection process for sequences that confer RNA stability and enhance total protein expression (see WO2017 / 060314, incorporated herein by reference). Alternatively, the 3'-UTR may be two repeated 3'-UTRs of human beta-globin mRNA.

[0032] Additionally or alternatively, in some embodiments, the RNA comprises a poly(A) tail comprising a length of at least 90 adenosine nucleotides (e.g., at least about 100 adenosine nucleotides, at least about 110 adenosine nucleotides, at least about 120 adenosine nucleotides, at least about 130 adenosine nucleotides, or more). In some embodiments, the poly(A) tail may comprise a length of about 90 to about 150 adenosine nucleotides (e.g., about 100 to about 150 adenosine nucleotides). In some embodiments, the poly(A) tail may comprise an interrupted poly(A) tail. For example, in some such embodiments, a poly(A) tail measuring about 90 to about 120 nucleotides (e.g., about 110 nucleotides in length) may be used (e.g., a poly(A) tail comprising SEQ ID NO: 14), consisting of a stretch of about 30 adenosine residues (e.g., about 28, about 29, about 30, about 31, or about 32 adenosine residues), followed by about 10 nucleotides (e.g., about 9, about 10, or about 11 random nucleotides) and a linker sequence of another about 70 adenosine nucleotides (e.g., about 65, about 66, about 67, about 68, about 69, about 70, about 71, about 72, about 73, about 74, or about 75 adenosine nucleotides). This poly(A) tail sequence is designed to improve RNA stability and translation efficiency.

[0033] Furthermore, in some embodiments, a nucleotide sequence encoding a secretory signal peptide (sec) can be fused to the antigen-coding region, preferably in such a way that the sec is translated as an N-terminal tag. In one embodiment, the sec corresponds to the secretory signal peptide of the SARS-CoV-2 S protein. In some embodiments, a sequence encoding a short linker peptide consisting mainly of the amino acids glycine (G) and serine (S), commonly used in fusion proteins, can be used as a GS / linker to join the secretory signal and the antigenic polypeptide.

[0034] The vaccine RNAs described herein can be complexed with proteins and / or lipids, preferably lipids, to form RNA particles for administration. When different RNA combinations are used, the RNAs can be complexed together or can be complexed separately with proteins and / or lipids to form RNA particles for administration.

[0035] In one aspect, the disclosure features a composition or medical preparation including RNA encoding a SARS-CoV-2 S polypeptide or immunogenic fragment thereof, wherein the SARS-CoV-2 S polypeptide or fragment includes: (a) an amino acid sequence having at least 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identity to SEQ ID NO:1 and including one of the following sets of amino acid substitutions relative to SEQ ID NO:1: (1) D985P, V987P, F817P, A892P, A899P, and A942P, (2) K986P, V987P, F817P, A892P, A899P, and A942P, (3) D985P, V987P. , F817P, A892P, A899P, A942P, D614G, R682G, R683S, and R685S, (4) K986P, V987P, F817P, A892P, A899P, A942P, D614G, R682G, R683S, and R685S, (5) D985P, K986P, V987P, F817P, A892P, A899P, A942P, D614G, R682G, R683S, and R685S, (6) D985P, V987P, F817P, A892P, A899P, A942P, R682 (7) K986P, V987P, F817P, A892P, A899P, A942P, R682G, R683S, and R685S; or (8) D985P, K986P, V987P, F817P, A892P, A899P, A942P, R682G, R683S, and R685S; (b) at least 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identical to SEQ ID NO: 69 and having at least 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identity to SEQ ID NO: 69. and an amino acid sequence containing one of the following sets of amino acid substitutions: (1) D982P, V984P, F814P, A889P, A896P, and A939P, (2) K983P, V984P, F814P, A889P, A896P, and A939P, (3) D982P, V984P, F814P, A889P, A896P, A939P, R679G, R680S, and R682S, (4) K983P, V984P, F814P, A889P, A896P, A939P, R679G, R680S, and R682S,or (5) D982P, K983P, V984P, F814P, A889P, A896P, A939P, R679G, R680S, and R682S, (c) an amino acid sequence having at least 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identity to SEQ ID NO:70 and including one of the following sets of amino acid substitutions relative to SEQ ID NO:70: (1) D982P, V984P, F814P, A889P, A896P, A939P, R679G, R680S, and R682S. 96P, and A939P, (2) K983P, V984P, F814P, A889P, A896P, and A939P, (3) D982P, V984P, F814P, A889P, A896P, A939P, R679G, R680S, and R682S, (4) K983P, V984P, F814P, A889P, A896P, A939P, R679G, R680S, and R682S, or (5) D982P, K983P, V984P, F814P, A889P, A896P, A or (d) an amino acid sequence having at least 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identity to SEQ ID NO: 104 or 105 and including one of the following sets of amino acid substitutions relative to SEQ ID NO: 104 or 105: (1) D980P, V982P, F812P, A887P, A894P, and A937P; (2) K981P, V982P, F812P, A887P, A894P, and A937P; P, F812P, A887P, A894P, and A937P, (3) D980P, V982P, F812P, A887P, A894P, A937P, R677G, R678S, and R680S, (4) K981P, V982P, F812P, A887P, A894P, A937P, R677G, R678S, and R680S, or (5) D980P, K981P, V982P, F812P, A887P, A894P, A937P, R677G, R678S, and R680S.

[0036] In some embodiments, the RNA comprises modified nucleosides in place of uridines. In some embodiments, the RNA comprises modified uridines in place of all uridines. In some embodiments, the RNA comprises N1-methyl-pseudouridine (m1ψ) in place of all uridines. In some embodiments, the RNA comprises a 5' cap. In some embodiments, the 5' cap is m2 7,3’-O Gppp(m1 2’-O ) ApG or containing it.

[0037] In some embodiments, the RNA comprises a 5'-UTR that is or comprises a modified human alpha-globin 5'-UTR. In some embodiments, the RNA comprises a 5'-UTR that comprises the nucleotide sequence of SEQ ID NO: 12, or a nucleotide sequence that is at least 99%, 98%, 97%, 96%, 95%, 90%, 85%, or 80% identical to the nucleotide sequence of SEQ ID NO: 12.

[0038] In some embodiments, the RNA comprises a 3'-UTR that is or includes a first sequence from a split amino-terminal enhancer (AES) messenger RNA and a second sequence from a mitochondrially encoded 12S ribosomal RNA. In some embodiments, the RNA comprises a 3'-UTR that includes the nucleotide sequence of SEQ ID NO: 13, or a nucleotide sequence that is at least 99%, 98%, 97%, 96%, 95%, 90%, 85%, or 80% identical to the nucleotide sequence of SEQ ID NO: 13.

[0039] In some embodiments, the RNA comprises a polyA sequence. In some embodiments, the polyA sequence comprises at least 100 nucleotides. In some embodiments, the polyA sequence comprises 30 adenine nucleotides followed by 70 adenine nucleotides, wherein the 30 adenine nucleotides and the 70 adenine nucleotides are separated by a linker sequence. In some embodiments, the polyA sequence comprises or consists of the nucleotide sequence of SEQ ID NO: 14, or a nucleotide sequence that is at least 99%, 98%, 97%, 96%, 95%, 90%, 85%, or 80% identical to the nucleotide sequence of SEQ ID NO: 14.

[0040] In some embodiments, the RNA is or will be formulated for intramuscular administration. In some embodiments, the RNA is or will be formulated as a particle. In some embodiments, the particle is a lipid nanoparticle (LNP) or lipoplex (LPX) particle. In some embodiments, the LNP comprises ((4-hydroxybutyl)azanediyl)bis(hexane-6,1-diyl)bis(2-hexyldecanoate), 2-[(polyethylene glycol)-2000]-N,N-ditetradecylacetamide, 1,2-distearoyl-sn-glycero-3-phosphocholine, and cholesterol. In some embodiments, lipoplex particles can be obtained by mixing RNA with liposomes.

[0041] In some embodiments, the RNA is mRNA or saRNA.

[0042] In some embodiments, the composition or medical preparation is a pharmaceutical composition. In some embodiments, the composition or medical preparation is a vaccine.

[0043] In some embodiments, the pharmaceutical composition further comprises one or more pharmaceutically acceptable carriers, diluents, and / or excipients.

[0044] In another aspect, the disclosure provides a composition or medical preparation comprising a first RNA encoding a first SARS-CoV-2 S polypeptide or immunogenic fragment thereof and a second RNA encoding a second SARS-CoV-2 S polypeptide or immunogenic fragment thereof, wherein (a) the first SARS-CoV-2 S polypeptide or fragment comprises an amino acid sequence having at least 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identity to SEQ ID NO: 1; and (b) the second SARS-CoV-2 The S polypeptide or fragment has at least 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identity to SEQ ID NO:1 and comprises an amino acid sequence containing one of the following sets of amino acid substitutions relative to SEQ ID NO:1: (1) D985P, V987P, F817P, A 892P, A899P, and A942P, (2) K986P, V987P, F817P, A892P, A899P, and A942P, (3) D985P, V987P, F817P, A892P, A899P, A942P, D614G, R682G, R683S, and R685S, (4) K986P, V987P, F817P, A89 2P, A899P, A942P, D614G, R682G, R683S, and R685S, (5) D985P, K986P, V987P, F817P, A892P, A899P, A942P, D614G, R682G, R683S, and R685S, (6) D985P, V987P, F817P, A892P, A899P, A942 P, R682G, R683S, and R685S, (7) K986P, V987P, F817P, A892P, A899P, A942P, R682G, R683S, and R685S, or (8) D985P, K986P, V987P, F817P, A892P, A899P, A942P, R682G, R683S, and R685S.

[0045] In one aspect, the disclosure features a composition or medical preparation including a first RNA encoding a first SARS-CoV-2 S polypeptide or immunogenic fragment thereof and a second RNA encoding a second SARS-CoV-2 S polypeptide or immunogenic fragment thereof, wherein (a) the first SARS-CoV-2 S polypeptide or fragment comprises an amino acid sequence having at least 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identity to SEQ ID NO: 1; and (b) the second SARS-CoV-2 The S polypeptide or fragment has at least 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identity to SEQ ID NO:69 and comprises an amino acid sequence containing one of the following sets of amino acid substitutions relative to SEQ ID NO:69: (1) D982P, V984P, F814P, A889P, A896P, and A939P; (2) K983P, V984P, F814P, A889P, A939P; 896P, and A939P, (3) D982P, V984P, F814P, A889P, A896P, A939P, R679G, R680S, and R682S, (4) K983P, V984P, F814P, A889P, A896P, A939P, R679G, R680S, and R682S, or (5) D982P, K983P, V984P, F814P, A889P, A896P, A939P, R679G, R680S, and R682S.

[0046] Another aspect of the disclosure features a composition or medical preparation including a first RNA encoding a first SARS-CoV-2 S polypeptide or immunogenic fragment thereof and a second RNA encoding a second SARS-CoV-2 S polypeptide or immunogenic fragment thereof, wherein (a) the first SARS-CoV-2 S polypeptide or fragment comprises an amino acid sequence having at least 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identity to SEQ ID NO:1, and (b) the second SARS-CoV-2 The S polypeptide or fragment has at least 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identity to SEQ ID NO:70 and comprises an amino acid sequence containing one of the following sets of amino acid substitutions relative to SEQ ID NO:70: (1) D982P, V984P, F814P, A889P, A896P, and A939P; (2) K983P, V984P, F814P, A889P, A939P; 896P, and A939P, (3) D982P, V984P, F814P, A889P, A896P, A939P, R679G, R680S, and R682S, (4) K983P, V984P, F814P, A889P, A896P, A939P, R679G, R680S, and R682S, or (5) D982P, K983P, V984P, F814P, A889P, A896P, A939P, R679G, R680S, and R682S.

[0047] Another aspect of the disclosure features a composition or medical preparation including a first RNA encoding a first SARS-CoV-2 S polypeptide or immunogenic fragment thereof and a second RNA encoding a second SARS-CoV-2 S polypeptide or immunogenic fragment thereof, wherein (a) the first SARS-CoV-2 S polypeptide or fragment comprises an amino acid sequence having at least 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identity to SEQ ID NO:1, and (b) the second SARS-CoV-2 The S polypeptide or fragment has at least 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identity to SEQ ID NO: 104 or 105 and comprises an amino acid sequence containing one of the following sets of amino acid substitutions relative to SEQ ID NO: 104 or 105: (1) D980P, V982P, F812P, A887P, A894P, and A937P; (2) K981P, V982P, F812P; A887P, A894P, and A937P, (3) D980P, V982P, F812P, A887P, A894P, A937P, R677G, R678S, and R680S, (4) K981P, V982P, F812P, A887P, A894P, A937P, R677G, R678S, and R680S, or (5) D980P, K981P, V982P, F812P, A887P, A894P, A937P, R677G, R678S, and R680S.

[0048] Another aspect of the present disclosure provides a composition or medical preparation comprising a first RNA encoding a first SARS-CoV-2 S polypeptide or immunogenic fragment thereof and a second RNA encoding a second SARS-CoV-2 S polypeptide or immunogenic fragment thereof, wherein (a) the first SARS-CoV-2 S polypeptide or fragment comprises an amino acid sequence having at least 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identity to SEQ ID NO:69; and (b) the second SARS-CoV-2 The S polypeptide or fragment has at least 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identity to SEQ ID NO:1 and comprises an amino acid sequence containing one of the following sets of amino acid substitutions relative to SEQ ID NO:1: (1) D985P, V987P, F817P, A 892P, A899P, and A942P, (2) K986P, V987P, F817P, A892P, A899P, and A942P, (3) D985P, V987P, F817P, A892P, A899P, A942P, D614G, R682G, R683S, and R685S, (4) K986P, V987P, F817P, A89 2P, A899P, A942P, D614G, R682G, R683S, and R685S, (5) D985P, K986P, V987P, F817P, A892P, A899P, A942P, D614G, R682G, R683S, and R685S, (6) D985P, V987P, F817P, A892P, A899P, A942 P, R682G, R683S, and R685S, (7) K986P, V987P, F817P, A892P, A899P, A942P, R682G, R683S, and R685S, or (8) D985P, K986P, V987P, F817P, A892P, A899P, A942P, R682G, R683S, and R685S.

[0049] Another aspect of the disclosure features a composition or medical preparation including a first RNA encoding a first SARS-CoV-2 S polypeptide or immunogenic fragment thereof and a second RNA encoding a second SARS-CoV-2 S polypeptide or immunogenic fragment thereof, wherein (a) the first SARS-CoV-2 S polypeptide or fragment comprises an amino acid sequence having at least 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identity to SEQ ID NO:69; and (b) the second SARS-CoV-2 The S polypeptide or fragment has at least 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identity to SEQ ID NO:69 and comprises an amino acid sequence containing one of the following sets of amino acid substitutions relative to SEQ ID NO:69: (1) D982P, V984P, F814P, A889P, A896P, and A939P; (2) K983P, V984P, F814P, A889P, A939P; 896P, and A939P, (3) D982P, V984P, F814P, A889P, A896P, A939P, R679G, R680S, and R682S, (4) K983P, V984P, F814P, A889P, A896P, A939P, R679G, R680S, and R682S, or (5) D982P, K983P, V984P, F814P, A889P, A896P, A939P, R679G, R680S, and R682S.

[0050] Another aspect of the disclosure features a composition or medical preparation including a first RNA encoding a first SARS-CoV-2 S polypeptide or immunogenic fragment thereof and a second RNA encoding a second SARS-CoV-2 S polypeptide or immunogenic fragment thereof, wherein (a) the first SARS-CoV-2 S polypeptide or fragment comprises an amino acid sequence having at least 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identity to SEQ ID NO:69; and (b) the second SARS-CoV-2 The S polypeptide or fragment has at least 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identity to SEQ ID NO:70 and comprises an amino acid sequence containing one of the following sets of amino acid substitutions relative to SEQ ID NO:70: (1) D982P, V984P, F814P, A889P, A896P, and A939P; (2) K983P, V984P, F814P, A889P, A939P; 896P, and A939P, (3) D982P, V984P, F814P, A889P, A896P, A939P, R679G, R680S, and R682S, (4) K983P, V984P, F814P, A889P, A896P, A939P, R679G, R680S, and R682S, or (5) D982P, K983P, V984P, F814P, A889P, A896P, A939P, R679G, R680S, and R682S.

[0051] Another aspect of the disclosure features a composition or medical preparation including a first RNA encoding a first SARS-CoV-2 S polypeptide or immunogenic fragment thereof and a second RNA encoding a second SARS-CoV-2 S polypeptide or immunogenic fragment thereof, wherein (a) the first SARS-CoV-2 S polypeptide or fragment comprises an amino acid sequence having at least 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identity to SEQ ID NO:69; and (b) the second SARS-CoV-2 The S polypeptide or fragment has at least 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identity to SEQ ID NO: 104 or 105 and comprises an amino acid sequence containing one of the following sets of amino acid substitutions relative to SEQ ID NO: 104 or 105: (1) D980P, V982P, F812P, A887P, A894P, and A937P; (2) K981P, V982P, F812P; A887P, A894P, and A937P, (3) D980P, V982P, F812P, A887P, A894P, A937P, R677G, R678S, and R680S, (4) K981P, V982P, F812P, A887P, A894P, A937P, R677G, R678S, and R680S, or (5) D980P, K981P, V982P, F812P, A887P, A894P, A937P, R677G, R678S, and R680S.

[0052] Another aspect of the disclosure features a composition or medical preparation including a first RNA encoding a first SARS-CoV-2 S polypeptide or immunogenic fragment thereof and a second RNA encoding a second SARS-CoV-2 S polypeptide or immunogenic fragment thereof, wherein (a) the first SARS-CoV-2 S polypeptide or fragment comprises an amino acid sequence having at least 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identity to SEQ ID NO: 70; and (b) the second SARS-CoV-2 The S polypeptide or fragment has at least 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identity to SEQ ID NO:1 and comprises an amino acid sequence containing one of the following sets of amino acid substitutions relative to SEQ ID NO:1: (1) D985P, V987P, F817P, A 892P, A899P, and A942P, (2) K986P, V987P, F817P, A892P, A899P, and A942P, (3) D985P, V987P, F817P, A892P, A899P, A942P, D614G, R682G, R683S, and R685S, (4) K986P, V987P, F817P, A89 2P, A899P, A942P, D614G, R682G, R683S, and R685S, (5) D985P, K986P, V987P, F817P, A892P, A899P, A942P, D614G, R682G, R683S, and R685S, (6) D985P, V987P, F817P, A892P, A899P, A942 P, R682G, R683S, and R685S, (7) K986P, V987P, F817P, A892P, A899P, A942P, R682G, R683S, and R685S, or (8) D985P, K986P, V987P, F817P, A892P, A899P, A942P, R682G, R683S, and R685S.

[0053] Another aspect of the disclosure features a composition or medical preparation including a first RNA encoding a first SARS-CoV-2 S polypeptide or immunogenic fragment thereof and a second RNA encoding a second SARS-CoV-2 S polypeptide or immunogenic fragment thereof, wherein (a) the first SARS-CoV-2 S polypeptide or fragment comprises an amino acid sequence having at least 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identity to SEQ ID NO: 70; and (b) the second SARS-CoV-2 The S polypeptide or fragment has at least 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identity to SEQ ID NO:69 and comprises an amino acid sequence containing one of the following sets of amino acid substitutions relative to SEQ ID NO:69: (1) D982P, V984P, F814P, A889P, A896P, and A939P; (2) K983P, V984P, F814P, A889P, A939P; 896P, and A939P, (3) D982P, V984P, F814P, A889P, A896P, A939P, R679G, R680S, and R682S, (4) K983P, V984P, F814P, A889P, A896P, A939P, R679G, R680S, and R682S, or (5) D982P, K983P, V984P, F814P, A889P, A896P, A939P, R679G, R680S, and R682S.

[0054] Another aspect of the disclosure features a composition or medical preparation including a first RNA encoding a first SARS-CoV-2 S polypeptide or immunogenic fragment thereof and a second RNA encoding a second SARS-CoV-2 S polypeptide or immunogenic fragment thereof, wherein (a) the first SARS-CoV-2 S polypeptide or fragment comprises an amino acid sequence having at least 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identity to SEQ ID NO: 70; and (b) the second SARS-CoV-2 The S polypeptide or fragment has at least 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identity to SEQ ID NO:70 and comprises an amino acid sequence containing one of the following sets of amino acid substitutions relative to SEQ ID NO:70: (1) D982P, V984P, F814P, A889P, A896P, and A939P; (2) K983P, V984P, F814P, A889P, A939P; 896P, and A939P, (3) D982P, V984P, F814P, A889P, A896P, A939P, R679G, R680S, and R682S, (4) K983P, V984P, F814P, A889P, A896P, A939P, R679G, R680S, and R682S, or (5) D982P, K983P, V984P, F814P, A889P, A896P, A939P, R679G, R680S, and R682S.

[0055] Another aspect of the disclosure features a composition or medical preparation including a first RNA encoding a first SARS-CoV-2 S polypeptide or immunogenic fragment thereof and a second RNA encoding a second SARS-CoV-2 S polypeptide or immunogenic fragment thereof, wherein (a) the first SARS-CoV-2 S polypeptide or fragment comprises an amino acid sequence having at least 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identity to SEQ ID NO: 70; and (b) the second SARS-CoV-2 The S polypeptide or fragment has at least 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identity to SEQ ID NO: 104 or 105 and comprises an amino acid sequence containing one of the following sets of amino acid substitutions relative to SEQ ID NO: 104 or 105: (1) D980P, V982P, F812P, A887P, A894P, and A937P; (2) K981P, V982P, F812P; A887P, A894P, and A937P, (3) D980P, V982P, F812P, A887P, A894P, A937P, R677G, R678S, and R680S, (4) K981P, V982P, F812P, A887P, A894P, A937P, R677G, R678S, and R680S, or (5) D980P, K981P, V982P, F812P, A887P, A894P, A937P, R677G, R678S, and R680S.

[0056] In one aspect, the disclosure provides a composition or medical preparation comprising a first RNA encoding a first SARS-CoV-2 S polypeptide or immunogenic fragment thereof and a second RNA encoding a second SARS-CoV-2 S polypeptide or immunogenic fragment thereof, wherein (a) the first SARS-CoV-2 S polypeptide or fragment comprises an amino acid sequence having at least 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identity to SEQ ID NO: 104 or SEQ ID NO: 105; and (b) the second SARS-CoV-2 The S polypeptide or fragment has at least 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identity to SEQ ID NO:1 and comprises an amino acid sequence containing one of the following sets of amino acid substitutions relative to SEQ ID NO:1: (1) D985P, V987P, F817P, A 892P, A899P, and A942P, (2) K986P, V987P, F817P, A892P, A899P, and A942P, (3) D985P, V987P, F817P, A892P, A899P, A942P, D614G, R682G, R683S, and R685S, (4) K986P, V987P, F817P, A89 2P, A899P, A942P, D614G, R682G, R683S, and R685S, (5) D985P, K986P, V987P, F817P, A892P, A899P, A942P, D614G, R682G, R683S, and R685S, (6) D985P, V987P, F817P, A892P, A899P, A942 P, R682G, R683S, and R685S, (7) K986P, V987P, F817P, A892P, A899P, A942P, R682G, R683S, and R685S, or (8) D985P, K986P, V987P, F817P, A892P, A899P, A942P, R682G, R683S, and R685S.

[0057] Another aspect of the disclosure features a composition or medical preparation including a first RNA encoding a first SARS-CoV-2 S polypeptide or immunogenic fragment thereof and a second RNA encoding a second SARS-CoV-2 S polypeptide or immunogenic fragment thereof, wherein (a) the first SARS-CoV-2 S polypeptide or fragment comprises an amino acid sequence having at least 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identity to SEQ ID NO: 104 or SEQ ID NO: 105; and (b) the second SARS-CoV-2 The S polypeptide or fragment has at least 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identity to SEQ ID NO:69 and comprises an amino acid sequence containing one of the following sets of amino acid substitutions relative to SEQ ID NO:69: (1) D982P, V984P, F814P, A889P, A896P, and A939P; (2) K983P, V984P, F814P, A889P, A939P; 896P, and A939P, (3) D982P, V984P, F814P, A889P, A896P, A939P, R679G, R680S, and R682S, (4) K983P, V984P, F814P, A889P, A896P, A939P, R679G, R680S, and R682S, or (5) D982P, K983P, V984P, F814P, A889P, A896P, A939P, R679G, R680S, and R682S.

[0058] Another aspect of the disclosure features a composition or medical preparation including a first RNA encoding a first SARS-CoV-2 S polypeptide or immunogenic fragment thereof and a second RNA encoding a second SARS-CoV-2 S polypeptide or immunogenic fragment thereof, wherein (a) the first SARS-CoV-2 S polypeptide or fragment comprises an amino acid sequence having at least 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identity to SEQ ID NO: 104 or SEQ ID NO: 105; and (b) the second SARS-CoV-2 The S polypeptide or fragment has at least 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identity to SEQ ID NO:70 and comprises an amino acid sequence containing one of the following sets of amino acid substitutions relative to SEQ ID NO:70: (1) D982P, V984P, F814P, A889P, A896P, and A939P; (2) K983P, V984P, F814P, A889P, A939P; 896P, and A939P, (3) D982P, V984P, F814P, A889P, A896P, A939P, R679G, R680S, and R682S, (4) K983P, V984P, F814P, A889P, A896P, A939P, R679G, R680S, and R682S, or (5) D982P, K983P, V984P, F814P, A889P, A896P, A939P, R679G, R680S, and R682S.

[0059] Another aspect of the disclosure features a composition or medical preparation including a first RNA encoding a first SARS-CoV-2 S polypeptide or immunogenic fragment thereof and a second RNA encoding a second SARS-CoV-2 S polypeptide or immunogenic fragment thereof, wherein (a) the first SARS-CoV-2 S polypeptide or fragment comprises an amino acid sequence having at least 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identity to SEQ ID NO: 104 or SEQ ID NO: 105; and (b) the second SARS-CoV-2 The S polypeptide or fragment has at least 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identity to SEQ ID NO: 104 or 105 and comprises an amino acid sequence containing one of the following sets of amino acid substitutions relative to SEQ ID NO: 104 or 105: (1) D980P, V982P, F812P, A887P, A894P, and A937P; (2) K981P, V982P, F812P; A887P, A894P, and A937P, (3) D980P, V982P, F812P, A887P, A894P, A937P, R677G, R678S, and R680S, (4) K981P, V982P, F812P, A887P, A894P, A937P, R677G, R678S, and R680S, or (5) D980P, K981P, V982P, F812P, A887P, A894P, A937P, R677G, R678S, and R680S.

[0060] Another aspect of the disclosure features a composition or medical preparation including a first RNA encoding a first SARS-CoV-2 S polypeptide or immunogenic fragment thereof and a second RNA encoding a second SARS-CoV-2 S polypeptide or immunogenic fragment thereof, wherein (a) the first SARS-CoV-2 S polypeptide or fragment has at least 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identity to SEQ ID NO: 1 and comprises an amino acid sequence that includes the following substitutions relative to SEQ ID NO: 1: D985P, V987P, F817P, A892P, A899P, and A942P; and (b) the second SARS-CoV-2 The S polypeptide or fragment has at least 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identity to SEQ ID NO:69 and comprises an amino acid sequence containing one of the following sets of amino acid substitutions relative to SEQ ID NO:69: (1) D982P, V984P, F814P, A889P, A896P, and A939P; (2) K983P, V984P, F814P, A889P, A939P; 896P, and A939P, (3) D982P, V984P, F814P, A889P, A896P, A939P, R679G, R680S, and R682S, (4) K983P, V984P, F814P, A889P, A896P, A939P, R679G, R680S, and R682S, or (5) D982P, K983P, V984P, F814P, A889P, A896P, A939P, R679G, R680S, and R682S.

[0061] Another aspect of the disclosure features a composition or medical preparation including a first RNA encoding a first SARS-CoV-2 S polypeptide or immunogenic fragment thereof and a second RNA encoding a second SARS-CoV-2 S polypeptide or immunogenic fragment thereof, wherein (a) the first SARS-CoV-2 S polypeptide or fragment has at least 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identity to SEQ ID NO: 1 and comprises an amino acid sequence that includes the following substitutions relative to SEQ ID NO: 1: K986P, V987P, F817P, A892P, A899P, and A942P; and (b) the second SARS-CoV-2 The S polypeptide or fragment has at least 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identity to SEQ ID NO:69 and comprises an amino acid sequence containing one of the following sets of amino acid substitutions relative to SEQ ID NO:69: (1) D982P, V984P, F814P, A889P, A896P, and A939P; (2) K983P, V984P, F814P, A889P, A939P; 896P, and A939P, (3) D982P, V984P, F814P, A889P, A896P, A939P, R679G, R680S, and R682S, (4) K983P, V984P, F814P, A889P, A896P, A939P, R679G, R680S, and R682S, or (5) D982P, K983P, V984P, F814P, A889P, A896P, A939P, R679G, R680S, and R682S.

[0062] Another aspect of the disclosure features a composition or medical preparation including a first RNA encoding a first SARS-CoV-2 S polypeptide or immunogenic fragment thereof and a second RNA encoding a second SARS-CoV-2 S polypeptide or immunogenic fragment thereof, wherein (a) the first SARS-CoV-2 S polypeptide or fragment has at least 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identity to SEQ ID NO: 1 and comprises an amino acid sequence that includes the following substitutions relative to SEQ ID NO: 1: D985P, V987P, F817P, A892P, A899P, A942P, D614G, R682G, R683S, and R685S; and (b) the second SARS-CoV-2 The S polypeptide or fragment has at least 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identity to SEQ ID NO:69 and comprises an amino acid sequence containing one of the following sets of amino acid substitutions relative to SEQ ID NO:69: (1) D982P, V984P, F814P, A889P, A896P, and A939P; (2) K983P, V984P, F814P, A889P, A939P; 896P, and A939P, (3) D982P, V984P, F814P, A889P, A896P, A939P, R679G, R680S, and R682S, (4) K983P, V984P, F814P, A889P, A896P, A939P, R679G, R680S, and R682S, or (5) D982P, K983P, V984P, F814P, A889P, A896P, A939P, R679G, R680S, and R682S.

[0063] Another aspect of the disclosure features a composition or medical preparation including a first RNA encoding a first SARS-CoV-2 S polypeptide or immunogenic fragment thereof and a second RNA encoding a second SARS-CoV-2 S polypeptide or immunogenic fragment thereof, wherein (a) the first SARS-CoV-2 S polypeptide or fragment has at least 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identity to SEQ ID NO: 1 and comprises an amino acid sequence that includes the following substitutions relative to SEQ ID NO: 1: K986P, V987P, F817P, A892P, A899P, A942P, D614G, R682G, R683S, and R685S; and (b) the second SARS-CoV-2 The S polypeptide or fragment has at least 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identity to SEQ ID NO:69 and comprises an amino acid sequence containing one of the following sets of amino acid substitutions relative to SEQ ID NO:69: (1) D982P, V984P, F814P, A889P, A896P, and A939P; (2) K983P, V984P, F814P, A889P, A939P; 896P, and A939P, (3) D982P, V984P, F814P, A889P, A896P, A939P, R679G, R680S, and R682S, (4) K983P, V984P, F814P, A889P, A896P, A939P, R679G, R680S, and R682S, or (5) D982P, K983P, V984P, F814P, A889P, A896P, A939P, R679G, R680S, and R682S.

[0064] Another aspect of the disclosure features a composition or medical preparation including a first RNA encoding a first SARS-CoV-2 S polypeptide or immunogenic fragment thereof and a second RNA encoding a second SARS-CoV-2 S polypeptide or immunogenic fragment thereof, wherein (a) the first SARS-CoV-2 S polypeptide or fragment has at least 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identity to SEQ ID NO: 1 and comprises an amino acid sequence that includes the following substitutions relative to SEQ ID NO: 1: D985P, K986P, V987P, F817P, A892P, A899P, A942P, D614G, R682G, R683S, and R685S; and (b) the second SARS-CoV-2 The S polypeptide or fragment has at least 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identity to SEQ ID NO:69 and comprises an amino acid sequence containing one of the following sets of amino acid substitutions relative to SEQ ID NO:69: (1) D982P, V984P, F814P, A889P, A896P, and A939P; (2) K983P, V984P, F814P, A889P, A939P; 896P, and A939P, (3) D982P, V984P, F814P, A889P, A896P, A939P, R679G, R680S, and R682S, (4) K983P, V984P, F814P, A889P, A896P, A939P, R679G, R680S, and R682S, or (5) D982P, K983P, V984P, F814P, A889P, A896P, A939P, R679G, R680S, and R682S.

[0065] Another aspect of the disclosure features a composition or medical preparation including a first RNA encoding a first SARS-CoV-2 S polypeptide or immunogenic fragment thereof and a second RNA encoding a second SARS-CoV-2 S polypeptide or immunogenic fragment thereof, wherein (a) the first SARS-CoV-2 S polypeptide or fragment has at least 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identity to SEQ ID NO: 1 and comprises an amino acid sequence that includes the following substitutions relative to SEQ ID NO: 1: D985P, V987P, F817P, A892P, A899P, A942P, R682G, R683S, and R685S; and (b) the second SARS-CoV-2 The S polypeptide or fragment has at least 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identity to SEQ ID NO:69 and comprises an amino acid sequence containing one of the following sets of amino acid substitutions relative to SEQ ID NO:69: (1) D982P, V984P, F814P, A889P, A896P, and A939P; (2) K983P, V984P, F814P, A889P, A939P; 896P, and A939P, (3) D982P, V984P, F814P, A889P, A896P, A939P, R679G, R680S, and R682S, (4) K983P, V984P, F814P, A889P, A896P, A939P, R679G, R680S, and R682S, or (5) D982P, K983P, V984P, F814P, A889P, A896P, A939P, R679G, R680S, and R682S.

[0066] Another aspect of the disclosure features a composition or medical preparation including a first RNA encoding a first SARS-CoV-2 S polypeptide or immunogenic fragment thereof and a second RNA encoding a second SARS-CoV-2 S polypeptide or immunogenic fragment thereof, wherein (a) the first SARS-CoV-2 S polypeptide or fragment has at least 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identity to SEQ ID NO: 1 and comprises an amino acid sequence that includes the following substitutions relative to SEQ ID NO: 1: K986P, V987P, F817P, A892P, A899P, A942P, R682G, R683S, and R685S; and (b) the second SARS-CoV-2 The S polypeptide or fragment has at least 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identity to SEQ ID NO:69 and comprises an amino acid sequence containing one of the following sets of amino acid substitutions relative to SEQ ID NO:69: (1) D982P, V984P, F814P, A889P, A896P, and A939P; (2) K983P, V984P, F814P, A889P, A939P; 896P, and A939P, (3) D982P, V984P, F814P, A889P, A896P, A939P, R679G, R680S, and R682S, (4) K983P, V984P, F814P, A889P, A896P, A939P, R679G, R680S, and R682S, or (5) D982P, K983P, V984P, F814P, A889P, A896P, A939P, R679G, R680S, and R682S.

[0067] Another aspect of the disclosure features a composition or medical preparation including a first RNA encoding a first SARS-CoV-2 S polypeptide or immunogenic fragment thereof and a second RNA encoding a second SARS-CoV-2 S polypeptide or immunogenic fragment thereof, wherein (a) the first SARS-CoV-2 S polypeptide or fragment has at least 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identity to SEQ ID NO: 1 and comprises an amino acid sequence that includes the following substitutions relative to SEQ ID NO: 1: D985P, K986P, V987P, F817P, A892P, A899P, A942P, R682G, R683S, R685S; and (b) the second SARS-CoV-2 The S polypeptide or fragment has at least 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identity to SEQ ID NO:69 and comprises an amino acid sequence containing one of the following sets of amino acid substitutions relative to SEQ ID NO:69: (1) D982P, V984P, F814P, A889P, A896P, and A939P; (2) K983P, V984P, F814P, A889P, A939P; 896P, and A939P, (3) D982P, V984P, F814P, A889P, A896P, A939P, R679G, R680S, and R682S, (4) K983P, V984P, F814P, A889P, A896P, A939P, R679G, R680S, and R682S, or (5) D982P, K983P, V984P, F814P, A889P, A896P, A939P, R679G, R680S, and R682S.

[0068] Another aspect of the disclosure features a composition or medical preparation including a first RNA encoding a first SARS-CoV-2 S polypeptide or immunogenic fragment thereof and a second RNA encoding a second SARS-CoV-2 S polypeptide or immunogenic fragment thereof, wherein (a) the first SARS-CoV-2 S polypeptide or fragment has at least 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identity to SEQ ID NO: 1 and comprises an amino acid sequence that includes the following substitutions relative to SEQ ID NO: 1: D985P, V987P, F817P, A892P, A899P, and A942P; and (b) the second SARS-CoV-2 The S polypeptide or fragment has at least 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identity to SEQ ID NO:70 and comprises an amino acid sequence containing one of the following sets of amino acid substitutions relative to SEQ ID NO:70: (1) D982P, V984P, F814P, A889P, A896P, and A939P; (2) K983P, V984P, F814P, A889P, A939P; 896P, and A939P, (3) D982P, V984P, F814P, A889P, A896P, A939P, R679G, R680S, and R682S, (4) K983P, V984P, F814P, A889P, A896P, A939P, R679G, R680S, and R682S, or (5) D982P, K983P, V984P, F814P, A889P, A896P, A939P, R679G, R680S, and R682S.

[0069] Another aspect of the disclosure features a composition or medical preparation including a first RNA encoding a first SARS-CoV-2 S polypeptide or immunogenic fragment thereof and a second RNA encoding a second SARS-CoV-2 S polypeptide or immunogenic fragment thereof, wherein (a) the first SARS-CoV-2 S polypeptide or fragment has at least 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identity to SEQ ID NO: 1 and comprises an amino acid sequence that includes the following substitutions relative to SEQ ID NO: 1: K986P, V987P, F817P, A892P, A899P, and A942P; and (b) the second SARS-CoV-2 The S polypeptide or fragment has at least 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identity to SEQ ID NO:70 and comprises an amino acid sequence containing one of the following sets of amino acid substitutions relative to SEQ ID NO:70: (1) D982P, V984P, F814P, A889P, A896P, and A939P; (2) K983P, V984P, F814P, A889P, A939P; 896P, and A939P, (3) D982P, V984P, F814P, A889P, A896P, A939P, R679G, R680S, and R682S, (4) K983P, V984P, F814P, A889P, A896P, A939P, R679G, R680S, and R682S, or (5) D982P, K983P, V984P, F814P, A889P, A896P, A939P, R679G, R680S, and R682S.

[0070] Another aspect of the disclosure features a composition or medical preparation including a first RNA encoding a first SARS-CoV-2 S polypeptide or immunogenic fragment thereof and a second RNA encoding a second SARS-CoV-2 S polypeptide or immunogenic fragment thereof, wherein (a) the first SARS-CoV-2 S polypeptide or fragment has at least 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identity to SEQ ID NO: 1 and comprises an amino acid sequence that includes the following substitutions relative to SEQ ID NO: 1: D985P, V987P, F817P, A892P, A899P, A942P, D614G, R682G, R683S, and R685S; and (b) the second SARS-CoV-2 The S polypeptide or fragment has at least 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identity to SEQ ID NO:70 and comprises an amino acid sequence containing one of the following sets of amino acid substitutions relative to SEQ ID NO:70: (1) D982P, V984P, F814P, A889P, A896P, and A939P; (2) K983P, V984P, F814P, A889P, A939P; 896P, and A939P, (3) D982P, V984P, F814P, A889P, A896P, A939P, R679G, R680S, and R682S, (4) K983P, V984P, F814P, A889P, A896P, A939P, R679G, R680S, and R682S, or (5) D982P, K983P, V984P, F814P, A889P, A896P, A939P, R679G, R680S, and R682S.

[0071] Another aspect of the disclosure features a composition or medical preparation including a first RNA encoding a first SARS-CoV-2 S polypeptide or immunogenic fragment thereof and a second RNA encoding a second SARS-CoV-2 S polypeptide or immunogenic fragment thereof, wherein (a) the first SARS-CoV-2 S polypeptide or fragment has at least 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identity to SEQ ID NO: 1 and comprises an amino acid sequence that includes the following substitutions relative to SEQ ID NO: 1: K986P, V987P, F817P, A892P, A899P, A942P, D614G, R682G, R683S, and R685S; and (b) the second SARS-CoV-2 The S polypeptide or fragment has at least 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identity to SEQ ID NO:70 and comprises an amino acid sequence containing one of the following sets of amino acid substitutions relative to SEQ ID NO:70: (1) D982P, V984P, F814P, A889P, A896P, and A939P; (2) K983P, V984P, F814P, A889P, A939P; 896P, and A939P, (3) D982P, V984P, F814P, A889P, A896P, A939P, R679G, R680S, and R682S, (4) K983P, V984P, F814P, A889P, A896P, A939P, R679G, R680S, and R682S, or (5) D982P, K983P, V984P, F814P, A889P, A896P, A939P, R679G, R680S, and R682S.

[0072] Another aspect of the disclosure features a composition or medical preparation including a first RNA encoding a first SARS-CoV-2 S polypeptide or immunogenic fragment thereof and a second RNA encoding a second SARS-CoV-2 S polypeptide or immunogenic fragment thereof, wherein (a) the first SARS-CoV-2 S polypeptide or fragment has at least 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identity to SEQ ID NO: 1 and comprises an amino acid sequence that includes the following substitutions relative to SEQ ID NO: 1: D985P, K986P, V987P, F817P, A892P, A899P, A942P, D614G, R682G, R683S, and R685S; and (b) the second SARS-CoV-2 The S polypeptide or fragment has at least 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identity to SEQ ID NO:70 and comprises an amino acid sequence containing one of the following sets of amino acid substitutions relative to SEQ ID NO:70: (1) D982P, V984P, F814P, A889P, A896P, and A939P; (2) K983P, V984P, F814P, A889P, A939P; 896P, and A939P, (3) D982P, V984P, F814P, A889P, A896P, A939P, R679G, R680S, and R682S, (4) K983P, V984P, F814P, A889P, A896P, A939P, R679G, R680S, and R682S, or (5) D982P, K983P, V984P, F814P, A889P, A896P, A939P, R679G, R680S, and R682S.

[0073] Another aspect of the disclosure features a composition or medical preparation including a first RNA encoding a first SARS-CoV-2 S polypeptide or immunogenic fragment thereof and a second RNA encoding a second SARS-CoV-2 S polypeptide or immunogenic fragment thereof, wherein (a) the first SARS-CoV-2 S polypeptide or fragment has at least 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identity to SEQ ID NO: 1 and comprises an amino acid sequence that includes the following substitutions relative to SEQ ID NO: 1: D985P, V987P, F817P, A892P, A899P, A942P, R682G, R683S, and R685S; and (b) the second SARS-CoV-2 The S polypeptide or fragment has at least 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identity to SEQ ID NO:70 and comprises an amino acid sequence containing one of the following sets of amino acid substitutions relative to SEQ ID NO:70: (1) D982P, V984P, F814P, A889P, A896P, and A939P; (2) K983P, V984P, F814P, A889P, A939P; 896P, and A939P, (3) D982P, V984P, F814P, A889P, A896P, A939P, R679G, R680S, and R682S, (4) K983P, V984P, F814P, A889P, A896P, A939P, R679G, R680S, and R682S, or (5) D982P, K983P, V984P, F814P, A889P, A896P, A939P, R679G, R680S, and R682S.

[0074] Another aspect of the disclosure features a composition or medical preparation including a first RNA encoding a first SARS-CoV-2 S polypeptide or immunogenic fragment thereof and a second RNA encoding a second SARS-CoV-2 S polypeptide or immunogenic fragment thereof, wherein (a) the first SARS-CoV-2 S polypeptide or fragment has at least 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identity to SEQ ID NO: 1 and comprises an amino acid sequence that includes the following substitutions relative to SEQ ID NO: 1: K986P, V987P, F817P, A892P, A899P, A942P, R682G, R683S, and R685S; and (b) the second SARS-CoV-2 The S polypeptide or fragment has at least 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identity to SEQ ID NO:70 and comprises an amino acid sequence containing one of the following sets of amino acid substitutions relative to SEQ ID NO:70: (1) D982P, V984P, F814P, A889P, A896P, and A939P; (2) K983P, V984P, F814P, A889P, A939P; 896P, and A939P, (3) D982P, V984P, F814P, A889P, A896P, A939P, R679G, R680S, and R682S, (4) K983P, V984P, F814P, A889P, A896P, A939P, R679G, R680S, and R682S, or (5) D982P, K983P, V984P, F814P, A889P, A896P, A939P, R679G, R680S, and R682S.

[0075] Another aspect of the disclosure features a composition or medical preparation including a first RNA encoding a first SARS-CoV-2 S polypeptide or immunogenic fragment thereof and a second RNA encoding a second SARS-CoV-2 S polypeptide or immunogenic fragment thereof, wherein (a) the first SARS-CoV-2 S polypeptide or fragment has at least 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identity to SEQ ID NO: 1 and comprises an amino acid sequence that includes the following substitutions relative to SEQ ID NO: 1: D985P, K986P, V987P, F817P, A892P, A899P, A942P, R682G, R683S, and R685S; and (b) the second SARS-CoV-2 The S polypeptide or fragment has at least 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identity to SEQ ID NO:70 and comprises an amino acid sequence containing one of the following sets of amino acid substitutions relative to SEQ ID NO:70: (1) D982P, V984P, F814P, A889P, A896P, and A939P; (2) K983P, V984P, F814P, A889P, A939P; 896P, and A939P, (3) D982P, V984P, F814P, A889P, A896P, A939P, R679G, R680S, and R682S, (4) K983P, V984P, F814P, A889P, A896P, A939P, R679G, R680S, and R682S, or (5) D982P, K983P, V984P, F814P, A889P, A896P, A939P, R679G, R680S, and R682S.

[0076] Another aspect of the disclosure features a composition or medical preparation including a first RNA encoding a first SARS-CoV-2 S polypeptide or immunogenic fragment thereof and a second RNA encoding a second SARS-CoV-2 S polypeptide or immunogenic fragment thereof, wherein (a) the first SARS-CoV-2 S polypeptide or fragment comprises an amino acid sequence having at least 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identity to SEQ ID NO: 1 and including the following substitutions relative to SEQ ID NO: 1: D985P, V987P, F817P, A892P, A899P, and A942P; and (b) the second SARS-CoV-2 The S polypeptide or fragment has at least 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identity to SEQ ID NO: 104 or 105 and comprises an amino acid sequence containing one of the following sets of amino acid substitutions relative to SEQ ID NO: 104 or 105: (1) D980P, V982P, F812P, A887P, A894P, and A937P; (2) K981P, V982P, F812P; A887P, A894P, and A937P, (3) D980P, V982P, F812P, A887P, A894P, A937P, R677G, R678S, and R680S, (4) K981P, V982P, F812P, A887P, A894P, A937P, R677G, R678S, and R680S, or (5) D980P, K981P, V982P, F812P, A887P, A894P, A937P, R677G, R678S, and R680S.

[0077] Another aspect of the disclosure features a composition or medical preparation including a first RNA encoding a first SARS-CoV-2 S polypeptide or immunogenic fragment thereof and a second RNA encoding a second SARS-CoV-2 S polypeptide or immunogenic fragment thereof, wherein (a) the first SARS-CoV-2 S polypeptide or fragment comprises an amino acid sequence having at least 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identity to SEQ ID NO: 1 and including the following substitutions relative to SEQ ID NO: 1: K986P, V987P, F817P, A892P, A899P, and A942P; and (b) the second SARS-CoV-2 The S polypeptide or fragment has at least 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identity to SEQ ID NO: 104 or 105 and comprises an amino acid sequence containing one of the following sets of amino acid substitutions relative to SEQ ID NO: 104 or 105: (1) D980P, V982P, F812P, A887P, A894P, and A937P; (2) K981P, V982P, F812P; A887P, A894P, and A937P, (3) D980P, V982P, F812P, A887P, A894P, A937P, R677G, R678S, and R680S, (4) K981P, V982P, F812P, A887P, A894P, A937P, R677G, R678S, and R680S, or (5) D980P, K981P, V982P, F812P, A887P, A894P, A937P, R677G, R678S, and R680S.

[0078] Another aspect of the disclosure features a composition or medical preparation including a first RNA encoding a first SARS-CoV-2 S polypeptide or immunogenic fragment thereof and a second RNA encoding a second SARS-CoV-2 S polypeptide or immunogenic fragment thereof, wherein (a) the first SARS-CoV-2 S polypeptide or fragment has at least 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identity to SEQ ID NO: 1 and comprises an amino acid sequence containing the following substitutions relative to SEQ ID NO: 1: D985P, V987P, F817P, A892P, A899P, A942P, D614G, R682G, R683S, and R685S; and (b) the second SARS-CoV-2 The S polypeptide or fragment has at least 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identity to SEQ ID NO: 104 or 105 and comprises an amino acid sequence containing one of the following sets of amino acid substitutions relative to SEQ ID NO: 104 or 105: (1) D980P, V982P, F812P, A887P, A894P, and A937P; (2) K981P, V982P, F812P; A887P, A894P, and A937P, (3) D980P, V982P, F812P, A887P, A894P, A937P, R677G, R678S, and R680S, (4) K981P, V982P, F812P, A887P, A894P, A937P, R677G, R678S, and R680S, or (5) D980P, K981P, V982P, F812P, A887P, A894P, A937P, R677G, R678S, and R680S.

[0079] Another aspect of the disclosure features a composition or medical preparation including a first RNA encoding a first SARS-CoV-2 S polypeptide or immunogenic fragment thereof and a second RNA encoding a second SARS-CoV-2 S polypeptide or immunogenic fragment thereof, wherein (a) the first SARS-CoV-2 S polypeptide or fragment has at least 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identity to SEQ ID NO: 1 and comprises an amino acid sequence that includes the following substitutions relative to SEQ ID NO: 1: K986P, V987P, F817P, A892P, A899P, A942P, D614G, R682G, R683S, and R685S; and (b) the second SARS-CoV-2 The S polypeptide or fragment has at least 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identity to SEQ ID NO: 104 or 105 and comprises an amino acid sequence containing one of the following sets of amino acid substitutions relative to SEQ ID NO: 104 or 105: (1) D980P, V982P, F812P, A887P, A894P, and A937P; (2) K981P, V982P, F812P; A887P, A894P, and A937P, (3) D980P, V982P, F812P, A887P, A894P, A937P, R677G, R678S, and R680S, (4) K981P, V982P, F812P, A887P, A894P, A937P, R677G, R678S, and R680S, or (5) D980P, K981P, V982P, F812P, A887P, A894P, A937P, R677G, R678S, and R680S.

[0080] Another aspect of the disclosure features a composition or medical preparation including a first RNA encoding a first SARS-CoV-2 S polypeptide or immunogenic fragment thereof and a second RNA encoding a second SARS-CoV-2 S polypeptide or immunogenic fragment thereof, wherein (a) the first SARS-CoV-2 S polypeptide or fragment has at least 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identity to SEQ ID NO: 1 and comprises an amino acid sequence that includes the following substitutions relative to SEQ ID NO: 1: D985P, K986P, V987P, F817P, A892P, A899P, A942P, D614G, R682G, R683S, and R685S; and (b) the second SARS-CoV-2 The S polypeptide or fragment has at least 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identity to SEQ ID NO: 104 or 105 and comprises an amino acid sequence containing one of the following sets of amino acid substitutions relative to SEQ ID NO: 104 or 105: (1) D980P, V982P, F812P, A887P, A894P, and A937P; (2) K981P, V982P, F812P; A887P, A894P, and A937P, (3) D980P, V982P, F812P, A887P, A894P, A937P, R677G, R678S, and R680S, (4) K981P, V982P, F812P, A887P, A894P, A937P, R677G, R678S, and R680S, or (5) D980P, K981P, V982P, F812P, A887P, A894P, A937P, R677G, R678S, and R680S.

[0081] Another aspect of the disclosure features a composition or medical preparation including a first RNA encoding a first SARS-CoV-2 S polypeptide or immunogenic fragment thereof and a second RNA encoding a second SARS-CoV-2 S polypeptide or immunogenic fragment thereof, wherein (a) the first SARS-CoV-2 S polypeptide or fragment has at least 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identity to SEQ ID NO: 1 and comprises an amino acid sequence that includes the following substitutions relative to SEQ ID NO: 1: D985P, V987P, F817P, A892P, A899P, A942P, R682G, R683S, and R685S; and (b) the second SARS-CoV-2 The S polypeptide or fragment has at least 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identity to SEQ ID NO: 104 or 105 and comprises an amino acid sequence containing one of the following sets of amino acid substitutions relative to SEQ ID NO: 104 or 105: (1) D980P, V982P, F812P, A887P, A894P, and A937P; (2) K981P, V982P, F812P; A887P, A894P, and A937P, (3) D980P, V982P, F812P, A887P, A894P, A937P, R677G, R678S, and R680S, (4) K981P, V982P, F812P, A887P, A894P, A937P, R677G, R678S, and R680S, or (5) D980P, K981P, V982P, F812P, A887P, A894P, A937P, R677G, R678S, and R680S.

[0082] Another aspect of the disclosure features a composition or medical preparation including a first RNA encoding a first SARS-CoV-2 S polypeptide or immunogenic fragment thereof and a second RNA encoding a second SARS-CoV-2 S polypeptide or immunogenic fragment thereof, wherein (a) the first SARS-CoV-2 S polypeptide or fragment has at least 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identity to SEQ ID NO: 1 and comprises an amino acid sequence that includes the following substitutions relative to SEQ ID NO: 1: K986P, V987P, F817P, A892P, A899P, A942P, R682G, R683S, and R685S; and (b) the second SARS-CoV-2 The S polypeptide or fragment has at least 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identity to SEQ ID NO: 104 or 105 and comprises an amino acid sequence containing one of the following sets of amino acid substitutions relative to SEQ ID NO: 104 or 105: (1) D980P, V982P, F812P, A887P, A894P, and A937P; (2) K981P, V982P, F812P; A887P, A894P, and A937P, (3) D980P, V982P, F812P, A887P, A894P, A937P, R677G, R678S, and R680S, (4) K981P, V982P, F812P, A887P, A894P, A937P, R677G, R678S, and R680S, or (5) D980P, K981P, V982P, F812P, A887P, A894P, A937P, R677G, R678S, and R680S.

[0083] Another aspect of the disclosure features a composition or medical preparation including a first RNA encoding a first SARS-CoV-2 S polypeptide or immunogenic fragment thereof and a second RNA encoding a second SARS-CoV-2 S polypeptide or immunogenic fragment thereof, wherein (a) the first SARS-CoV-2 S polypeptide or fragment has at least 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identity to SEQ ID NO: 1 and comprises an amino acid sequence that includes the following substitutions relative to SEQ ID NO: 1: D985P, K986P, V987P, F817P, A892P, A899P, A942P, R682G, R683S, and R685S; and (b) the second SARS-CoV-2 The S polypeptide or fragment has at least 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identity to SEQ ID NO: 104 or 105 and comprises an amino acid sequence containing one of the following sets of amino acid substitutions relative to SEQ ID NO: 104 or 105: (1) D980P, V982P, F812P, A887P, A894P, and A937P; (2) K981P, V982P, F812P; A887P, A894P, and A937P, (3) D980P, V982P, F812P, A887P, A894P, A937P, R677G, R678S, and R680S, (4) K981P, V982P, F812P, A887P, A894P, A937P, R677G, R678S, and R680S, or (5) D980P, K981P, V982P, F812P, A887P, A894P, A937P, R677G, R678S, and R680S.

[0084] Another aspect of the disclosure features a composition or medical preparation including a first RNA encoding a first SARS-CoV-2 S polypeptide or immunogenic fragment thereof and a second RNA encoding a second SARS-CoV-2 S polypeptide or immunogenic fragment thereof, wherein (a) the first SARS-CoV-2 S polypeptide or fragment comprises an amino acid sequence having at least 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identity to SEQ ID NO: 69 and including the following substitutions relative to SEQ ID NO: 69: D982P, V984P, F814P, A889P, A896P, and A939P; and (b) the second SARS-CoV-2 The S polypeptide or fragment has at least 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identity to SEQ ID NO:70 and comprises an amino acid sequence containing one of the following sets of amino acid substitutions relative to SEQ ID NO:70: (1) D982P, V984P, F814P, A889P, A896P, and A939P; (2) K983P, V984P, F814P, A889P, A939P; 896P, and A939P, (3) D982P, V984P, F814P, A889P, A896P, A939P, R679G, R680S, and R682S, (4) K983P, V984P, F814P, A889P, A896P, A939P, R679G, R680S, and R682S, or (5) D982P, K983P, V984P, F814P, A889P, A896P, A939P, R679G, R680S, and R682S.

[0085] Another aspect of the disclosure features a composition or medical preparation including a first RNA encoding a first SARS-CoV-2 S polypeptide or immunogenic fragment thereof and a second RNA encoding a second SARS-CoV-2 S polypeptide or immunogenic fragment thereof, wherein (a) the first SARS-CoV-2 S polypeptide or fragment comprises an amino acid sequence having at least 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identity to SEQ ID NO: 69 and including the following substitutions relative to SEQ ID NO: 69: K983P, V984P, F814P, A889P, A896P, and A939P; and (b) the second SARS-CoV-2 The S polypeptide or fragment has at least 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identity to SEQ ID NO:70 and comprises an amino acid sequence containing one of the following sets of amino acid substitutions relative to SEQ ID NO:70: (1) D982P, V984P, F814P, A889P, A896P, and A939P; (2) K983P, V984P, F814P, A889P, A939P; 896P, and A939P, (3) D982P, V984P, F814P, A889P, A896P, A939P, R679G, R680S, and R682S, (4) K983P, V984P, F814P, A889P, A896P, A939P, R679G, R680S, and R682S, or (5) D982P, K983P, V984P, F814P, A889P, A896P, A939P, R679G, R680S, and R682S.

[0086] Another aspect of the disclosure features a composition or medical preparation including a first RNA encoding a first SARS-CoV-2 S polypeptide or immunogenic fragment thereof and a second RNA encoding a second SARS-CoV-2 S polypeptide or immunogenic fragment thereof, wherein (a) the first SARS-CoV-2 S polypeptide or fragment comprises an amino acid sequence having at least 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identity to SEQ ID NO: 69 and including the following substitutions relative to SEQ ID NO: 69: D982P, V984P, F814P, A889P, A896P, A939P, R679G, R680S, and R682S; and (b) the second SARS-CoV-2 The S polypeptide or fragment has at least 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identity to SEQ ID NO:70 and comprises an amino acid sequence containing one of the following sets of amino acid substitutions relative to SEQ ID NO:70: (1) D982P, V984P, F814P, A889P, A896P, and A939P; (2) K983P, V984P, F814P, A889P, A939P; 896P, and A939P, (3) D982P, V984P, F814P, A889P, A896P, A939P, R679G, R680S, and R682S, (4) K983P, V984P, F814P, A889P, A896P, A939P, R679G, R680S, and R682S, or (5) D982P, K983P, V984P, F814P, A889P, A896P, A939P, R679G, R680S, and R682S.

[0087] Another aspect of the disclosure features a composition or medical preparation including a first RNA encoding a first SARS-CoV-2 S polypeptide or immunogenic fragment thereof and a second RNA encoding a second SARS-CoV-2 S polypeptide or immunogenic fragment thereof, wherein (a) the first SARS-CoV-2 S polypeptide or fragment comprises an amino acid sequence having at least 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identity to SEQ ID NO: 69 and including the following substitutions relative to SEQ ID NO: 69: K983P, V984P, F814P, A889P, A896P, A939P, R679G, R680S, and R682S; and (b) the second SARS-CoV-2 The S polypeptide or fragment has at least 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identity to SEQ ID NO:70 and comprises an amino acid sequence containing one of the following sets of amino acid substitutions relative to SEQ ID NO:70: (1) D982P, V984P, F814P, A889P, A896P, and A939P; (2) K983P, V984P, F814P, A889P, A939P; 896P, and A939P, (3) D982P, V984P, F814P, A889P, A896P, A939P, R679G, R680S, and R682S, (4) K983P, V984P, F814P, A889P, A896P, A939P, R679G, R680S, and R682S, or (5) D982P, K983P, V984P, F814P, A889P, A896P, A939P, R679G, R680S, and R682S.

[0088] Another aspect of the disclosure features a composition or medical preparation including a first RNA encoding a first SARS-CoV-2 S polypeptide or immunogenic fragment thereof and a second RNA encoding a second SARS-CoV-2 S polypeptide or immunogenic fragment thereof, wherein (a) the first SARS-CoV-2 S polypeptide or fragment has at least 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identity to SEQ ID NO: 69 and comprises an amino acid sequence that includes the following substitutions relative to SEQ ID NO: 69: D982P, K983P, V984P, F814P, A889P, A896P, A939P, R679G, R680S, and R682S; and (b) the second SARS-CoV-2 The S polypeptide or fragment has at least 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identity to SEQ ID NO:70 and comprises an amino acid sequence containing one of the following sets of amino acid substitutions relative to SEQ ID NO:70: (1) D982P, V984P, F814P, A889P, A896P, and A939P; (2) K983P, V984P, F814P, A889P, A939P; 896P, and A939P, (3) D982P, V984P, F814P, A889P, A896P, A939P, R679G, R680S, and R682S, (4) K983P, V984P, F814P, A889P, A896P, A939P, R679G, R680S, and R682S, or (5) D982P, K983P, V984P, F814P, A889P, A896P, A939P, R679G, R680S, and R682S.

[0089] Another aspect of the disclosure features a composition or medical preparation including a first RNA encoding a first SARS-CoV-2 S polypeptide or immunogenic fragment thereof and a second RNA encoding a second SARS-CoV-2 S polypeptide or immunogenic fragment thereof, wherein (a) the first SARS-CoV-2 S polypeptide or fragment comprises an amino acid sequence having at least 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identity to SEQ ID NO: 69 and including the following substitutions relative to SEQ ID NO: 69: D982P, V984P, F814P, A889P, A896P, and A939P; and (b) the second SARS-CoV-2 The S polypeptide or fragment has at least 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identity to SEQ ID NO: 104 or 105 and comprises an amino acid sequence containing one of the following sets of amino acid substitutions relative to SEQ ID NO: 104 or 105: (1) D980P, V982P, F812P, A887P, A894P, and A937P; (2) K981P, V982P, F812P; A887P, A894P, and A937P, (3) D980P, V982P, F812P, A887P, A894P, A937P, R677G, R678S, and R680S, (4) K981P, V982P, F812P, A887P, A894P, A937P, R677G, R678S, and R680S, or (5) D980P, K981P, V982P, F812P, A887P, A894P, A937P, R677G, R678S, and R680S.

[0090] Another aspect of the disclosure features a composition or medical preparation including a first RNA encoding a first SARS-CoV-2 S polypeptide or immunogenic fragment thereof and a second RNA encoding a second SARS-CoV-2 S polypeptide or immunogenic fragment thereof, wherein (a) the first SARS-CoV-2 S polypeptide or fragment comprises an amino acid sequence having at least 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identity to SEQ ID NO: 69 and including the following substitutions relative to SEQ ID NO: 69: K983P, V984P, F814P, A889P, A896P, and A939P; and (b) the second SARS-CoV-2 The S polypeptide or fragment has at least 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identity to SEQ ID NO: 104 or 105 and comprises an amino acid sequence containing one of the following sets of amino acid substitutions relative to SEQ ID NO: 104 or 105: (1) D980P, V982P, F812P, A887P, A894P, and A937P; (2) K981P, V982P, F812P; A887P, A894P, and A937P, (3) D980P, V982P, F812P, A887P, A894P, A937P, R677G, R678S, and R680S, (4) K981P, V982P, F812P, A887P, A894P, A937P, R677G, R678S, and R680S, or (5) D980P, K981P, V982P, F812P, A887P, A894P, A937P, R677G, R678S, and R680S.

[0091] Another aspect of the disclosure features a composition or medical preparation including a first RNA encoding a first SARS-CoV-2 S polypeptide or immunogenic fragment thereof and a second RNA encoding a second SARS-CoV-2 S polypeptide or immunogenic fragment thereof, wherein (a) the first SARS-CoV-2 S polypeptide or fragment comprises an amino acid sequence having at least 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identity to SEQ ID NO: 69 and including the following substitutions relative to SEQ ID NO: 69: D982P, V984P, F814P, A889P, A896P, A939P, R679G, R680S, and R682S; and (b) the second SARS-CoV-2 The S polypeptide or fragment has at least 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identity to SEQ ID NO: 104 or 105 and comprises an amino acid sequence containing one of the following sets of amino acid substitutions relative to SEQ ID NO: 104 or 105: (1) D980P, V982P, F812P, A887P, A894P, and A937P; (2) K981P, V982P, F812P; A887P, A894P, and A937P, (3) D980P, V982P, F812P, A887P, A894P, A937P, R677G, R678S, and R680S, (4) K981P, V982P, F812P, A887P, A894P, A937P, R677G, R678S, and R680S, or (5) D980P, K981P, V982P, F812P, A887P, A894P, A937P, R677G, R678S, and R680S.

[0092] Another aspect of the disclosure features a composition or medical preparation including a first RNA encoding a first SARS-CoV-2 S polypeptide or immunogenic fragment thereof and a second RNA encoding a second SARS-CoV-2 S polypeptide or immunogenic fragment thereof, wherein (a) the first SARS-CoV-2 S polypeptide or fragment comprises an amino acid sequence having at least 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identity to SEQ ID NO: 69 and including the following substitutions relative to SEQ ID NO: 69: K983P, V984P, F814P, A889P, A896P, A939P, R679G, R680S, and R682S; and (b) the second SARS-CoV-2 The S polypeptide or fragment has at least 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identity to SEQ ID NO: 104 or 105 and comprises an amino acid sequence containing one of the following sets of amino acid substitutions relative to SEQ ID NO: 104 or 105: (1) D980P, V982P, F812P, A887P, A894P, and A937P; (2) K981P, V982P, F812P; A887P, A894P, and A937P, (3) D980P, V982P, F812P, A887P, A894P, A937P, R677G, R678S, and R680S, (4) K981P, V982P, F812P, A887P, A894P, A937P, R677G, R678S, and R680S, or (5) D980P, K981P, V982P, F812P, A887P, A894P, A937P, R677G, R678S, and R680S.

[0093] Another aspect of the disclosure features a composition or medical preparation including a first RNA encoding a first SARS-CoV-2 S polypeptide or immunogenic fragment thereof and a second RNA encoding a second SARS-CoV-2 S polypeptide or immunogenic fragment thereof, wherein (a) the first SARS-CoV-2 S polypeptide or fragment has at least 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identity to SEQ ID NO: 69 and comprises an amino acid sequence that includes the following substitutions relative to SEQ ID NO: 69: D982P, K983P, V984P, F814P, A889P, A896P, A939P, R679G, R680S, and R682S; and (b) the second SARS-CoV-2 The S polypeptide or fragment has at least 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identity to SEQ ID NO: 104 or 105 and comprises an amino acid sequence containing one of the following sets of amino acid substitutions relative to SEQ ID NO: 104 or 105: (1) D980P, V982P, F812P, A887P, A894P, and A937P; (2) K981P, V982P, F812P; A887P, A894P, and A937P, (3) D980P, V982P, F812P, A887P, A894P, A937P, R677G, R678S, and R680S, (4) K981P, V982P, F812P, A887P, A894P, A937P, R677G, R678S, and R680S, or (5) D980P, K981P, V982P, F812P, A887P, A894P, A937P, R677G, R678S, and R680S.

[0094] Another aspect of the disclosure features a composition or medical preparation including a first RNA encoding a first SARS-CoV-2 S polypeptide or immunogenic fragment thereof and a second RNA encoding a second SARS-CoV-2 S polypeptide or immunogenic fragment thereof, wherein (a) the first SARS-CoV-2 S polypeptide or fragment comprises an amino acid sequence having at least 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identity to SEQ ID NO: 70 and including the following substitutions relative to SEQ ID NO: 70: D982P, V984P, F814P, A889P, A896P, and A939P; and (b) the second SARS-CoV-2 The S polypeptide or fragment has at least 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identity to SEQ ID NO: 104 or 105 and comprises an amino acid sequence containing one of the following sets of amino acid substitutions relative to SEQ ID NO: 104 or 105: (1) D980P, V982P, F812P, A887P, A894P, and A937P; (2) K981P, V982P, F812P; A887P, A894P, and A937P, (3) D980P, V982P, F812P, A887P, A894P, A937P, R677G, R678S, and R680S, (4) K981P, V982P, F812P, A887P, A894P, A937P, R677G, R678S, and R680S, or (5) D980P, K981P, V982P, F812P, A887P, A894P, A937P, R677G, R678S, and R680S.

[0095] Another aspect of the disclosure features a composition or medical preparation including a first RNA encoding a first SARS-CoV-2 S polypeptide or immunogenic fragment thereof and a second RNA encoding a second SARS-CoV-2 S polypeptide or immunogenic fragment thereof, wherein (a) the first SARS-CoV-2 S polypeptide or fragment comprises an amino acid sequence having at least 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identity to SEQ ID NO: 70 and including the following substitutions relative to SEQ ID NO: 70: K983P, V984P, F814P, A889P, A896P, and A939P; and (b) the second SARS-CoV-2 The S polypeptide or fragment has at least 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identity to SEQ ID NO: 104 or 105 and comprises an amino acid sequence containing one of the following sets of amino acid substitutions relative to SEQ ID NO: 104 or 105: (1) D980P, V982P, F812P, A887P, A894P, and A937P; (2) K981P, V982P, F812P; A887P, A894P, and A937P, (3) D980P, V982P, F812P, A887P, A894P, A937P, R677G, R678S, and R680S, (4) K981P, V982P, F812P, A887P, A894P, A937P, R677G, R678S, and R680S, or (5) D980P, K981P, V982P, F812P, A887P, A894P, A937P, R677G, R678S, and R680S.

[0096] Another aspect of the disclosure features a composition or medical preparation including a first RNA encoding a first SARS-CoV-2 S polypeptide or immunogenic fragment thereof and a second RNA encoding a second SARS-CoV-2 S polypeptide or immunogenic fragment thereof, wherein (a) the first SARS-CoV-2 S polypeptide or fragment comprises an amino acid sequence having at least 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identity to SEQ ID NO: 70 and including the following substitutions relative to SEQ ID NO: 70: D982P, V984P, F814P, A889P, A896P, A939P, R679G, R680S, and R682S; and (b) the second SARS-CoV-2 The S polypeptide or fragment has at least 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identity to SEQ ID NO: 104 or 105 and comprises an amino acid sequence containing one of the following sets of amino acid substitutions relative to SEQ ID NO: 104 or 105: (1) D980P, V982P, F812P, A887P, A894P, and A937P; (2) K981P, V982P, F812P; A887P, A894P, and A937P, (3) D980P, V982P, F812P, A887P, A894P, A937P, R677G, R678S, and R680S, (4) K981P, V982P, F812P, A887P, A894P, A937P, R677G, R678S, and R680S, or (5) D980P, K981P, V982P, F812P, A887P, A894P, A937P, R677G, R678S, and R680S.

[0097] Another aspect of the disclosure features a composition or medical preparation including a first RNA encoding a first SARS-CoV-2 S polypeptide or immunogenic fragment thereof and a second RNA encoding a second SARS-CoV-2 S polypeptide or immunogenic fragment thereof, wherein (a) the first SARS-CoV-2 S polypeptide or fragment comprises an amino acid sequence having at least 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identity to SEQ ID NO: 70 and including the following substitutions relative to SEQ ID NO: 70: K983P, V984P, F814P, A889P, A896P, A939P, R679G, R680S, and R682S; and (b) the second SARS-CoV-2 The S polypeptide or fragment has at least 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identity to SEQ ID NO: 104 or 105 and comprises an amino acid sequence containing one of the following sets of amino acid substitutions relative to SEQ ID NO: 104 or 105: (1) D980P, V982P, F812P, A887P, A894P, and A937P; (2) K981P, V982P, F812P; A887P, A894P, and A937P, (3) D980P, V982P, F812P, A887P, A894P, A937P, R677G, R678S, and R680S, (4) K981P, V982P, F812P, A887P, A894P, A937P, R677G, R678S, and R680S, or (5) D980P, K981P, V982P, F812P, A887P, A894P, A937P, R677G, R678S, and R680S.

[0098] Another aspect of the disclosure features a composition or medical preparation including a first RNA encoding a first SARS-CoV-2 S polypeptide or immunogenic fragment thereof and a second RNA encoding a second SARS-CoV-2 S polypeptide or immunogenic fragment thereof, wherein (a) the first SARS-CoV-2 S polypeptide or fragment comprises an amino acid sequence having at least 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identity to SEQ ID NO: 70 and including the following substitutions relative to SEQ ID NO: 70: D982P, K983P, V984P, F814P, A889P, A896P, A939P, R679G, R680S, and R682S; and (b) the second SARS-CoV-2 The S polypeptide or fragment has at least 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identity to SEQ ID NO: 104 or 105 and comprises an amino acid sequence containing one of the following sets of amino acid substitutions relative to SEQ ID NO: 104 or 105: (1) D980P, V982P, F812P, A887P, A894P, and A937P; (2) K981P, V982P, F812P; A887P, A894P, and A937P, (3) D980P, V982P, F812P, A887P, A894P, A937P, R677G, R678S, and R680S, (4) K981P, V982P, F812P, A887P, A894P, A937P, R677G, R678S, and R680S, or (5) D980P, K981P, V982P, F812P, A887P, A894P, A937P, R677G, R678S, and R680S.

[0099] The present disclosure provides, among other things, the insight that incorporation of the D985P mutation rather than the K986P mutation can improve protein expression and / or immunogenicity (e.g., improve neutralization response). In some embodiments, incorporation of D985P rather than K986P can provide such improvement when combined with one or more other proline mutations (e.g., one or more proline mutations disclosed herein). In some embodiments, incorporation of D985P rather than K986P can provide such improvement when combined with V987P (e.g., one or more proline mutations disclosed herein). In some embodiments, incorporation of D985P rather than K986P can provide such improvement when combined with one or more (e.g., all) of F817P, A892P, A899P, A942P, and V987P.

[0100] In some embodiments, the present disclosure provides the insight that RNAs encoding SARS-CoV-2 S proteins comprising one or more proline mutations (e.g., one or more of the proline mutations disclosed herein and / or a combination of proline mutations) and a mutated furin cleavage site can provide an improved immune response (e.g., an improved immune response compared to a similar or identical construct comprising an intact furin cleavage site).

[0101] In some embodiments, the first RNA and the second RNA each include a modified nucleoside in place of a uridine. In some embodiments, the first RNA and the second RNA each include a modified uridine in place of every uridine. In some embodiments, the first RNA and the second RNA each include N1-methyl-pseudouridine (m1ψ) in place of every uridine.

[0102] In some embodiments, the first RNA and the second RNA each comprise a 5' cap. In some embodiments, the 5' cap is m2 7,3’- O Gppp(m1 2’-O )Contains ApG.

[0103] In some embodiments, the first RNA and the second RNA each comprise a 5'-UTR that is or comprises a modified human alpha-globin 5'-UTR. In some embodiments, the first RNA and the second RNA each comprise a 5'-UTR that comprises the nucleotide sequence of SEQ ID NO: 12, or a nucleotide sequence that is at least 99%, 98%, 97%, 96%, 95%, 90%, 85%, or 80% identical to the nucleotide sequence of SEQ ID NO: 12.

[0104] In some embodiments, the first RNA and the second RNA each comprise a 3'-UTR that is or includes a first sequence from a split amino-terminal enhancer (AES) messenger RNA and a second sequence from a mitochondrially encoded 12S ribosomal RNA. In some embodiments, the first RNA and the second RNA each comprise a 3'-UTR that includes the nucleotide sequence of SEQ ID NO: 13, or a nucleotide sequence that is at least 99%, 98%, 97%, 96%, 95%, 90%, 85%, or 80% identical to the nucleotide sequence of SEQ ID NO: 13.

[0105] In some embodiments, the first RNA and the second RNA each comprise a polyA sequence. In some embodiments, the first RNA and the second RNA each comprise a polyA sequence comprising at least 100 nucleotides. In some embodiments, the first RNA and the second RNA each comprise 30 adenine nucleotides followed by 70 adenine nucleotides, wherein the 30 adenine nucleotides and the 70 adenine nucleotides are separated by a linker sequence. In some embodiments, the first RNA and the second RNA each comprise a polyA sequence comprising or consisting of the nucleotide sequence of SEQ ID NO: 14, or a nucleotide sequence that is at least 99%, 98%, 97%, 96%, 95%, 90%, 85%, or 80% identical to the nucleotide sequence of SEQ ID NO: 14.

[0106] In some embodiments, the first RNA and the second RNA are each formulated or to be formulated for intramuscular administration.

[0107] In some embodiments, the first RNA and the second RNA are each formulated or to be formulated as particles. In some embodiments, the first RNA and the second RNA are each formulated as lipid nanoparticles (LNPs) or lipoplex (LPX) particles. In some embodiments, the LNPs comprise ((4-hydroxybutyl)azanediyl)bis(hexane-6,1-diyl)bis(2-hexyldecanoate), 2-[(polyethylene glycol)-2000]-N,N-ditetradecylacetamide, 1,2-distearoyl-sn-glycero-3-phosphocholine, and cholesterol.

[0108] In some embodiments, the first RNA and the second RNA are formulated in separate LNPs. In some embodiments, the first RNA and the second RNA are formulated in the same LNP. In some embodiments, lipoplex particles can be obtained by mixing RNA with liposomes.

[0109] In some embodiments, the first RNA and the second RNA are each mRNA. In some embodiments, the first RNA and the second RNA are each saRNA.

[0110] In some embodiments, the composition or medical preparation is a pharmaceutical composition. In some embodiments, the composition or medical preparation is a vaccine.

[0111] In some embodiments, the pharmaceutical composition further comprises one or more pharmaceutically acceptable carriers, diluents, and / or excipients.

[0112] In another aspect, the present disclosure provides a method of inducing an immune response in a subject, the method comprising inducing an immune response in the subject by administering to the subject a composition or pharmaceutical preparation described herein.

[0113] In some embodiments, the SARS-CoV-2 S polypeptide comprises an amino acid sequence that does not contain a D985P substitution relative to SEQ ID NO: 1, an amino acid sequence that does not contain a D982P substitution relative to SEQ ID NO: 69 or SEQ ID NO: 70, or an amino acid sequence that does not contain a D980P substitution relative to SEQ ID NO: 104 or SEQ ID NO: 105.

[0114] In some embodiments, the method further comprises administering a second RNA encoding a second SARS-CoV-2 S polypeptide or immunogenic fragment thereof, wherein the second SARS-CoV-2 S polypeptide or immunogenic fragment is an omicron-variant SARS-CoV-2 S polypeptide that is not the BA.1 omicron variant.

[0115] In some embodiments, the method further comprises administering a second, different RNA encoding a second SARS-CoV-2 S polypeptide or immunogenic fragment thereof, wherein the second SARS-CoV-2 S polypeptide or fragment is selected from the SARS-CoV-2 S polypeptides or fragments described herein.

[0116] Another aspect of the present disclosure provides a method of inducing an immune response in a subject, the method comprising inducing an immune response in the subject by administering to the subject a composition or pharmaceutical formulation described herein. In some embodiments, the SARS-CoV-2 S polypeptide comprises an amino acid sequence that does not contain a D985P substitution relative to SEQ ID NO: 1, an amino acid sequence that does not contain a D982P substitution relative to SEQ ID NO: 69 or SEQ ID NO: 70, or an amino acid sequence that does not contain a D980P substitution relative to SEQ ID NO: 104 or SEQ ID NO: 105.

[0117] In some embodiments, the method further comprises administering a second composition or medical preparation, wherein the second composition or medical preparation comprises RNA encoding a SARS-CoV-2 S polypeptide or immunogenic fragment of an omicron variant that is not the BA.1 omicron variant.

[0118] In some embodiments, the method further comprises administering a second composition or medical preparation comprising a third RNA encoding a third SARS-CoV-2 S polypeptide or immunogenic fragment thereof, and a fourth RNA encoding a fourth SARS-CoV-2 S polypeptide or immunogenic fragment thereof.

[0119] In some embodiments, the third RNA encodes a SARS-CoV-2 S polypeptide or immunogenic fragment thereof that is a first or second SARS-CoV-2 S polypeptide or immunogenic fragment thereof recited in any one of claims 24-102, and the third RNA encodes a SARS-CoV-2 S polypeptide or fragment that is different from the first SARS-CoV-2 S polypeptide or fragment encoded by the first RNA and / or different from the second SARS-CoV-2 S polypeptide or fragment encoded by the second RNA.

[0120] In some embodiments, the fourth RNA encodes a SARS-CoV-2 S polypeptide or immunogenic fragment thereof that is a first or second SARS-CoV-2 S polypeptide or immunogenic fragment thereof recited in any one of claims 24-102, and the fourth RNA encodes a SARS-CoV-2 S polypeptide or fragment that is different from the first SARS-CoV-2 S polypeptide or fragment encoded by the first RNA and / or different from the second SARS-CoV-2 S polypeptide or fragment encoded by the second RNA.

[0121] In some embodiments, the third RNA encodes a SARS-CoV-2 S polypeptide or immunogenic fragment thereof that is a first or second SARS-CoV-2 S polypeptide or immunogenic fragment thereof recited in any one of claims 24-102, and the third RNA encodes a SARS-CoV-2 S polypeptide or fragment that is different from the first SARS-CoV-2 S polypeptide or fragment encoded by the first RNA and that is different from the second SARS-CoV-2 S polypeptide or fragment encoded by the second RNA.

[0122] In some embodiments, the fourth RNA encodes a SARS-CoV-2 S polypeptide or immunogenic fragment thereof that is a first or second SARS-CoV-2 S polypeptide or immunogenic fragment thereof recited in any one of claims 24-102, and the fourth RNA encodes a SARS-CoV-2 S polypeptide or fragment that is different from the first SARS-CoV-2 S polypeptide or fragment encoded by the first RNA and / or different from the second SARS-CoV-2 S polypeptide or fragment encoded by the second RNA.

[0123] the third RNA encodes a SARS-CoV-2 S polypeptide or immunogenic fragment thereof that is a first or second SARS-CoV-2 S polypeptide or immunogenic fragment thereof recited in any one of claims 24 to 102, wherein the third RNA encodes a SARS-CoV-2 S polypeptide or fragment that is different from the first SARS-CoV-2 S polypeptide or fragment encoded by the first RNA and that is different from the second SARS-CoV-2 S polypeptide or fragment encoded by the second RNA; the fourth RNA encodes a SARS-CoV-2 S polypeptide or immunogenic fragment thereof that is a first or second SARS-CoV-2 S polypeptide or immunogenic fragment thereof recited in any one of claims 24 to 102, wherein the fourth RNA encodes a SARS-CoV-2 S polypeptide or fragment that is different from the first SARS-CoV-2 S polypeptide or fragment encoded by the first RNA and that is different from the second SARS-CoV-2 S polypeptide or fragment encoded by the second RNA.

[0124] In some embodiments, each of the first, second, third, and fourth RNAs encodes a different SARS-CoV-2 S polypeptide or immunogenic fragment thereof.

[0125] In another aspect, a monovalent vaccine described herein can be administered in conjunction with a bivalent vaccine described herein. For example, in some embodiments, a method for inducing an immune response comprises administering to a subject (i) a composition or medical preparation described herein comprising RNA encoding a SARS-CoV-2 S polypeptide or immunogenic fragment thereof described herein, and (ii) a composition or medical preparation comprising at least a first RNA encoding a first SARS-CoV-2 S polypeptide or immunogenic fragment thereof described herein and a second RNA encoding a second SARS-CoV-2 S polypeptide or immunogenic fragment thereof. [DDM1] In some embodiments, the monovalent vaccine and the bivalent vaccine can be administered at least three weeks apart, including, for example, at least four weeks, at least five weeks, at least six weeks, at least seven weeks, at least eight weeks, or more. In some embodiments, the monovalent vaccine and the bivalent vaccine can be administered at least three months apart, including, for example, at least four months, at least five months, at least six months, at least seven months, at least eight months, or more. In some embodiments, the monovalent and bivalent vaccines may be administered in different arms in a single session, hi some embodiments, the monovalent and bivalent vaccines may be administered in a single injection as a trivalent vaccine (e.g., the monovalent and bivalent vaccines are mixed together prior to administration). [Brief explanation of the drawings]

[0126] [Figure 1] Overview of the S protein organization of the SARS-CoV-2 S protein. The sequence within the S1 subunit consists of a signal sequence (SS) and a receptor-binding domain (RBD), which is a key subunit within the S protein involved in binding to the human cell receptor ACE2. The S2 subunit contains the S2 protease cleavage site (S2'), followed by a fusion peptide (FP) for membrane fusion, heptad repeats (HR1 and HR2) with a central helix (CH) domain, a transmembrane domain (TM), and a cytoplasmic tail (CT). [Figure 2]Exemplary SARS-CoV-2 Vaccine Constructs. Based on the complete and wild-type S protein, we designed different constructs encoding (1) the complete protein with mutations close to the first heptad repeat (HRP1) containing stabilizing mutations that preserve the neutralization-sensitive site, (2) the S1 domain, or (3) only the RB domain (RBD). To further stabilize the protein fragment, a fibritin domain (F) was fused to the C-terminus. All constructs begin with a signal peptide (SP) to ensure Golgi transport to the plasma membrane. [Figure 3] General structure of a particular RNA vaccine. Schematic of the general structure of a particular RNA vaccine with a 5' cap, 5' and 3' untranslated regions, coding sequence with essential secretory signal peptide and GS linker, and poly(A) tail. Note that individual elements are not drawn exactly to scale compared to their respective sequence lengths. UTR = untranslated region, sec = secretory signal peptide, RBD = receptor binding domain, GS = glycine-serine linker. [Figure 4] General structure of a particular RNA vaccine. Schematic of the general structure of a particular RNA drug substance with a 5' cap, 5' and 3' untranslated regions, coding sequence with essential secretory signal peptide and GS linker, and poly(A) tail. Note that individual elements are not drawn exactly to scale compared to their respective sequence lengths. GS = glycine-serine linker, UTR = untranslated region, Sec = secretory signal peptide, RBD = receptor binding domain. [Figure 5]General structure of a particular RNA vaccine. Schematic of the general structure of an RNA vaccine with a 5' cap, 5' and 3' untranslated regions, coding sequence for Venezuelan equine encephalitis virus (VEEV) RNA-dependent RNA polymerase replicase, and SARS-CoV-2 antigen with a unique secretory signal peptide and GS linker, and poly(A) tail. Note that individual elements are not drawn exactly to scale compared to their respective sequence lengths. UTR = untranslated region, Sec = secretory signal peptide, RBD = receptor-binding domain, GS = glycine-serine linker. [Figure 6] Schematic diagram of the SARS-CoV-2 S protein organization and constructs for SARS-CoV-2 vaccine development. Based on the wild-type S protein, two different transmembrane-anchored RBD-based vaccine constructs were designed, encoding RBD fragments fused to the T4 fibritin trimerization domain (F) and a unique transmembrane domain (TM). Construct (1) starts with the SARS-CoV-2-S signal peptide (SP, AA 1-19 of the S protein), whereas construct (2) starts with the human Ig heavy chain signal peptide (huSec) to ensure Golgi transport to the plasma membrane. [Figure 7] Anti-S protein IgG responses at days 6, 14, and 21 after immunization with LNP-C12-formulated modRNA encoding a transmembrane-anchored RBD-based vaccine construct. BALB / c mice were immunized IM once with 4 μg of LNP-C12-formulated transmembrane-anchored RBD-based vaccine construct (a surrogate for BNT162b3c / BNT162b3d). At days 6, 14, and 21 after immunization, animals were bled, and serum samples were analyzed for total anti-S1 (left) and anti-RBD (right) antigen-specific immunoglobulin G (IgG), as measured via ELISA. Different serum dilutions were included on the graph for days 6 (1:50), 14 (1:300), and 21 (1:900). Each point on the graph represents one mouse, and all mouse samples were measured in duplicate (group size n = 8, mean + SEM included in each group). [Figure 8]Neutralization of SARS-CoV-2 pseudovirus on days 6, 14, and 21 after immunization with LNP-C12-formulated modRNA encoding a transmembrane-anchored RBD-based vaccine construct. BALB / c mice were immunized IM once with 4 μg of LNP-C12-formulated transmembrane-anchored RBD-based vaccine construct (a surrogate for BNT162b3c / BNT162b3d). On days 6, 14, and 21 after immunization, animals were bled and serum was tested for SARS-CoV-2 pseudovirus neutralization. The graph shows the pVN50 serum dilution (50% reduction in infectious events compared to a positive control without serum). One point on the graph represents one mouse. All mouse samples were measured in duplicate. Group size n=8. The mean + SEM is indicated by the horizontal bar with whiskers for each group. LLOQ, lower limit of quantitation. ULOQ, upper limit of quantitation. [Figure 9] Pseudovirus neutralization titers (pVNT50) of sera collected 21 days after the second dose and 1 month after the third dose of BNT162b2 against VSV-SARS-CoV-2-S pseudoviruses bearing the Wuhan Hu-1 reference or Omicron BA.1 lineage spike protein were measured. N = 19–20 sera from immunized subjects collected either 21 days after the second BNT162b2 dose or 1 month after the third BNT162b2 dose were tested. For values ​​below the limit of detection (LOD, 10), the LOD / 2 value was plotted. Group GMTs (values ​​above the bars) with 95% confidence intervals are shown. [Figure 10] CD8+ T cell epitopes in the BNT162b2 vaccine remain largely unaffected by Omicron variant mutations. Figure 1 shows the number of previously identified MHC-I epitopes affected in various variants of concern (VOCs). Approximately 80% of previously identified CD8+ epitopes are unaffected by Omicron BA.1 variant mutations, suggesting that two doses of BNT162b2 can still induce protection against severe disease. [Figure 11]Neutralization of Omicron BA.1 after two doses of BNT162b2 and a variant-specific booster. Shown is neutralization of Omicron BA.1 variants from serum of patients who received two doses of BNT162b2 and (i) a third booster dose of BNT162b2, or (ii) a third booster dose of RNA encoding a spike protein with an alpha or delta variant mutation, or a third booster dose of both a spike protein containing an alpha mutation and a spike protein containing a delta mutation. These values ​​are derived from separate neutralization GMTs from pseudovirus testing. Also shown is a schematic illustrating the process for developing new SARS-CoV-2 variant-specific vaccines. [Figure 12] Longitudinal analysis of neutralizing antibody responses to VSV-SARS-CoV-2-S pseudoviruses bearing Wuhan or Omicron BA.1 variant spike proteins in a subset of study participants. Sera from n=9 participants were tested, extracted 21 days after dose 2, before dose 3, 1 month after dose 3, and 3 months after dose 3. Each serum was tested in duplicate, and individual geometric mean 50% pseudovirus neutralization titers (GMTs) were calculated. For values ​​below the limit of detection (LOD), a value of LOD / 2 was assigned. Group GMTs (table values) and 95% confidence intervals per time point are shown. [Figure 13] Analysis of HLA class I cell epitope conservation between the Wuhan and Omicron BA.1 variants. HLA class I-restricted spike protein epitopes with T cell reactivity identified based on their recognition by CD8+ T cells and reported in the IEDB (n = 244) are plotted by their position (top) along the spike protein (bottom). Epitope adaptation is mapped by the amino acid position of the epitope's center. Epitopes conserved in both variants are marked in light gray (n = 208), while epitopes spanning the Omicron BA.1 mutation site are marked in dark gray (n = 36). NTD = N-terminal domain; RBD = receptor-binding domain; FCS = furin cleavage site. The S1 and S2 regions of the spike protein are indicated. [Figure 14]Schematic of an exemplary vaccination regimen. [Figure 15] Cohorts, sample collection, and experimental settings for characterizing immune responses in Omicron BA.1 breakthrough cases. Blood samples were collected from four cohorts: Omicron-naive individuals vaccinated two or three times with BNT162b2, and individuals vaccinated two or three times with BNT162b2 who subsequently developed breakthrough infection with Omicron BA.1. PBMCs and serum were isolated within the Omicron-naive cohort at the indicated time points after most recent vaccination. For the convalescent cohort, the time from most recent vaccination to Omicron BA.1 infection and infection in PBMC and serum isolation are shown (all values ​​are specified as median ranges). Serum neutralization capacity was assessed using pseudovirus and live virus neutralization tests. SARS-CoV-2 spike-specific BMEM cells were assessed by a flow cytometry-based B cell phenotyping assay using bulk PBMCs. N / A, not applicable. [Figure 16-1]Omicron BA.1 breakthrough infection in individuals vaccinated twice and three times with BNT162b2 induces broad neutralization of Omicron BA.1, BA.2, and other VOCs. Sera were collected 22 days after the second dose from a twice-vaccinated individual (BNT162b22) (open circles), 28 days after the third dose from a three-vaccinated individual (BNT162b23) (filled circles), 46 days post-infection from a twice-vaccinated individual with Omicron BA.1 breakthrough infection (BNT162b22 + Omi) (open triangles), and 44 days post-infection from a three-vaccinated individual with Omicron BA.1 breakthrough infection (BNT162b23 + Omi) (filled triangles). Sera were tested in duplicate. (A) 50% pseudovirus neutralization (pVN50) geometric mean titers (GMTs) are shown. (B) 50% virus neutralization (VN50) GMTs are shown. (C) Geometric mean ratios of SARS-CoV-2 variants of concern (VOC) and Wuhan VN50 GMTs are shown. For values ​​below the limit of detection (LOD), the LOD / 2 value was plotted. Values ​​above the violin plot represent group GMTs. Wuhan neutralization group GMTs were compared to titers against the indicated variants and SARS-CoV-1 using a nonparametric Friedman test with Dunn's multiple comparisons correction. Multiplicity-adjusted p-values ​​are shown. (A) pVN50 GMTs against Wuhan, VOC, and SARS-CoV-1 pseudoviruses. (B) VN50 GMTs against live SARS-CoV-2 Wuhan, beta, delta, and omicron BA.1. (C) Group geometric mean ratios with 95% confidence intervals for all cohorts shown in (B). [Figure 16-2] Same as above. [Figure 17]BMEM cells from individuals vaccinated two and three times with BNT162b2 broadly recognize VOCs and are further boosted by Omicron BA.1 breakthrough infection. PBMC samples from a twice-vaccinated individual (BNT162b22) 22 days after the second dose (open squares) and 5 months after the second dose (open circles), a sample from a three-vaccinated individual (BNT162b23) 84 days after the third dose (filled circles), a sample from a twice-vaccinated, Omicron BA.1 breakthrough-infected individual (BNT162b22+Omi) 46 days after infection (open triangles), and a sample from a three-vaccinated, Omicron BA.1 breakthrough-infected individual (BNT162b23+Omi) 44 days after infection (filled triangles) were analyzed by flow cytometry for SARS-CoV-2-specific BMEM cell (BMEM-CD3-CD19+CD20+IgD-CD38 intermediate / low) frequency via B cell bait staining. (A) Schematic of one-dimensional staining of BMEM cells with fluorescent dye-labeled SARS-CoV-2 S protein tetramer bait, enabling differentiation of variant recognition. The frequencies of Wuhan or VOC full-length S protein- (B) and RBD- (C) specific BMEM cells in the four populations were analyzed. The frequencies of variant-specific BMEM cells were normalized to the Wuhan frequencies for S protein (D) and RBD- (E) binding. (F) The frequency ratio of RBD protein-specific BMEM cells to full-length S protein-specific BMEM cells is shown. [Figure 18-1]Omicron BA.1 breakthrough infection in individuals vaccinated two and three times with BNT162b2 primarily boosts BMEM against conserved epitopes that are extensively shared between the Wuhan S protein and other VOCs, rather than strictly Omicron S-specific epitopes. PBMC samples from a twice-vaccinated individual (BNT162b22) 22 days after the second dose (open squares) and 5 months after the second dose (open circles), a sample from a three-vaccinated individual (BNT162b23) 84 days after the third dose (filled circles), a sample from a twice-vaccinated, Omicron BA.1 breakthrough-infected individual (BNT162b22+Omi) 46 days after infection (open triangles), and a sample from a three-vaccinated, Omicron BA.1 breakthrough-infected individual (BNT162b23+Omi) 44 days after infection (filled triangles) were analyzed by flow cytometry for SARS-CoV-2-specific memory B cell (BMEM-CD3-CD19+CD20+IgD-CD38 intermediate / low) frequency via B cell bait staining (schematic diagram shown in (A)). (B) Representative flow diagrams of Omicron BA.1 and Wuhan S protein binding and RBD binding for each of the four populations studied. The frequencies of BMEM binding to Omicron BA.1, Wuhan, or both (shared) are shown for the full-length S protein in (C), and the frequencies of RBD are shown for Omicron BA.1-experienced and Omicron BA.1-naive individuals vaccinated with BNT162b2 two and three times in (D). (E) Venn diagram visualizing the combinatorial (Boolean) gating strategy to identify all four variants (All 4+ve) that recognize only the Omicron BA.1 S protein (Omicron-only) or only the Wuhan S protein (Wuhan-only) and cross-reactive BMEMs that simultaneously recognize BMEM. The frequencies of these three BMEM subgroups were compared for the full-length S protein (F) and RBD (G) in the four different populations studied.RBD variant recognition patterns by BMEMs were assessed through a Boolean flow cytometry gating strategy, and the frequency of recognition of Wuhan and / or variant RBD combinations is displayed in (H) for all Omicron convalescent subjects (pooled 2- and 3-dose vaccinated individuals, n=13). (I) Conserved sites within the RBD domain recognized by RBD-specific BMEMs after Omicron BA.1 breakthrough infection. Mean values ​​are shown in C, D, F, and G. n=1 individuals per group. [Figure 18-2] Same as above. [Figure 18-3] Same as above. [Figure 18-4] Same as above. [Figure 19] Omicron BA.1 breakthrough infection in individuals vaccinated with other approved COVID-19 vaccines or combination regimens resulted in immune sera that broadly neutralized SARS-CoV-1 in addition to Omicron BA.1, BA.2, and other VOCs. Sera were collected from 10 individuals vaccinated with other approved COVID-19 vaccines or combination regimens at a median of 43 days postinfection (gray diamonds). Sera were tested in duplicate, and individual 50% pseudovirus neutralization (pVN50) geometric mean titers (GMTs) against SARS-CoV-2 Wuhan, alpha, beta, delta, and Omicron BA.1 and BA.2 variants, as well as SARS-CoV-1, were plotted. For values ​​below the limit of detection (LOD), the LOD / 2 value was plotted. Values ​​above the violin plot represent group GMTs. The Wuhan neutralization group GMTs were compared with titers against the indicated variants and SARS-CoV-1 using a nonparametric Friedman test with Dunn's multiple comparison correction. Multiplicity-adjusted p-values ​​are shown. Approved vaccines include AZD1222, BNT162b2 (in some embodiments, as part of a 4-dose series), Ad26.COV2.S, mRNA-1273 (administered as a 2-dose or 3-dose series), and combinations thereof. [Figure 20-1]50% neutralization titers of sera collected one month after the fourth dose of BNT162b2 or an Omicron-specific booster. Subjects who previously received two doses of BNT162b2 and a third (booster) dose of BNT162b2 (30 ug) received a dose (30 ug) of either (i) RNA encoding a SARS-CoV-2 S protein from an Omicron variant (e.g., as described herein (referred to herein as an "Omicron BA.1-specific RNA vaccine"), or (ii) BNT162b2 as the fourth (booster) dose. Subject sera were collected one month after administration of the fourth (booster) dose. Group GMTs (values ​​above the bars) with 95% confidence intervals are shown. "b2" refers to sera from subjects who received a Wuhan-specific RNA vaccine as the fourth (booster) dose of BNT162b2. "OMI" refers to sera from subjects who received an Omicron BA.1-specific fourth (booster) dose. "FFRNT" refers to serum from subjects who received the fourth dose of the Omicron BA.1-specific RNA vaccine. Also shown are the fold change in titer from before to after the fourth dose (pre-vaccination / post-vaccination fold increase), and the ratio of the geometric mean ratio (GMR) and geometric mean fold increase (GMFR) observed in subjects who received the fourth dose of the Omicron BA.1-specific RNA vaccine as the fourth dose compared to subjects who received BNT162b2 as the fourth dose of the Omicron BA.1-specific RNA vaccine. "FFRNT" refers to fluorescent focus reduction neutralization assay. SARS-CoV-2 from the variants indicated in the figure. Neutralization data were obtained using the FFRNT assay with virus particles containing the S protein. (A) Comparison of neutralizing antibody titers against SARS-CoV-2-S pseudoviruses containing SARS-CoV-2 S proteins with mutations characteristic of the Omicron BA.1 variant, excluding sera from subjects previously or currently infected with SARS-CoV-2. (B) Comparison of neutralizing antibody titers against SARS-CoV-2 pseudoviruses containing SARS-CoV-2 S proteins with mutations characteristic of the Omicron BA.1 variant in sera from populations including subjects previously or currently infected with SARS-CoV-2 (determined by antigen assay or PCR assay, respectively).(C) Comparison of neutralizing antibody titers against SARS-CoV-2 pseudoviruses containing the SARS-CoV-2 S protein from the Wuhan strain, excluding sera from subjects previously or currently infected with SARS-CoV-2. (D) Comparison of neutralizing antibody titers against SARS-CoV-2 pseudoviruses containing the SARS-CoV-2 S protein from the Wuhan strain in sera from populations including individuals previously or currently infected with SARS-CoV-2 (determined by antigen assay or PCR assay, respectively). (E) Comparison of neutralizing antibody titers against SARS-CoV-2 pseudoviruses containing the SARS-CoV-2 S protein with a mutation characteristic of the delta variant, excluding sera from subjects previously or currently infected with SARS-CoV-2. (F) Comparison of neutralizing antibody titers (determined by antigen assay or PCR assay, respectively) against SARS-CoV-2 pseudoviruses containing SARS-CoV-2 proteins with mutations characteristic of the delta variant in serum from populations including subjects previously or currently infected with SARS-CoV-2. [Figure 20-2] Same as above. [Figure 20-3] Same as above. [Figure 20-4] Same as above. [Figure 20-5] Same as above. [Figure 20-6] Same as above. [Figure 21]Neutralization of SARS-CoV-2 pseudovirus 7 days after immunization with modRNA encoding variant-specific S proteins. Mice were immunized twice with LNP-formulated vaccines containing (i) BNT162b2 (encoding the SARS-CoV-2 S protein from the Wuhan strain), (ii) RNA encoding a SARS-CoV-2 S protein with mutations characteristic of the Omicron BA.1 variant (Omi), (iii) RNA encoding an S protein with mutations characteristic of the delta variant, (iv) a combination of BNT162b2 and RNA encoding a protein with mutations characteristic of the Omicron BA.1 variant (B2 + Omi), or (v) RNA encoding a SARS-CoV-2 S protein with mutations characteristic of the delta variant and RNA encoding a SARS-CoV-2 S protein with mutations characteristic of the Omicron BA.1 variant (Delta + Omi). Seven days after the second immunization, animals were bled and sera were tested for neutralization of SARS-CoV-2-S pseudoviruses containing SARS-CoV-2 S protein from the Wuhan strain or SARS-CoV-2 S protein with mutations characteristic of the beta, delta, or omicron BA.1 variants. Graphs show the pVN50 serum dilution (50% reduction in infectious events compared to the positive control without serum). Each point on the graph represents one mouse. All mouse samples were measured in duplicate. The mean + SEM is indicated by the whiskered horizontal bar for each group. LLOD, lower limit of detection. ULOD, upper limit of detection. [Figure 22-1]RNA encoding a SARS-CoV-2 S protein with mutations characteristic of the beta variant increases neutralizing antibody titers against SARS-CoV-2 when administered to patients who had previously received two doses of a vaccine encoding the Wuhan strain of SARS-CoV-2 S protein. Subjects who had previously received two doses of an RNA vaccine encoding the Wuhan strain of SARS-CoV-2 S protein were administered a third and fourth dose of an RNA vaccine encoding a SARS-CoV-2 S protein with mutations characteristic of the beta variant. Neutralizing antibody titers were measured before administration of an RNA vaccine encoding the Wuhan strain of SARS-CoV-2 S protein (D1-PreVax), one month after administration of the second dose of an RNA vaccine encoding the Wuhan strain of SARS-CoV-2 S protein (M1PD2), one month after administration of the third dose of an RNA vaccine encoding a SARS-CoV-2 S protein with mutations characteristic of the SARS-CoV-2 beta variant, and one month after administration of the fourth dose of an RNA vaccine encoding a SARS-CoV-2 S protein with mutations characteristic of the SARS-CoV-2 beta variant. The third and fourth doses were administered one month apart. GMFR refers to the geometric mean fold increase and is a measure of the increase in neutralizing antibody titers since the previous vaccine dose (e.g., GMFR after dose 2 (PD2) is a measure of the increase in neutralizing antibody titers compared to before any vaccine administration (pre-vax)). (A) Neutralizing antibody titers measured in a virus neutralization assay using virus particles containing the Wuhan strain SARS-CoV-2 S protein. (B) Neutralizing antibody titers measured in a virus neutralization assay using virus particles containing the SARS-CoV-2 S protein with mutations characteristic of beta variants. [Figure 22-2] Same as above. [Figure 23-1]50% neutralization titers of sera collected 7 days after the fourth dose of BNT162b2, Omicron BA.1-specific booster, or bivalent vaccine. Subjects who previously received two doses of BNT162b2 (30ug) and a third (booster) dose of BNT162b2 (30ug) may receive a second booster vaccine containing: (i) a 30ug dose of BNT162b2 (encoding the SARS-CoV-2 S protein from the Wuhan strain); (ii) a 60ug dose of BNT162b2; (iii) a 30ug dose of RNA encoding a SARS-CoV-2 S protein with a mutation characteristic of the Omicron BA.1 variant (e.g., as described herein (referred to herein as an "Omicron BA.1-specific RNA vaccine"); (iii) a 60ug dose of RNA encoding a SARS-CoV-2 S protein with a mutation characteristic of the Omicron BA.1 variant; (iv) 15ug of BNT162b2 and 15ug of SARS-CoV-2 S protein containing a mutation characteristic of the Omicron BA.1 variant. Subjects received a 30ug dose of a bivalent vaccine containing BNT162b2 and RNA encoding the S protein, or (v) a 60ug dose of a bivalent vaccine containing 30ug of BNT162b2 and 30ug of RNA encoding a SARS-CoV-2 S protein containing mutations characteristic of the Omicron BA.1 variant. Geometric mean ratios (GMRs) of titers in serum from subjects were collected 7 days after the fourth dose. "b2" refers to serum from subjects who received the Wuhan-specific RNA vaccine as the fourth dose of BNT162b2. "OMI" refers to serum from subjects who received the Omicron BA.1-specific fourth dose. "Bivalent" refers to serum from subjects who received BNT162b2 and a SARS-CoV-2 S protein containing mutations characteristic of the Omicron BA.1 variant as the fourth dose. Refers to serum from subjects administered a composition containing RNA encoding the S protein. Also shown is the fold increase in titer from before administration of the fourth dose to 7 days after administration of the fourth dose (*fold increase). "FFRNT" refers to fluorescent focus reduction neutralization assay. Neutralization data were obtained using the FFRNT assay with viral particles containing SARS-CoV-2 S proteins with mutations characteristic of the variants shown in the figure. LLOQ refers to the lower limit of quantitation, and ULOQ refers to the upper limit of quantitation.(A) Comparison of neutralizing antibody titers against SARS-CoV-2 pseudoviruses containing SARS-CoV-2 S proteins with mutations characteristic of the Omicron BA.1 variant, excluding sera from subjects previously or currently infected with SARS-CoV-2. (B) Comparison of neutralizing antibody titers against SARS-CoV-2 pseudoviruses containing SARS-CoV-2 S proteins with mutations characteristic of the Omicron BA.1 variant in sera from populations including subjects previously or currently infected with SARS-CoV-2 (as determined by antibody testing or PCR assay, respectively). (C) Comparison of neutralizing antibody titers against SARS-CoV-2 pseudoviruses containing the Wuhan strain SARS-CoV-2 S protein, excluding sera from subjects previously or currently infected with SARS-CoV-2. (D) Comparison of neutralizing antibody titers against SARS-CoV-2 pseudoviruses containing the Wuhan strain of SARS-CoV-2 S protein in sera from populations including individuals previously infected or currently infected with SARS-CoV-2. (E) Comparison of neutralizing antibody titers against SARS-CoV-2 pseudoviruses containing the SARS-CoV-2 S protein with mutations characteristic of the delta variant. Sera from subjects previously infected or currently infected with SARS-CoV-2 are excluded. (F) Comparison of neutralizing antibody titers against SARS-CoV-2 pseudoviruses containing the SARS-CoV-2 S protein with mutations characteristic of the delta variant in sera from populations including subjects previously infected or currently infected with SARS-CoV-2.(G) The geometric mean rise (GMR) in neutralizing antibodies observed in subjects receiving 60ug of BNT162b2, 30ug of RNA encoding a SARS-CoV-2 S protein with mutations characteristic of the Omicron BA.1 variant (OMI 30ug), 60ug of RNA encoding a SARS-CoV-2 S protein with mutations characteristic of the Omicron BA.1 variant (OMI 60ug), 30ug of a bivalent vaccine comprising 15ug of BNT162b2 and 15ug of RNA encoding a SARS-CoV-2 S protein with mutations characteristic of the Omicron BA.1 variant (Bivalent 30ug), or 60ug of a bivalent vaccine comprising 30ug of BNT162b2 and 30ug of RNA encoding a SARS-CoV-2 S protein with mutations characteristic of the Omicron BA.1 variant (Bivalent 60ug) compared to subjects receiving 30ug of BNT162b2 as the fourth dose. Results are presented for both population pools that exclude subjects previously or currently infected with SARS-CoV-2 and population pools that include these subjects. [Figure 23-2] Same as above. [Figure 23-3] Same as above. [Figure 23-4] Same as above. [Figure 23-5] Same as above. [Figure 23-6] Same as above. [Figure 23-7] Same as above. [Figure 24-1]Reactogenicity of certain exemplary RNAs (formulated in LNPs) at a given dose: Subjects receiving a 60ug dose of RNA encoding the SARS-CoV-2 S protein are more likely to experience injection site pain and a similar systemic reaction than subjects receiving a 30ug dose of RNA. Subjects were administered 30ug or 60ug of RNA encoding the SARS-CoV-2 S protein from the Wuhan strain (BNT162b2, corresponding to groups G1 and G2, respectively), 30ug or 60ug of RNA encoding a SARS-CoV-2 S protein with mutations characteristic of the Omicron BA.1 variant (BNT162b2 OMI, corresponding to groups G3 and G4, respectively), 30ug of a bivalent vaccine containing 15ug of RNA encoding the SARS-CoV-2 S protein from the Wuhan strain and 15ug of RNA encoding a SARS-CoV-2 S protein with mutations characteristic of the Omicron BA.1 variant (BNT162B2 (15ug) + BNT162b2 OMI (15ug), corresponding to group G5), or 30ug of RNA encoding the SARS-CoV-2 S protein from the Wuhan strain and 30ug of SARS-CoV-2 S protein with mutations characteristic of the Omicron BA.1 variant. Subjects were administered 60 μg of a bivalent vaccine (BNT162b2 (30 μg) + BNT162b2 OMI (30 μg), corresponding to group G6) containing RNA encoding the S protein. (A) Local reactions observed within 7 days of injection, including redness, swelling, and pain at the injection site. Increased pain at the injection site was observed in subjects receiving 60 μg of the Omicron BA.1 variant or RNA encoding the SARS-CoV-2 S protein containing mutations characteristic of the bivalent vaccine compared to the other doses tested. (B) Systemic reactions observed within 7 days of injection, including fever, fatigue, headache, chills, vomiting, diarrhea, muscle pain, joint pain, and medication use. Systemic reactions up to 7 days were observed to be broadly similar across the different groups. Fatigue tended to be higher after administration of the 60 μg dose compared to the 30 μg dose. [Figure 24-2] Same as above. [Figure 25-1]Omicron BA.1 breakthrough infection in individuals vaccinated two and three times with BNT162b2 induces broad neutralization of Omicron BA.1, BA.2, and other VOCs, but to a lesser extent BA.4 and BA.5. This figure is an extension of Figure 16, which includes data on neutralization activity against Omicron BA.4 and BA.5. As depicted in Figure 16, sera were tested in duplicate, and the 50% pseudovirus neutralization (pVN50) geometric mean titers (GMTs) (in A and B) and the geometric mean ratios of SARS-CoV-1 pVN50 GMTs normalized to SARS-CoV-2 variants of concern (VOCs) and Wuhan pVN50 GMTs were plotted. For values ​​below the limit of detection (LOD), the LOD / 2 value was plotted. The values ​​above the violin plot represent group GMTs. Wuhan neutralization group GMTs were compared with titers against the indicated variants and SARS-CoV-1 using a nonparametric Friedman test with Dunn's multiple comparison correction. Multiplicity-adjusted p-values ​​are shown. (A) pVN50 GMTs against Wuhan, VOC, and SARS-CoV-1 pseudoviruses in patients who received two or three doses of BNT162b2. (B) pVN50 GMTs against Wuhan, VOC, and SARS-CoV-1 pseudoviruses in patients who received two or three doses of BNT162b2 and were previously infected with the Omicron BA.1 variant of SARS-CoV-2. (C) Group geometric mean ratios with 95% confidence intervals for all cohorts shown in (A) and (B). [Figure 25-2] Same as above. [Figure 26]Omicron BA.1 breakthrough infection in individuals vaccinated with other approved COVID-19 vaccines or combination regimens results in immune sera that broadly neutralize Omicron BA.1, BA.2, and other VOCs, but to a lesser extent BA.4 and BA.5. This figure is an extension of Figure 19, which includes data on neutralization activity against Omicron BA.4 and BA.5. As described in Figure 19, sera were tested in duplicate, and individual 50% pseudovirus neutralization (pVN50) geometric mean titers (GMTs) against SARS-CoV-2 Wuhan, alpha, beta, delta, and Omicron BA.1, BA.2, and BA.4 / 5 variants, as well as SARS-CoV-1, were plotted. For values ​​below the limit of detection (LOD), the LOD / 2 value was plotted. The values ​​above the violin plot represent the group GMTs. Wuhan neutralization group GMTs were compared with titers against the indicated variants and SARS-CoV-1 using a nonparametric Friedman test with Dunn's multiple comparison correction. Multiplicity-adjusted p-values ​​are shown. [Figure 27] Sequences of the RBD of the SARS-CoV-2 Wuhan strain and its variants. The top sequence corresponds to the Wuhan strain, the second sequence corresponds to the alpha variant, the third sequence corresponds to the delta variant, the fourth sequence corresponds to the Omicron BA.1 variant, and the fifth sequence corresponds to the Omicron BA.4 / 5 variant. Variant-specific amino acid alterations are shown in bold. [Figure 28]Cohorts and sample collection for the study described in Example 14. A schematic diagram is shown for testing immune responses in three-vaccinated patients who were (i) Omicron-naive, (ii) infected with the BA.1 Omicron variant, or (iii) infected with the BA.2 Omicron variant. Blood samples were collected from three cohorts: Omicron-naive individuals vaccinated three times with BNT162b2 (BNT162b23), and individuals vaccinated with a homologous or heterologous three-dose regimen who subsequently had a breakthrough infection with Omicron during either the BA.1-dominant period in Germany (November 2021-January 2022, all Vax+BA.1) or the BA.2-dominant period (March-May 2022, all Vax+BA.2, blue). Serum (droplets) was isolated in the Omicron-naive cohort at the indicated time points after most recent vaccination, and for the convalescent cohort, the time from most recent vaccination to Omicron infection and the time from infection to serum isolation are shown. All values ​​are specified as median ranges. Serum neutralization capacity was assessed using a pseudovirus neutralization test. [Figure 29]50% pseudovirus neutralization (pVN50) geometric mean titers (GMTs) from BNT162b23 and All Vax+Omi BA.1 breakthrough infection cohorts. Serum was collected from Omicron-naive BNT162b2-vaccinated individuals 28 days after the third dose (BNT162b23, circles) and from vaccinated individuals who later developed Omicron BA.1 breakthrough infection (all Vax+Omi BA.1, triangles) at a median of 43 days post-infection. 50% pseudovirus neutralization (pVN50) geometric mean titers (GMTs) for Omicron-naive individuals are plotted in (A) and for BA.1-breakthrough infected individuals in (B). This data, with the exception of the BA.2.12.1 neutralization data, was previously published in Quandt et al. (e.g., "Omicron BA.1 breakthrough infection drives cross-variant neutralization and memory B cell formation against conserved epitopes." Science immunology, eabq2427 (2022), doi:10.1126 / sciimmunol.abq2427). Sera were tested in duplicate. For values ​​below the limit of detection (LOD), the LOD / 2 value was plotted. Values ​​above the violin plot represent group GMTs. Wuhan neutralization group GMTs were compared to titers against the indicated variants and SARS-CoV-1 using a nonparametric Friedman test with Dunn's multiple comparisons correction. Multiplicity-adjusted p-values ​​are shown. [Figure 30-1]Omicron BA.2 breakthrough infection in previously vaccinated individuals refocuses neutralization against Omicron BA.2 and BA.2-derived subvariants BA.2.12.1 and BA.4 / BA.5. Sera were collected from individuals vaccinated three times with BNT162b2 who subsequently underwent Omicron BA.1 breakthrough infection at a median of 44 days post-infection (BNT162b23+Omi BA.1, triangles) and from individuals vaccinated three times with BNT162b2 who subsequently underwent Omicron BA.2 breakthrough infection at 38 days post-infection (BNT162b23+Omi BA.2, squares). Plots of 50% pseudovirus neutralization (pVN50) geometric mean titers (GMTs) (A, B) and geometric mean ratios of SARS-CoV-1 pVN50 GMTs normalized to SARS-CoV-2 variants of concern (VOCs) and Wuhan pVN50 GMTs (C). pVN50 GMT and geometric mean ratio data for Omicron-naive BNT162b2-vaccinated individuals (BNT162b23, circle) and Omicron BA.1 breakthrough-infected BNT162b2-vaccinated individuals (BNT162b23, circle) were previously published in Quandt et al. ("Omicron BA.1 breakthrough infection drives cross-variant neutralization and memory B cell formation against conserved epitopes." Science immunology, eabq2427 (2022), doi:10.1126 / sciimmunol.abq2427), except for BA.2.12.1 neutralization data. Sera were tested in duplicate. For values ​​below the limit of detection (LOD), the LOD / 2 value was plotted. Values ​​above the violin plot represent group GMTs. Wuhan neutralization group GMTs were compared with titers against the indicated variants and SARS-CoV-1 using a nonparametric Friedman test with Dunn's multiple comparison correction. Multiplicity-adjusted p-values ​​are shown. (A, B) pVN50 GMTs against Wuhan, VOC, and SARS-CoV-1 pseudoviruses. (C) Group geometric mean ratios with 95% confidence intervals. [Figure 30-2] Same as above. [Figure 31] Characteristics of the SARS-CoV-2 S glycoprotein used in the VSV-SARS-CoV-2 pseudovirus-based neutralization assay. The Wuhan-Hu-1 isolate SARS-CoV-2 S glycoprotein (GenBank: QHD43416.1) was used as a reference. The amino acid position, amino acid description (single-letter code), and type of mutation (substitution, deletion, insertion) are indicated. NTD, N-terminal domain; RBD, receptor-binding domain; Δ, deletion; ins, insertion; *, cytoplasmic domain truncated for the C-terminal 19 amino acids. [Figure 32] Alterations in the spike glycoprotein amino acid sequence of SARS-CoV-2 Omicron sublineages. The amino acid position, amino acid description (single-letter code), and type of mutation (substitution, deletion, or insertion) are shown. White text in boxes indicates amino acid substitutions by sublineage; Δ, deletion; ins, insertion; NTD, N-terminal domain; RBD, receptor-binding domain. [Figure 33] Immunization Protocol for Studies with VOC Boosters. BALB / c mice were immunized according to the indicated schedule with two doses (1 μg each) of the original BNT162b2 vaccine, followed by at least one dose (1 μg total) of a monovalent, bivalent, or trivalent booster dose of either: (a) original BNT162b2 ("BNT162b2"), (b) BNT162b2 OMI BA.1 ("OMI BA.1"), (c) BNT162b2 OMI BA.4 / 5 ("OMI BA.4 / 5"), or combinations thereof. [Figure 34] Baseline grouped neutralization GMTs. Sera collected from mice immunized as shown in Figure 33 (day 104, pre-boost) were evaluated for geometric mean titers of neutralizing antibodies against various strains. Data are presented grouped by cohort. [Figure 35] Baseline staggered neutralization GMTs. Sera collected from mice immunized as shown in Figure 33 (day 104, pre-boost) were evaluated for geometric mean titers of neutralizing antibodies against the various strains. Data are presented in a staggered format (i.e., for each strain for which neutralization was evaluated). [Figure 36] Baseline cross-neutralization. Sera collected from mice immunized as shown in Figure 33 (day 104, pre-boost) were evaluated for geometric mean titers of neutralizing antibodies against the various strains. Cross-neutralization results are presented as the calculated variant / Wuhan reference GMT ratio. [Figure 37] Geometric mean fold increase in GMT after boost. Sera collected from mice immunized as shown in Figure 33 (day 111, 7 days after boost) were evaluated for the geometric mean fold increase in GMT of neutralizing antibodies against various strains. [Figure 38] Grouped Neutralization GMTs After Boost. Sera collected from mice immunized as shown in Figure 33 (day 111, 7 days after boost) were evaluated for the geometric mean fold increase in GMTs of neutralizing antibodies against the various strains. Data are presented grouped by cohort. [Figure 39] Cross-neutralization after boost. Sera collected from mice immunized as shown in Figure 33 (day 111, 7 days post-boost) were evaluated for the geometric mean fold increase in GMT of neutralizing antibodies against various strains. Cross-neutralization results are presented as the calculated variant / Wuhan reference GMT ratio. [Figure 40] Exemplary spike protein amino acid mutations. Modified amino acid residues are shown and used to produce RNA vaccines encoding variant coronavirus spike proteins. In some examples, such amino acid modifications can be combined with other amino acid residue modifications, as shown under "Mutation" and "Mutation Type" in Figure 41. Amino acid positions are numbered relative to the S protein sequence from the Wuhan sequence (SEQ ID NO: 1). In some embodiments, various combinations of amino acid mutations as described herein can be applied to different coronavirus S proteins or immunogenic fragments thereof. [Figure 41]Exemplary spike protein variants. Exemplary combinations of spike protein mutations, including modified amino acid residues, mutation types, and furin mutations (682 / 683 / 684 / 685 RRAR to GSAS), are shown. RNA constructs encoding exemplary combinations of spike protein mutations were evaluated for S protein expression, CR3022 epitope response, and ACE2 response. Amino acid positions are numbered relative to the S protein sequence from the Wuhan sequence (SEQ ID NO: 1). In some embodiments, various combinations of amino acid mutations as described herein can be applied to different coronavirus S proteins or immunogenic fragments thereof. [Figure 42] Effect of RNAs encoding exemplary spike protein variants on neutralization against various coronavirus strains and / or variants. RNAs encoding exemplary spike protein variants (e.g., containing a P6' backbone as shown in Figure 40, D614G, and a furin site mutation (682 / 683 / 684 / 685 RRAR to GSAS)) stimulated higher neutralization titers across a variety of coronaviruses. [Figure 43]The BNT162b5 format bivalent (Wuhan + BA.4 / 5) is more immunogenic than the BNT162b2 format bivalent (Wuhan + BA.4 / 5). Mice were administered two doses of BNT162b2 21 days apart, followed by a bivalent vaccine comprising: (i) BNT162b2; (ii) a first RNA encoding the Wuhan spike protein and a second RNA encoding a SARS-CoV-2 spike protein comprising mutations characteristic of the BA.4 / 5 omicron variant, wherein the spike protein encoded by each of the first and second RNAs also comprises the mutations K986P and V987P ("BNT162b2 Bivalent (BA.4 / 5)"). or (iii) a bivalent vaccine comprising a first RNA encoding the Wuhan spike protein and a second RNA encoding a SARS-CoV-2 spike protein comprising mutations characteristic of the BA.4 / 5 omicron variant, wherein the S protein encoded by each of the first and second RNAs also comprises P6' mutations (D985P, V987P, F817P, A892P, A899P, and A942P), D614G, and a furin cleavage site mutation (682 / 683 / 684 / 685 RRAR vs. GSAS) ("BNT162b5 Bivalent (BA.4 / 5)"). Serum samples were collected one month after the third dose, and neutralization titers were determined for Wuhan and Omicron variants BA.1, BA.2, BA.2.12.1, and BA.4 / 5 using a pseudovirus neutralization assay (50% pVNT titer). [Figure 44-1]Bivalent BNT162b5 provides improved immune responses in vaccine-experienced human subjects. Human subjects who previously received three doses of BNT162b2 (encoding the SARS-CoV-2 S protein of the Wuhan strain and including the K986P and V987P mutations) were administered a bivalent vaccine comprising: (i) a first RNA encoding the Wuhan spike protein and a second RNA encoding a SARS-CoV-2 spike protein including mutations characteristic of the BA.1 omicron variant, wherein the spike protein encoded by each of the first and second RNAs contains the K986P and V987P mutations ("BNT162b2 bivalent Omi"). or (ii) a bivalent vaccine comprising a first RNA encoding the Wuhan spike protein and a second RNA encoding a SARS-CoV-2 spike protein comprising mutations characteristic of the BA.2 omicron variant, wherein the S protein encoded by each of the first and second RNAs also comprises P6' mutations (D985P, V987P, F817P, A892P, A899P, and A942P), D614G, and a mutation at the furin cleavage site (682 / 683 / 684 / 685 RRAR to GSAS) ("BNT162b5 Bivalent (BA.2)"). Serum was collected one month after administration of the RNA vaccine, and neutralization titers were collected for the Wuhan ("WT"), Omicron BA.1 ("BA.1"), or Omicron BA.2 ("BA.2") SARS-CoV-2 variants. Titers are shown for (A) all subjects, (B) subjects who showed evidence of prior SARS-CoV-2 infection at the time of administration of the SARS-CoV-2 vaccine, and (C) subjects who showed no evidence of prior SARS-CoV-2 infection at the time of administration of the SARS-CoV-2 vaccine. Titer values ​​are indicated above each bar. Titers were collected using a fluorescent focus reduction neutralization titer (FFRNT) assay. "1MPD4" refers to one month after dose 4. "WT" refers to the Wuhan strain. "LLOQ" stands for lower limit of quantitation. [Figure 44-2] Same as above. [Figure 44-3] Same as above. [Figure 45]Bivalent BNT162b5 and BNT162b6 provide improved immune responses when administered as boosters to vaccine-experienced mice. Mice administered two doses of BNT162b2 (encoding the SARS-CoV-2 S protein of the Wuhan strain and containing the K986P and V987P mutations) were administered a bivalent vaccine comprising: (i) a first RNA encoding the Wuhan spike protein and a second RNA encoding a SARS-CoV-2 spike protein containing mutations characteristic of the BA.4 / 5 omicron variant, wherein the spike proteins encoded by each of the first and second RNAs contain the K986P and V987P mutations ("BNT162b2 bivalent"). (ii) a bivalent vaccine comprising a first RNA encoding the Wuhan spike protein and a second RNA encoding a SARS-CoV-2 spike protein comprising mutations characteristic of the BA.4 / 5 omicron variant, wherein the S protein encoded by each of the first and second RNAs contains P6' mutations (D985P, V987P, F817P, A892P, A899P, and A942P), D614G, and mutations at the furin cleavage site (682 / 683 / 684 / 685 or (iii) a bivalent vaccine comprising a first RNA encoding the Wuhan spike protein and a second RNA encoding a SARS-CoV-2 spike protein comprising mutations characteristic of the BA.4 / 5 omicron variant, wherein the S protein encoded by each of the first and second RNAs also comprises P6 mutations (K986P, V987P, F817P, A892P, A899P, and A942P), D614G, and mutations at the furin cleavage site (682 / 683 / 684 / 685 RRAR to GSAS), and mutations introducing disulfide bonds (T547C, N978C) (“BNT162b6 Bivalent (BA.4 / 5)”). Serum was collected one month after administration of the RNA vaccine, and neutralization titers were determined for a collection of variants of concern (shown in the legend). 50% neutralization titers ("50% pVNT titer") are shown. "LOD" represents the limit of detection. [Figure 46-1] Figure 46. In vitro characterization of exemplary SARS-CoV-2 variants. Protein expression, ACE2 binding, and CR3022 binding (neutralizing antibodies) are shown for certain exemplary SARS-CoV-2 variants containing mutations described herein (mutations present in each SARS-CoV-2 variant listed in Table 34). Figure 47. In vitro characterization of exemplary SARS-CoV-2 variants. Protein expression, ACE2 binding, and CR3022 binding (neutralizing antibodies) are shown for certain exemplary SARS-CoV-2 variants containing mutations described herein (mutations present in each SARS-CoV-2 variant listed in Table 34). [Figure 46-2] Same as above. [Figure 46-3] Same as above. [Figure 47-1]Figure 45. Bivalent BNT162b6 and BNT162b7 provide improved immune responses when administered as boosters to unvaccinated mice. Mice were administered (i) a bivalent vaccine comprising a first RNA encoding the Wuhan spike protein and a second RNA encoding a SARS-CoV-2 spike protein comprising mutations characteristic of the BA.4 / 5 omicron variant, wherein the spike protein encoded by each of the first and second RNAs also comprises the K986P and V987P mutations ("BNT162b2 bivalent BA.4 / 5"); (ii) a bivalent vaccine comprising a first RNA encoding the Wuhan spike protein and a second RNA encoding a SARS-CoV-2 spike protein comprising mutations characteristic of the BA.4 / 5 omicron variant, wherein the S protein encoded by each of the first and second RNAs also comprises the P6' mutations (D985P, V987P, F817P, A892P, A899P, and A942P), D614G, and mutations at the furin cleavage site (682 / 683 / 684 / 685P). (iii) a bivalent vaccine comprising a first RNA encoding the Wuhan spike protein and a second RNA encoding a SARS-CoV-2 spike protein containing mutations characteristic of the BA.4 / 5 omicron variant, wherein the S protein encoded by each of the first and second RNAs also contains P6 mutations (K986P, V987P, F817P, A892P, A899P, and A942P), D614G, a furin cleavage site mutation (682 / 683 / 684 / 685 RRAR to GSAS), and mutations to introduce disulfide bonds (T547C, N978C) ("BNT162b6 Bivalent (BA.2)"); or (iv) a bivalent vaccine comprising a first RNA encoding the Wuhan spike protein and a second RNA encoding a SARS-CoV-2 spike protein containing mutations characteristic of the BA.4 / 5 omicron variant, wherein the S protein encoded by each of the first and second RNAs also contains P6 mutations (K986P, V987P, F817P, A892P, A899P, and A942P), D614G, a furin cleavage site mutation (682 / 683 / 684 / 685 RRAR to GSAS), and mutations to introduce disulfide bonds (T547C, N978C) ("BNT162b6 Bivalent (BA.2)").A bivalent vaccine ("BNT162b7 Bivalent (BA.4 / 5")) containing a second RNA encoding a SARS-CoV-2 spike protein containing mutations characteristic of the 4 / 5 omicron variant, where the S protein encoded by each of the first and second RNAs also contained P6 mutations (K986P, V987P, F817P, A892P, A899P, and A942P), D614G, a furin cleavage site mutation (682 / 683 / 684 / 685 RRAR to GSAS), and mutations to introduce a disulfide bond (T547C, S982C) was administered. Serum was collected one month after administration of the RNA vaccine, and neutralization titers were determined for a collection of variants of concern (indicated in the legend). 50% neutralization titers ("50%"). pVNT titers are shown in (A) and (B), and neutralization titers relative to BNT162b2 titers are shown in (C). [Figure 47-2] Same as above. DETAILED DESCRIPTION OF THE INVENTION

[0127] Although the present disclosure is described in detail below, it should be understood that the disclosure is not limited to the specific methodology, protocols, and reagents described herein, as these may vary. It should also be understood that the terminology used herein is for the purpose of describing particular embodiments only, and is not intended to limit the scope of the present disclosure, which is limited only by the appended claims. Unless otherwise defined, all technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art.

[0128] Preferably, the terms used herein are defined as set forth in "A multilingual glossary of biotechnological terms: (IUPAC Recommendations)", H.G.W. Leuenberger, B. Nagel, and H. Kolbl, Eds., Helvetica Chimica Acta, CH-4010 Basel, Switzerland, (1995).

[0129] The practice of the present disclosure will employ, unless otherwise indicated, conventional methods of chemistry, biochemistry, cell biology, immunology, and recombinant DNA techniques described in the art (see, e.g., Molecular Cloning: A Laboratory Manual, 2nd Edition, J. Sambrook et al. eds., Cold Spring Harbor Laboratory Press, Cold Spring Harbor 1989).

[0130] The following describes elements of the present disclosure. While these elements are listed with specific embodiments, it should be understood that they can be combined in any manner and in any number to create additional embodiments. The various described examples and embodiments should not be construed as limiting the disclosure to only the explicitly described embodiments. This description should be understood to disclose and encompass embodiments combining the explicitly described embodiments with any number of the disclosed elements. Furthermore, any permutation and combination of all described elements should be considered disclosed by this description unless the context dictates otherwise.

[0131] Several documents are cited throughout the text of this specification. Each document cited herein (including all patents, patent applications, scientific publications, manufacturer's specifications, instructions, etc.), whether supra or infra, is hereby incorporated by reference in its entirety. Nothing herein should be construed as an admission that the present disclosure is not entitled to antedate such disclosure.

[0132] definition The following provides definitions that apply to all aspects of this disclosure. The following terms have the following meanings unless otherwise indicated. Any undefined terms have their art-recognized meanings.

[0133] The term "about," in the context of numerical values ​​or ranges described herein, means approximately or approximately, and in one embodiment, means ±20%, ±10%, ±5%, or ±3% of the recited or claimed numerical value or range.

[0134] As used in the context of describing this disclosure (particularly in the context of the claims), the terms "a," "an," and "the," and similar referents, should be construed to cover both the singular and the plural unless otherwise indicated herein or clearly contradicted by context. The recitation of ranges of values ​​herein is merely intended to serve as a shorthand method of referring individually to each separate value falling within the range. Unless otherwise indicated herein, each separate value is incorporated herein as if it were individually recited herein. All methods described herein can be performed in any suitable order unless otherwise indicated herein or clearly contradicted by context. The use of any and all examples, or exemplary language (e.g., "etc.") provided herein is intended merely to better illustrate the disclosure and does not limit the scope of the claims. No language in the specification should be construed as indicating any non-claimed element essential to the practice of the disclosure.

[0135] Unless expressly specified otherwise, the term "comprising" is used in the context of this document to indicate that further members may optionally be present in addition to the members of the list introduced by "comprising." However, it is contemplated as a specific embodiment of the present disclosure that the term "comprising" encompasses the possibility that no further members are present, i.e., for the purposes of this embodiment, "comprising" should be understood to have the meaning of "consisting of" or "consisting essentially of."

[0136] As used herein, terms such as "reduce," "diminish," "inhibit," or "impair" refer to an overall reduction or ability to cause an overall reduction in levels, preferably of at least 5%, at least 10%, at least 20%, at least 50%, at least 75%, or even more. These terms include complete or essentially complete inhibition, i.e., reduction to zero or essentially to zero.

[0137] Terms such as "increase," "enhance," or "exceed" preferably relate to an increase or enhancement of at least 10%, at least 20%, at least 30%, at least 40%, at least 50%, at least 80%, at least 100%, at least 200%, at least 500%, or even more.

[0138] According to the present disclosure, the term "peptide" includes oligo- and polypeptides and refers to a substance containing about 2 or more, about 3 or more, about 4 or more, about 6 or more, about 8 or more, about 10 or more, about 13 or more, about 16 or more, about 20 or more, and up to about 50, about 100, or about 150 consecutive amino acids linked together via peptide bonds. The terms "protein" or "polypeptide" refer to large peptides, particularly peptides having at least about 150 amino acids, although the terms "peptide," "protein," and "polypeptide" are generally used interchangeably herein.

[0139] A "therapeutic protein" when provided to a subject in a therapeutically effective amount has a positive or beneficial effect on a condition or disease state of the subject. In one embodiment, a therapeutic protein has curative or palliative properties and can be administered to improve, alleviate, reduce, reverse, delay the onset, or reduce the severity of one or more symptoms of a disease or disorder. A therapeutic protein can have prophylactic properties and can be used to delay the onset of a disease or reduce the severity of such a disease or pathological condition. The term "therapeutic protein" includes whole proteins or peptides and can also refer to therapeutically active fragments thereof. It can also include therapeutically active variants of proteins. Examples of therapeutically active proteins include, but are not limited to, antigens for vaccination and immune stimulants such as cytokines.

[0140] "Fragment" refers to a portion of an amino acid sequence (peptide or protein), i.e., a sequence representing an amino acid sequence truncated at the N-terminus and / or C-terminus. C-terminally truncated fragments (N-terminal fragments) can be obtained, for example, by translating a truncated open reading frame lacking the 3' end of the open reading frame. N-terminally truncated fragments (C-terminal fragments) can be obtained, for example, by translating a truncated open reading frame lacking the 5' end of the open reading frame, as long as the truncated open reading frame contains the initiation codon responsible for initiating translation. A fragment of an amino acid sequence comprises, for example, at least 50%, at least 60%, at least 70%, at least 80%, or at least 90% of the amino acid residues from the amino acid sequence. A fragment of an amino acid sequence preferably comprises at least 6, in particular at least 8, at least 12, at least 15, at least 20, at least 30, at least 50, or at least 100 consecutive amino acids from the amino acid sequence.

[0141] As used herein, "variant" refers to an amino acid sequence that differs from a parent amino acid sequence by at least one amino acid modification. The parent amino acid sequence can be a naturally occurring or wild-type (WT) amino acid sequence, or can be a modified version of the wild-type amino acid sequence. Preferably, the variant amino acid sequence has at least one amino acid modification compared to the parent amino acid sequence, e.g., 1 to about 20 amino acid modifications compared to the parent, preferably 1 to about 10 or 1 to about 5 amino acid modifications.

[0142] As used herein, "wild-type" or "WT" or "native" refers to an amino acid sequence found in nature, including allelic variants. A wild-type amino acid sequence, peptide, or protein has an amino acid sequence that has not been intentionally modified.

[0143] For purposes of this disclosure, a "variant" of an amino acid sequence (peptide, protein, or polypeptide) includes amino acid insertion variants, amino acid addition variants, amino acid deletion variants, and / or amino acid substitution variants. The term "variant" includes all mutants, splice variants, post-translationally modified variants, conformations, isoforms, allelic variants, species variants, and species homologs, particularly those that occur naturally. The term "variant" particularly includes fragments of an amino acid sequence.

[0144] Amino acid insertion variants include the insertion of a single or two or more amino acids into a specific amino acid sequence. In amino acid sequence variants with insertions, one or more amino acid residues are inserted at a specific site in the amino acid sequence, although random insertion is also possible with appropriate screening of the resulting product. Amino acid addition variants include amino- and / or carboxy-terminal fusions of one or more amino acids, such as 1, 2, 3, 5, 10, 20, 30, 50, or more amino acids. Amino acid deletion variants are characterized by the removal of one or more amino acids from the sequence, for example, the removal of 1, 2, 3, 5, 10, 20, 30, 50, or more amino acids. The deletion may occur at any position in the protein. Amino acid deletion variants, including deletions at the N- and / or C-termini of a protein, are also referred to as N- and / or C-terminal truncation variants. Amino acid substitution variants are characterized by at least one residue in the sequence being removed and another residue being inserted in its place. Modifications at positions within the amino acid sequence that are not conserved between homologous proteins or peptides and / or replacement of amino acids with other amino acids with similar properties are preferred. Preferably, the amino acid changes in peptide and protein variants are conservative amino acid changes, i.e., substitutions of similarly charged or uncharged amino acids. Conservative amino acid changes involve substitutions within a family of amino acids related by their side chains. Naturally occurring amino acids are generally classified into four families: acidic (aspartate, glutamate), basic (lysine, arginine, histidine), nonpolar (alanine, valine, leucine, isoleucine, proline, phenylalanine, methionine, tryptophan), and uncharged polar (glycine, asparagine, glutamine, cysteine, serine, threonine, tyrosine) amino acids. Phenylalanine, tryptophan, and tyrosine are sometimes jointly classified as aromatic amino acids. In one embodiment, conservative amino acid substitutions include substitutions within the following groups: Glycine, Alanine; valine, isoleucine, leucine; Aspartic acid, glutamic acid; Asparagine, glutamine; Serine, threonine; lysine, arginine; and Phenylalanine, tyrosine.

[0145] Preferably, the degree of similarity, preferably identity, between a given amino acid sequence and an amino acid sequence that is a variant of the given amino acid sequence will be at least about 60%, 70%, 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99%. The degree of similarity or identity is preferably given over an amino acid region that is at least about 10%, at least about 20%, at least about 30%, at least about 40%, at least about 50%, at least about 60%, at least about 70%, at least about 80%, at least about 90%, or about 100% of the entire length of the reference amino acid sequence. For example, if the reference amino acid sequence consists of 200 amino acids, the degree of similarity or identity is, in some embodiments, preferably given over at least about 20, at least about 40, at least about 60, at least about 80, at least about 100, at least about 120, at least about 140, at least about 160, at least about 180, or about 200 consecutive amino acids. In some embodiments, the degree of similarity or identity is given over the entire length of the reference amino acid sequence. Alignment to determine sequence similarity, preferably sequence identity, can be performed using tools known in the art, preferably using best sequence alignment, for example, Align using standard settings, preferably EMBOSS::Needle, Matrix:Blosum62, Gap Open 10.0, Gap Extend 0.5.

[0146] "Sequence similarity" refers to the percentage of amino acids that are identical or represent conservative amino acid substitutions. "Sequence identity" between two amino acid sequences refers to the percentage of amino acids that are identical between the sequences. "Sequence identity" between two nucleic acid sequences refers to the percentage of nucleotides that are identical between the sequences.

[0147] "% identical," "% identity," or similar terms are intended to refer, inter alia, to the percentage of nucleotides or amino acids that are identical in optimal alignment between the compared sequences. The percentage is purely statistical; the differences between the two sequences may, but are not necessarily, randomly distributed over the entire length of the compared sequences. Comparison of two sequences is usually performed by comparing the sequences after optimal alignment over a segment or "comparison window" to identify local regions of corresponding sequences. Optimal alignment for comparison can be performed manually or with the aid of the local homology algorithm of Smith and Waterman, 1981, Ads App. Math. 2, 482, with the aid of the local homology algorithm of Neddleman and Wunsch, 1970, J. Mol. Biol. 48, 443, with the aid of the similarity search algorithm of Pearson and Lipman, 1988, Proc. Natl. Acad. Sci. USA 88, 2444, or with the aid of computer programs that use such algorithms (GAP, BESTFIT, FASTA, BLAST P, BLAST N, and TFASTA in the Wisconsin Genetics software package (Genetics Computer Group, 575 Science Drive, Madison, Wis.)). In some embodiments, the percent identity of two sequences is determined using the BLASTN or BLASTP algorithm, available at the National Center for Biotechnology Information (NCBI) website (e.g., blast.ncbi.nlm.nih.gov / Blast.cgi?PAGE_TYPE=BlastSearch&BLAST_SPEC=blast2seq&LINK_LOC=align2seq).In some embodiments, the algorithm parameters used for the BLASTN algorithm on the NCBI website include: (i) expectation threshold set to 10, (ii) word size set to 28, (iii) maximum match within query range set to 0, (iv) match / mismatch score set to 1, -2, (v) gap cost set to linear, and (vi) a filter for low complexity regions is used. In some embodiments, the algorithm parameters used for the BLASTP algorithm on the NCBI website include: (i) expectation threshold set to 10, (ii) word size set to 3, (iii) maximum match within query range set to 0, (iv) matrix set to BLOSUM62, (v) gap costs set to Existence:11 Extension:1, and (vi) conditional composition score matrix adjustment.

[0148] Percentage identity is obtained by determining the number of identical positions where the compared sequences correspond, dividing this number by the number of positions being compared (e.g., the number of positions in the reference sequence), and multiplying this result by 100.

[0149] In some embodiments, the degree of similarity or identity is given for a region that is at least about 50%, at least about 60%, at least about 70%, at least about 80%, at least about 90%, or about 100% of the entire length of the reference sequence. For example, if the reference nucleic acid sequence consists of 200 nucleotides, the degree of identity is given in some embodiments for at least about 100, at least about 120, at least about 140, at least about 160, at least about 180, or about 200 consecutive nucleotides. In some embodiments, the degree of similarity or identity is given for the entire length of the reference sequence.

[0150] Homologous amino acid sequences according to the present disclosure exhibit an identity of at least 40%, particularly at least 50%, at least 60%, at least 70%, at least 80%, at least 90%, and preferably at least 95%, at least 98, or at least 99% of the amino acid residues.

[0151] The amino acid sequence variants described herein may be readily prepared by one skilled in the art, for example, by recombinant DNA manipulation. The manipulation of DNA sequences to prepare peptides or proteins with substitutions, additions, insertions, or deletions is described in detail, for example, in Sambrook et al. (1989). Furthermore, the peptides and amino acid variants described herein may be readily prepared with the aid of known peptide synthesis techniques, such as, for example, solid-phase synthesis and similar methods.

[0152] In one embodiment, a fragment or variant of an amino acid sequence (peptide or protein) is preferably a "functional fragment" or "functional variant." The term "functional fragment" or "functional variant" of an amino acid sequence refers to any fragment or variant that exhibits one or more functional properties identical or similar to those of the amino acid sequence from which it is derived, i.e., is functionally equivalent. With respect to an antigen or antigen sequence, one particular function is one or more immunogenic activities exhibited by the amino acid sequence from which the fragment or variant is derived. As used herein, the term "functional fragment" or "functional variant" particularly refers to a variant molecule or sequence that includes an amino acid sequence that is modified by one or more amino acids compared to the amino acid sequence of the parent molecule or sequence, and that can still perform one or more functions of the parent molecule or sequence, such as eliciting an immune response. In one embodiment, modifications in the amino acid sequence of the parent molecule or sequence do not significantly affect or alter the characteristics of the molecule or sequence. In different embodiments, the function of the functional fragment or functional variant may be reduced but still significantly present, for example, the immunogenicity of the functional variant may be at least 50%, at least 60%, at least 70%, at least 80%, or at least 90% of that of the parent molecule or sequence, however, in other embodiments, the immunogenicity of the functional fragment or functional variant may be enhanced compared to the parent molecule or sequence.

[0153] An amino acid sequence (peptide, protein, or polypeptide) "derived from" a specified amino acid sequence (peptide, protein, or polypeptide) refers to the origin of the first amino acid sequence. Preferably, an amino acid sequence derived from a particular amino acid sequence has an amino acid sequence that is identical, essentially identical, or homologous to the particular sequence or a fragment thereof. An amino acid sequence derived from a particular amino acid sequence may be a variant of the particular sequence or a fragment thereof. For example, it will be understood by those skilled in the art that antigens suitable for use herein can be modified such that the sequence varies from the naturally occurring sequence or the native sequence from which they are derived while retaining the desired activity of the native sequence.

[0154] As used herein, "educational materials" or "instructions" include publications, records, drawings, or any other medium of expression that can be used to communicate the usefulness of the disclosed compositions and methods. The educational materials of the disclosed kits can be, for example, affixed to a container containing the disclosed compositions or shipped together with a container containing the composition. Alternatively, the educational materials can be shipped separately from the container with the intention that the educational materials and the composition be used cooperatively by the recipient.

[0155] "Isolated" means altered or removed from the natural state. For example, a nucleic acid or peptide that is naturally present in a living animal is not "isolated," but the same nucleic acid or peptide that has been partially or completely separated from the coexisting materials of its natural state is "isolated." An isolated nucleic acid or protein can exist in a substantially purified form, or can exist in a non-native environment, such as, for example, a host cell.

[0156] The term "recombinant" in the context of the present disclosure means "made by genetic engineering." Preferably, a "recombinant entity," such as a recombinant nucleic acid, in the context of the present disclosure is not naturally occurring.

[0157] As used herein, the term "naturally occurring" refers to the fact that an entity can be found in nature. For example, a peptide or nucleic acid that is present in an organism (including viruses), can be isolated from a natural source, and has not been intentionally modified by humans in a laboratory is naturally occurring.

[0158] "Physiological pH," as used herein, refers to a pH of about 7.5.

[0159] The term "genetic modification" or simply "modification" includes the transfection of cells with nucleic acids. The term "transfection" refers to the introduction of nucleic acids, particularly RNA, into cells. For purposes of this disclosure, the term "transfection" also includes the introduction of nucleic acids into cells or the uptake of nucleic acids by such cells, which may be present in a subject, e.g., a patient. Thus, according to the present disclosure, cells for transfection of nucleic acids described herein can be present in vitro or in vivo; for example, the cells can form part of an organ, tissue, and / or organism of a patient. According to the present disclosure, transfection can be transient or stable. For some applications of transfection, transient expression of the transfected genetic material is sufficient. RNA can be transfected into cells to transiently express its encoded protein. Because nucleic acids introduced during the transfection process are typically not integrated into the nuclear genome, the exogenous nucleic acid is diluted or degraded through mitosis. Cells that allow episomal amplification of nucleic acids significantly reduce the dilution rate. If it is desired that the transfected nucleic acid actually remains in the genome of the cell and its daughter cells, stable transfection must be performed. Such stable transfection can be achieved by using a virus-based system or a transposon-based system for transfection. Generally, the nucleic acid encoding the antigen is transiently transfected into the cell. RNA can be transfected into the cell to transiently express the encoded protein.

[0160] The term "seroconversion" includes a 4-fold or greater increase from pre-vaccination to 1 month after dose 2.

[0161] coronavirus Coronaviruses are enveloped, positive-sense, single-stranded RNA ((+)ssRNA) viruses. They have the largest genomes (26–32 kb) of any known RNA virus and are phylogenetically divided into four genera (α, β, γ, and δ), with betacoronaviruses further subdivided into four lineages (A, B, C, and D). Coronaviruses infect a wide range of avian and mammalian species, including humans. Some human coronaviruses generally cause mild respiratory illness, although the disease may be more severe in infants, the elderly, and immunocompromised patients. Middle East respiratory syndrome coronavirus (MERS-CoV) and severe acute respiratory syndrome coronavirus (SARS-CoV), which belong to betacoronavirus lineages C and B, respectively, are highly pathogenic. Both viruses emerged from animal reservoirs into human populations within the past 15 years and caused outbreaks with high case fatality rates. The outbreak of severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2), which causes atypical pneumonia (coronavirus disease 2019, COVID-19), has been raging in China since mid-December 2019 and has developed into a public health emergency of international concern. SARS-CoV-2 (MN908947.3) belongs to the betacoronavirus lineage B and shares at least 70% sequence similarity with SARS-CoV.

[0162] Generally, coronaviruses have four structural proteins: envelope (E), membrane (M), nucleocapsid (N), and spike (S). The E and M proteins have important functions in virus assembly, while the N protein is required for viral RNA synthesis. The key glycoprotein, S, is involved in virus binding and entry into target cells. The S protein is synthesized as a single-chain inactive precursor that is cleaved into two noncovalently associated subunits, S1 and S2, by a furin-like host protease in the producing cell. The S1 subunit contains a receptor-binding domain (RBD) that recognizes host cell receptors. The S2 subunit contains a fusion peptide, two heptad repeats, and a transmembrane domain, all of which undergo extensive conformational rearrangements to mediate fusion of the viral and host cell membranes. The S1 and S2 subunits trimerize to form the large prefusion spike.

[0163] The SARS-CoV-2 S precursor protein can be proteolytically cleaved into S1 (685 aa) and S2 (588 aa) subunits. The S1 subunit contains the receptor-binding domain (RBD), which mediates viral entry into susceptible cells via the host angiotensin-converting enzyme 2 (ACE2) receptor.

[0164] antigen Many embodiments of the present disclosure include the use of RNA encoding an amino acid sequence comprising the SARS-CoV-2 S protein, an immunogenic variant thereof, or an immunogenic fragment of the SARS-CoV-2 S protein or its immunogenic variant. Thus, the RNA encodes a peptide or protein comprising at least an epitope of the SARS-CoV-2 S protein or its immunogenic variant to induce an immune response in a subject against the coronavirus S protein, particularly the SARS-CoV-2 S protein. Amino acid sequences comprising the SARS-CoV-2 S protein, an immunogenic variant thereof, or an immunogenic fragment of the SARS-CoV-2 S protein or its immunogenic variant (i.e., antigenic peptides or proteins) are also referred to herein as "vaccine antigens," "peptide and protein antigens," "antigenic molecules," or simply "antigens." The SARS-CoV-2 S protein, an immunogenic variant thereof, or an immunogenic fragment of the SARS-CoV-2 S protein or its immunogenic variant are also referred to herein as "antigenic peptides or proteins" or "antigenic sequences."

[0165] The SARS-CoV-2 coronavirus full-length spike (S) protein from the first detected SARS-CoV-2 strain (referred to herein as the Wuhan strain) consists of 1273 amino acids and has the amino acid sequence according to SEQ ID NO: 1:

[0166] For purposes of this disclosure, the above sequences are considered to be wild-type or Wuhan SARS-CoV-2 S protein amino acid sequences. Unless otherwise indicated, position numbering in the SARS-CoV-2 S protein given herein is relative to the amino acid sequence according to SEQ ID NO: 1. One of skill in the art reading this disclosure will be able to determine the location of corresponding positions in SARS-CoV-2 S protein variants.

[0167] Table 1 below includes additional exemplary S proteins from various coronavirus variants, including alpha, beta, gamma, delta, and Omicron variants (including Omicron BA.1, BA.2, and BA.4 / 5). Unless otherwise specified, "Omicron variant," as used herein, refers to any Omicron variant, including, for example, the Omicron variants described herein and their progeny.

[0168] The amino acid sequences were obtained from the UniProt database, accessible on the World Wide Web at uniprot.org, or the GenBank database, accessible on the World Wide Web at ncbi.nlm.nih.gov; the UniProt or GenBank database accession numbers for each spike protein sequence are included in Table 1. These amino acid sequences correspond to the amino acid sequences of naturally occurring coronavirus spike proteins. In some embodiments, the amino acid sequences of naturally occurring coronavirus spike proteins encoded by the RNA constructs described herein can be modified, as described herein, to produce immunogenic polypeptides comprising variant coronavirus spike proteins that are modifications of naturally occurring coronavirus spike proteins or fragments thereof. For example, in some embodiments, the amino acid sequences of naturally occurring coronavirus spike proteins encoded by the RNA constructs described herein are substituted to produce immunogenic polypeptides comprising variant coronavirus spike proteins that are modifications of naturally occurring coronavirus spike proteins or fragments thereof, as described herein.

[0169] Similar to the amino acid sequences of naturally occurring coronavirus spike proteins, the amino acid sequences of the spike proteins of these SARS-CoV-2 variants (e.g., including alpha, beta, gamma, delta, and omicron variants (including omicron BA.1, BA.2, BA.4 / 5)) encoded by the RNA constructs described herein can be modified at corresponding positions as described herein to produce immunogenic polypeptides comprising variant coronavirus spike proteins that are modifications of naturally occurring variant coronavirus spike proteins or fragments thereof. For example, in some embodiments, the amino acid sequences of the spike proteins of these SARS-CoV-2 variants encoded by the RNA constructs described herein are substituted to produce immunogenic polypeptides comprising variant coronavirus spike proteins that are modifications of variant coronavirus spike proteins or fragments thereof, as described herein. Additional variants not specifically described below are also contemplated. For example, any variant coronavirus spike protein having 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, or 99% or greater sequence identity to a native coronavirus spike protein sequence encoded by an RNA construct described herein can be modified (e.g., substituted) at the corresponding positions described herein to produce an immunogenic polypeptide comprising a variant coronavirus spike protein that is a modification of the native coronavirus spike protein or a fragment thereof. [Table 1-1] [Table 1-2] [Table 1-3] [Table 1-4] [Table 1-5] [Table 1-6] [Table 1-7] [Table 1-8] [Table 1-9]

[0170] In some embodiments, the coronavirus spike protein sequence comprises SEQ ID NO: 105, shown below, which is the SARS-CoV-2 (Omicron BA.4 / 5) sequence represented by SEQ ID NO: 104 (see Table 1), but differs by one amino acid at position 403 and contains the R403S mutation.

[0171] Coronavirus spike protein modification In certain embodiments, a full-length spike (S) protein (e.g., the full-length S protein of SEQ ID NO: 1) is modified to stabilize the native pre-fusion conformation. Certain mutations that stabilize pre-fusion conformation are known in the art, as disclosed, for example, in WO2022 / 266010A1, WO2021 / 243122A2, and Hsieh, Ching-Lin, et al. ("Structure-based design of prefusion-stabilized SARS-CoV-2 spikes," Science 369.6510(2020):1501-1505), the contents of each of which are incorporated herein by reference in their entirety. In some embodiments, the SARS-CoV-2 S protein may be stabilized by introducing one or more glycine mutations (e.g., one or more glycine mutations at the crown of the helix turn region of the S protein, the 12 amino acids between heptad region 1 (HR1) and the central helix (CH) or heptad region 2 (HR2) region of the S2 subunit, and / or at one or more of L984, D985, K986, and V987 (positions relative to SEQ ID NO:1). In some embodiments, the spike protein comprises a glycine mutation at each of L984, D985, K986, and V987 (i.e., at positions corresponding to these residues in SEQ ID NO:1). In some embodiments, the SARS-CoV-2 S protein may be stabilized by introducing one or more proline mutations. In some embodiments, the SARS-CoV-2 S protein comprises a proline substitution at residues 986 and / or 987 of SEQ ID NO:1. In some embodiments, the SARS-CoV-2 S protein comprises a proline substitution at one or more of residues 817, 892, 899, and 942 of SEQ ID NO: 1. In some embodiments, the SARS-CoV-2 S protein comprises a proline substitution at each of residues 817, 892, 899, and 942 of SEQ ID NO: 1. In some embodiments, the SARS-CoV-2 S protein comprises a proline substitution at each of residues 817, 892, 899, 942, 986, and 987 of SEQ ID NO: 1.

[0172] In some embodiments, the SARS-CoV-2 S protein comprises a proline substitution at residue 985 and / or 987 of SEQ ID NO: 1. In some embodiments, the SARS-CoV-2 S protein comprises a proline substitution at each of residues 817, 892, 899, 942, 985, and 987 of SEQ ID NO: 1.

[0173] In some embodiments, stabilization of the pre-fusion conformation can be achieved by introducing two consecutive proline substitutions at AS residues 986 and 987 in the full-length spike protein. Specifically, a spike (S) protein-stabilizing protein variant is obtained in such a way that the amino acid residue at position 986 is replaced with a proline and the amino acid residue at position 987 is also replaced with a proline. In one embodiment, a SARS-CoV-2 S protein variant in which the native pre-fusion conformation is stabilized comprises the amino acid sequence set forth in SEQ ID NO:7. (SEQ ID NO: 7)

[0174] In some embodiments, the spike protein can be modified to block pre- to post-fusion conformational changes (referred to herein as a "pre- and post-fusion block"). In some embodiments, the pre- and post-fusion block can be introduced by introducing two cysteine ​​mutations at residues close to each other in the folded protein (e.g., at positions close to each other in the pre-fusion conformation of the spike protein). Examples of pre- and post-fusion block mutations include L984C-A989C and I980C-Q992C.

[0175] In some embodiments, the spike protein can be modified to reduce "shedding" (i.e., reduce separation of the S1 and S2 subunits). In some embodiments, the spike protein can be modified to reduce shedding by introducing a mutation at the furin cleavage site (e.g., one or more mutations at residues 682-685 of SEQ ID NO: 1) such that the furin protease can no longer bind to and / or cleave the S protein. In some embodiments, the S protein can be modified to reduce shedding by introducing a mutation at each of residues 682, 683, and 685 (e.g., introducing mutations at (i) R682G, R683S, and R685S, or (ii) R682Q, R683Q, and R685Q).

[0176] In some embodiments, the S protein can be modified to reduce shedding by introducing cysteine ​​mutations that can form disulfide bonds (e.g., by introducing cysteine ​​mutations at positions close to each other in the folded conformation of the S protein, e.g., at residues A570 and N960).

[0177] In some embodiments, one or more modifications may be introduced into the spike protein to stabilize the "up" conformation (referred to herein as an "RBD-up" mutation). Without wishing to be bound by theory, the up conformation of the SARS-CoV-2 spike protein is thought to increase the exposure of neutralization-sensitive residues. Therefore, mutations that stabilize the up conformation may produce more immunogenic vaccines.

[0178] Table 2 below lists various combinations of amino acid modifications that can be introduced into the coronavirus spike protein sequences disclosed above, thereby producing polynucleotides (e.g., RNA) encoding immunogenic polypeptides comprising coronavirus spike proteins that are variants of native coronavirus spike proteins or fragments thereof. The "+" symbol indicates the inclusion of a particular modification in a particular S protein sequence from a coronavirus strain or variant (e.g., a SARS-CoV-2 strain and / or variant listed in Table 1). In some examples, a spike protein sequence may contain any combination of modifications in Table 2 below. The amino acid positions listed in Table 2 are numbered relative to SEQ ID NO: 1 (Wuhan), SEQ ID NO: 69 (Omicron BA.1), SEQ ID NO: 70 (Omicron BA.2), and SEQ ID NO: 104 (Omicron BA.4 / 5). Corresponding amino acid positions in spike protein sequences from other coronavirus variants (e.g., alpha, beta, or delta variants) can be determined by alignment with SEQ ID NO: 1 (see, e.g., Table 5). [Table 2-1] [Table 2-2] [Table 2-3] [Table 2-4] [Table 2-5] [Table 2-6] [Table 2-7] [Table 2-8]

[0179] Table 3 below lists various combinations of amino acid modifications that can be introduced into the coronavirus spike protein sequences disclosed above, thereby enabling the production of polynucleotides (e.g., RNA) encoding immunogenic polypeptides comprising coronavirus spike proteins that are variants of native coronavirus spike proteins or fragments thereof. Similar to Table 2, Table 3 lists the positions of amino acid modifications (with respect to the Wuhan spike protein sequence according to SEQ ID NO: 1), and Table 3 also includes specific amino acid residues that are substituted with native amino acid residues. A "+" symbol indicates the inclusion of a specific modification in a particular S protein sequence from a coronavirus strain or variant (e.g., a SARS-CoV-2 strain and / or variant listed in Table 1). In some examples, coronavirus spike protein variants encoded by RNA vaccines may include any combination of the modifications in Table 2 above, such as any of the specific substitutions listed in Table 3. The amino acid positions listed in Table 2 are numbered relative to SEQ ID NO: 1 (Wuhan), SEQ ID NO: 69 (Omicron BA.1), SEQ ID NO: 70 (Omicron BA.2), and SEQ ID NO: 104 (Omicron BA.4 / 5). The amino acid positions corresponding to spike protein sequences from other coronavirus variants (e.g., alpha, beta, or delta variants) can be determined by alignment with SEQ ID NO: 1 (see, e.g., Table 5). [Table 3-1] [Table 3-2] [Table 3-3] [Table 3-4] [Table 3-5] [Table 3-6] [Table 3-7] [Table 3-8]

[0180] Table 4 below lists various combinations of amino acid modifications that can be introduced into the coronavirus spike protein sequences disclosed herein (see, e.g., Table 1), thus enabling the production of polynucleotides (e.g., RNA) encoding immunogenic polypeptides comprising coronavirus spike proteins that are variants of native coronavirus spike proteins or fragments thereof. The "+" symbol indicates the inclusion of a particular modification in a particular S protein sequence from a coronavirus strain or variant (e.g., a SARS-CoV-2 strain and / or variant listed in Table 1). In some examples, a spike protein sequence may contain any combination of the modifications in Table 4 below. The amino acid positions shown in Table 4 correspond to SEQ ID NO: 69 (SARS-CoV-2 omicron UFO69279.1 (BA.1, formerly B.1.1.529) in Table 1), and corresponding amino acid positions in other coronavirus spike proteins can be determined by sequence alignment (see, e.g., the alignment of various coronavirus spike protein sequences in Table 5). [Table 4-1] [Table 4-2] [Table 4-3] [Table 4-4] [Table 4-5] [Table 4-6] [Table 4-7]

[0181] In some embodiments, the amino acid corresponding to amino acid 326 of SEQ ID NO:69 (SARS-CoV-2 omicron UFO69279.1 (BA.1, formerly B.1.1.529) can be substituted to produce a variant coronavirus spike protein encoded by an RNA as described herein. In some embodiments, the amino acid corresponding to amino acid 326 of SEQ ID NO:69 can be substituted with a serine residue to produce a variant coronavirus spike protein encoded by an RNA as described herein. The substitution at 326 with a serine residue can be referred to herein as 326S.

[0182] In some embodiments, the amino acid corresponding to amino acid position 364 of SEQ ID NO:69 (SARS-CoV-2 omicron UFO69279.1 (BA.1, formerly B.1.1.529) can be substituted to produce a variant coronavirus spike protein encoded by an RNA as described herein. In some embodiments, the amino acid corresponding to amino acid position 364 of SEQ ID NO:69 can be substituted with a phenylalanine residue to produce a variant coronavirus spike protein encoded by an RNA as described herein. The substitution at 364 with a phenylalanine residue can be referred to herein as 364F.

[0183] In some embodiments, the amino acid corresponding to amino acid 567 of SEQ ID NO:69 can be substituted to produce a variant coronavirus spike protein encoded by the RNA described herein. In some embodiments, the amino acid corresponding to amino acid 567 of SEQ ID NO:69 can be substituted with a cysteine ​​residue to produce a variant coronavirus spike protein encoded by the RNA described herein. The substitution of 567 with a cysteine ​​residue can be referred to herein as 567C.

[0184] In some embodiments, the amino acid corresponding to amino acid 611 of SEQ ID NO:69 can be substituted to produce a variant coronavirus spike protein encoded by the RNA described herein. In some embodiments, the amino acid corresponding to amino acid 611 of SEQ ID NO:69 can be substituted with a glycine residue to produce a variant coronavirus spike protein encoded by the RNA described herein. The substitution of glycine at 611 can be referred to herein as 611G.

[0185] In some embodiments, the amino acid corresponding to amino acid 814 of SEQ ID NO:69 can be substituted to produce a variant coronavirus spike protein encoded by the RNA described herein. In some embodiments, the amino acid corresponding to amino acid 814 of SEQ ID NO:69 can be substituted with a phenylalanine residue to produce a variant coronavirus spike protein encoded by the RNA described herein. The substitution of phenylalanine at 814 can be referred to herein as 814P.

[0186] In some embodiments, the amino acid corresponding to amino acid 840 of SEQ ID NO:69 can be substituted to produce a variant coronavirus spike protein encoded by the RNA described herein. In some embodiments, the amino acid corresponding to amino acid 840 of SEQ ID NO:69 can be substituted with an asparagine residue to produce a variant coronavirus spike protein encoded by the RNA described herein. The substitution of an asparagine residue at 840 can be referred to herein as 840N.

[0187] In some embodiments, the amino acid corresponding to amino acid 851 of SEQ ID NO: 69 can be substituted to produce a variant coronavirus spike protein encoded by the RNA described herein. In some embodiments, the amino acid corresponding to amino acid 851 of SEQ ID NO: 69 can be substituted with a phenylalanine residue to produce a variant coronavirus spike protein encoded by the RNA described herein. The substitution of phenylalanine at 851 can be referred to herein as 851F.

[0188] In some embodiments, the amino acid corresponding to amino acid 889 of SEQ ID NO:69 can be substituted to produce a variant coronavirus spike protein encoded by the RNA described herein. In some embodiments, the amino acid corresponding to amino acid 889 of SEQ ID NO:69 can be substituted with a proline residue to produce a variant coronavirus spike protein encoded by the RNA described herein. The substitution of a proline residue at 889 can be referred to herein as 889P.

[0189] In some embodiments, the amino acid corresponding to amino acid 896 of SEQ ID NO:69 can be substituted to produce a variant coronavirus spike protein encoded by the RNA described herein. In some embodiments, the amino acid corresponding to amino acid 896 of SEQ ID NO:69 can be substituted with a proline residue to produce a variant coronavirus spike protein encoded by the RNA described herein. The substitution of 896 with a proline residue can be referred to herein as 896P.

[0190] In some embodiments, the amino acid corresponding to amino acid 939 of SEQ ID NO:69 can be substituted to produce a variant coronavirus spike protein encoded by the RNA described herein. In some embodiments, the amino acid corresponding to amino acid 939 of SEQ ID NO:69 can be substituted with a proline residue to produce a variant coronavirus spike protein encoded by the RNA described herein. The substitution of a proline residue at 939 can be referred to herein as 939P.

[0191] In some embodiments, the amino acid corresponding to amino acid 957 of SEQ ID NO:69 can be substituted to produce a variant coronavirus spike protein encoded by the RNA described herein. In some embodiments, the amino acid corresponding to amino acid 957 of SEQ ID NO:69 can be substituted with a cysteine ​​residue to produce a variant coronavirus spike protein encoded by the RNA described herein. The substitution of 957 with a cysteine ​​residue can be referred to herein as 957C.

[0192] In some embodiments, the amino acid corresponding to amino acid 977 of SEQ ID NO:69 can be substituted to produce a variant coronavirus spike protein encoded by the RNA described herein. In some embodiments, the amino acid corresponding to amino acid 977 of SEQ ID NO:69 can be substituted with a cysteine ​​residue to produce a variant coronavirus spike protein encoded by the RNA described herein. The substitution of 977 with a cysteine ​​residue can be referred to herein as 977C.

[0193] In some embodiments, the amino acid corresponding to amino acid 981 of SEQ ID NO:69 can be substituted to produce a variant coronavirus spike protein encoded by the RNA described herein. In some embodiments, the amino acid corresponding to amino acid 981 of SEQ ID NO:69 can be substituted with a cysteine ​​residue to produce a variant coronavirus spike protein encoded by the RNA described herein. The substitution of 981 with a cysteine ​​residue can be referred to herein as 981C.

[0194] In some embodiments, the amino acid corresponding to amino acid 982 of SEQ ID NO:69 can be substituted to produce a variant coronavirus spike protein encoded by the RNA described herein. In some embodiments, the amino acid corresponding to amino acid 982 of SEQ ID NO:69 can be substituted with a proline residue to produce a variant coronavirus spike protein encoded by the RNA described herein. The substitution of 982 with a proline residue can be referred to herein as 982P.

[0195] In some embodiments, the amino acid corresponding to amino acid 983 of SEQ ID NO:69 can be substituted to produce a variant coronavirus spike protein encoded by the RNA described herein. In some embodiments, the amino acid corresponding to amino acid 983 of SEQ ID NO:69 can be substituted with a proline residue to produce a variant coronavirus spike protein encoded by the RNA described herein. The substitution of 983 with a proline residue can be referred to herein as 983P.

[0196] In some embodiments, the amino acid corresponding to amino acid 984 of SEQ ID NO:69 can be substituted to produce a variant coronavirus spike protein encoded by the RNA described herein. In some embodiments, the amino acid corresponding to amino acid 984 of SEQ ID NO:69 can be substituted with a proline residue to produce a variant coronavirus spike protein encoded by the RNA described herein. The substitution of 983 with a proline residue can be referred to herein as 984P.

[0197] In some embodiments, the amino acid corresponding to amino acid 986 of SEQ ID NO:69 can be substituted to produce a variant coronavirus spike protein encoded by the RNA described herein. In some embodiments, the amino acid corresponding to amino acid 986 of SEQ ID NO:69 can be substituted with a cysteine ​​residue to produce a variant coronavirus spike protein encoded by the RNA described herein. The substitution of 986 with a cysteine ​​residue can be referred to herein as 986C.

[0198] In some embodiments, the amino acid corresponding to amino acid 989 of SEQ ID NO:69 can be substituted to produce a variant coronavirus spike protein encoded by the RNA described herein. In some embodiments, the amino acid corresponding to amino acid 989 of SEQ ID NO:69 can be substituted with a cysteine ​​residue to produce a variant coronavirus spike protein encoded by the RNA described herein. The substitution of 989 with a cysteine ​​residue can be referred to herein as 989C.

[0199] In some embodiments, the variant spike proteins encoded by the RNA described herein have at least or up to 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, and / or 18 of the following modifications at positions 326, 364, 567, 611, 814, 840, 851, 889, 896, 939 as set forth in the corresponding amino acids of the SARS-CoV-2 Omicron (BA.1, formerly B.1.1.529) spike protein, UniProt Accession Number UFO69279.1, or the spike protein of another coronavirus: in some embodiments, the modifications at that position or the corresponding position 326 are serine, 364 is phenylalanine, 567 is cysteine, 611 is glycine, 814 is proline, 840 is asparagine, and 851 is phenylalanine. nin, 889 is proline, 896 is proline, 939 is proline, 957 is cysteine, 977 is cysteine, 981 is cysteine, 982 is proline, 983 is proline, 984 is proline, 986 is cysteine, and 989 is cysteine, and in a further embodiment, the variant spike protein is SARS-CoV-2 omicron (BA.1, formerly B.1. 1.529) spike protein UniProt accession number UFO69279.1 has at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, 99.1%, 99.2%, 99.3%, 99.4%, 99.5%, 99.6%, 99.7%, 99.8%, or 99.9% sequence identity to spike protein UniProt accession number UFO69279.1. In some examples, the modifications described herein, alone or in combination with any one or more additional modifications described herein, may produce an RNA (e.g., as described herein) encoding an immunogenic polypeptide comprising a variant coronavirus spike protein that is a variant of a native coronavirus spike protein or a fragment thereof.In some embodiments, these modifications can (a) increase recruitment by the RBD of the variant coronavirus spike protein of the RBD-up conformation, exposing more neutralization-sensitive epitopes on the spike protein; (b) decrease recruitment by the RBD of the variant coronavirus spike protein of the RBD-down conformation; (c) increase expression of the variant coronavirus spike protein compared to the native coronavirus spike protein; (d) increase adoption of the pre-fusion conformation; (e) decrease shedding of the S1 subunit of the variant coronavirus spike protein; and / or (f) improve localization of the variant coronavirus spike protein to the host cell membrane.

[0200] The mutations described herein and in, for example, Tables 2A, 2B, and 2C may be introduced into the S protein sequences of other coronavirus strains or variant sequences, or immunogenic fragments thereof, and the corresponding positions may be determined by sequence alignment with SEQ ID NO: 69 (see, for example, Table 5).

[0201] In some embodiments, the variant spike protein encoded by the RNA described herein comprises at least one of the following modifications at positions 326, 364, 567, 611, 814, 840, 851, 889, 896, 939, 957, 977, 981, 982, 983, 984, 986, 989 as set forth in the corresponding amino acids of the SARS-CoV-2 Omicron (BA.1, formerly B.1.1.529) spike protein, UniProt Accession Number UFO69279.1, or the spike protein of another coronavirus: or at most 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, and / or 18, and in some embodiments the modification at that position or the corresponding 326 position is to any amino acid except phenylalanine, 364 is any amino acid except valine, 567 is any amino acid except alanine, 611 is any amino acid except glycine, 814 is any amino acid except phenylalanine, 840 is any amino acid except aspartic acid, and 851 is any amino acid except lysine. 981 is any amino acid except leucine, 982 is any amino acid except aspartic acid, 983 is any amino acid except lysine, 984 is any amino acid except valine, 986 is any amino acid except alanine, 989 is any amino acid except alanine, 990 is any amino acid except alanine, 991 is any amino acid except leucine, 992 is any amino acid except aspartic acid, 993 is any amino acid except leucine, 994 is any amino acid except valine, 995 is any amino acid except alanine, 996 is any amino acid except alanine, 997 is any amino acid except isoleucine, 998 is any amino acid except leucine, 999 is any amino acid except leucine, 996 is any amino acid except alanine, 997 is any amino acid except isoleucine, 998 is any amino acid except isoleucine, 999 is any amino acid except leucine, 999 is any amino acid except isole ... is any amino acid except glutamine, and in further embodiments, the variant spike protein has a sequence identity of at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, 99.1%, 99.2%, 99.3%, 99.4%, 99.5%, 99.6%, 99.7%, 99.8% or 99.9% to the SARS-CoV-2 Omicron (BA.1, formerly B.1.1.529) spike protein, UniProt accession number UFO69279.1.8%, or 99.9% sequence identity. In some examples, the modifications described herein, alone or in combination with any one or more additional modifications described herein, can produce RNA encoding an isolated immunogenic polypeptide comprising a variant coronavirus spike protein that is a variant of a native coronavirus spike protein or a fragment thereof. In some embodiments, these modifications can (a) increase recruitment by the RBD of the variant coronavirus spike protein of the RBD-up conformation, exposing more neutralization-sensitive epitopes on the spike protein; (b) decrease recruitment by the RBD of the variant coronavirus spike protein of the RBD-down conformation; (c) increase expression of the variant coronavirus spike protein compared to the native coronavirus spike protein; (d) increase adoption of the prefusion conformation; (e) decrease shedding of the S1 subunit of the variant coronavirus spike protein; and / or (f) improve localization of the variant coronavirus spike protein to the host cell membrane.

[0202] The amino acid in each human coronavirus spike protein sequence and its corresponding position relative to SEQ ID NO: 1 can be determined based on an alignment of the protein sequences. Table 5 below is an alignment of human coronavirus spike protein sequences (e.g., the spike protein sequences in Table 1). The highlighted positions in the alignment below correspond to the positions of the modified amino acids identified in Table 2 above. [Table 5-1] [Table 5-2] [Table 5-3] [Table 5-4] [Table 5-5] [Table 5-6]

[0203] coronavirus variants Those of skill in the art will be aware of various spike variants and / or resources documenting them. For example, the following strains, their SARS-CoV-2 S protein amino acid sequences, and particularly their modifications compared to the wild-type SARS-CoV-2 S protein amino acid sequence, e.g., compared to SEQ ID NO: 1, are useful herein:

[0204] B.1.1.7 ("Variant of Concern 202012 / 01" (VOC-202012 / 01) B.1.1.7 is a SARS-CoV-2 variant first detected in October 2020 during the COVID-19 pandemic in the UK in samples collected the previous month and quickly began spreading by mid-December. This correlated with a significant increase in COVID-19 infection rates in the UK, an increase that is thought to be at least in part due to an N501Y mutation in the receptor-binding domain of the spike glycoprotein, which is required for binding to ACE2 on human cells. The B.1.1.7 variant is defined by 23 mutations: 13 nonsynonymous mutations, 4 deletions, and 6 synonymous mutations (i.e., 17 protein-altering mutations and 6 non-altering mutations). Spike protein changes in B.1.1.7 include deletions 69-70, 144, N501Y, A570D, D614G, P681H, T716I, S982A, and D1118H.

[0205] B.1.351(501.V2) The B.1.351 lineage, colloquially known as the South African COVID-19 variant, is a variant of SARS-CoV-2. Preliminary results indicate that this variant may have increased transmissibility. The B.1.351 variant is defined by multiple spike protein changes, including L18F, D80A, D215G, deletions 242-244, R246I, K417N, E484K, N501Y, D614G, and A701V. Three mutations of particular interest reside in the spike region of the B.1.351 genome: K417N, E484K, and N501Y.

[0206] B.1.1.298 (Cluster 5) B.1.1.298 was discovered in North Denmark and is believed to have been transmitted from minks to humans via mink farms. Several different mutations in the virus's spike protein have been identified. Specific mutations include deletions 69-70, Y453F, D614G, I692V, M1229I, and optionally S1147L.

[0207] P.1(B.1.1.248) The lineage B.1.1.248, known as the Brazil (Brazilian) variant, is one of the SARS-CoV-2 variants designated the P.1 lineage. P.1 has several S protein modifications [L18F, T20N, P26S, D138Y, R190S, K417T, E484K, N501Y, D614G, H655Y, T1027I, V1176F] and is similar to the South African variant B.1.351 at certain critical RBD positions (K417, E484, N501).

[0208] B.1.427 / B.1.429(CAL.20C) Lineage B.1.427 / B.1.429, also known as CAL.20C, is defined by the following modifications in the S-protein: S13I, W152C, L452R, and D614G (of which the L452R modification is of particular concern). The CDC lists B.1.427 / B.1.429 as a "variant of concern."

[0209] B.1.525 B.1.525 has the same E484K modification found in the P.1 and B.1.351 variants, and the same ΔH69 / ΔV70 deletion found in B.1.1.7 and B.1.1.298. It also has the modifications D614G, Q677H, and F888L.

[0210] B.1.526 B.1.526 was detected as an emerging lineage of virus isolates in the New York area that shares mutations with previously reported variants. The most common set of spike mutations in this lineage are L5F, T95I, D253G, E484K, D614G, and A701V.

[0211] The table below provides an overview of circulating SARS-CoV-2 strains that are VOI / VOC. [Table 6]

[0212] B.1.1.529 B.1.529 ("Omicron") was first detected in South Africa in November 2021. Omicron was found to grow approximately 70 times faster than the delta variant and quickly became the dominant strain of SARS-CoV-2 worldwide. Since its initial detection, several sublineages have emerged. Current Omicron variants of concern, along with certain characteristic mutations associated with their respective S proteins (mutation positions shown relative to SEQ ID NO: 1), are listed in Table 3A below. In some embodiments, the BA.4 and BA.5 variants have the same S protein amino acid sequence, in which case the term "BA.4 / 5" may be used to refer to the amino acid sequence of the S protein that can be found in either BA.4 or BA.5. Similarly, in some embodiments, the BA.4.6 and BF.7 variants have the same protein amino acid sequence, in which case the term "BA.4.6 / BF.7" may be used to refer to the amino acid sequence of the S protein present in either BA.4.6 or BF.7. [Table 7-1] [Table 7-2] [Table 7-3]

[0213] In addition to the Omicron variants described above, additional variants of BA.5 (e.g., such variants including BF.14) have been observed that contain one or more of the following mutations in the S protein (positions shown relative to SEQ ID NO: 1): E340X (e.g., E340K), R346X (e.g., R346T, R346I, or R346S), K444X (e.g., K444N or K444T), V445X, N450D, and S:N460X (e.g., N460K).

[0214] In some embodiments, the RNA described herein comprises a nucleotide sequence encoding a SARS-CoV-2 S protein that includes one or more (e.g., including 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, or more) mutations characteristic of an Omicron variant (e.g., one or more mutations of an Omicron variant listed in Table 3A). In some embodiments, the RNA comprises a nucleotide sequence encoding a SARS-CoV-2 S protein that includes one or more (e.g., including 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, or more) mutations listed in Table 3A. In some such embodiments, the one or more mutations may be derived from two or more variants as listed in Table 3A. In some embodiments, the RNA comprises a nucleotide sequence encoding a SARS-CoV-2 S protein that includes each of the mutations identified in Table 3A as being characteristic of a particular Omicron variant (e.g., in some embodiments, the RNA comprises a nucleotide sequence encoding a SARS-CoV-2 S protein that includes each of the mutations listed in Table 3A as being characteristic of Omicron BA.1, BA.2, BA.2.12.1, BA.4 / 5, BA.2.75, BA.2.75.1, BA.4.6, BQ.1.1, XBB, XBB.1, XBB.2, or XBB.1.3).

[0215] In some embodiments, the RNA disclosed herein comprises a nucleotide sequence encoding an immunogenic fragment of the SARS-CoV-2 S protein (e.g., the RBD) and includes one or more mutations characteristic of a SARS-CoV-2 variant (e.g., the Omicron variant described herein). For example, in some embodiments, the RNA includes a nucleotide sequence encoding the RBD of the S protein of a SARS-CoV-2 variant (e.g., the region of the S protein corresponding to amino acids 327-528 of SEQ ID NO: 1, which region includes one or more mutations within this region that are characteristic of a variant of concern).

[0216] In some embodiments, the RNA encodes a SARS-CoV-2 S protein that includes a subset of mutations listed in Table 3A. In some embodiments, the RNA encodes a SARS-CoV-2 S protein that includes the mutations listed in Table 3A that are most common for a particular variant (e.g., mutations detected in at least 30%, at least 40%, at least 50%, at least 60%, at least 70%, at least 80%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% of the sequences collected to date for a given variant sequenced). Mutation prevalence can be determined, for example, based on published sequences (e.g., sequences collected and made publicly available by GISAID).

[0217] In some embodiments, the RNA described herein encodes a SARS-CoV-2 S protein that includes one or more mutations characteristic of the BA.4 / 5 variant. In some embodiments, the one or more mutations characteristic of the BA.4 / 5 variant include T19I, Δ24-26, A27S, Δ024-26, A27S, Δ69 / 70, G142D, G142D, V213G, G339D, S371F, S373P, S375F, T376A, D405N, K417N, N440K, L452R, S477N, T478K, E484A, F486V, Q498R, N501Y, Y505H, D614G, H655Y, N679K, P681H, N764K, D796Y, Q954H, and N969K. In some embodiments, the RNA described herein encodes a SARS-CoV-2 S protein that includes one or more mutations characteristic of the BA.4 / 5 variant, excluding R408S. In some embodiments, the RNA described herein encodes a SARS-CoV-2 S protein that includes one or more (e.g., including 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, or more) mutations characteristic of the BA.4 / 5 variant, excluding R408S.

[0218] In some embodiments, the RNA described herein encodes a SARS-CoV-2 S protein that includes one or more (e.g., including 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, or more) mutations characteristic of the BA.2.75 variant. In some embodiments, one or more mutations characteristic of the BA.2.75 variant include T19I, Δ24-26, A27S, G142D, K147E, W152R, F157L, I210V, V213G, G257S, G339H, S371F, S373P, S375F, T376A, D405N, R408S, K417N, N440K, G446S, N460K, S477N, T478K, E484A, F486V, Q498R, N501Y, Y505H D614G, H655Y, N679K, P681H, N764K, D796Y, Q954H, and N969K. In some embodiments, the RNAs described herein encode a SARS-CoV-2 S protein that includes one or more mutations characteristic of the BA.4 / 5 variant, excluding R408S. In some embodiments, the RNAs described herein encode a SARS-CoV-2 S protein that includes one or more (e.g., including 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, or more) mutations characteristic of the BA.4 / BA.5 variant, excluding R408S and N354D. In some embodiments, the RNAs described herein encode a SARS-CoV-2 S protein that includes one or more (e.g., including 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, or more) mutations characteristic of the BA.2.75 variant.In some embodiments, one or more mutations characteristic of the BA.2.75 variant include T19I, Δ24-26, A27S, G142D, K147E, W152R, F157L, I210V, V213G, G257S, G339H, S371F, S373P, S375F, T376A, D405N, R408S, K417N, N440K, G446S, N460K, S477N, T478K, E484A, Q498R, N501Y, Y505H D614G, H655Y, N679K, P681H, N764K, Q954H, and N969K. In some embodiments, the RNAs described herein encode a SARS-CoV-2 S protein that includes one or more (e.g., including 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, or more) mutations characteristic of the BA.2.75 variant, excluding N354D. In some embodiments, the RNAs described herein encode a SARS-CoV-2 S protein that includes one or more (e.g., including 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, or more) mutations characteristic of the BA.2.75 variant, excluding D796Y. In some embodiments, the RNA described herein encodes a SARS-CoV-2 S protein that contains one or more (e.g., including 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, or more) mutations characteristic of the BA.2.75 variant, excluding D796Y and N354D.

[0219] In some embodiments, the RNA described herein encodes a SARS-CoV-2 S protein that includes one or more mutations characteristic of the BA.2.75.2 variant. In some embodiments, the one or more mutations characteristic of the BA.2.75.2 variant are T19I, Δ24-26, A27S, G142D, K147E, W152R, F157L, I210V, V213G, G257S, G339H, R346T, N354D, S371F, S373P, S375F, T376 A, D405N, R408S, K417N, N440K, G446S, N460K, S477N, T478K, E484A, F486S, Q498R, N501Y, Y505H, D614G, H655Y, N679K, P681H, N764K, D796Y, Q954H, N969K, and D1199N. In some embodiments, the RNA described herein encodes a SARS-CoV-2 S protein that includes one or more (e.g., including 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, or more) mutations characteristic of the BA.2.75.2 variant, excluding R346T.

[0220] In some embodiments, the RNA described herein encodes a SARS-CoV-2 S protein that includes one or more mutations characteristic of the BA.4.6 or BF.7 variant. In some embodiments, one or more mutations characteristic of a BA.4.6 or BF.7 variant include T19I, Δ24-26, A27S, Δ69 / 70, G142D, V213G, G339D, R346T, S371F, S373P, S375F, T376A, D405N, K417N, N440K, L452R, S477N, T478K, E484A, F486V, Q498R, N501Y, Y505H, D614G, H655Y, N679K, P681H, N764K, D796Y, Q954H, and N969K. In some embodiments, the RNAs described herein encode a SARS-CoV-2 S protein that includes one or more (e.g., including 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, or more) mutations characteristic of the BA.4.6 or BF.7 variant, excluding R408S. In some embodiments, the RNAs described herein encode a SARS-CoV-2 S protein that includes one or more (e.g., including 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, or more) mutations characteristic of the BA.4.6 or BF.7 variant, excluding N658S. In some embodiments, the RNA described herein encodes a SARS-CoV-2 S protein that contains one or more (e.g., including 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, or more) mutations characteristic of the BA.4.6 or BF.7 variant, excluding N658S and R408S.

[0221] In some embodiments, the RNA described herein encodes a SARS-CoV-2 S protein that includes one or more mutations characteristic of an Omicron XBB variant. In some embodiments, the one or more mutations characteristic of an Omicron XBB variant are T19I, Δ24-26, A27S, V83A, G142D, Δ144, H146Q, Q183E, V213E, G339H, R346T, L368I, S371F, S373P, S375F, T376A, D405N , R408S, K417N, N440K, V445P, G446S, N460K, S477N, T478K, E484A, F486S, F490S, Q498R, N501Y, Y505H, D614G, H655Y, N679K, P681H, N764K, D796Y, Q954H, and N969K.

[0222] In some embodiments, the RNA described herein encodes a SARS-CoV-2 S protein that includes one or more mutations characteristic of an Omicron XBB.1 variant. In some embodiments, the one or more mutations characteristic of an Omicron XBB.1 variant include G252V. In some embodiments, the one or more mutations characteristic of an Omicron XBB.1 variant include T19I, Δ24-26, A27S, V83A, G142D, Δ144, H146Q, Q183E, V213E, G252V, G339H, R346T, L368I, S371F, S373P, S375F, T376A, D 405N, R408S, K417N, N440K, V445P, G446S, N460K, S477N, T478K, E484A, F486S, F490S, Q498R, N501Y, Y505H, D614G, H655Y, N679K, P681H, N764K, D796Y, Q954H, and N969K. In some embodiments, the RNAs described herein encode a SARS-CoV-2 S protein that includes one or more (e.g., including 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, or more) mutations characteristic of Omicron XBB.1 variants, excluding Q493R. In some embodiments, the RNAs described herein encode a SARS-CoV-2 S protein that contains one or more (e.g., including 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, or more) mutations characteristic of the Omicron XBB variant, excluding Q493R and G252V.

[0223] In some embodiments, the RNA described herein encodes a SARS-CoV-2 S protein that includes one or more mutations characteristic of Omicron XBB.2 variants. In some embodiments, the one or more mutations characteristic of Omicron XBB.2 variants include D253G. In some embodiments, the one or more mutations characteristic of Omicron XBB.2 variants include T19I, Δ24-26, A27S, V83A, G142D, Δ144, H146Q, Q183E, V213E, D253G, G339H, R346T, L368I, S371F, S373P, S375F, T376A, D405 N, R408S, K417N, N440K, V445P, G446S, N460K, S477N, T478K, E484A, F486S, F490S, Q493R, Q498R, N501Y, Y505H, D614G, H655Y, N679K, P681H, N764K, D796Y, Q954H, and N969K.

[0224] In some embodiments, the RNA described herein encodes a SARS-CoV-2 S protein that includes one or more mutations characteristic of Omicron XBB.1.3 variants. In some embodiments, the one or more mutations characteristic of Omicron XBB.1.3 variants include G252V and A484T. In some embodiments, the one or more mutations characteristic of Omicron XBB.1.3 variants include T19I, Δ24-26, A27S, V83A, G142D, Δ144, H146Q, Q183E, V213E, G252V, G339H, R346T, L368I, S371F, S373P, S375F, T376A, D40 5N, R408S, K417N, N440K, V445P, G446S, N460K, S477N, T478K, A484T, F486S, F490S, Q493R, Q498R, N501Y, Y505H, D614G, H655Y, N679K, P681H, N764K, D796Y, Q954H, and N969K.

[0225] In some embodiments, the RNA described herein encodes a SARS-CoV-2 S protein that includes one or more mutations characteristic of the BQ.1.1 variant. In some embodiments, one or more mutations characteristic of a BQ.1.1 variant include T19I, Δ24-26, A27S, Δ69 / 70, G142D, V213G, G339D, S371F, S373P, S375F, T376A, D405N, R408S, K417N, N440K, K444T, L452R, N463K, S477N, T478K, E484A, F486V, Q498R, N501Y, Y505H, D614G, H655Y, N679K, P681H, N764K, D796Y, Q954H, and N969K. In some embodiments, the RNA described herein encodes a SARS-CoV-2 S protein that includes one or more (e.g., including 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, or more) mutations characteristic of the BQ.1.1 variant.

[0226] In one embodiment, the vaccine antigen described herein comprises, consists essentially of, or consists of the spike protein (S) of SARS-CoV-2, a variant thereof, or a fragment thereof, and includes one or more mutations characteristic of SARS-CoV-2 variants (e.g., one or more of the mutations associated with the omicron variant listed in Table 3A).

[0227] In one embodiment, the vaccine antigen comprises (a) the amino acid sequence of amino acids 17 to 1273 of SEQ ID NO: 1 or 7, an amino acid sequence having at least 99%, 98%, 97%, 96%, 95%, 90%, 85%, or 80% identity to the amino acid sequence of amino acids 17 to 1273 of SEQ ID NO: 1 or 7, or an immunogenic fragment of the amino acid sequence of amino acids 17 to 1273 of SEQ ID NO: 1 or 7, or an amino acid sequence having at least 99%, 98%, 97%, 96%, 95%, 90%, 85%, or 80% identity to an immunogenic fragment of the amino acid sequence of amino acids 17 to 1273 of SEQ ID NO: 1 or 7, and (b) one or more mutations associated with a SARS-CoV-2 variant of concern (e.g., one or more mutations listed in Table 3A). In one embodiment, the vaccine antigen comprises the amino acid sequence of amino acids 17 to 1273 of SEQ ID NO: 1 or 7, and includes one or more mutations associated with a SARS-CoV-2 variant of concern (e.g., one or more mutations listed in Table 3A).

[0228] In one embodiment, the vaccine antigen comprises (a) the amino acid sequence of amino acids 17 to 1273 of SEQ ID NO: 80, an amino acid sequence having at least 99%, 98%, 97%, 96%, 95%, 90%, 85%, or 80% identity to the amino acid sequence of amino acids 17 to 1273 of SEQ ID NO: 80, or an immunogenic fragment of the amino acid sequence of amino acids 17 to 1273 of SEQ ID NO: 80, or an amino acid sequence having at least 99%, 98%, 97%, 96%, 95%, 90%, 85%, or 80% identity to an immunogenic fragment of the amino acid sequence of amino acids 17 to 1273 of SEQ ID NO: 80, and (b) one or more of the mutations listed in Table 3A. In one embodiment, the vaccine antigen comprises the amino acid sequence of amino acids 17 to 1273 of SEQ ID NO: 80, and includes one or more mutations associated with a SARS-CoV-2 variant of concern (e.g., one or more mutations listed in Table 3A).

[0229] In one embodiment, the RNA encoding the vaccine antigen is selected from the group consisting of (i) the nucleotide sequence of nucleotides 49 to 3819 of SEQ ID NO: 2, 8, or 9, a nucleotide sequence having at least 99%, 98%, 97%, 96%, 95%, 90%, 85%, or 80% identity to the nucleotide sequence of nucleotides 49 to 3819 of SEQ ID NO: 2, 8, or 9, or a fragment of the nucleotide sequence of nucleotides 49 to 3819 of SEQ ID NO: 2, 8, or 9, or a nucleotide sequence having at least 99%, 98%, 97%, 96%, 95%, 90%, 85%, or 80% identity to a fragment of the nucleotide sequence of nucleotides 49 to 3819 of SEQ ID NO: 2, 8, or 9, and / or (ii) the nucleotide sequence of SEQ ID NO: 1 or 7. 7, or an immunogenic fragment of the amino acid sequence of amino acids 17-1273 of SEQ ID NO: 1 or 7, or an amino acid sequence having at least 99%, 98%, 97%, 96%, 95%, 90%, 85%, or 80% identity to the amino acid sequence of amino acids 17-1273 of SEQ ID NO: 1 or 7; and (b) a nucleotide sequence encoding a SARS-CoV-2 S protein containing one or more mutations associated with a SARS-CoV-2 variant of concern (e.g., one or more mutations listed in Table 3A). In one embodiment, the RNA encoding the vaccine antigen (a) encodes an amino acid sequence comprising (i) the nucleotide sequence of nucleotides 49 to 3819 of SEQ ID NO: 2, 8, or 9, and / or (ii) the amino acid sequence of amino acids 17 to 1273 of SEQ ID NO: 1 or 7, and (b) comprises one or more mutations characteristic of SARS-CoV-2 of concern (e.g., one or more mutations listed in Table 3A).

[0230] In one embodiment, the RNA encoding the vaccine antigen is selected from the group consisting of (i) the nucleotide sequence of nucleotides 49 to 3819 of SEQ ID NO: 81, a nucleotide sequence having at least 99%, 98%, 97%, 96%, 95%, 90%, 85%, or 80% identity to the nucleotide sequence of nucleotides 49 to 3819 of SEQ ID NO: 81, or a fragment of the nucleotide sequence of nucleotides 49 to 3819 of SEQ ID NO: 81, or a nucleotide sequence having at least 99%, 98%, 97%, 96%, 95%, 90%, 85%, or 80% identity to a fragment of the nucleotide sequence of nucleotides 49 to 3819 of SEQ ID NO: 81, and / or (ii) amino acids 17 to 19 of SEQ ID NO: 80. 273 amino acid sequence, an amino acid sequence having at least 99%, 98%, 97%, 96%, 95%, 90%, 85%, or 80% identity to the amino acid sequence of amino acids 17 to 1273 of SEQ ID NO: 80, or an immunogenic fragment of the amino acid sequence of amino acids 17 to 1273 of SEQ ID NO: 80, or an amino acid sequence having at least 99%, 98%, 97%, 96%, 95%, 90%, 85%, or 80% identity to the amino acid sequence of amino acids 17 to 1273 of SEQ ID NO: 80, and (b) comprises one or more mutations associated with a SARS-CoV-2 variant of concern (e.g., one or more mutations listed in Table 3A). In one embodiment, the RNA encoding the vaccine antigen (a) comprises a nucleotide sequence encoding an amino acid sequence comprising (i) the nucleotide sequence of nucleotides 49 to 3819 of SEQ ID NO: 81 and / or (ii) the amino acid sequence of amino acids 17 to 1273 of SEQ ID NO: 80 or 7, and (b) one or more mutations characteristic of a SARS-CoV-2 variant of concern (e.g., one or more mutations listed in Table 3A).

[0231] In one embodiment, the vaccine antigen comprises, consists essentially of, or consists of the SARS-CoV-2 spike S1 fragment (S1) (the S1 subunit of the spike protein (S) of SARS-CoV-2), a variant thereof, or a fragment thereof, including one or more mutations of the SARS-CoV-2 variants described herein.

[0232] In one embodiment, the vaccine antigen comprises the amino acid sequence of amino acids 17-683 of SEQ ID NO: 1, an amino acid sequence having at least 99%, 98%, 97%, 96%, 95%, 90%, 85%, or 80% identity to the amino acid sequence of amino acids 17-683 of SEQ ID NO: 1, or an immunogenic fragment of the amino acid sequence of amino acids 17-683 of SEQ ID NO: 1, or an amino acid sequence having at least 99%, 98%, 97%, 96%, 95%, 90%, 85%, or 80% identity to an immunogenic fragment of the amino acid sequence of amino acids 17-683 of SEQ ID NO: 1, and comprising one or more mutations characteristic of SARS-CoV-2 variants (e.g., one or more mutations listed in Table 3A). In one embodiment, the vaccine antigen comprises the amino acid sequence of amino acids 17-683 of SEQ ID NO: 1, and comprising one or more mutations characteristic of SARS-CoV-2 variants (e.g., one or more mutations listed in Table 3A).

[0233] In one embodiment, the vaccine antigen comprises the amino acid sequence of amino acids 17-683 of SEQ ID NO: 80, an amino acid sequence having at least 99%, 98%, 97%, 96%, 95%, 90%, 85%, or 80% identity to the amino acid sequence of amino acids 17-683 of SEQ ID NO: 1, an immunogenic fragment of the amino acid sequence of amino acids 17-683 of SEQ ID NO: 1, or an amino acid sequence having at least 99%, 98%, 97%, 96%, 95%, 90%, 85%, or 80% identity to an immunogenic fragment of the amino acid sequence of amino acids 17-683 of SEQ ID NO: 80, and one or more mutations characteristic of SARS-CoV-2 variants (e.g., one or more mutations listed in Table 3A). In one embodiment, the vaccine antigen comprises the amino acid sequence of amino acids 17-683 of SEQ ID NO: 80, and includes one or more mutations characteristic of SARS-CoV-2 variants (e.g., one or more mutations listed in Table 3A).

[0234] Vaccine antigens and their combinations In one embodiment, the vaccine antigen described herein comprises, consists essentially of, or consists of the spike protein (S) of SARS-CoV-2, a variant thereof, or a fragment thereof.

[0235] In one embodiment, the vaccine antigen comprises the amino acid sequence of amino acids 17 to 1273 of SEQ ID NO: 1 or 7, an amino acid sequence having at least 99%, 98%, 97%, 96%, 95%, 90%, 85%, or 80% identity to the amino acid sequence of amino acids 17 to 1273 of SEQ ID NO: 1 or 7, or an immunogenic fragment of the amino acid sequence of amino acids 17 to 1273 of SEQ ID NO: 1 or 7, or an amino acid sequence having at least 99%, 98%, 97%, 96%, 95%, 90%, 85%, or 80% identity to the amino acid sequence of amino acids 17 to 1273 of SEQ ID NO: 1 or 7. In one embodiment, the vaccine antigen comprises the amino acid sequence of amino acids 17 to 1273 of SEQ ID NO: 1 or 7.

[0236] In one embodiment, the RNA encoding the vaccine antigen is (i) the nucleotide sequence of nucleotides 49 to 3819 of SEQ ID NO: 2, 8, or 9, a nucleotide sequence having at least 99%, 98%, 97%, 96%, 95%, 90%, 85%, or 80% identity to the nucleotide sequence of nucleotides 49 to 3819 of SEQ ID NO: 2, 8, or 9, or a fragment of the nucleotide sequence of nucleotides 49 to 3819 of SEQ ID NO: 2, 8, or 9, or a nucleotide sequence having at least 99%, 98%, 97%, 96%, 95%, 90%, 85%, or 80% identity to the nucleotide sequence of nucleotides 49 to 3819 of SEQ ID NO: 2, 8, or 9. and / or (ii) encodes an amino acid sequence comprising the amino acid sequence of amino acids 17 to 1273 of SEQ ID NO: 1 or 7, an amino acid sequence having at least 99%, 98%, 97%, 96%, 95%, 90%, 85%, or 80% identity to the amino acid sequence of amino acids 17 to 1273 of SEQ ID NO: 1 or 7, or an immunogenic fragment of the amino acid sequence of amino acids 17 to 1273 of SEQ ID NO: 1 or 7, or an amino acid sequence having at least 99%, 98%, 97%, 96%, 95%, 90%, 85%, or 80% identity to the amino acid sequence of amino acids 17 to 1273 of SEQ ID NO: 1 or 7. In one embodiment, the RNA encoding the vaccine antigen (i) comprises the nucleotide sequence of nucleotides 49 to 3819 of SEQ ID NO: 2, 8, or 9, and / or (ii) encodes an amino acid sequence comprising the amino acid sequence of amino acids 17 to 1273 of SEQ ID NO: 1 or 7.

[0237] In one embodiment, the vaccine antigen comprises, consists essentially of, or consists of the SARS-CoV-2 spike S1 fragment (S1) (the S1 subunit of the spike protein (S) of SARS-CoV-2), a variant thereof, or a fragment thereof.

[0238] In one embodiment, the vaccine antigen comprises the amino acid sequence of amino acids 17 to 683 of SEQ ID NO: 1, an amino acid sequence having at least 99%, 98%, 97%, 96%, 95%, 90%, 85%, or 80% identity to the amino acid sequence of amino acids 17 to 683 of SEQ ID NO: 1, or an immunogenic fragment of the amino acid sequence of amino acids 17 to 683 of SEQ ID NO: 1, or an amino acid sequence having at least 99%, 98%, 97%, 96%, 95%, 90%, 85%, or 80% identity to the amino acid sequence of amino acids 17 to 683 of SEQ ID NO: 1. In one embodiment, the vaccine antigen comprises the amino acid sequence of amino acids 17 to 683 of SEQ ID NO: 1.

[0239] In one embodiment, the RNA encoding the vaccine antigen is (i) the nucleotide sequence of nucleotides 49 to 2049 of SEQ ID NO: 2, 8, or 9, a nucleotide sequence having at least 99%, 98%, 97%, 96%, 95%, 90%, 85%, or 80% identity to the nucleotide sequence of nucleotides 49 to 2049 of SEQ ID NO: 2, 8, or 9, or a fragment of the nucleotide sequence of nucleotides 49 to 2049 of SEQ ID NO: 2, 8, or 9, or a fragment of the nucleotide sequence of nucleotides 49 to 2049 of SEQ ID NO: 2, 8, or 9, or a fragment of the nucleotide sequence of nucleotides 49 to 2049 of SEQ ID NO: 2, 8, or 9 95%, 90%, 85%, or 80% identity to the amino acid sequence of amino acids 17 to 683 of SEQ ID NO: 1, and / or (ii) encodes an amino acid sequence comprising the amino acid sequence of amino acids 17 to 683 of SEQ ID NO: 1, an amino acid sequence having at least 99%, 98%, 97%, 96%, 95%, 90%, 85%, or 80% identity to the amino acid sequence of amino acids 17 to 683 of SEQ ID NO: 1, or an immunogenic fragment of the amino acid sequence of amino acids 17 to 683 of SEQ ID NO: 1, or an amino acid sequence having at least 99%, 98%, 97%, 96%, 95%, 90%, 85%, or 80% identity to the amino acid sequence of amino acids 17 to 683 of SEQ ID NO: 1. In one embodiment, the RNA encoding the vaccine antigen (i) comprises the nucleotide sequence of nucleotides 49 to 2049 of SEQ ID NO: 2, 8, or 9, and / or (ii) encodes an amino acid sequence comprising the amino acid sequence of amino acids 17 to 683 of SEQ ID NO: 1.

[0240] In one embodiment, the vaccine antigen comprises the amino acid sequence of amino acids 17 to 685 of SEQ ID NO: 1, an amino acid sequence having at least 99%, 98%, 97%, 96%, 95%, 90%, 85%, or 80% identity to the amino acid sequence of amino acids 17 to 685 of SEQ ID NO: 1, or an immunogenic fragment of the amino acid sequence of amino acids 17 to 685 of SEQ ID NO: 1, or an amino acid sequence having at least 99%, 98%, 97%, 96%, 95%, 90%, 85%, or 80% identity to the amino acid sequence of amino acids 17 to 685 of SEQ ID NO: 1. In one embodiment, the vaccine antigen comprises the amino acid sequence of amino acids 17 to 685 of SEQ ID NO: 1.

[0241] In one embodiment, the RNA encoding the vaccine antigen is (i) the nucleotide sequence of nucleotides 49 to 2055 of SEQ ID NO: 2, 8, or 9, a nucleotide sequence having at least 99%, 98%, 97%, 96%, 95%, 90%, 85%, or 80% identity to the nucleotide sequence of nucleotides 49 to 2055 of SEQ ID NO: 2, 8, or 9, or a fragment of the nucleotide sequence of nucleotides 49 to 2055 of SEQ ID NO: 2, 8, or 9, or a fragment of the nucleotide sequence of nucleotides 49 to 2055 of SEQ ID NO: 2, 8, or 9, or a fragment of the nucleotide sequence of nucleotides 49 to 2055 of SEQ ID NO: 2, 8, or 9, and / or (ii) encodes an amino acid sequence comprising the amino acid sequence of amino acids 17 to 685 of SEQ ID NO: 1, an amino acid sequence having at least 99%, 98%, 97%, 96%, 95%, 90%, 85%, or 80% identity to the amino acid sequence of amino acids 17 to 685 of SEQ ID NO: 1, or an immunogenic fragment of the amino acid sequence of amino acids 17 to 685 of SEQ ID NO: 1, or an amino acid sequence having at least 99%, 98%, 97%, 96%, 95%, 90%, 85%, or 80% identity to the amino acid sequence of amino acids 17 to 685 of SEQ ID NO: 1. In one embodiment, the RNA encoding the vaccine antigen (i) comprises the nucleotide sequence of nucleotides 49 to 2055 of SEQ ID NO: 2, 8, or 9, and / or (ii) encodes an amino acid sequence comprising the amino acid sequence of amino acids 17 to 685 of SEQ ID NO: 1.

[0242] In one embodiment, the vaccine antigen comprises, consists essentially of, or consists of the receptor binding domain (RBD) of the S1 subunit of the spike protein (S) of SARS-CoV-2, a variant thereof, or a fragment thereof. The amino acid sequence of amino acids 327-528 of SEQ ID NO: 1, a variant thereof, or a fragment thereof is also referred to herein as the "RBD" or "RBD domain."

[0243] In one embodiment, the vaccine antigen comprises the amino acid sequence of amino acids 327 to 528 of SEQ ID NO: 1, an amino acid sequence having at least 99%, 98%, 97%, 96%, 95%, 90%, 85%, or 80% identity to the amino acid sequence of amino acids 327 to 528 of SEQ ID NO: 1, or an immunogenic fragment of the amino acid sequence of amino acids 327 to 528 of SEQ ID NO: 1, or an amino acid sequence having at least 99%, 98%, 97%, 96%, 95%, 90%, 85%, or 80% identity to the amino acid sequence of amino acids 327 to 528 of SEQ ID NO: 1. In one embodiment, the vaccine antigen comprises the amino acid sequence of amino acids 327 to 528 of SEQ ID NO: 1.

[0244] In one embodiment, the RNA encoding the vaccine antigen is (i) the nucleotide sequence of nucleotides 979-1584 of SEQ ID NO: 2, 8, or 9, a nucleotide sequence having at least 99%, 98%, 97%, 96%, 95%, 90%, 85%, or 80% identity to the nucleotide sequence of nucleotides 979-1584 of SEQ ID NO: 2, 8, or 9, or a fragment of the nucleotide sequence of nucleotides 979-1584 of SEQ ID NO: 2, 8, or 9, or a fragment of the nucleotide sequence of nucleotides 979-1584 of SEQ ID NO: 2, 8, or 9, or a fragment of the nucleotide sequence of nucleotides 979-1584 of SEQ ID NO: 2, 8, or 9, and / or (ii) encodes an amino acid sequence comprising the amino acid sequence of amino acids 327 to 528 of SEQ ID NO: 1, an amino acid sequence having at least 99%, 98%, 97%, 96%, 95%, 90%, 85%, or 80% identity to the amino acid sequence of amino acids 327 to 528 of SEQ ID NO: 1, or an immunogenic fragment of the amino acid sequence of amino acids 327 to 528 of SEQ ID NO: 1, or an amino acid sequence having at least 99%, 98%, 97%, 96%, 95%, 90%, 85%, or 80% identity to the amino acid sequence of amino acids 327 to 528 of SEQ ID NO: 1. In one embodiment, the RNA encoding the vaccine antigen (i) comprises the nucleotide sequence of nucleotides 979 to 1584 of SEQ ID NO: 2, 8, or 9, and / or (ii) encodes an amino acid sequence comprising the amino acid sequence of amino acids 327 to 528 of SEQ ID NO: 1.

[0245] According to certain embodiments, the signal peptide is fused directly or via a linker to the SARS-CoV-2 S protein, its variants, or fragments thereof, i.e., antigenic peptide or protein. Thus, in one embodiment, the signal peptide is fused to the above-mentioned amino acid sequence derived from the SARS-CoV-2 S protein or its immunogenic fragment (antigenic peptide or protein) constituted by the above-mentioned vaccine antigen.

[0246] Such signal peptides are typically about 15-30 amino acids in length and are preferably, but not limited to, sequences located at the N-terminus of an antigenic peptide or protein. The signal peptides defined herein preferably enable the transport of the antigenic peptide or protein encoded by the RNA to a specific cellular compartment, preferably the cell surface, the endoplasmic reticulum (ER), or the endosomal-lysosomal compartment. In one embodiment, the signal peptide sequence defined herein includes, but is not limited to, the signal peptide sequence of the SARS-CoV-2 S protein, particularly a sequence comprising the amino acid sequence of amino acids 1-16 or 1-19 of SEQ ID NO: 1, or a functional variant thereof.

[0247] In one embodiment, the signal sequence comprises the amino acid sequence of amino acids 1-16 of SEQ ID NO:1, an amino acid sequence having at least 99%, 98%, 97%, 96%, 95%, 90%, 85%, or 80% identity to the amino acid sequence of amino acids 1-16 of SEQ ID NO:1, or a functional fragment of the amino acid sequence of amino acids 1-16 of SEQ ID NO:1, or an amino acid sequence having at least 99%, 98%, 97%, 96%, 95%, 90%, 85%, or 80% identity to the amino acid sequence of amino acids 1-16 of SEQ ID NO:1. In one embodiment, the signal sequence comprises the amino acid sequence of amino acids 1-16 of SEQ ID NO:1.

[0248] In one embodiment, (i) the RNA encoding the signal sequence is a nucleotide sequence having at least 99%, 98%, 97%, 96%, 95%, 90%, 85%, or 80% identity to the nucleotide sequence of nucleotides 1 to 48 of SEQ ID NO: 2, 8, or 9, or to a fragment of the nucleotide sequence of nucleotides 1 to 48 of SEQ ID NO: 2, 8, or 9, or to the nucleotide sequence of nucleotides 1 to 48 of SEQ ID NO: 2, 8, or 9. and / or (ii) encodes an amino acid sequence comprising the amino acid sequence of amino acids 1 to 16 of SEQ ID NO: 1, an amino acid sequence having at least 99%, 98%, 97%, 96%, 95%, 90%, 85%, or 80% identity to the amino acid sequence of amino acids 1 to 16 of SEQ ID NO: 1, or a functional fragment of the amino acid sequence of amino acids 1 to 16 of SEQ ID NO: 1, or an amino acid sequence having at least 99%, 98%, 97%, 96%, 95%, 90%, 85%, or 80% identity to the amino acid sequence of amino acids 1 to 16 of SEQ ID NO: 1. In one embodiment, the RNA encoding the signal sequence (i) comprises the nucleotide sequence of nucleotides 1 to 48 of SEQ ID NO: 2, 8, or 9, and / or (ii) encodes an amino acid sequence comprising the amino acid sequence of amino acids 1 to 16 of SEQ ID NO: 1.

[0249] In one embodiment, the signal sequence comprises the amino acid sequence of amino acids 1-19 of SEQ ID NO:1, an amino acid sequence having at least 99%, 98%, 97%, 96%, 95%, 90%, 85%, or 80% identity to the amino acid sequence of amino acids 1-19 of SEQ ID NO:1, or a functional fragment of the amino acid sequence of amino acids 1-19 of SEQ ID NO:1, or an amino acid sequence having at least 99%, 98%, 97%, 96%, 95%, 90%, 85%, or 80% identity to the amino acid sequence of amino acids 1-19 of SEQ ID NO:1. In one embodiment, the signal sequence comprises the amino acid sequence of amino acids 1-19 of SEQ ID NO:1.

[0250] In one embodiment, (i) the RNA encoding the signal sequence is a nucleotide sequence having at least 99%, 98%, 97%, 96%, 95%, 90%, 85%, or 80% identity to the nucleotide sequence of nucleotides 1 to 57 of SEQ ID NO: 2, 8, or 9, or a fragment of the nucleotide sequence of nucleotides 1 to 57 of SEQ ID NO: 2, 8, or 9, or a nucleotide sequence having at least 99%, 98%, 97%, 96%, 95%, 90%, 85%, or 80% identity to the nucleotide sequence of nucleotides 1 to 57 of SEQ ID NO: 2, 8, or 9. and / or (ii) encodes an amino acid sequence comprising the amino acid sequence of amino acids 1-19 of SEQ ID NO: 1, an amino acid sequence having at least 99%, 98%, 97%, 96%, 95%, 90%, 85%, or 80% identity to the amino acid sequence of amino acids 1-19 of SEQ ID NO: 1, or a functional fragment of the amino acid sequence of amino acids 1-19 of SEQ ID NO: 1, or an amino acid sequence having at least 99%, 98%, 97%, 96%, 95%, 90%, 85%, or 80% identity to the amino acid sequence of amino acids 1-19 of SEQ ID NO: 1. In one embodiment, the RNA encoding the signal sequence (i) comprises the nucleotide sequence of nucleotides 1-57 of SEQ ID NO: 2, 8, or 9, and / or (ii) encodes an amino acid sequence comprising the amino acid sequence of amino acids 1-19 of SEQ ID NO: 1.

[0251] In some embodiments, the RNA comprises a sequence encoding a signal peptide. In some embodiments, a signal peptide sequence as defined herein includes, but is not limited to, an immunoglobulin signal peptide sequence, e.g., the signal peptide sequence of an immunoglobulin heavy chain variable region, and the immunoglobulin may be a human immunoglobulin. Specifically, in some embodiments, a signal peptide sequence as defined herein includes a sequence comprising the amino acid sequence of amino acids 1-22 of SEQ ID NO: 31, or a functional variant thereof.

[0252] In some embodiments, the signal peptide sequence is functional in mammalian cells. In some embodiments, the signal sequence utilized is "endogenous" in that it is naturally associated (e.g., linked) with the encoded polypeptide. In some embodiments, the signal sequence utilized is heterologous to the encoded polypeptide, e.g., it is not a native part of the polypeptide (e.g., protein) in which it is included. In some embodiments, the signal peptide is typically a sequence characterized by a length of about 15-30 amino acids. In many embodiments, the signal peptide is located at the N-terminus of the encoded polypeptide described herein, but is not limited thereto. In some embodiments, the signal peptide preferably enables transport of the polypeptide encoded by the RNA of the present disclosure with which it is associated to a defined cellular compartment, preferably the cell surface, the endoplasmic reticulum (ER), or an endosomal-lysosomal compartment. In some embodiments, the signal sequence is selected from an S1S2 signal peptide (aa1-16 or aa1-19), an immunoglobulin secretory signal peptide (aa1-22), an HSV-1 gD signal peptide (MGGAAARLGAVILFVVIVGLHGVRSKY), an HSV-2 gD signal peptide (MGRLTSGVGTAALLVVAVGLRVVCA), a human SPARC signal peptide, a human insulin isoform 1 signal peptide, a human albumin signal peptide, or the like. One of skill in the art will recognize other secretory signal peptides, for example, as disclosed in WO2017 / 0810822220 (e.g., SEQ ID NOS: 1-1115 and 1728, or fragment variants thereof) and WO2019 / 008001. In some embodiments, the RNA sequence encodes an epitope that includes or can otherwise be linked to a signal sequence (e.g., a secretory sequence), such as those listed in Table A, or a sequence having at least 1, 2, 3, 4, or 5 amino acid differences therefrom. In some embodiments, a signal sequence such as MFVFLVLLPLVSSQCVNLT, or a sequence that differs therefrom by at least 1, 2, 3, 4, or up to 5 amino acids, is utilized.In some embodiments, a sequence such as MFVFLVLLPLVSSQCVNLT, or a sequence having 1, 2, 3, 4, or up to 5 amino acid differences thereto, is utilized. In some embodiments, the signal sequence is selected from those included in Table A below and / or encoded by the sequences in Table B below. [Table 8] [Table 9]

[0253] In one embodiment, the signal sequence comprises the amino acid sequence of amino acids 1-22 of SEQ ID NO: 31, an amino acid sequence having at least 99%, 98%, 97%, 96%, 95%, 90%, 85%, or 80% identity to the amino acid sequence of amino acids 1-22 of SEQ ID NO: 31, or a functional fragment of the amino acid sequence of amino acids 1-22 of SEQ ID NO: 31, or an amino acid sequence having at least 99%, 98%, 97%, 96%, 95%, 90%, 85%, or 80% identity to the amino acid sequence of amino acids 1-22 of SEQ ID NO: 31. In one embodiment, the signal sequence comprises the amino acid sequence of amino acids 1-22 of SEQ ID NO: 31.

[0254] In one embodiment, the RNA encoding the signal sequence is (i) the nucleotide sequence of nucleotides 54-119 of SEQ ID NO: 32, a nucleotide sequence having at least 99%, 98%, 97%, 96%, 95%, 90%, 85%, or 80% identity to the nucleotide sequence of nucleotides 54-119 of SEQ ID NO: 32, or a fragment of the nucleotide sequence of nucleotides 54-119 of SEQ ID NO: 32, or a nucleotide sequence having at least 99%, 98%, 97%, 96%, 95%, 90%, 85%, or 80% identity to the nucleotide sequence of nucleotides 54-119 of SEQ ID NO: 32. and / or (ii) encodes an amino acid sequence comprising the amino acid sequence of amino acids 1-22 of SEQ ID NO: 31, an amino acid sequence having at least 99%, 98%, 97%, 96%, 95%, 90%, 85%, or 80% identity to the amino acid sequence of amino acids 1-22 of SEQ ID NO: 31, or a functional fragment of the amino acid sequence of amino acids 1-22 of SEQ ID NO: 31, or an amino acid sequence having at least 99%, 98%, 97%, 96%, 95%, 90%, 85%, or 80% identity to the amino acid sequence of amino acids 1-22 of SEQ ID NO: 31. In one embodiment, the RNA encoding the signal sequence (i) comprises the nucleotide sequence of nucleotides 54-119 of SEQ ID NO: 32, and / or (ii) encodes an amino acid sequence comprising the amino acid sequence of amino acids 1-22 of SEQ ID NO: 31.

[0255] Such a signal peptide is preferably used to facilitate secretion of the encoded antigenic peptide or protein. More preferably, a signal peptide as defined herein is fused to the encoded antigenic peptide or protein as defined herein.

[0256] Thus, in a particularly preferred embodiment, the RNA described herein comprises at least one coding region encoding an antigenic peptide or protein and a signal peptide, the signal peptide being preferably fused to the antigenic peptide or protein, more preferably to the N-terminus of the antigenic peptide or protein described herein.

[0257] In one embodiment, the vaccine antigen comprises the amino acid sequence of SEQ ID NO: 1 or 7, an amino acid sequence having at least 99%, 98%, 97%, 96%, 95%, 90%, 85%, or 80% identity to the amino acid sequence of SEQ ID NO: 1 or 7, or an immunogenic fragment of the amino acid sequence of SEQ ID NO: 1 or 7, or an amino acid sequence having at least 99%, 98%, 97%, 96%, 95%, 90%, 85%, or 80% identity to the amino acid sequence of SEQ ID NO: 1 or 7. In one embodiment, the vaccine antigen comprises the amino acid sequence of SEQ ID NO: 1 or 7.

[0258] In one embodiment, the RNA encoding the vaccine antigen is (i) a nucleotide sequence of SEQ ID NO: 2, 8, or 9, a nucleotide sequence having at least 99%, 98%, 97%, 96%, 95%, 90%, 85%, or 80% identity to the nucleotide sequence of SEQ ID NO: 2, 8, or 9, or a fragment of the nucleotide sequence of SEQ ID NO: 2, 8, or 9, or a nucleotide sequence having at least 99%, 98%, 97%, 96%, 95%, 90%, 85%, or 80% identity to the nucleotide sequence of SEQ ID NO: 2, 8, or 9. and / or (ii) encodes an amino acid sequence comprising the amino acid sequence of SEQ ID NO: 1 or 7, an amino acid sequence having at least 99%, 98%, 97%, 96%, 95%, 90%, 85%, or 80% identity to the amino acid sequence of SEQ ID NO: 1 or 7, or an immunogenic fragment of the amino acid sequence of SEQ ID NO: 1 or 7, or an amino acid sequence having at least 99%, 98%, 97%, 96%, 95%, 90%, 85%, or 80% identity to the amino acid sequence of SEQ ID NO: 1 or 7. In one embodiment, the RNA encoding the vaccine antigen (i) comprises the nucleotide sequence of SEQ ID NO: 2, 8, or 9, and / or (ii) encodes an amino acid sequence comprising the amino acid sequence of SEQ ID NO: 1 or 7.

[0259] In one embodiment, the vaccine antigen comprises the amino acid sequence of SEQ ID NO:7, an amino acid sequence having at least 99%, 98%, 97%, 96%, 95%, 90%, 85%, or 80% identity to the amino acid sequence of SEQ ID NO:7, or an immunogenic fragment of the amino acid sequence of SEQ ID NO:7, or an amino acid sequence having at least 99%, 98%, 97%, 96%, 95%, 90%, 85%, or 80% identity to the amino acid sequence of SEQ ID NO:7. In one embodiment, the vaccine antigen comprises the amino acid sequence of SEQ ID NO:7.

[0260] In one embodiment, the RNA encoding the vaccine antigen is (i) a nucleotide sequence of SEQ ID NO: 15, 16, 19, 20, 24, or 25, a nucleotide sequence having at least 99%, 98%, 97%, 96%, 95%, 90%, 85%, or 80% identity to the nucleotide sequence of SEQ ID NO: 15, 16, 19, 20, 24, or 25, or a fragment of ... and / or (ii) encodes an amino acid sequence comprising the amino acid sequence of SEQ ID NO: 7, an amino acid sequence having at least 99%, 98%, 97%, 96%, 95%, 90%, 85%, or 80% identity to the amino acid sequence of SEQ ID NO: 7, or an immunogenic fragment of the amino acid sequence of SEQ ID NO: 7, or an amino acid sequence having at least 99%, 98%, 97%, 96%, 95%, 90%, 85%, or 80% identity to the amino acid sequence of SEQ ID NO: 7. In one embodiment, the RNA encoding the vaccine antigen (i) comprises the nucleotide sequence of SEQ ID NO: 15, 16, 19, 20, 24, or 25, and / or (ii) encodes an amino acid sequence comprising the amino acid sequence of SEQ ID NO: 7.

[0261] In one embodiment, the vaccine antigen comprises the amino acid sequence of amino acids 1 to 683 of SEQ ID NO: 1, an amino acid sequence having at least 99%, 98%, 97%, 96%, 95%, 90%, 85%, or 80% identity to the amino acid sequence of amino acids 1 to 683 of SEQ ID NO: 1, or an immunogenic fragment of the amino acid sequence of amino acids 1 to 683 of SEQ ID NO: 1, or an amino acid sequence having at least 99%, 98%, 97%, 96%, 95%, 90%, 85%, or 80% identity to the amino acid sequence of amino acids 1 to 683 of SEQ ID NO: 1. In one embodiment, the vaccine antigen comprises the amino acid sequence of amino acids 1 to 683 of SEQ ID NO: 1.

[0262] In one embodiment, the RNA encoding the vaccine antigen is (i) a nucleotide sequence of nucleotides 1 to 2049 of SEQ ID NO: 2, 8, or 9, a nucleotide sequence having at least 99%, 98%, 97%, 96%, 95%, 90%, 85%, or 80% identity to the nucleotide sequence of nucleotides 1 to 2049 of SEQ ID NO: 2, 8, or 9, or a fragment of the nucleotide sequence of nucleotides 1 to 2049 of SEQ ID NO: 2, 8, or 9, or a fragment of the nucleotide sequence of nucleotides 1 to 2049 of SEQ ID NO: 2, 8, or 9, or a fragment of the nucleotide sequence of nucleotides 1 to 2049 of SEQ ID NO: 2, 8, or 9, 95%, 90%, 85%, or 80% identity to the amino acid sequence of amino acids 1 to 683 of SEQ ID NO: 1, and / or (ii) encodes an amino acid sequence comprising the amino acid sequence of amino acids 1 to 683 of SEQ ID NO: 1, an amino acid sequence having at least 99%, 98%, 97%, 96%, 95%, 90%, 85%, or 80% identity to the amino acid sequence of amino acids 1 to 683 of SEQ ID NO: 1, or an immunogenic fragment of the amino acid sequence of amino acids 1 to 683 of SEQ ID NO: 1, or an amino acid sequence having at least 99%, 98%, 97%, 96%, 95%, 90%, 85%, or 80% identity to the amino acid sequence of amino acids 1 to 683 of SEQ ID NO: 1. In one embodiment, the RNA encoding the vaccine antigen (i) comprises the nucleotide sequence of nucleotides 1 to 2049 of SEQ ID NO: 2, 8, or 9, and / or (ii) encodes an amino acid sequence comprising the amino acid sequence of amino acids 1 to 683 of SEQ ID NO: 1.

[0263] In one embodiment, the vaccine antigen comprises the amino acid sequence of amino acids 1 to 685 of SEQ ID NO: 1, an amino acid sequence having at least 99%, 98%, 97%, 96%, 95%, 90%, 85%, or 80% identity to the amino acid sequence of amino acids 1 to 685 of SEQ ID NO: 1, or an immunogenic fragment of the amino acid sequence of amino acids 1 to 685 of SEQ ID NO: 1, or an amino acid sequence having at least 99%, 98%, 97%, 96%, 95%, 90%, 85%, or 80% identity to the amino acid sequence of amino acids 1 to 685 of SEQ ID NO: 1. In one embodiment, the vaccine antigen comprises the amino acid sequence of amino acids 1 to 685 of SEQ ID NO: 1.

[0264] In one embodiment, the RNA encoding the vaccine antigen is (i) a nucleotide sequence of nucleotides 1 to 2055 of SEQ ID NO: 2, 8, or 9, a nucleotide sequence having at least 99%, 98%, 97%, 96%, 95%, 90%, 85%, or 80% identity to the nucleotide sequence of nucleotides 1 to 2055 of SEQ ID NO: 2, 8, or 9, or a fragment of the nucleotide sequence of nucleotides 1 to 2055 of SEQ ID NO: 2, 8, or 9, or a nucleotide sequence having at least 99%, 98%, 97%, 96%, 95%, 90%, 85%, or 80% identity to the nucleotide sequence of nucleotides 1 to 2055 of SEQ ID NO: 2, 8, or 9 95%, 96%, 95%, 90%, 85%, or 80% identity to the amino acid sequence of amino acids 1 to 685 of SEQ ID NO: 1, and / or (ii) encodes an amino acid sequence comprising the amino acid sequence of amino acids 1 to 685 of SEQ ID NO: 1, an amino acid sequence having at least 99%, 98%, 97%, 96%, 95%, 90%, 85%, or 80% identity to the amino acid sequence of amino acids 1 to 685 of SEQ ID NO: 1, or an immunogenic fragment of the amino acid sequence of amino acids 1 to 685 of SEQ ID NO: 1, or an amino acid sequence having at least 99%, 98%, 97%, 96%, 95%, 90%, 85%, or 80% identity to the amino acid sequence of amino acids 1 to 685 of SEQ ID NO: 1. In one embodiment, the RNA encoding the vaccine antigen (i) comprises the nucleotide sequence of nucleotides 1 to 2055 of SEQ ID NO: 2, 8, or 9, and / or (ii) encodes an amino acid sequence comprising the amino acid sequence of amino acids 1 to 685 of SEQ ID NO: 1.

[0265] In one embodiment, the vaccine antigen comprises the amino acid sequence of SEQ ID NO: 3, an amino acid sequence having at least 99%, 98%, 97%, 96%, 95%, 90%, 85%, or 80% identity to the amino acid sequence of SEQ ID NO: 3, or an immunogenic fragment of the amino acid sequence of SEQ ID NO: 3, or an amino acid sequence having at least 99%, 98%, 97%, 96%, 95%, 90%, 85%, or 80% identity to the amino acid sequence of SEQ ID NO: 3. In one embodiment, the vaccine antigen comprises the amino acid sequence of SEQ ID NO: 3.

[0266] In one embodiment, the RNA encoding the vaccine antigen (i) comprises the nucleotide sequence of SEQ ID NO: 4, a nucleotide sequence having at least 99%, 98%, 97%, 96%, 95%, 90%, 85%, or 80% identity to the nucleotide sequence of SEQ ID NO: 4, or a fragment of the nucleotide sequence of SEQ ID NO: 4, or a nucleotide sequence having at least 99%, 98%, 97%, 96%, 95%, 90%, 85%, or 80% identity to the nucleotide sequence of SEQ ID NO: 4, and / or (ii) encodes an amino acid sequence comprising the amino acid sequence of SEQ ID NO: 3, an amino acid sequence having at least 99%, 98%, 97%, 96%, 95%, 90%, 85%, or 80% identity to the amino acid sequence of SEQ ID NO: 3, or an immunogenic fragment of the amino acid sequence of SEQ ID NO: 3, or an amino acid sequence having at least 99%, 98%, 97%, 96%, 95%, 90%, 85%, or 80% identity to the amino acid sequence of SEQ ID NO: 3. In one embodiment, the RNA encoding the vaccine antigen (i) comprises the nucleotide sequence of SEQ ID NO:4, and / or (ii) encodes an amino acid sequence comprising the amino acid sequence of SEQ ID NO:3.

[0267] In one embodiment, the vaccine antigen comprises the amino acid sequence of amino acids 1 to 221 of SEQ ID NO: 29, an amino acid sequence having at least 99%, 98%, 97%, 96%, 95%, 90%, 85%, or 80% identity to the amino acid sequence of amino acids 1 to 221 of SEQ ID NO: 29, or an immunogenic fragment of the amino acid sequence of amino acids 1 to 221 of SEQ ID NO: 29, or an amino acid sequence having at least 99%, 98%, 97%, 96%, 95%, 90%, 85%, or 80% identity to the amino acid sequence of amino acids 1 to 221 of SEQ ID NO: 29. In one embodiment, the vaccine antigen comprises the amino acid sequence of amino acids 1 to 221 of SEQ ID NO: 29.

[0268] In one embodiment, the RNA encoding the vaccine antigen is (i) a fragment of the nucleotide sequence of nucleotides 54-716 of SEQ ID NO: 30, a fragment of the nucleotide sequence of nucleotides 54-716 of SEQ ID NO: 30, or a fragment of the nucleotide sequence of nucleotides 54-716 of SEQ ID NO: 30, or a fragment of the nucleotide sequence of nucleotides 54-716 of SEQ ID NO: 30, or a fragment of the nucleotide sequence of nucleotides 54-716 of SEQ ID NO: 30, or a fragment of the nucleotide sequence of nucleotides 54-716 of SEQ ID NO: 30, and / or (ii) encodes an amino acid sequence comprising the amino acid sequence of amino acids 1 to 221 of SEQ ID NO: 29, an amino acid sequence having at least 99%, 98%, 97%, 96%, 95%, 90%, 85%, or 80% identity to the amino acid sequence of amino acids 1 to 221 of SEQ ID NO: 29, or an immunogenic fragment of the amino acid sequence of amino acids 1 to 221 of SEQ ID NO: 29, or an amino acid sequence having at least 99%, 98%, 97%, 96%, 95%, 90%, 85%, or 80% identity to the amino acid sequence of amino acids 1 to 221 of SEQ ID NO: 29. In one embodiment, the RNA encoding the vaccine antigen (i) comprises the nucleotide sequence of nucleotides 54 to 716 of SEQ ID NO: 30, and / or (ii) encodes an amino acid sequence comprising the amino acid sequence of amino acids 1 to 221 of SEQ ID NO: 29.

[0269] In one embodiment, the vaccine antigen comprises the amino acid sequence of amino acids 1 to 224 of SEQ ID NO: 31, an amino acid sequence having at least 99%, 98%, 97%, 96%, 95%, 90%, 85%, or 80% identity to the amino acid sequence of amino acids 1 to 224 of SEQ ID NO: 31, or an immunogenic fragment of the amino acid sequence of amino acids 1 to 224 of SEQ ID NO: 31, or an amino acid sequence having at least 99%, 98%, 97%, 96%, 95%, 90%, 85%, or 80% identity to the amino acid sequence of amino acids 1 to 224 of SEQ ID NO: 31. In one embodiment, the vaccine antigen comprises the amino acid sequence of amino acids 1 to 224 of SEQ ID NO: 31.

[0270] In one embodiment, the RNA encoding the vaccine antigen is (i) a fragment of the nucleotide sequence of nucleotides 54-725 of SEQ ID NO: 32, a fragment of the nucleotide sequence of nucleotides 54-725 of SEQ ID NO: 32, or a fragment of the nucleotide sequence of nucleotides 54-725 of SEQ ID NO: 32, or a fragment of the nucleotide sequence of nucleotides 54-725 of SEQ ID NO: 32, or a fragment of the nucleotide sequence of nucleotides 54-725 of SEQ ID NO: 32, and / or (ii) encodes an amino acid sequence comprising the amino acid sequence of amino acids 1 to 224 of SEQ ID NO: 31, an amino acid sequence having at least 99%, 98%, 97%, 96%, 95%, 90%, 85%, or 80% identity to the amino acid sequence of amino acids 1 to 224 of SEQ ID NO: 31, or an immunogenic fragment of the amino acid sequence of amino acids 1 to 224 of SEQ ID NO: 31, or an amino acid sequence having at least 99%, 98%, 97%, 96%, 95%, 90%, 85%, or 80% identity to the amino acid sequence of amino acids 1 to 224 of SEQ ID NO: 31. In one embodiment, the RNA encoding the vaccine antigen (i) comprises the nucleotide sequence of nucleotides 54 to 725 of SEQ ID NO: 32, and / or (ii) encodes an amino acid sequence comprising the amino acid sequence of amino acids 1 to 224 of SEQ ID NO: 31.

[0271] Multimerization domain In some embodiments, the RNA utilized as described herein comprises a sequence encoding a multimerization element (e.g., a heterologous multimerization element). In some embodiments, the heterologous multimerization element comprises a dimerization, trimerization, or tetramerization element. In some embodiments, the multimerization element is one described in WO2017 / 081082 (e.g., SEQ ID NOS: 1116-1167, or a fragment or variant thereof). Exemplary trimerization and tetramerization elements include, but are not limited to, engineered leucine zippers, the fibritin Foldon domain from Enterobacteriaceae phage T4, GCN4pll, GCN4-pll, and p53. In some embodiments, the provided encoded polypeptide(s) are capable of forming a trimeric complex. For example, the utilized encoded polypeptide(s) may comprise a domain that allows for the formation of a multimeric complex, such as, for example, a trimeric complex of an amino acid sequence comprising the encoded polypeptide(s) described herein. In some embodiments, the domain that allows for the formation of a multimeric complex comprises a trimerization domain, e.g., a trimerization domain described herein. In some embodiments, the encoded polypeptide(s) can be modified, e.g., by adding the "Foldon" trimerization domain from T4-fibritin to increase its immunogenicity.

[0272] According to certain embodiments, the trimerization domain is fused to the SARS-CoV-2 S protein, its variants, or fragments thereof, i.e., antigenic peptide or protein, either directly or via a linker, e.g., a glycine / serine linker. Thus, in one embodiment, the trimerization domain is fused to the above-mentioned amino acid sequence derived from the SARS-CoV-2 S protein or an immunogenic fragment thereof (antigenic peptide or protein) constituted by the above-mentioned vaccine antigen (which can optionally be fused to a signal peptide as described above).

[0273] Such a trimerization domain is preferably located at, but is not limited to, the C-terminus of an antigenic peptide or protein. The trimerization domain defined herein preferably enables trimerization of antigenic peptides or proteins encoded by RNA. Examples of trimerization domains defined herein include, but are not limited to, Foldon, the natural trimerization domain of T4 fibritin. The C-terminal domain of T4 fibritin (Foldon) is essential for the formation of the fibritin trimer structure and can be used as an artificial trimerization domain. In one embodiment, the trimerization domain defined herein includes, but is not limited to, a sequence comprising the amino acid sequence of amino acids 3 to 29 of SEQ ID NO: 10, or a functional variant thereof. In one embodiment, the trimerization domain defined herein includes, but is not limited to, a sequence comprising the amino acid sequence of SEQ ID NO: 10, or a functional variant thereof.

[0274] In one embodiment, the trimerization domain comprises the amino acid sequence of amino acids 3 to 29 of SEQ ID NO: 10, an amino acid sequence having at least 99%, 98%, 97%, 96%, 95%, 90%, 85%, or 80% identity to the amino acid sequence of amino acids 3 to 29 of SEQ ID NO: 10, or a functional fragment of the amino acid sequence of amino acids 3 to 29 of SEQ ID NO: 10, or an amino acid sequence having at least 99%, 98%, 97%, 96%, 95%, 90%, 85%, or 80% identity to the amino acid sequence of amino acids 3 to 29 of SEQ ID NO: 10. In one embodiment, the trimerization domain comprises the amino acid sequence of amino acids 3 to 29 of SEQ ID NO: 10.

[0275] In one embodiment, the RNA encoding the trimerization domain is (i) the nucleotide sequence of nucleotides 7-87 of SEQ ID NO:11, a nucleotide sequence having at least 99%, 98%, 97%, 96%, 95%, 90%, 85%, or 80% identity to the nucleotide sequence of nucleotides 7-87 of SEQ ID NO:11, or a fragment of the nucleotide sequence of nucleotides 7-87 of SEQ ID NO:11, or a nucleotide sequence having at least 99%, 98%, 97%, 96%, 95%, 90%, 85%, or 80% identity to the nucleotide sequence of nucleotides 7-87 of SEQ ID NO:11. and / or (ii) encodes an amino acid sequence comprising the amino acid sequence of amino acids 3 to 29 of SEQ ID NO: 10, an amino acid sequence having at least 99%, 98%, 97%, 96%, 95%, 90%, 85%, or 80% identity to the amino acid sequence of amino acids 3 to 29 of SEQ ID NO: 10, or a functional fragment of the amino acid sequence of amino acids 3 to 29 of SEQ ID NO: 10, or an amino acid sequence having at least 99%, 98%, 97%, 96%, 95%, 90%, 85%, or 80% identity to the amino acid sequence of amino acids 3 to 29 of SEQ ID NO: 10. In one embodiment, the RNA encoding the trimerization domain (i) comprises the nucleotide sequence of nucleotides 7 to 87 of SEQ ID NO: 11, and / or (ii) encodes an amino acid sequence comprising the amino acid sequence of amino acids 3 to 29 of SEQ ID NO: 10.

[0276] In one embodiment, the trimerization domain comprises the amino acid sequence of SEQ ID NO: 10, an amino acid sequence having at least 99%, 98%, 97%, 96%, 95%, 90%, 85%, or 80% identity to the amino acid sequence of SEQ ID NO: 10, or a functional fragment of the amino acid sequence of SEQ ID NO: 10, or an amino acid sequence having at least 99%, 98%, 97%, 96%, 95%, 90%, 85%, or 80% identity to the amino acid sequence of SEQ ID NO: 10. In one embodiment, the trimerization domain comprises the amino acid sequence of SEQ ID NO: 10.

[0277] In one embodiment, the RNA encoding the trimerization domain is (i) a nucleotide sequence of SEQ ID NO: 11, a nucleotide sequence having at least 99%, 98%, 97%, 96%, 95%, 90%, 85%, or 80% identity to the nucleotide sequence of SEQ ID NO: 11, or a fragment of the nucleotide sequence of SEQ ID NO: 11, or a nucleotide sequence having at least 99%, 98%, 97%, 96%, 95%, 90%, 85%, or 80% identity to the nucleotide sequence of SEQ ID NO: 11. and / or (ii) encodes an amino acid sequence comprising the amino acid sequence of SEQ ID NO: 10, an amino acid sequence having at least 99%, 98%, 97%, 96%, 95%, 90%, 85%, or 80% identity to the amino acid sequence of SEQ ID NO: 10, or a functional fragment of the amino acid sequence of SEQ ID NO: 10, or an amino acid sequence having at least 99%, 98%, 97%, 96%, 95%, 90%, 85%, or 80% identity to the amino acid sequence of SEQ ID NO: 10. In one embodiment, the RNA encoding the trimerization domain (i) comprises the nucleotide sequence of SEQ ID NO: 11 and / or (ii) encodes an amino acid sequence comprising the amino acid sequence of SEQ ID NO: 10.

[0278] Such trimerization domains are preferably used to promote trimerization of the encoded antigenic peptide or protein. More preferably, a trimerization domain as defined herein is fused to an antigenic peptide or protein as defined herein.

[0279] Thus, in a particularly preferred embodiment, the RNA described herein comprises at least one coding region encoding an antigenic peptide or protein and a trimerization domain as defined herein, which trimerization domain is preferably fused to the antigenic peptide or protein, more preferably to the C-terminus of the antigenic peptide or protein.

[0280] In one embodiment, the vaccine antigen comprises the amino acid sequence of SEQ ID NO:5, an amino acid sequence having at least 99%, 98%, 97%, 96%, 95%, 90%, 85%, or 80% identity to the amino acid sequence of SEQ ID NO:5, or an immunogenic fragment of the amino acid sequence of SEQ ID NO:5, or an amino acid sequence having at least 99%, 98%, 97%, 96%, 95%, 90%, 85%, or 80% identity to the amino acid sequence of SEQ ID NO:5. In one embodiment, the vaccine antigen comprises the amino acid sequence of SEQ ID NO:5.

[0281] In one embodiment, the RNA encoding the vaccine antigen (i) comprises the nucleotide sequence of SEQ ID NO: 6, a nucleotide sequence having at least 99%, 98%, 97%, 96%, 95%, 90%, 85%, or 80% identity to the nucleotide sequence of SEQ ID NO: 6, or a fragment of the nucleotide sequence of SEQ ID NO: 6, or a nucleotide sequence having at least 99%, 98%, 97%, 96%, 95%, 90%, 85%, or 80% identity to the nucleotide sequence of SEQ ID NO: 6, and / or (ii) encodes an amino acid sequence comprising the amino acid sequence of SEQ ID NO: 5, an amino acid sequence having at least 99%, 98%, 97%, 96%, 95%, 90%, 85%, or 80% identity to the amino acid sequence of SEQ ID NO: 5, or an immunogenic fragment of the amino acid sequence of SEQ ID NO: 5, or an amino acid sequence having at least 99%, 98%, 97%, 96%, 95%, 90%, 85%, or 80% identity to the amino acid sequence of SEQ ID NO: 5. In one embodiment, the RNA encoding the vaccine antigen (i) comprises the nucleotide sequence of SEQ ID NO:6 and / or (ii) encodes an amino acid sequence comprising the amino acid sequence of SEQ ID NO:5.

[0282] In one embodiment, the RNA encoding the vaccine antigen is (i) a nucleotide sequence of SEQ ID NO: 17, 21, or 26, a nucleotide sequence having at least 99%, 98%, 97%, 96%, 95%, 90%, 85%, or 80% identity to the nucleotide sequence of SEQ ID NO: 17, 21, or 26, or a fragment of the nucleotide sequence of SEQ ID NO: 17, 21, or 26, or a nucleotide sequence having at least 99%, 98%, 97%, 96%, 95%, 90%, 85%, or 80% identity to the nucleotide sequence of SEQ ID NO: 17, 21, or 26 95%, 90%, 85%, or 80% identity to the amino acid sequence of SEQ ID NO: 5, and / or (ii) encodes an amino acid sequence comprising the amino acid sequence of SEQ ID NO: 5, an amino acid sequence having at least 99%, 98%, 97%, 96%, 95%, 90%, 85%, or 80% identity to the amino acid sequence of SEQ ID NO: 5, or an immunogenic fragment of the amino acid sequence of SEQ ID NO: 5, or an amino acid sequence having at least 99%, 98%, 97%, 96%, 95%, 90%, 85%, or 80% identity to the amino acid sequence of SEQ ID NO: 5. In one embodiment, the RNA encoding the vaccine antigen (i) comprises the nucleotide sequence of SEQ ID NO: 17, 21, or 26, and / or (ii) encodes an amino acid sequence comprising the amino acid sequence of SEQ ID NO: 5.

[0283] In one embodiment, the vaccine antigen comprises the amino acid sequence of SEQ ID NO: 18, an amino acid sequence having at least 99%, 98%, 97%, 96%, 95%, 90%, 85%, or 80% identity to the amino acid sequence of SEQ ID NO: 18, or an immunogenic fragment of the amino acid sequence of SEQ ID NO: 18, or an amino acid sequence having at least 99%, 98%, 97%, 96%, 95%, 90%, 85%, or 80% identity to the amino acid sequence of SEQ ID NO: 18. In one embodiment, the vaccine antigen comprises the amino acid sequence of SEQ ID NO: 18.

[0284] In one embodiment, the vaccine antigen comprises the amino acid sequence of amino acids 1 to 257 of SEQ ID NO: 29, an amino acid sequence having at least 99%, 98%, 97%, 96%, 95%, 90%, 85%, or 80% identity to the amino acid sequence of amino acids 1 to 257 of SEQ ID NO: 29, or an immunogenic fragment of the amino acid sequence of amino acids 1 to 257 of SEQ ID NO: 29, or an amino acid sequence having at least 99%, 98%, 97%, 96%, 95%, 90%, 85%, or 80% identity to the amino acid sequence of amino acids 1 to 257 of SEQ ID NO: 29. In one embodiment, the vaccine antigen comprises the amino acid sequence of amino acids 1 to 257 of SEQ ID NO: 29.

[0285] In one embodiment, the RNA encoding the vaccine antigen is (i) a fragment of the nucleotide sequence of nucleotides 54 to 824 of SEQ ID NO: 30, a fragment of the nucleotide sequence of nucleotides 54 to 824 of SEQ ID NO: 30, or a fragment of the nucleotide sequence of nucleotides 54 to 824 of SEQ ID NO: 30, or a fragment of the nucleotide sequence of nucleotides 54 to 824 of SEQ ID NO: 30, or a fragment of the nucleotide sequence of nucleotides 54 to 824 of SEQ ID NO: 30, or a fragment of the nucleotide sequence of nucleotides 54 to 824 of SEQ ID NO: 30, and / or (ii) encodes an amino acid sequence comprising the amino acid sequence of amino acids 1 to 257 of SEQ ID NO: 29, an amino acid sequence having at least 99%, 98%, 97%, 96%, 95%, 90%, 85%, or 80% identity to the amino acid sequence of amino acids 1 to 257 of SEQ ID NO: 29, or an immunogenic fragment of the amino acid sequence of amino acids 1 to 257 of SEQ ID NO: 29, or an amino acid sequence having at least 99%, 98%, 97%, 96%, 95%, 90%, 85%, or 80% identity to the amino acid sequence of amino acids 1 to 257 of SEQ ID NO: 29. In one embodiment, the RNA encoding the vaccine antigen (i) comprises the nucleotide sequence of nucleotides 54 to 824 of SEQ ID NO: 30, and / or (ii) encodes an amino acid sequence comprising the amino acid sequence of amino acids 1 to 257 of SEQ ID NO: 29.

[0286] In one embodiment, the vaccine antigen comprises the amino acid sequence of amino acids 1 to 260 of SEQ ID NO: 31, an amino acid sequence having at least 99%, 98%, 97%, 96%, 95%, 90%, 85%, or 80% identity to the amino acid sequence of amino acids 1 to 260 of SEQ ID NO: 31, or an immunogenic fragment of the amino acid sequence of amino acids 1 to 260 of SEQ ID NO: 31, or an amino acid sequence having at least 99%, 98%, 97%, 96%, 95%, 90%, 85%, or 80% identity to the amino acid sequence of amino acids 1 to 260 of SEQ ID NO: 31. In one embodiment, the vaccine antigen comprises the amino acid sequence of amino acids 1 to 260 of SEQ ID NO: 31.

[0287] In one embodiment, the RNA encoding the vaccine antigen is (i) a fragment of the nucleotide sequence of nucleotides 54 to 833 of SEQ ID NO: 32, a fragment of the nucleotide sequence of nucleotides 54 to 833 of SEQ ID NO: 32, or a fragment of the nucleotide sequence of nucleotides 54 to 833 of SEQ ID NO: 32, or a fragment of the nucleotide sequence of nucleotides 54 to 833 of SEQ ID NO: 32, or a fragment of the nucleotide sequence of nucleotides 54 to 833 of SEQ ID NO: 32, or a fragment of the nucleotide sequence of nucleotides 54 to 833 of SEQ ID NO: 32, and / or (ii) encodes an amino acid sequence comprising the amino acid sequence of amino acids 1 to 260 of SEQ ID NO: 31, an amino acid sequence having at least 99%, 98%, 97%, 96%, 95%, 90%, 85%, or 80% identity to the amino acid sequence of amino acids 1 to 260 of SEQ ID NO: 31, or an immunogenic fragment of the amino acid sequence of amino acids 1 to 260 of SEQ ID NO: 31, or an amino acid sequence having at least 99%, 98%, 97%, 96%, 95%, 90%, 85%, or 80% identity to the amino acid sequence of amino acids 1 to 260 of SEQ ID NO: 31. In one embodiment, the RNA encoding the vaccine antigen (i) comprises the nucleotide sequence of nucleotides 54 to 833 of SEQ ID NO: 32, and / or (ii) encodes an amino acid sequence comprising the amino acid sequence of amino acids 1 to 260 of SEQ ID NO: 31.

[0288] In one embodiment, the vaccine antigen comprises the amino acid sequence of amino acids 20 to 257 of SEQ ID NO: 29, an amino acid sequence having at least 99%, 98%, 97%, 96%, 95%, 90%, 85%, or 80% identity to the amino acid sequence of amino acids 20 to 257 of SEQ ID NO: 29, or an immunogenic fragment of the amino acid sequence of amino acids 20 to 257 of SEQ ID NO: 29, or an amino acid sequence having at least 99%, 98%, 97%, 96%, 95%, 90%, 85%, or 80% identity to the amino acid sequence of amino acids 20 to 257 of SEQ ID NO: 29. In one embodiment, the vaccine antigen comprises the amino acid sequence of amino acids 20 to 257 of SEQ ID NO: 29.

[0289] In one embodiment, the RNA encoding the vaccine antigen is (i) a fragment of the nucleotide sequence of nucleotides 111 to 824 of SEQ ID NO: 30, a fragment of the nucleotide sequence of nucleotides 111 to 824 of SEQ ID NO: 30, or a fragment of the nucleotide sequence of nucleotides 111 to 824 of SEQ ID NO: 30, or a fragment of the nucleotide sequence of nucleotides 111 to 824 of SEQ ID NO: 30, or a fragment of the nucleotide sequence of nucleotides 111 to 824 of SEQ ID NO: 30, or a fragment of the nucleotide sequence of nucleotides 111 to 824 of SEQ ID NO: 30, and / or (ii) encodes an amino acid sequence comprising the amino acid sequence of amino acids 20 to 257 of SEQ ID NO: 29, an amino acid sequence having at least 99%, 98%, 97%, 96%, 95%, 90%, 85%, or 80% identity to the amino acid sequence of amino acids 20 to 257 of SEQ ID NO: 29, or an immunogenic fragment of the amino acid sequence of amino acids 20 to 257 of SEQ ID NO: 29, or an amino acid sequence having at least 99%, 98%, 97%, 96%, 95%, 90%, 85%, or 80% identity to the amino acid sequence of amino acids 20 to 257 of SEQ ID NO: 29. In one embodiment, the RNA encoding the vaccine antigen (i) comprises the nucleotide sequence of nucleotides 111 to 824 of SEQ ID NO: 30, and / or (ii) encodes an amino acid sequence comprising the amino acid sequence of amino acids 20 to 257 of SEQ ID NO: 29.

[0290] In one embodiment, the vaccine antigen comprises the amino acid sequence of amino acids 23 to 260 of SEQ ID NO: 31, an amino acid sequence having at least 99%, 98%, 97%, 96%, 95%, 90%, 85%, or 80% identity to the amino acid sequence of amino acids 23 to 260 of SEQ ID NO: 31, or an immunogenic fragment of the amino acid sequence of amino acids 23 to 260 of SEQ ID NO: 31, or an amino acid sequence having at least 99%, 98%, 97%, 96%, 95%, 90%, 85%, or 80% identity to the amino acid sequence of amino acids 23 to 260 of SEQ ID NO: 31. In one embodiment, the vaccine antigen comprises the amino acid sequence of amino acids 23 to 260 of SEQ ID NO: 31.

[0291] In one embodiment, the RNA encoding the vaccine antigen is (i) a fragment of the nucleotide sequence of nucleotides 120 to 833 of SEQ ID NO: 32, a fragment of the nucleotide sequence of nucleotides 120 to 833 of SEQ ID NO: 32, or a fragment of the nucleotide sequence of nucleotides 120 to 833 of SEQ ID NO: 32, or a fragment of the nucleotide sequence of nucleotides 120 to 833 of SEQ ID NO: 32, or a fragment of the nucleotide sequence of nucleotides 120 to 833 of SEQ ID NO: 32, and / or (ii) encodes an amino acid sequence comprising the amino acid sequence of amino acids 23 to 260 of SEQ ID NO: 31, an amino acid sequence having at least 99%, 98%, 97%, 96%, 95%, 90%, 85%, or 80% identity to the amino acid sequence of amino acids 23 to 260 of SEQ ID NO: 31, or an immunogenic fragment of the amino acid sequence of amino acids 23 to 260 of SEQ ID NO: 31, or an amino acid sequence having at least 99%, 98%, 97%, 96%, 95%, 90%, 85%, or 80% identity to the amino acid sequence of amino acids 23 to 260 of SEQ ID NO: 31. In one embodiment, the RNA encoding the vaccine antigen (i) comprises the nucleotide sequence of nucleotides 120 to 833 of SEQ ID NO: 32, and / or (ii) encodes an amino acid sequence comprising the amino acid sequence of amino acids 23 to 260 of SEQ ID NO: 31.

[0292] Transmembrane domain In some embodiments, the RNA described herein comprises a sequence encoding a membrane-associated element (e.g., a heterologous membrane-associated element), such as a transmembrane domain. The transmembrane domain can be N-terminal, C-terminal, or internal to the encoded polypeptide. The coding sequence for the transmembrane element is typically positioned in-frame (i.e., in the same reading frame), 5', 3', or internal to the coding sequence of the sequence to which it is linked (e.g., a sequence encoding a polypeptide(s)). In some embodiments, the transmembrane domain comprises or is the transmembrane domain of influenza virus hemagglutinin (HA), HIV-1 Env, equine infectious anemia virus (EIAV), murine leukemia virus (MLV), mouse mammary tumor virus, vesicular stomatitis virus (VSV) G protein, rabies virus, or a seven transmembrane domain receptor.

[0293] According to certain embodiments, the transmembrane domain is fused to the SARS-CoV-2 S protein, its variants, or fragments thereof, i.e., antigenic peptide or protein, either directly or via a linker, e.g., a glycine / serine linker. Thus, in one embodiment, the transmembrane domain is fused to a SARS-CoV-2 S polypeptide or an immunogenic fragment thereof (antigenic peptide or protein), which can optionally be fused to a signal peptide and / or trimerization domain as described above.

[0294] secretion signal Such a transmembrane domain is preferably, but not limited to, located at the C-terminus of the antigenic peptide or protein. Preferably, such a transmembrane domain, if present, is located C-terminal to the trimerization domain. In one embodiment, the trimerization domain is located between the SARS-CoV-2 S protein, variant thereof, or fragment thereof, i.e., the antigenic peptide or protein, and the transmembrane domain.

[0295] A transmembrane domain as defined herein preferably allows anchoring of the antigenic peptide or protein encoded by the RNA into the cell membrane.

[0296] In one embodiment, the transmembrane domain sequence defined herein includes, but is not limited to, the transmembrane domain sequence of the SARS-CoV-2 S protein, particularly a sequence comprising the amino acid sequence of amino acids 1207 to 1254 of SEQ ID NO: 1, or a functional variant thereof.

[0297] In one embodiment, the transmembrane domain sequence comprises the amino acid sequence of amino acids 1207 to 1254 of SEQ ID NO: 1, an amino acid sequence having at least 99%, 98%, 97%, 96%, 95%, 90%, 85%, or 80% identity to the amino acid sequence of amino acids 1207 to 1254 of SEQ ID NO: 1, or a functional fragment of the amino acid sequence of amino acids 1207 to 1254 of SEQ ID NO: 1, or an amino acid sequence having at least 99%, 98%, 97%, 96%, 95%, 90%, 85%, or 80% identity to the amino acid sequence of amino acids 1207 to 1254 of SEQ ID NO: 1. In one embodiment, the transmembrane domain sequence comprises the amino acid sequence of amino acids 1207 to 1254 of SEQ ID NO: 1.

[0298] In one embodiment, the RNA encoding the transmembrane domain sequence is (i) the nucleotide sequence of nucleotides 36...

Claims

1. 1. A composition or medical preparation comprising RNA encoding a SARS-CoV-2 S polypeptide or an immunogenic fragment thereof, said SARS-CoV-2 S polypeptide or fragment comprising: (a) an amino acid sequence having at least 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identity to SEQ ID NO:1 and containing one of the following sets of amino acid substitutions relative to SEQ ID NO:1: (1) D985P, V987P, F817P, A892P, A899P, and A942P; (2) K986P, V987P, F817P, A892P, A899P, and A942P; (3) D985P, V987P, F817P, A892P, A899P, A942P, D614G, R682G, R683S, and R685S; (4) K986P, V987P, F817P, A892P, A899P, A942P, D614G, R682G, R683S, and R685S; (5) D985P, K986P, V987P, F817P, A892P, A899P, A942P, D614G, R682G, R683S, and R685S; (6) D985P, V987P, F817P, A892P, A899P, A942P, R682G, R683S, and R685S; (7) K986P, V987P, F817P, A892P, A899P, A942P, R682G, R683S, and R685S; or (8) D985P, K986P, V987P, F817P, A892P, A899P, A942P, R682G, R683S, and R685S; (b) an amino acid sequence having at least 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identity to SEQ ID NO:69 and containing one of the following sets of amino acid substitutions relative to SEQ ID NO:69: (1) D982P, V984P, F814P, A889P, A896P, and A939P; (2) K983P, V984P, F814P, A889P, A896P, and A939P; (3) D982P, V984P, F814P, A889P, A896P, A939P, R679G, R680S, and R682S; (4) K983P, V984P, F814P, A889P, A896P, A939P, R679G, R680S, and R682S; or (5) D982P, K983P, V984P, F814P, A889P, A896P, A939P, R679G, R680S, and R682S; (c) an amino acid sequence having at least 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identity to SEQ ID NO: 70 and containing one of the following sets of amino acid substitutions relative to SEQ ID NO: 70: (1) D982P, V984P, F814P, A889P, A896P, and A939P; (2) K983P, V984P, F814P, A889P, A896P, and A939P; (3) D982P, V984P, F814P, A889P, A896P, A939P, R679G, R680S, and R682S; (4) K983P, V984P, F814P, A889P, A896P, A939P, R679G, R680S, and R682S; or (5) D982P, K983P, V984P, F814P, A889P, A896P, A939P, R679G, R680S, and R682S; or (d) an amino acid sequence having at least 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identity to SEQ ID NO: 104 or 105 and comprising one of the following sets of amino acid substitutions relative to SEQ ID NO: 104 or 105: (1) D980P, V982P, F812P, A887P, A894P, and A937P; (2) K981P, V982P, F812P, A887P, A894P, and A937P; (3) D980P, V982P, F812P, A887P, A894P, A937P, R677G, R678S, and R680S; (4) K981P, V982P, F812P, A887P, A894P, A937P, R677G, R678S, and R680S; or (5) The composition or medical preparation comprising D980P, K981P, V982P, F812P, A887P, A894P, A937P, R677G, R678S, and R680S.

2. 2. The composition or medical preparation of claim 1, wherein the RNA comprises modified nucleosides in place of uridine.

3. 3. The composition or medical preparation of claim 1 or 2, wherein the RNA comprises modified uridines in place of all uridines.

4. The composition or medical preparation of any one of claims 1 to 3, wherein the RNA comprises N1-methyl-pseudouridines (m1ψ) in place of all uridines.

5. The composition or medical preparation of any one of claims 1 to 4, wherein the RNA comprises a 5' cap.

6. The 5' cap is ２ ７，３’－Ｏ Gppp(m １ ２’－Ｏ 6. The composition or medical preparation of claim 5, which is or comprises ApG.

7. The composition or medical preparation of any one of claims 1 to 6, wherein the RNA comprises a 5'-UTR that is or comprises a modified human alpha-globin 5'-UTR.

8. 8. The composition or medical preparation of any one of claims 1 to 7, wherein the RNA comprises a 5'UTR comprising a nucleotide sequence of SEQ ID NO:12, or a nucleotide sequence that is at least 99%, 98%, 97%, 96%, 95%, 90%, 85%, or 80% identical to the nucleotide sequence of SEQ ID NO:

12.

9. 9. The composition or medical preparation of any one of claims 1 to 8, wherein the RNA comprises a 3'-UTR that is or includes a first sequence from a split amino-terminal enhancer (AES) messenger RNA and a second sequence from a mitochondrially encoded 12S ribosomal RNA.

10. 10. The composition or medical preparation of any one of claims 1 to 9, wherein the RNA comprises a 3'UTR comprising a nucleotide sequence of SEQ ID NO:13, or a nucleotide sequence that is at least 99%, 98%, 97%, 96%, 95%, 90%, 85%, or 80% identical to the nucleotide sequence of SEQ ID NO:

13.

11. The composition or medical preparation according to any one of claims 1 to 10, wherein the RNA comprises a polyA sequence.

12. The composition or medical preparation of claim 11, wherein the polyA sequence comprises at least 100 nucleotides.

13. 13. The composition or medical preparation of claim 11 or 12, wherein the polyA sequence comprises 30 adenine nucleotides followed by 70 adenine nucleotides, the 30 adenine nucleotides and the 70 adenine nucleotides being separated by a linker sequence.

14. 14. The composition or medical preparation according to any one of claims 11 to 13, wherein said polyA sequence comprises or consists of a nucleotide sequence of SEQ ID NO:14 or a nucleotide sequence which is at least 99%, 98%, 97%, 96%, 95%, 90%, 85%, or 80% identical to said nucleotide sequence of SEQ ID NO:

14.

15. A composition or medical preparation according to any one of claims 1 to 14, wherein said RNA is formulated or is to be formulated for intramuscular administration.

16. A composition or medical preparation according to any one of claims 1 to 15, wherein the RNA is or is to be formulated as a particle.

17. 17. The composition or medical preparation of claim 16, wherein the particle is a lipid nanoparticle (LNP) or a lipoplex (LPX) particle.

18. 18. The composition or medical preparation of claim 17, wherein the LNP comprises a cationically ionizable lipid, a neutral lipid, a sterol, and a polymer-lipid conjugate.

19. 18. The composition or medical preparation according to claim 17, wherein said lipoplex particles are obtainable by mixing said RNA with liposomes.

20. The composition or medical preparation according to any one of claims 1 to 19, wherein said RNA is mRNA or saRNA.

21. A composition or medical preparation according to any one of claims 1 to 20 which is a pharmaceutical composition.

22. A composition or medical preparation according to any one of claims 1 to 21 which is a vaccine.

23. 23. The composition or medical preparation of claim 21 or 22, wherein the pharmaceutical composition further comprises one or more pharma- ceutically acceptable carriers, diluents, and / or excipients.

24. 1. A composition or medical preparation comprising a first RNA encoding a first SARS-CoV-2 S polypeptide or an immunogenic fragment thereof, and a second RNA encoding a second SARS-CoV-2 S polypeptide or an immunogenic fragment thereof, (a) the first SARS-CoV-2 S polypeptide or fragment comprises an amino acid sequence having at least 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identity to SEQ ID NO:1; (b) the second SARS-CoV-2 S polypeptide or fragment has at least 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identity to SEQ ID NO:1 and the following set of amino acid substitutions relative to SEQ ID NO:1: (1) D985P, V987P, F817P, A892P, A899P, and A942P; (2) K986P, V987P, F817P, A892P, A899P, and A942P; (3) D985P, V987P, F817P, A892P, A899P, A942P, D614G, R682G, R683S, and R685S; (4) K986P, V987P, F817P, A892P, A899P, A942P, D614G, R682G, R683S, and R685S; (5) D985P, K986P, V987P, F817P, A892P, A899P, A942P, D614G, R682G, R683S, and R685S; (6) D985P, V987P, F817P, A892P, A899P, A942P, R682G, R683S, and R685S; (7) K986P, V987P, F817P, A892P, A899P, A942P, R682G, R683S, and R685S; or (8) The composition or medical preparation, comprising an amino acid sequence including one of D985P, K986P, V987P, F817P, A892P, A899P, A942P, R682G, R683S, and R685S.

25. 1. A composition or medical preparation comprising a first RNA encoding a first SARS-CoV-2 S polypeptide or an immunogenic fragment thereof, and a second RNA encoding a second SARS-CoV-2 S polypeptide or an immunogenic fragment thereof, (a) the first SARS-CoV-2 S polypeptide or fragment comprises an amino acid sequence having at least 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identity to SEQ ID NO:1; (b) the second SARS-CoV-2 S polypeptide or fragment has at least 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identity to SEQ ID NO:69 and has the following set of amino acid substitutions relative to SEQ ID NO:69: (1) D982P, V984P, F814P, A889P, A896P, and A939P; (2) K983P, V984P, F814P, A889P, A896P, and A939P; (3) D982P, V984P, F814P, A889P, A896P, A939P, R679G, R680S, and R682S; (4) K983P, V984P, F814P, A889P, A896P, A939P, R679G, R680S, and R682S; or (5) The composition or medical preparation, comprising an amino acid sequence including one of D982P, K983P, V984P, F814P, A889P, A896P, A939P, R679G, R680S, and R682S.

26. 1. A composition or medical preparation comprising a first RNA encoding a first SARS-CoV-2 S polypeptide or an immunogenic fragment thereof, and a second RNA encoding a second SARS-CoV-2 S polypeptide or an immunogenic fragment thereof, (a) the first SARS-CoV-2 S polypeptide or fragment comprises an amino acid sequence having at least 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identity to SEQ ID NO:1; (b) the second SARS-CoV-2 S polypeptide or fragment has at least 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identity to SEQ ID NO:70 and has the following set of amino acid substitutions relative to SEQ ID NO:70: (1) D982P, V984P, F814P, A889P, A896P, and A939P; (2) K983P, V984P, F814P, A889P, A896P, and A939P; (3) D982P, V984P, F814P, A889P, A896P, A939P, R679G, R680S, and R682S; (4) K983P, V984P, F814P, A889P, A896P, A939P, R679G, R680S, and R682S; or (5) The composition or medical preparation, comprising an amino acid sequence including one of D982P, K983P, V984P, F814P, A889P, A896P, A939P, R679G, R680S, and R682S.

27. 1. A composition or medical preparation comprising a first RNA encoding a first SARS-CoV-2 S polypeptide or an immunogenic fragment thereof, and a second RNA encoding a second SARS-CoV-2 S polypeptide or an immunogenic fragment thereof, (a) the first SARS-CoV-2 S polypeptide or fragment comprises an amino acid sequence having at least 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identity to SEQ ID NO:1; (b) the second SARS-CoV-2 S polypeptide or fragment has at least 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identity to SEQ ID NO: 104 or 105 and has the following set of amino acid substitutions relative to SEQ ID NO: 104 or 105: (1) D980P, V982P, F812P, A887P, A894P, and A937P; (2) K981P, V982P, F812P, A887P, A894P, and A937P; (3) D980P, V982P, F812P, A887P, A894P, A937P, R677G, R678S, and R680S; (4) K981P, V982P, F812P, A887P, A894P, A937P, R677G, R678S, and R680S; or (5) The composition or medical preparation, comprising an amino acid sequence including one of D980P, K981P, V982P, F812P, A887P, A894P, A937P, R677G, R678S, and R680S.

28. 1. A composition or medical preparation comprising a first RNA encoding a first SARS-CoV-2 S polypeptide or an immunogenic fragment thereof, and a second RNA encoding a second SARS-CoV-2 S polypeptide or an immunogenic fragment thereof, (a) the first SARS-CoV-2 S polypeptide or fragment comprises an amino acid sequence having at least 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identity to SEQ ID NO:69; (b) the second SARS-CoV-2 S polypeptide or fragment has at least 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identity to SEQ ID NO:1 and the following set of amino acid substitutions relative to SEQ ID NO:1: (1) D985P, V987P, F817P, A892P, A899P, and A942P; (2) K986P, V987P, F817P, A892P, A899P, and A942P; (3) D985P, V987P, F817P, A892P, A899P, A942P, D614G, R682G, R683S, and R685S; (4) K986P, V987P, F817P, A892P, A899P, A942P, D614G, R682G, R683S, and R685S; (5) D985P, K986P, V987P, F817P, A892P, A899P, A942P, D614G, R682G, R683S, and R685S; (6) D985P, V987P, F817P, A892P, A899P, A942P, R682G, R683S, and R685S; (7) K986P, V987P, F817P, A892P, A899P, A942P, R682G, R683S, and R685S; or (8) The composition or medical preparation, comprising an amino acid sequence including one of D985P, K986P, V987P, F817P, A892P, A899P, A942P, R682G, R683S, and R685S.

29. 1. A composition or medical preparation comprising a first RNA encoding a first SARS-CoV-2 S polypeptide or an immunogenic fragment thereof, and a second RNA encoding a second SARS-CoV-2 S polypeptide or an immunogenic fragment thereof, (a) the first SARS-CoV-2 S polypeptide or fragment comprises an amino acid sequence having at least 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identity to SEQ ID NO:69; (b) the second SARS-CoV-2 S polypeptide or fragment has at least 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identity to SEQ ID NO:69 and has the following set of amino acid substitutions relative to SEQ ID NO:69: (1) D982P, V984P, F814P, A889P, A896P, and A939P; (2) K983P, V984P, F814P, A889P, A896P, and A939P; (3) D982P, V984P, F814P, A889P, A896P, A939P, R679G, R680S, and R682S; (4) K983P, V984P, F814P, A889P, A896P, A939P, R679G, R680S, and R682S; or (5) The composition or medical preparation, comprising an amino acid sequence including one of D982P, K983P, V984P, F814P, A889P, A896P, A939P, R679G, R680S, and R682S.

30. 1. A composition or medical preparation comprising a first RNA encoding a first SARS-CoV-2 S polypeptide or an immunogenic fragment thereof, and a second RNA encoding a second SARS-CoV-2 S polypeptide or an immunogenic fragment thereof, (a) the first SARS-CoV-2 S polypeptide or fragment comprises an amino acid sequence having at least 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identity to SEQ ID NO:69; (b) the second SARS-CoV-2 S polypeptide or fragment has at least 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identity to SEQ ID NO:70 and has the following set of amino acid substitutions relative to SEQ ID NO:70: (1) D982P, V984P, F814P, A889P, A896P, and A939P; (2) K983P, V984P, F814P, A889P, A896P, and A939P; (3) D982P, V984P, F814P, A889P, A896P, A939P, R679G, R680S, and R682S; (4) K983P, V984P, F814P, A889P, A896P, A939P, R679G, R680S, and R682S; or (5) The composition or medical preparation, comprising an amino acid sequence including one of D982P, K983P, V984P, F814P, A889P, A896P, A939P, R679G, R680S, and R682S.

31. 1. A composition or medical preparation comprising a first RNA encoding a first SARS-CoV-2 S polypeptide or an immunogenic fragment thereof, and a second RNA encoding a second SARS-CoV-2 S polypeptide or an immunogenic fragment thereof, (a) the first SARS-CoV-2 S polypeptide or fragment comprises an amino acid sequence having at least 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identity to SEQ ID NO:69; (b) the second SARS-CoV-2 S polypeptide or fragment has at least 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identity to SEQ ID NO: 104 or 105 and has the following set of amino acid substitutions relative to SEQ ID NO: 104 or 105: (1) D980P, V982P, F812P, A887P, A894P, and A937P; (2) K981P, V982P, F812P, A887P, A894P, and A937P; (3) D980P, V982P, F812P, A887P, A894P, A937P, R677G, R678S, and R680S; (4) K981P, V982P, F812P, A887P, A894P, A937P, R677G, R678S, and R680S; or (5) The composition or medical preparation, comprising an amino acid sequence including one of D980P, K981P, V982P, F812P, A887P, A894P, A937P, R677G, R678S, and R680S.

32. 1. A composition or medical preparation comprising a first RNA encoding a first SARS-CoV-2 S polypeptide or an immunogenic fragment thereof, and a second RNA encoding a second SARS-CoV-2 S polypeptide or an immunogenic fragment thereof, (a) the first SARS-CoV-2 S polypeptide or fragment comprises an amino acid sequence having at least 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identity to SEQ ID NO:70; (b) the second SARS-CoV-2 S polypeptide or fragment has at least 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identity to SEQ ID NO:1 and the following set of amino acid substitutions relative to SEQ ID NO:1: (1) D985P, V987P, F817P, A892P, A899P, and A942P; (2) K986P, V987P, F817P, A892P, A899P, and A942P; (3) D985P, V987P, F817P, A892P, A899P, A942P, D614G, R682G, R683S, and R685S; (4) K986P, V987P, F817P, A892P, A899P, A942P, D614G, R682G, R683S, and R685S; (5) D985P, K986P, V987P, F817P, A892P, A899P, A942P, D614G, R682G, R683S, and R685S; (6) D985P, V987P, F817P, A892P, A899P, A942P, R682G, R683S, and R685S; (7) K986P, V987P, F817P, A892P, A899P, A942P, R682G, R683S, and R685S; or (8) The composition or medical preparation, comprising an amino acid sequence including one of D985P, K986P, V987P, F817P, A892P, A899P, A942P, R682G, R683S, and R685S.

33. 1. A composition or medical preparation comprising a first RNA encoding a first SARS-CoV-2 S polypeptide or an immunogenic fragment thereof, and a second RNA encoding a second SARS-CoV-2 S polypeptide or an immunogenic fragment thereof, (a) the first SARS-CoV-2 S polypeptide or fragment comprises an amino acid sequence having at least 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identity to SEQ ID NO:70; (b) the second SARS-CoV-2 S polypeptide or fragment has at least 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identity to SEQ ID NO:69 and has the following set of amino acid substitutions relative to SEQ ID NO:69: (1) D982P, V984P, F814P, A889P, A896P, and A939P; (2) K983P, V984P, F814P, A889P, A896P, and A939P; (3) D982P, V984P, F814P, A889P, A896P, A939P, R679G, R680S, and R682S; (4) K983P, V984P, F814P, A889P, A896P, A939P, R679G, R680S, and R682S; or (5) The composition or medical preparation, comprising an amino acid sequence including one of D982P, K983P, V984P, F814P, A889P, A896P, A939P, R679G, R680S, and R682S.

34. 1. A composition or medical preparation comprising a first RNA encoding a first SARS-CoV-2 S polypeptide or an immunogenic fragment thereof, and a second RNA encoding a second SARS-CoV-2 S polypeptide or an immunogenic fragment thereof, (a) the first SARS-CoV-2 S polypeptide or fragment comprises an amino acid sequence having at least 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identity to SEQ ID NO:70; (b) the second SARS-CoV-2 S polypeptide or fragment has at least 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identity to SEQ ID NO:70 and has the following set of amino acid substitutions relative to SEQ ID NO:70: (1) D982P, V984P, F814P, A889P, A896P, and A939P; (2) K983P, V984P, F814P, A889P, A896P, and A939P; (3) D982P, V984P, F814P, A889P, A896P, A939P, R679G, R680S, and R682S; (4) K983P, V984P, F814P, A889P, A896P, A939P, R679G, R680S, and R682S; or (5) The composition or medical preparation, comprising an amino acid sequence including one of D982P, K983P, V984P, F814P, A889P, A896P, A939P, R679G, R680S, and R682S.

35. 1. A composition or medical preparation comprising a first RNA encoding a first SARS-CoV-2 S polypeptide or an immunogenic fragment thereof, and a second RNA encoding a second SARS-CoV-2 S polypeptide or an immunogenic fragment thereof, (a) the first SARS-CoV-2 S polypeptide or fragment comprises an amino acid sequence having at least 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identity to SEQ ID NO:70; (b) the second SARS-CoV-2 S polypeptide or fragment has at least 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identity to SEQ ID NO: 104 or 105 and has the following set of amino acid substitutions relative to SEQ ID NO: 104 or 105: (1) D980P, V982P, F812P, A887P, A894P, and A937P; (2) K981P, V982P, F812P, A887P, A894P, and A937P; (3) D980P, V982P, F812P, A887P, A894P, A937P, R677G, R678S, and R680S; (4) K981P, V982P, F812P, A887P, A894P, A937P, R677G, R678S, and R680S; or (5) The composition or medical preparation, comprising an amino acid sequence including one of D980P, K981P, V982P, F812P, A887P, A894P, A937P, R677G, R678S, and R680S.

36. 1. A composition or medical preparation comprising a first RNA encoding a first SARS-CoV-2 S polypeptide or an immunogenic fragment thereof, and a second RNA encoding a second SARS-CoV-2 S polypeptide or an immunogenic fragment thereof, (a) the first SARS-CoV-2 S polypeptide or fragment comprises an amino acid sequence having at least 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identity to SEQ ID NO:104 or SEQ ID NO:105; (b) the second SARS-CoV-2 S polypeptide or fragment has at least 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identity to SEQ ID NO:1 and the following set of amino acid substitutions relative to SEQ ID NO:1: (1) D985P, V987P, F817P, A892P, A899P, and A942P; (2) K986P, V987P, F817P, A892P, A899P, and A942P; (3) D985P, V987P, F817P, A892P, A899P, A942P, D614G, R682G, R683S, and R685S; (4) K986P, V987P, F817P, A892P, A899P, A942P, D614G, R682G, R683S, and R685S; (5) D985P, K986P, V987P, F817P, A892P, A899P, A942P, D614G, R682G, R683S, and R685S; (6) D985P, V987P, F817P, A892P, A899P, A942P, R682G, R683S, and R685S; (7) K986P, V987P, F817P, A892P, A899P, A942P, R682G, R683S, and R685S; or (8) The composition or medical preparation, comprising an amino acid sequence including one of D985P, K986P, V987P, F817P, A892P, A899P, A942P, R682G, R683S, and R685S.

37. 1. A composition or medical preparation comprising a first RNA encoding a first SARS-CoV-2 S polypeptide or an immunogenic fragment thereof, and a second RNA encoding a second SARS-CoV-2 S polypeptide or an immunogenic fragment thereof, (a) the first SARS-CoV-2 S polypeptide or fragment comprises an amino acid sequence having at least 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identity to SEQ ID NO:104 or SEQ ID NO:105; (b) the second SARS-CoV-2 S polypeptide or fragment has at least 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identity to SEQ ID NO:69 and has the following set of amino acid substitutions relative to SEQ ID NO:69: (1) D982P, V984P, F814P, A889P, A896P, and A939P; (2) K983P, V984P, F814P, A889P, A896P, and A939P; (3) D982P, V984P, F814P, A889P, A896P, A939P, R679G, R680S, and R682S; (4) K983P, V984P, F814P, A889P, A896P, A939P, R679G, R680S, and R682S; or (5) The composition or medical preparation, comprising an amino acid sequence including one of D982P, K983P, V984P, F814P, A889P, A896P, A939P, R679G, R680S, and R682S.

38. 1. A composition or medical preparation comprising a first RNA encoding a first SARS-CoV-2 S polypeptide or an immunogenic fragment thereof, and a second RNA encoding a second SARS-CoV-2 S polypeptide or an immunogenic fragment thereof, (a) the first SARS-CoV-2 S polypeptide or fragment comprises an amino acid sequence having at least 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identity to SEQ ID NO:104 or SEQ ID NO:105; (b) the second SARS-CoV-2 S polypeptide or fragment has at least 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identity to SEQ ID NO:70 and has the following set of amino acid substitutions relative to SEQ ID NO:70: (1) D982P, V984P, F814P, A889P, A896P, and A939P; (2) K983P, V984P, F814P, A889P, A896P, and A939P; (3) D982P, V984P, F814P, A889P, A896P, A939P, R679G, R680S, and R682S; (4) K983P, V984P, F814P, A889P, A896P, A939P, R679G, R680S, and R682S; or (5) The composition or medical preparation, comprising an amino acid sequence including one of D982P, K983P, V984P, F814P, A889P, A896P, A939P, R679G, R680S, and R682S.

39. 1. A composition or medical preparation comprising a first RNA encoding a first SARS-CoV-2 S polypeptide or an immunogenic fragment thereof, and a second RNA encoding a second SARS-CoV-2 S polypeptide or an immunogenic fragment thereof, (a) the first SARS-CoV-2 S polypeptide or fragment comprises an amino acid sequence having at least 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identity to SEQ ID NO:104 or SEQ ID NO:105; (b) the second SARS-CoV-2 S polypeptide or fragment has at least 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identity to SEQ ID NO: 104 or 105 and has the following set of amino acid substitutions relative to SEQ ID NO: 104 or 105: (1) D980P, V982P, F812P, A887P, A894P, and A937P; (2) K981P, V982P, F812P, A887P, A894P, and A937P; (3) D980P, V982P, F812P, A887P, A894P, A937P, R677G, R678S, and R680S; (4) K981P, V982P, F812P, A887P, A894P, A937P, R677G, R678S, and R680S; or (5) The composition or medical preparation, comprising an amino acid sequence including one of D980P, K981P, V982P, F812P, A887P, A894P, A937P, R677G, R678S, and R680S.

40. 1. A composition or medical preparation comprising a first RNA encoding a first SARS-CoV-2 S polypeptide or an immunogenic fragment thereof, and a second RNA encoding a second SARS-CoV-2 S polypeptide or an immunogenic fragment thereof, (a) the first SARS-CoV-2 S polypeptide or fragment comprises an amino acid sequence having at least 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identity to SEQ ID NO:1 and including the following substitutions relative to SEQ ID NO:1: D985P, V987P, F817P, A892P, A899P, and A942P; (b) the second SARS-CoV-2 S polypeptide or fragment has at least 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identity to SEQ ID NO:69 and has the following set of amino acid substitutions relative to SEQ ID NO:69: (1) D982P, V984P, F814P, A889P, A896P, and A939P; (2) K983P, V984P, F814P, A889P, A896P, and A939P; (3) D982P, V984P, F814P, A889P, A896P, A939P, R679G, R680S, and R682S; (4) K983P, V984P, F814P, A889P, A896P, A939P, R679G, R680S, and R682S; or (5) The composition or medical preparation, comprising an amino acid sequence including one of D982P, K983P, V984P, F814P, A889P, A896P, A939P, R679G, R680S, and R682S.

41. 1. A composition or medical preparation comprising a first RNA encoding a first SARS-CoV-2 S polypeptide or an immunogenic fragment thereof, and a second RNA encoding a second SARS-CoV-2 S polypeptide or an immunogenic fragment thereof, (a) the first SARS-CoV-2 S polypeptide or fragment comprises an amino acid sequence having at least 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identity to SEQ ID NO:1 and including the following substitutions relative to SEQ ID NO:1: K986P, V987P, F817P, A892P, A899P, and A942P; (b) the second SARS-CoV-2 S polypeptide or fragment has at least 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identity to SEQ ID NO:69 and has the following set of amino acid substitutions relative to SEQ ID NO:69: (1) D982P, V984P, F814P, A889P, A896P, and A939P; (2) K983P, V984P, F814P, A889P, A896P, and A939P; (3) D982P, V984P, F814P, A889P, A896P, A939P, R679G, R680S, and R682S; (4) K983P, V984P, F814P, A889P, A896P, A939P, R679G, R680S, and R682S; or (5) The composition or medical preparation, comprising an amino acid sequence including one of D982P, K983P, V984P, F814P, A889P, A896P, A939P, R679G, R680S, and R682S.

42. 1. A composition or medical preparation comprising a first RNA encoding a first SARS-CoV-2 S polypeptide or an immunogenic fragment thereof, and a second RNA encoding a second SARS-CoV-2 S polypeptide or an immunogenic fragment thereof, (a) the first SARS-CoV-2 S polypeptide or fragment comprises an amino acid sequence having at least 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identity to SEQ ID NO:1 and including the following substitutions relative to SEQ ID NO:1: D985P, V987P, F817P, A892P, A899P, A942P, D614G, R682G, R683S, and R685S; (b) the second SARS-CoV-2 S polypeptide or fragment has at least 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identity to SEQ ID NO:69 and has the following set of amino acid substitutions relative to SEQ ID NO:69: (1) D982P, V984P, F814P, A889P, A896P, and A939P; (2) K983P, V984P, F814P, A889P, A896P, and A939P; (3) D982P, V984P, F814P, A889P, A896P, A939P, R679G, R680S, and R682S; (4) K983P, V984P, F814P, A889P, A896P, A939P, R679G, R680S, and R682S; or (5) The composition or medical preparation, comprising an amino acid sequence including one of D982P, K983P, V984P, F814P, A889P, A896P, A939P, R679G, R680S, and R682S.

43. 1. A composition or medical preparation comprising a first RNA encoding a first SARS-CoV-2 S polypeptide or an immunogenic fragment thereof, and a second RNA encoding a second SARS-CoV-2 S polypeptide or an immunogenic fragment thereof, (a) the first SARS-CoV-2 S polypeptide or fragment comprises an amino acid sequence having at least 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identity to SEQ ID NO:1 and including the following substitutions relative to SEQ ID NO:1: K986P, V987P, F817P, A892P, A899P, A942P, D614G, R682G, R683S, and R685S; (b) the second SARS-CoV-2 S polypeptide or fragment has at least 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identity to SEQ ID NO:69 and has the following set of amino acid substitutions relative to SEQ ID NO:69: (1) D982P, V984P, F814P, A889P, A896P, and A939P; (2) K983P, V984P, F814P, A889P, A896P, and A939P; (3) D982P, V984P, F814P, A889P, A896P, A939P, R679G, R680S, and R682S; (4) K983P, V984P, F814P, A889P, A896P, A939P, R679G, R680S, and R682S; or (5) The composition or medical preparation, comprising an amino acid sequence including one of D982P, K983P, V984P, F814P, A889P, A896P, A939P, R679G, R680S, and R682S.

44. 1. A composition or medical preparation comprising a first RNA encoding a first SARS-CoV-2 S polypeptide or an immunogenic fragment thereof, and a second RNA encoding a second SARS-CoV-2 S polypeptide or an immunogenic fragment thereof, (a) the first SARS-CoV-2 S polypeptide or fragment comprises an amino acid sequence having at least 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identity to SEQ ID NO:1 and including the following substitutions relative to SEQ ID NO:1: D985P, K986P, V987P, F817P, A892P, A899P, A942P, D614G, R682G, R683S, and R685S; (b) the second SARS-CoV-2 S polypeptide or fragment has at least 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identity to SEQ ID NO:69 and has the following set of amino acid substitutions relative to SEQ ID NO:69: (1) D982P, V984P, F814P, A889P, A896P, and A939P; (2) K983P, V984P, F814P, A889P, A896P, and A939P; (3) D982P, V984P, F814P, A889P, A896P, A939P, R679G, R680S, and R682S; (4) K983P, V984P, F814P, A889P, A896P, A939P, R679G, R680S, and R682S; or (5) The composition or medical preparation, comprising an amino acid sequence including one of D982P, K983P, V984P, F814P, A889P, A896P, A939P, R679G, R680S, and R682S.

45. 1. A composition or medical preparation comprising a first RNA encoding a first SARS-CoV-2 S polypeptide or an immunogenic fragment thereof, and a second RNA encoding a second SARS-CoV-2 S polypeptide or an immunogenic fragment thereof, (a) the first SARS-CoV-2 S polypeptide or fragment comprises an amino acid sequence having at least 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identity to SEQ ID NO:1 and including the following substitutions relative to SEQ ID NO:1: D985P, V987P, F817P, A892P, A899P, A942P, R682G, R683S, and R685S; (b) the second SARS-CoV-2 S polypeptide or fragment has at least 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identity to SEQ ID NO:69 and has the following set of amino acid substitutions relative to SEQ ID NO:69: (1) D982P, V984P, F814P, A889P, A896P, and A939P; (2) K983P, V984P, F814P, A889P, A896P, and A939P; (3) D982P, V984P, F814P, A889P, A896P, A939P, R679G, R680S, and R682S; (4) K983P, V984P, F814P, A889P, A896P, A939P, R679G, R680S, and R682S; or (5) The composition or medical preparation, comprising an amino acid sequence including one of D982P, K983P, V984P, F814P, A889P, A896P, A939P, R679G, R680S, and R682S.

46. 1. A composition or medical preparation comprising a first RNA encoding a first SARS-CoV-2 S polypeptide or an immunogenic fragment thereof, and a second RNA encoding a second SARS-CoV-2 S polypeptide or an immunogenic fragment thereof, (a) the first SARS-CoV-2 S polypeptide or fragment comprises an amino acid sequence having at least 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identity to SEQ ID NO:1 and including the following substitutions relative to SEQ ID NO:1: K986P, V987P, F817P, A892P, A899P, A942P, R682G, R683S, and R685S; (b) the second SARS-CoV-2 S polypeptide or fragment has at least 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identity to SEQ ID NO:69 and has the following set of amino acid substitutions relative to SEQ ID NO:69: (1) D982P, V984P, F814P, A889P, A896P, and A939P; (2) K983P, V984P, F814P, A889P, A896P, and A939P; (3) D982P, V984P, F814P, A889P, A896P, A939P, R679G, R680S, and R682S; (4) K983P, V984P, F814P, A889P, A896P, A939P, R679G, R680S, and R682S; or (5) The composition or medical preparation, comprising an amino acid sequence including one of D982P, K983P, V984P, F814P, A889P, A896P, A939P, R679G, R680S, and R682S.

47. 1. A composition or medical preparation comprising a first RNA encoding a first SARS-CoV-2 S polypeptide or an immunogenic fragment thereof, and a second RNA encoding a second SARS-CoV-2 S polypeptide or an immunogenic fragment thereof, (a) the first SARS-CoV-2 S polypeptide or fragment comprises an amino acid sequence having at least 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identity to SEQ ID NO:1 and including the following substitutions relative to SEQ ID NO:1: D985P, K986P, V987P, F817P, A892P, A899P, A942P, R682G, R683S, R685S; (b) the second SARS-CoV-2 S polypeptide or fragment has at least 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identity to SEQ ID NO:69 and has the following set of amino acid substitutions relative to SEQ ID NO:69: (1) D982P, V984P, F814P, A889P, A896P, and A939P; (2) K983P, V984P, F814P, A889P, A896P, and A939P; (3) D982P, V984P, F814P, A889P, A896P, A939P, R679G, R680S, and R682S; (4) K983P, V984P, F814P, A889P, A896P, A939P, R679G, R680S, and R682S; or (5) The composition or medical preparation, comprising an amino acid sequence including one of D982P, K983P, V984P, F814P, A889P, A896P, A939P, R679G, R680S, and R682S.

48. 1. A composition or medical preparation comprising a first RNA encoding a first SARS-CoV-2 S polypeptide or an immunogenic fragment thereof, and a second RNA encoding a second SARS-CoV-2 S polypeptide or an immunogenic fragment thereof, (a) the first SARS-CoV-2 S polypeptide or fragment comprises an amino acid sequence having at least 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identity to SEQ ID NO:1 and including the following substitutions relative to SEQ ID NO:1: D985P, V987P, F817P, A892P, A899P, and A942P; (b) the second SARS-CoV-2 S polypeptide or fragment has at least 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identity to SEQ ID NO:70 and has the following set of amino acid substitutions relative to SEQ ID NO:70: (1) D982P, V984P, F814P, A889P, A896P, and A939P; (2) K983P, V984P, F814P, A889P, A896P, and A939P; (3) D982P, V984P, F814P, A889P, A896P, A939P, R679G, R680S, and R682S; (4) K983P, V984P, F814P, A889P, A896P, A939P, R679G, R680S, and R682S; or (5) The composition or medical preparation, comprising an amino acid sequence including one of D982P, K983P, V984P, F814P, A889P, A896P, A939P, R679G, R680S, and R682S.

49. 1. A composition or medical preparation comprising a first RNA encoding a first SARS-CoV-2 S polypeptide or an immunogenic fragment thereof, and a second RNA encoding a second SARS-CoV-2 S polypeptide or an immunogenic fragment thereof, (a) the first SARS-CoV-2 S polypeptide or fragment comprises an amino acid sequence having at least 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identity to SEQ ID NO:1 and including the following substitutions relative to SEQ ID NO:1: K986P, V987P, F817P, A892P, A899P, and A942P; (b) the second SARS-CoV-2 S polypeptide or fragment has at least 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identity to SEQ ID NO:70 and has the following set of amino acid substitutions relative to SEQ ID NO:70: (1) D982P, V984P, F814P, A889P, A896P, and A939P; (2) K983P, V984P, F814P, A889P, A896P, and A939P; (3) D982P, V984P, F814P, A889P, A896P, A939P, R679G, R680S, and R682S; (4) K983P, V984P, F814P, A889P, A896P, A939P, R679G, R680S, and R682S; or (5) The composition or medical preparation, comprising an amino acid sequence including one of D982P, K983P, V984P, F814P, A889P, A896P, A939P, R679G, R680S, and R682S.

50. 1. A composition or medical preparation comprising a first RNA encoding a first SARS-CoV-2 S polypeptide or an immunogenic fragment thereof, and a second RNA encoding a second SARS-CoV-2 S polypeptide or an immunogenic fragment thereof, (a) the first SARS-CoV-2 S polypeptide or fragment comprises an amino acid sequence having at least 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identity to SEQ ID NO:1 and including the following substitutions relative to SEQ ID NO:1: D985P, V987P, F817P, A892P, A899P, A942P, D614G, R682G, R683S, and R685S; (b) the second SARS-CoV-2 S polypeptide or fragment has at least 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identity to SEQ ID NO:70 and has the following set of amino acid substitutions relative to SEQ ID NO:70: (1) D982P, V984P, F814P, A889P, A896P, and A939P; (2) K983P, V984P, F814P, A889P, A896P, and A939P; (3) D982P, V984P, F814P, A889P, A896P, A939P, R679G, R680S, and R682S; (4) K983P, V984P, F814P, A889P, A896P, A939P, R679G, R680S, and R682S; or (5) The composition or medical preparation, comprising an amino acid sequence including one of D982P, K983P, V984P, F814P, A889P, A896P, A939P, R679G, R680S, and R682S.

51. 1. A composition or medical preparation comprising a first RNA encoding a first SARS-CoV-2 S polypeptide or an immunogenic fragment thereof, and a second RNA encoding a second SARS-CoV-2 S polypeptide or an immunogenic fragment thereof, (a) the first SARS-CoV-2 S polypeptide or fragment comprises an amino acid sequence having at least 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identity to SEQ ID NO:1 and including the following substitutions relative to SEQ ID NO:1: K986P, V987P, F817P, A892P, A899P, A942P, D614G, R682G, R683S, and R685S; (b) the second SARS-CoV-2 S polypeptide or fragment has at least 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identity to SEQ ID NO:70 and has the following set of amino acid substitutions relative to SEQ ID NO:70: (1) D982P, V984P, F814P, A889P, A896P, and A939P; (2) K983P, V984P, F814P, A889P, A896P, and A939P; (3) D982P, V984P, F814P, A889P, A896P, A939P, R679G, R680S, and R682S; (4) K983P, V984P, F814P, A889P, A896P, A939P, R679G, R680S, and R682S; or (5) The composition or medical preparation, comprising an amino acid sequence including one of D982P, K983P, V984P, F814P, A889P, A896P, A939P, R679G, R680S, and R682S.

52. 1. A composition or medical preparation comprising a first RNA encoding a first SARS-CoV-2 S polypeptide or an immunogenic fragment thereof, and a second RNA encoding a second SARS-CoV-2 S polypeptide or an immunogenic fragment thereof, (a) the first SARS-CoV-2 S polypeptide or fragment comprises an amino acid sequence having at least 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identity to SEQ ID NO:1 and including the following substitutions relative to SEQ ID NO:1: D985P, K986P, V987P, F817P, A892P, A899P, A942P, D614G, R682G, R683S, and R685S; (b) the second SARS-CoV-2 S polypeptide or fragment has at least 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identity to SEQ ID NO:70 and has the following set of amino acid substitutions relative to SEQ ID NO:70: (1) D982P, V984P, F814P, A889P, A896P, and A939P; (2) K983P, V984P, F814P, A889P, A896P, and A939P; (3) D982P, V984P, F814P, A889P, A896P, A939P, R679G, R680S, and R682S; (4) K983P, V984P, F814P, A889P, A896P, A939P, R679G, R680S, and R682S; or (5) The composition or medical preparation, comprising an amino acid sequence including one of D982P, K983P, V984P, F814P, A889P, A896P, A939P, R679G, R680S, and R682S.

53. 1. A composition or medical preparation comprising a first RNA encoding a first SARS-CoV-2 S polypeptide or an immunogenic fragment thereof, and a second RNA encoding a second SARS-CoV-2 S polypeptide or an immunogenic fragment thereof, (a) the first SARS-CoV-2 S polypeptide or fragment comprises an amino acid sequence having at least 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identity to SEQ ID NO:1 and including the following substitutions relative to SEQ ID NO:1: D985P, V987P, F817P, A892P, A899P, A942P, R682G, R683S, and R685S; (b) the second SARS-CoV-2 S polypeptide or fragment has at least 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identity to SEQ ID NO:70 and has the following set of amino acid substitutions relative to SEQ ID NO:70: (1) D982P, V984P, F814P, A889P, A896P, and A939P; (2) K983P, V984P, F814P, A889P, A896P, and A939P; (3) D982P, V984P, F814P, A889P, A896P, A939P, R679G, R680S, and R682S; (4) K983P, V984P, F814P, A889P, A896P, A939P, R679G, R680S, and R682S; or (5) The composition or medical preparation, comprising an amino acid sequence including one of D982P, K983P, V984P, F814P, A889P, A896P, A939P, R679G, R680S, and R682S.

54. 1. A composition or medical preparation comprising a first RNA encoding a first SARS-CoV-2 S polypeptide or an immunogenic fragment thereof, and a second RNA encoding a second SARS-CoV-2 S polypeptide or an immunogenic fragment thereof, (a) the first SARS-CoV-2 S polypeptide or fragment comprises an amino acid sequence having at least 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identity to SEQ ID NO:1 and including the following substitutions relative to SEQ ID NO:1: K986P, V987P, F817P, A892P, A899P, A942P, R682G, R683S, and R685S; (b) the second SARS-CoV-2 S polypeptide or fragment has at least 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identity to SEQ ID NO:70 and has the following set of amino acid substitutions relative to SEQ ID NO:70: (1) D982P, V984P, F814P, A889P, A896P, and A939P; (2) K983P, V984P, F814P, A889P, A896P, and A939P; (3) D982P, V984P, F814P, A889P, A896P, A939P, R679G, R680S, and R682S; (4) K983P, V984P, F814P, A889P, A896P, A939P, R679G, R680S, and R682S; or (5) The composition or medical preparation, comprising an amino acid sequence including one of D982P, K983P, V984P, F814P, A889P, A896P, A939P, R679G, R680S, and R682S.

55. 1. A composition or medical preparation comprising a first RNA encoding a first SARS-CoV-2 S polypeptide or an immunogenic fragment thereof, and a second RNA encoding a second SARS-CoV-2 S polypeptide or an immunogenic fragment thereof, (a) the first SARS-CoV-2 S polypeptide or fragment comprises an amino acid sequence having at least 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identity to SEQ ID NO:1 and including the following substitutions relative to SEQ ID NO:1: D985P, K986P, V987P, F817P, A892P, A899P, A942P, R682G, R683S, and R685S; (b) the second SARS-CoV-2 S polypeptide or fragment has at least 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identity to SEQ ID NO:70 and has the following set of amino acid substitutions relative to SEQ ID NO:70: (1) D982P, V984P, F814P, A889P, A896P, and A939P; (2) K983P, V984P, F814P, A889P, A896P, and A939P; (3) D982P, V984P, F814P, A889P, A896P, A939P, R679G, R680S, and R682S; (4) K983P, V984P, F814P, A889P, A896P, A939P, R679G, R680S, and R682S; or (5) The composition or medical preparation, comprising an amino acid sequence including one of D982P, K983P, V984P, F814P, A889P, A896P, A939P, R679G, R680S, and R682S.

56. 1. A composition or medical preparation comprising a first RNA encoding a first SARS-CoV-2 S polypeptide or an immunogenic fragment thereof, and a second RNA encoding a second SARS-CoV-2 S polypeptide or an immunogenic fragment thereof, (a) the first SARS-CoV-2 S polypeptide or fragment comprises an amino acid sequence having at least 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identity to SEQ ID NO:1 and including the following substitutions relative to SEQ ID NO:1: D985P, V987P, F817P, A892P, A899P, and A942P; (b) the second SARS-CoV-2 S polypeptide or fragment has at least 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identity to SEQ ID NO: 104 or 105 and has the following set of amino acid substitutions relative to SEQ ID NO: 104 or 105: (1) D980P, V982P, F812P, A887P, A894P, and A937P; (2) K981P, V982P, F812P, A887P, A894P, and A937P; (3) D980P, V982P, F812P, A887P, A894P, A937P, R677G, R678S, and R680S; (4) K981P, V982P, F812P, A887P, A894P, A937P, R677G, R678S, and R680S; or (5) The composition or medical preparation, comprising an amino acid sequence including one of D980P, K981P, V982P, F812P, A887P, A894P, A937P, R677G, R678S, and R680S.

57. 1. A composition or medical preparation comprising a first RNA encoding a first SARS-CoV-2 S polypeptide or an immunogenic fragment thereof, and a second RNA encoding a second SARS-CoV-2 S polypeptide or an immunogenic fragment thereof, (a) the first SARS-CoV-2 S polypeptide or fragment comprises an amino acid sequence having at least 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identity to SEQ ID NO:1 and including the following substitutions relative to SEQ ID NO:1: K986P, V987P, F817P, A892P, A899P, and A942P; (b) the second SARS-CoV-2 S polypeptide or fragment has at least 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identity to SEQ ID NO: 104 or 105 and has the following set of amino acid substitutions relative to SEQ ID NO: 104 or 105: (1) D980P, V982P, F812P, A887P, A894P, and A937P; (2) K981P, V982P, F812P, A887P, A894P, and A937P; (3) D980P, V982P, F812P, A887P, A894P, A937P, R677G, R678S, and R680S; (4) K981P, V982P, F812P, A887P, A894P, A937P, R677G, R678S, and R680S; or (5) The composition or medical preparation, comprising an amino acid sequence including one of D980P, K981P, V982P, F812P, A887P, A894P, A937P, R677G, R678S, and R680S.

58. 1. A composition or medical preparation comprising a first RNA encoding a first SARS-CoV-2 S polypeptide or an immunogenic fragment thereof, and a second RNA encoding a second SARS-CoV-2 S polypeptide or an immunogenic fragment thereof, (a) the first SARS-CoV-2 S polypeptide or fragment comprises an amino acid sequence having at least 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identity to SEQ ID NO:1 and including the following substitutions relative to SEQ ID NO:1: D985P, V987P, F817P, A892P, A899P, A942P, D614G, R682G, R683S, and R685S; (b) the second SARS-CoV-2 S polypeptide or fragment has at least 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identity to SEQ ID NO: 104 or 105 and has the following set of amino acid substitutions relative to SEQ ID NO: 104 or 105: (1) D980P, V982P, F812P, A887P, A894P, and A937P; (2) K981P, V982P, F812P, A887P, A894P, and A937P; (3) D980P, V982P, F812P, A887P, A894P, A937P, R677G, R678S, and R680S; (4) K981P, V982P, F812P, A887P, A894P, A937P, R677G, R678S, and R680S; or (5) The composition or medical preparation, comprising an amino acid sequence including one of D980P, K981P, V982P, F812P, A887P, A894P, A937P, R677G, R678S, and R680S.

59. 1. A composition or medical preparation comprising a first RNA encoding a first SARS-CoV-2 S polypeptide or an immunogenic fragment thereof, and a second RNA encoding a second SARS-CoV-2 S polypeptide or an immunogenic fragment thereof, (a) the first SARS-CoV-2 S polypeptide or fragment comprises an amino acid sequence having at least 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identity to SEQ ID NO:1 and including the following substitutions relative to SEQ ID NO:1: K986P, V987P, F817P, A892P, A899P, A942P, D614G, R682G, R683S, and R685S; (b) the second SARS-CoV-2 S polypeptide or fragment has at least 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identity to SEQ ID NO: 104 or 105 and has the following set of amino acid substitutions relative to SEQ ID NO: 104 or 105: (1) D980P, V982P, F812P, A887P, A894P, and A937P; (2) K981P, V982P, F812P, A887P, A894P, and A937P; (3) D980P, V982P, F812P, A887P, A894P, A937P, R677G, R678S, and R680S; (4) K981P, V982P, F812P, A887P, A894P, A937P, R677G, R678S, and R680S; or (5) The composition or medical preparation, comprising an amino acid sequence including one of D980P, K981P, V982P, F812P, A887P, A894P, A937P, R677G, R678S, and R680S.

60. 1. A composition or medical preparation comprising a first RNA encoding a first SARS-CoV-2 S polypeptide or an immunogenic fragment thereof, and a second RNA encoding a second SARS-CoV-2 S polypeptide or an immunogenic fragment thereof, (a) the first SARS-CoV-2 S polypeptide or fragment comprises an amino acid sequence having at least 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identity to SEQ ID NO:1 and including the following substitutions relative to SEQ ID NO:1: D985P, K986P, V987P, F817P, A892P, A899P, A942P, D614G, R682G, R683S, and R685S; (b) the second SARS-CoV-2 S polypeptide or fragment has at least 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identity to SEQ ID NO: 104 or 105 and has the following set of amino acid substitutions relative to SEQ ID NO: 104 or 105: (1) D980P, V982P, F812P, A887P, A894P, and A937P; (2) K981P, V982P, F812P, A887P, A894P, and A937P; (3) D980P, V982P, F812P, A887P, A894P, A937P, R677G, R678S, and R680S; (4) K981P, V982P, F812P, A887P, A894P, A937P, R677G, R678S, and R680S; or (5) The composition or medical preparation, comprising an amino acid sequence including one of D980P, K981P, V982P, F812P, A887P, A894P, A937P, R677G, R678S, and R680S.

61. 1. A composition or medical preparation comprising a first RNA encoding a first SARS-CoV-2 S polypeptide or an immunogenic fragment thereof, and a second RNA encoding a second SARS-CoV-2 S polypeptide or an immunogenic fragment thereof, (a) the first SARS-CoV-2 S polypeptide or fragment comprises an amino acid sequence having at least 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identity to SEQ ID NO:1 and including the following substitutions relative to SEQ ID NO:1: D985P, V987P, F817P, A892P, A899P, A942P, R682G, R683S, and R685S; (b) the second SARS-CoV-2 S polypeptide or fragment has at least 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identity to SEQ ID NO: 104 or 105 and has the following set of amino acid substitutions relative to SEQ ID NO: 104 or 105: (1) D980P, V982P, F812P, A887P, A894P, and A937P; (2) K981P, V982P, F812P, A887P, A894P, and A937P; (3) D980P, V982P, F812P, A887P, A894P, A937P, R677G, R678S, and R680S; (4) K981P, V982P, F812P, A887P, A894P, A937P, R677G, R678S, and R680S; or (5) The composition or medical preparation, comprising an amino acid sequence including one of D980P, K981P, V982P, F812P, A887P, A894P, A937P, R677G, R678S, and R680S.

62. 1. A composition or medical preparation comprising a first RNA encoding a first SARS-CoV-2 S polypeptide or an immunogenic fragment thereof, and a second RNA encoding a second SARS-CoV-2 S polypeptide or an immunogenic fragment thereof, (a) the first SARS-CoV-2 S polypeptide or fragment comprises an amino acid sequence having at least 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identity to SEQ ID NO:1 and including the following substitutions relative to SEQ ID NO:1: K986P, V987P, F817P, A892P, A899P, A942P, R682G, R683S, and R685S; (b) the second SARS-CoV-2 S polypeptide or fragment has at least 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identity to SEQ ID NO: 104 or 105 and has the following set of amino acid substitutions relative to SEQ ID NO: 104 or 105: (1) D980P, V982P, F812P, A887P, A894P, and A937P; (2) K981P, V982P, F812P, A887P, A894P, and A937P; (3) D980P, V982P, F812P, A887P, A894P, A937P, R677G, R678S, and R680S; (4) K981P, V982P, F812P, A887P, A894P, A937P, R677G, R678S, and R680S; or (5) The composition or medical preparation, comprising an amino acid sequence including one of D980P, K981P, V982P, F812P, A887P, A894P, A937P, R677G, R678S, and R680S.

63. 1. A composition or medical preparation comprising a first RNA encoding a first SARS-CoV-2 S polypeptide or an immunogenic fragment thereof, and a second RNA encoding a second SARS-CoV-2 S polypeptide or an immunogenic fragment thereof, (a) the first SARS-CoV-2 S polypeptide or fragment comprises an amino acid sequence having at least 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identity to SEQ ID NO:1 and including the following substitutions relative to SEQ ID NO:1: D985P, K986P, V987P, F817P, A892P, A899P, A942P, R682G, R683S, and R685S; (b) the second SARS-CoV-2 S polypeptide or fragment has at least 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identity to SEQ ID NO: 104 or 105 and has the following set of amino acid substitutions relative to SEQ ID NO: 104 or 105: (1) D980P, V982P, F812P, A887P, A894P, and A937P; (2) K981P, V982P, F812P, A887P, A894P, and A937P; (3) D980P, V982P, F812P, A887P, A894P, A937P, R677G, R678S, and R680S; (4) K981P, V982P, F812P, A887P, A894P, A937P, R677G, R678S, and R680S; or (5) The composition or medical preparation, comprising an amino acid sequence including one of D980P, K981P, V982P, F812P, A887P, A894P, A937P, R677G, R678S, and R680S.

64. 1. A composition or medical preparation comprising a first RNA encoding a first SARS-CoV-2 S polypeptide or an immunogenic fragment thereof, and a second RNA encoding a second SARS-CoV-2 S polypeptide or an immunogenic fragment thereof, (a) the first SARS-CoV-2 S polypeptide or fragment comprises an amino acid sequence having at least 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identity to SEQ ID NO:69 and including the following substitutions relative to SEQ ID NO:69: D982P, V984P, F814P, A889P, A896P, and A939P; (b) the second SARS-CoV-2 S polypeptide or fragment has at least 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identity to SEQ ID NO:70 and has the following set of amino acid substitutions relative to SEQ ID NO:70: (1) D982P, V984P, F814P, A889P, A896P, and A939P; (2) K983P, V984P, F814P, A889P, A896P, and A939P; (3) D982P, V984P, F814P, A889P, A896P, A939P, R679G, R680S, and R682S; (4) K983P, V984P, F814P, A889P, A896P, A939P, R679G, R680S, and R682S; or (5) The composition or medical preparation, comprising an amino acid sequence including one of D982P, K983P, V984P, F814P, A889P, A896P, A939P, R679G, R680S, and R682S.

65. 1. A composition or medical preparation comprising a first RNA encoding a first SARS-CoV-2 S polypeptide or an immunogenic fragment thereof, and a second RNA encoding a second SARS-CoV-2 S polypeptide or an immunogenic fragment thereof, (a) the first SARS-CoV-2 S polypeptide or fragment comprises an amino acid sequence having at least 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identity to SEQ ID NO:69 and including the following substitutions relative to SEQ ID NO:69: K983P, V984P, F814P, A889P, A896P, and A939P; (b) the second SARS-CoV-2 S polypeptide or fragment has at least 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identity to SEQ ID NO:70 and has the following set of amino acid substitutions relative to SEQ ID NO:70: (1) D982P, V984P, F814P, A889P, A896P, and A939P; (2) K983P, V984P, F814P, A889P, A896P, and A939P; (3) D982P, V984P, F814P, A889P, A896P, A939P, R679G, R680S, and R682S; (4) K983P, V984P, F814P, A889P, A896P, A939P, R679G, R680S, and R682S; or (5) The composition or medical preparation, comprising an amino acid sequence including one of D982P, K983P, V984P, F814P, A889P, A896P, A939P, R679G, R680S, and R682S.

66. 1. A composition or medical preparation comprising a first RNA encoding a first SARS-CoV-2 S polypeptide or an immunogenic fragment thereof, and a second RNA encoding a second SARS-CoV-2 S polypeptide or an immunogenic fragment thereof, (a) the first SARS-CoV-2 S polypeptide or fragment comprises an amino acid sequence having at least 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identity to SEQ ID NO:69 and including the following substitutions relative to SEQ ID NO:69: D982P, V984P, F814P, A889P, A896P, A939P, R679G, R680S, and R682S; (b) the second SARS-CoV-2 S polypeptide or fragment has at least 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identity to SEQ ID NO:70 and has the following set of amino acid substitutions relative to SEQ ID NO:70: (1) D982P, V984P, F814P, A889P, A896P, and A939P; (2) K983P, V984P, F814P, A889P, A896P, and A939P; (3) D982P, V984P, F814P, A889P, A896P, A939P, R679G, R680S, and R682S; (4) K983P, V984P, F814P, A889P, A896P, A939P, R679G, R680S, and R682S; or (5) The composition or medical preparation, comprising an amino acid sequence including one of D982P, K983P, V984P, F814P, A889P, A896P, A939P, R679G, R680S, and R682S.

67. 1. A composition or medical preparation comprising a first RNA encoding a first SARS-CoV-2 S polypeptide or an immunogenic fragment thereof, and a second RNA encoding a second SARS-CoV-2 S polypeptide or an immunogenic fragment thereof, (a) the first SARS-CoV-2 S polypeptide or fragment comprises an amino acid sequence having at least 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identity to SEQ ID NO:69 and including the following substitutions relative to SEQ ID NO:69: K983P, V984P, F814P, A889P, A896P, A939P, R679G, R680S, and R682S; (b) the second SARS-CoV-2 S polypeptide or fragment has at least 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identity to SEQ ID NO:70 and has the following set of amino acid substitutions relative to SEQ ID NO:70: (1) D982P, V984P, F814P, A889P, A896P, and A939P; (2) K983P, V984P, F814P, A889P, A896P, and A939P; (3) D982P, V984P, F814P, A889P, A896P, A939P, R679G, R680S, and R682S; (4) K983P, V984P, F814P, A889P, A896P, A939P, R679G, R680S, and R682S; or (5) The composition or medical preparation, comprising an amino acid sequence including one of D982P, K983P, V984P, F814P, A889P, A896P, A939P, R679G, R680S, and R682S.

68. 1. A composition or medical preparation comprising a first RNA encoding a first SARS-CoV-2 S polypeptide or an immunogenic fragment thereof, and a second RNA encoding a second SARS-CoV-2 S polypeptide or an immunogenic fragment thereof, (a) the first SARS-CoV-2 S polypeptide or fragment comprises an amino acid sequence having at least 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identity to SEQ ID NO:69 and including the following substitutions relative to SEQ ID NO:69: D982P, K983P, V984P, F814P, A889P, A896P, A939P, R679G, R680S, and R682S; (b) the second SARS-CoV-2 S polypeptide or fragment has at least 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identity to SEQ ID NO:70 and has the following set of amino acid substitutions relative to SEQ ID NO:70: (1) D982P, V984P, F814P, A889P, A896P, and A939P; (2) K983P, V984P, F814P, A889P, A896P, and A939P; (3) D982P, V984P, F814P, A889P, A896P, A939P, R679G, R680S, and R682S; (4) K983P, V984P, F814P, A889P, A896P, A939P, R679G, R680S, and R682S; or (5) The composition or medical preparation, comprising an amino acid sequence including one of D982P, K983P, V984P, F814P, A889P, A896P, A939P, R679G, R680S, and R682S.

69. 1. A composition or medical preparation comprising a first RNA encoding a first SARS-CoV-2 S polypeptide or an immunogenic fragment thereof, and a second RNA encoding a second SARS-CoV-2 S polypeptide or an immunogenic fragment thereof, (a) the first SARS-CoV-2 S polypeptide or fragment comprises an amino acid sequence having at least 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identity to SEQ ID NO:69 and including the following substitutions relative to SEQ ID NO:69: D982P, V984P, F814P, A889P, A896P, and A939P; (b) the second SARS-CoV-2 S polypeptide or fragment has at least 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identity to SEQ ID NO: 104 or 105 and has the following set of amino acid substitutions relative to SEQ ID NO: 104 or 105: (1) D980P, V982P, F812P, A887P, A894P, and A937P; (2) K981P, V982P, F812P, A887P, A894P, and A937P; (3) D980P, V982P, F812P, A887P, A894P, A937P, R677G, R678S, and R680S; (4) K981P, V982P, F812P, A887P, A894P, A937P, R677G, R678S, and R680S; or (5) The composition or medical preparation, comprising an amino acid sequence including one of D980P, K981P, V982P, F812P, A887P, A894P, A937P, R677G, R678S, and R680S.

70. 1. A composition or medical preparation comprising a first RNA encoding a first SARS-CoV-2 S polypeptide or an immunogenic fragment thereof, and a second RNA encoding a second SARS-CoV-2 S polypeptide or an immunogenic fragment thereof, (a) the first SARS-CoV-2 S polypeptide or fragment comprises an amino acid sequence having at least 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identity to SEQ ID NO:69 and including the following substitutions relative to SEQ ID NO:69: K983P, V984P, F814P, A889P, A896P, and A939P; (b) the second SARS-CoV-2 S polypeptide or fragment has at least 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identity to SEQ ID NO: 104 or 105 and has the following set of amino acid substitutions relative to SEQ ID NO: 104 or 105: (1) D980P, V982P, F812P, A887P, A894P, and A937P; (2) K981P, V982P, F812P, A887P, A894P, and A937P; (3) D980P, V982P, F812P, A887P, A894P, A937P, R677G, R678S, and R680S; (4) K981P, V982P, F812P, A887P, A894P, A937P, R677G, R678S, and R680S; or (5) The composition or medical preparation, comprising an amino acid sequence including one of D980P, K981P, V982P, F812P, A887P, A894P, A937P, R677G, R678S, and R680S.

71. 1. A composition or medical preparation comprising a first RNA encoding a first SARS-CoV-2 S polypeptide or an immunogenic fragment thereof, and a second RNA encoding a second SARS-CoV-2 S polypeptide or an immunogenic fragment thereof, (a) the first SARS-CoV-2 S polypeptide or fragment comprises an amino acid sequence having at least 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identity to SEQ ID NO:69 and including the following substitutions relative to SEQ ID NO:69: D982P, V984P, F814P, A889P, A896P, A939P, R679G, R680S, and R682S; (b) the second SARS-CoV-2 S polypeptide or fragment has at least 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identity to SEQ ID NO: 104 or 105 and has the following set of amino acid substitutions relative to SEQ ID NO: 104 or 105: (1) D980P, V982P, F812P, A887P, A894P, and A937P; (2) K981P, V982P, F812P, A887P, A894P, and A937P; (3) D980P, V982P, F812P, A887P, A894P, A937P, R677G, R678S, and R680S; (4) K981P, V982P, F812P, A887P, A894P, A937P, R677G, R678S, and R680S; or (5) The composition or medical preparation, comprising an amino acid sequence including one of D980P, K981P, V982P, F812P, A887P, A894P, A937P, R677G, R678S, and R680S.

72. 1. A composition or medical preparation comprising a first RNA encoding a first SARS-CoV-2 S polypeptide or an immunogenic fragment thereof, and a second RNA encoding a second SARS-CoV-2 S polypeptide or an immunogenic fragment thereof, (a) the first SARS-CoV-2 S polypeptide or fragment comprises an amino acid sequence having at least 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identity to SEQ ID NO:69 and including the following substitutions relative to SEQ ID NO:69: K983P, V984P, F814P, A889P, A896P, A939P, R679G, R680S, and R682S; (b) the second SARS-CoV-2 S polypeptide or fragment has at least 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identity to SEQ ID NO: 104 or 105 and has the following set of amino acid substitutions relative to SEQ ID NO: 104 or 105: (1) D980P, V982P, F812P, A887P, A894P, and A937P; (2) K981P, V982P, F812P, A887P, A894P, and A937P; (3) D980P, V982P, F812P, A887P, A894P, A937P, R677G, R678S, and R680S; (4) K981P, V982P, F812P, A887P, A894P, A937P, R677G, R678S, and R680S; or (5) The composition or medical preparation, comprising an amino acid sequence including one of D980P, K981P, V982P, F812P, A887P, A894P, A937P, R677G, R678S, and R680S.

73. 1. A composition or medical preparation comprising a first RNA encoding a first SARS-CoV-2 S polypeptide or an immunogenic fragment thereof, and a second RNA encoding a second SARS-CoV-2 S polypeptide or an immunogenic fragment thereof, (a) the first SARS-CoV-2 S polypeptide or fragment comprises an amino acid sequence having at least 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identity to SEQ ID NO:69 and including the following substitutions relative to SEQ ID NO:69: D982P, K983P, V984P, F814P, A889P, A896P, A939P, R679G, R680S, and R682S; (b) the second SARS-CoV-2 S polypeptide or fragment has at least 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identity to SEQ ID NO: 104 or 105 and has the following set of amino acid substitutions relative to SEQ ID NO: 104 or 105: (1) D980P, V982P, F812P, A887P, A894P, and A937P; (2) K981P, V982P, F812P, A887P, A894P, and A937P; (3) D980P, V982P, F812P, A887P, A894P, A937P, R677G, R678S, and R680S; (4) K981P, V982P, F812P, A887P, A894P, A937P, R677G, R678S, and R680S; or (5) The composition or medical preparation, comprising an amino acid sequence including one of D980P, K981P, V982P, F812P, A887P, A894P, A937P, R677G, R678S, and R680S.

74. 1. A composition or medical preparation comprising a first RNA encoding a first SARS-CoV-2 S polypeptide or an immunogenic fragment thereof, and a second RNA encoding a second SARS-CoV-2 S polypeptide or an immunogenic fragment thereof, (a) the first SARS-CoV-2 S polypeptide or fragment comprises an amino acid sequence having at least 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identity to SEQ ID NO:70 and including the following substitutions relative to SEQ ID NO:70: D982P, V984P, F814P, A889P, A896P, and A939P; (b) the second SARS-CoV-2 S polypeptide or fragment has at least 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identity to SEQ ID NO: 104 or 105 and has the following set of amino acid substitutions relative to SEQ ID NO: 104 or 105: (1) D980P, V982P, F812P, A887P, A894P, and A937P; (2) K981P, V982P, F812P, A887P, A894P, and A937P; (3) D980P, V982P, F812P, A887P, A894P, A937P, R677G, R678S, and R680S; (4) K981P, V982P, F812P, A887P, A894P, A937P, R677G, R678S, and R680S; or (5) The composition or medical preparation, comprising an amino acid sequence including one of D980P, K981P, V982P, F812P, A887P, A894P, A937P, R677G, R678S, and R680S.

75. 1. A composition or medical preparation comprising a first RNA encoding a first SARS-CoV-2 S polypeptide or an immunogenic fragment thereof, and a second RNA encoding a second SARS-CoV-2 S polypeptide or an immunogenic fragment thereof, (a) the first SARS-CoV-2 S polypeptide or fragment comprises an amino acid sequence having at least 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identity to SEQ ID NO:70 and including the following substitutions relative to SEQ ID NO:70: K983P, V984P, F814P, A889P, A896P, and A939P; (b) the second SARS-CoV-2 S polypeptide or fragment has at least 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identity to SEQ ID NO: 104 or 105 and has the following set of amino acid substitutions relative to SEQ ID NO: 104 or 105: (1) D980P, V982P, F812P, A887P, A894P, and A937P; (2) K981P, V982P, F812P, A887P, A894P, and A937P; (3) D980P, V982P, F812P, A887P, A894P, A937P, R677G, R678S, and R680S; (4) K981P, V982P, F812P, A887P, A894P, A937P, R677G, R678S, and R680S; or (5) The composition or medical preparation, comprising an amino acid sequence including one of D980P, K981P, V982P, F812P, A887P, A894P, A937P, R677G, R678S, and R680S.

76. 1. A composition or medical preparation comprising a first RNA encoding a first SARS-CoV-2 S polypeptide or an immunogenic fragment thereof, and a second RNA encoding a second SARS-CoV-2 S polypeptide or an immunogenic fragment thereof, (a) the first SARS-CoV-2 S polypeptide or fragment comprises an amino acid sequence having at least 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identity to SEQ ID NO:70 and including the following substitutions relative to SEQ ID NO:70: D982P, V984P, F814P, A889P, A896P, A939P, R679G, R680S, and R682S; (b) the second SARS-CoV-2 S polypeptide or fragment has at least 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identity to SEQ ID NO: 104 or 105 and has the following set of amino acid substitutions relative to SEQ ID NO: 104 or 105: (1) D980P, V982P, F812P, A887P, A894P, and A937P; (2) K981P, V982P, F812P, A887P, A894P, and A937P; (3) D980P, V982P, F812P, A887P, A894P, A937P, R677G, R678S, and R680S; (4) K981P, V982P, F812P, A887P, A894P, A937P, R677G, R678S, and R680S; or (5) The composition or medical preparation, comprising an amino acid sequence including one of D980P, K981P, V982P, F812P, A887P, A894P, A937P, R677G, R678S, and R680S.

77. 1. A composition or medical preparation comprising a first RNA encoding a first SARS-CoV-2 S polypeptide or an immunogenic fragment thereof, and a second RNA encoding a second SARS-CoV-2 S polypeptide or an immunogenic fragment thereof, (a) the first SARS-CoV-2 S polypeptide or fragment comprises an amino acid sequence having at least 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identity to SEQ ID NO:70 and including the following substitutions relative to SEQ ID NO:70: K983P, V984P, F814P, A889P, A896P, A939P, R679G, R680S, and R682S; (b) the second SARS-CoV-2 S polypeptide or fragment has at least 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identity to SEQ ID NO: 104 or 105 and has the following set of amino acid substitutions relative to SEQ ID NO: 104 or 105: (1) D980P, V982P, F812P, A887P, A894P, and A937P; (2) K981P, V982P, F812P, A887P, A894P, and A937P; (3) D980P, V982P, F812P, A887P, A894P, A937P, R677G, R678S, and R680S; (4) K981P, V982P, F812P, A887P, A894P, A937P, R677G, R678S, and R680S; or (5) The composition or medical preparation, comprising an amino acid sequence including one of D980P, K981P, V982P, F812P, A887P, A894P, A937P, R677G, R678S, and R680S.

78. 1. A composition or medical preparation comprising a first RNA encoding a first SARS-CoV-2 S polypeptide or an immunogenic fragment thereof, and a second RNA encoding a second SARS-CoV-2 S polypeptide or an immunogenic fragment thereof, (a) the first SARS-CoV-2 S polypeptide or fragment comprises an amino acid sequence having at least 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identity to SEQ ID NO:70 and including the following substitutions relative to SEQ ID NO:70: D982P, K983P, V984P, F814P, A889P, A896P, A939P, R679G, R680S, and R682S; (b) the second SARS-CoV-2 S polypeptide or fragment has at least 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identity to SEQ ID NO: 104 or 105 and has the following set of amino acid substitutions relative to SEQ ID NO: 104 or 105: (1) D980P, V982P, F812P, A887P, A894P, and A937P; (2) K981P, V982P, F812P, A887P, A894P, and A937P; (3) D980P, V982P, F812P, A887P, A894P, A937P, R677G, R678S, and R680S; (4) K981P, V982P, F812P, A887P, A894P, A937P, R677G, R678S, and R680S; or (5) The composition or medical preparation, comprising an amino acid sequence including one of D980P, K981P, V982P, F812P, A887P, A894P, A937P, R677G, R678S, and R680S.

79. 79. A composition or medical preparation according to any one of claims 24 to 78, wherein the first RNA and the second RNA each comprise a modified nucleoside in place of uridine.

80. 80. The composition or medical preparation of any one of claims 24 to 79, wherein the first RNA and the second RNA each comprise modified uridines in place of all uridines.

81. 81. The composition or medical preparation of any one of claims 24 to 80, wherein the first RNA and the second RNA each comprise N1-methyl-pseudouridine (m1ψ) in place of all uridines.

82. 82. The composition or medical preparation of any one of claims 24 to 81, wherein the first RNA and the second RNA each comprise a 5' cap.

83. The 5' cap is ２ ７，３’－Ｏ Gppp(m １ ２’－Ｏ 83. The composition or medical preparation of claim 82, comprising ApG.

84. 84. The composition or medical preparation of any one of claims 24 to 83, wherein the first RNA and the second RNA each comprise a 5'-UTR that is or comprises a modified human alpha-globin 5'-UTR.

85. 85. The composition or medical preparation of any one of claims 24 to 84, wherein the first RNA and the second RNA each comprise a 5'UTR comprising a nucleotide sequence of SEQ ID NO:12 or a nucleotide sequence that is at least 99%, 98%, 97%, 96%, 95%, 90%, 85%, or 80% identical to the nucleotide sequence of SEQ ID NO:

12.

86. 86. The composition or medical preparation of any one of claims 24 to 85, wherein the first RNA and the second RNA each comprise a 3'-UTR that is or includes a first sequence from a split amino-terminal enhancer (AES) messenger RNA and a second sequence from a mitochondrially encoded 12S ribosomal RNA.

87. 87. The composition or medical preparation of any one of claims 24 to 86, wherein the first RNA and the second RNA each comprise a 3'UTR comprising a nucleotide sequence of SEQ ID NO:13, or a nucleotide sequence that is at least 99%, 98%, 97%, 96%, 95%, 90%, 85%, or 80% identical to the nucleotide sequence of SEQ ID NO:

13.

88. A composition or medical preparation according to any one of claims 24 to 87, wherein the first RNA and the second RNA each comprise a polyA sequence.

89. 90. The composition or medical preparation of claim 88, wherein the first RNA and the second RNA each comprise a polyA sequence comprising at least 100 nucleotides.

90. 90. The composition or medical preparation of claim 88 or 89, wherein the first RNA and the second RNA each comprise 30 adenine nucleotides followed by 70 adenine nucleotides, the 30 adenine nucleotides and the 70 adenine nucleotides being separated by a linker sequence.

91. 91. The composition or medical preparation of any one of claims 88 to 90, wherein the first RNA and the second RNA each comprise a polyA sequence comprising or consisting of the nucleotide sequence of SEQ ID NO:14 or a nucleotide sequence that is at least 99%, 98%, 97%, 96%, 95%, 90%, 85%, or 80% identical to the nucleotide sequence of SEQ ID NO:

14.

92. 92. The composition or medical preparation of any one of claims 24 to 91, wherein the first RNA and the second RNA are each formulated for intramuscular administration or are intended to be formulated for intramuscular administration.

93. 93. A composition or medical preparation according to any one of claims 24 to 92, wherein said first RNA and said second RNA are each formulated as particles or are to be formulated as particles.

94. 94. The composition or medical preparation of claim 93, wherein the first RNA and the second RNA are each to be formulated as a lipid nanoparticle (LNP) or lipoplex (LPX) particle.

95. 95. The composition or medical preparation of claim 94, wherein the LNP comprises a cationically ionizable lipid, a neutral lipid, a sterol, and a polymer-lipid conjugate.

96. The immunogenic composition of claim 94 or 95, wherein the first RNA and the second RNA are formulated in separate LNPs.

97. The immunogenic composition of claim 94 or 95, wherein the first RNA and the second RNA are formulated in the same LNP.

98. 95. The composition or medical preparation of claim 94, wherein said lipoplex particles are obtainable by mixing said RNA with liposomes.

99. 99. The composition or medical preparation of any one of claims 24 to 98, wherein the first RNA and the second RNA are each mRNA, or the first RNA and the second RNA are each saRNA.

100. A composition or medical preparation according to any one of claims 24 to 99, which is a pharmaceutical composition.

101. A composition or medical preparation according to any one of claims 24 to 100 which is a vaccine.

102. 102. The composition or medical preparation of claim 100 or 101, wherein the pharmaceutical composition further comprises one or more pharma- ceutically acceptable carriers, diluents, and / or excipients.

103. A method of inducing an immune response in a subject, comprising inducing an immune response in the subject by administering to the subject a composition or medical preparation according to any one of claims 1 to 23.

104. 104. The method of claim 103, wherein the SARS-CoV-2 S polypeptide comprises an amino acid sequence that does not contain a D985P substitution relative to SEQ ID NO:1, an amino acid sequence that does not contain a D982P substitution relative to SEQ ID NO:69 or SEQ ID NO:70, or an amino acid sequence that does not contain a D980P substitution relative to SEQ ID NO:104 or SEQ ID NO:

105.

105. 105. The method of claim 103 or 104, further comprising administering a second RNA encoding a second SARS-CoV-2 S polypeptide or immunogenic fragment thereof, wherein said second SARS-CoV-2 S polypeptide or immunogenic fragment is a SARS-CoV-2 S polypeptide of the omicron variant that is not the BA.1 omicron variant.

106. 105. The method of claim 103 or 104, further comprising administering a second, different RNA encoding a second SARS-CoV-2 S polypeptide or immunogenic fragment thereof, wherein said second SARS-CoV-2 S polypeptide or fragment is selected from the SARS-CoV-2 S polypeptides or fragments listed in claim 1.

107. 103. A method of inducing an immune response in a subject, comprising inducing an immune response in the subject by administering to the subject a composition or medical preparation according to any one of claims 24 to 102.

108. 108. The method of claim 107, wherein the SARS-CoV-2 S polypeptide comprises an amino acid sequence that does not contain a D985P substitution relative to SEQ ID NO:1, an amino acid sequence that does not contain a D982P substitution relative to SEQ ID NO:69 or SEQ ID NO:70, or an amino acid sequence that does not contain a D980P substitution relative to SEQ ID NO:104 or SEQ ID NO:

105.

109. The method of claim 107 or 108, further comprising administering a second composition or medical preparation, wherein said second composition or medical preparation comprises RNA encoding a SARS-CoV-2 S polypeptide or immunogenic fragment of an omicron variant that is not the BA.1 omicron variant.

110. 109. The method of claim 107 or 108, further comprising administering a second composition or medical preparation, said second composition or medical preparation comprising a third RNA encoding a third SARS-CoV-2 S polypeptide or immunogenic fragment thereof, and a fourth RNA encoding a fourth SARS-CoV-2 S polypeptide or immunogenic fragment thereof.

111. 111. The method of claim 110, wherein the third RNA encodes a SARS-CoV-2 S polypeptide or an immunogenic fragment thereof that is a first or second SARS-CoV-2 S polypeptide or an immunogenic fragment thereof recited in any one of claims 24-102, and wherein the third RNA encodes a SARS-CoV-2 S polypeptide or fragment that is different from the first SARS-CoV-2 S polypeptide or fragment encoded by the first RNA and / or different from the second SARS-CoV-2 S polypeptide or fragment encoded by the second RNA.

112. 111. The method of claim 110, wherein the fourth RNA encodes a SARS-CoV-2 S polypeptide or an immunogenic fragment thereof that is a first or second SARS-CoV-2 S polypeptide or an immunogenic fragment thereof recited in any one of claims 24-102, and wherein the fourth RNA encodes a SARS-CoV-2 S polypeptide or fragment that is different from the first SARS-CoV-2 S polypeptide or fragment encoded by the first RNA and / or different from the second SARS-CoV-2 S polypeptide or fragment encoded by the second RNA.

113. 111. The method of claim 110, wherein the third RNA encodes a SARS-CoV-2 S polypeptide or immunogenic fragment thereof that is a first or second SARS-CoV-2 S polypeptide or immunogenic fragment thereof recited in any one of claims 24-102, and wherein the third RNA encodes a SARS-CoV-2 S polypeptide or fragment that is different from the first SARS-CoV-2 S polypeptide or fragment encoded by the first RNA and different from the second SARS-CoV-2 S polypeptide or fragment encoded by the second RNA.

114. 111. The method of claim 110, wherein the fourth RNA encodes a SARS-CoV-2 S polypeptide or immunogenic fragment thereof that is a first or second SARS-CoV-2 S polypeptide or immunogenic fragment thereof recited in any one of claims 24-102, and wherein the fourth RNA encodes a SARS-CoV-2 S polypeptide or fragment that is different from the first SARS-CoV-2 S polypeptide or fragment encoded by the first RNA and different from the second SARS-CoV-2 S polypeptide or fragment encoded by the second RNA.

115. The third RNA encodes a SARS-CoV-2 S polypeptide or an immunogenic fragment thereof that is a first or second SARS-CoV-2 S polypeptide or an immunogenic fragment thereof recited in any one of claims 24 to 102, the third RNA encodes a SARS-CoV-2 S polypeptide or fragment that is different from the first SARS-CoV-2 S polypeptide or fragment encoded by the first RNA and different from the second SARS-CoV-2 S polypeptide or fragment encoded by the second RNA, the fourth RNA encodes a SARS-CoV-2 S polypeptide or an immunogenic fragment thereof that is a first or second SARS-CoV-2 S polypeptide or an immunogenic fragment thereof recited in any one of claims 24 to 102, the fourth RNA encodes a SARS-CoV-2 S polypeptide or an immunogenic fragment thereof that is a first or second SARS-CoV-2 S polypeptide or an immunogenic fragment thereof recited in any one of claims 24 to 102, the fourth RNA encodes a SARS-CoV-2 S polypeptide or an immunogenic fragment thereof that is different from the first SARS-CoV-2 S polypeptide or fragment encoded by the first RNA.

111. The method of claim 110, wherein the second RNA encodes a SARS-CoV-2 S polypeptide or fragment that is distinct from the S polypeptide or fragment and that is distinct from the second SARS-CoV-2 S polypeptide or fragment encoded by the second RNA.

116. 116. The method of claim 115, wherein each of the first, second, third, and fourth RNAs encodes a different SARS-CoV-2 S polypeptide or an immunogenic fragment thereof.

117. 13. A method of inducing an immune response in a subject, comprising inducing an immune response in the subject by administering to the subject: (i) a composition or medical preparation according to any one of claims 1 to 23, and (ii) a composition or medical preparation according to any one of claims 24 to 102.

118. The method of claim 117, wherein the subject receiving the composition or medicinal preparation of (ii) has previously been administered the composition or medicinal preparation of (i).

119. The method of claim 117, wherein the subject receiving the composition or medicinal preparation of (i) has previously been administered the composition or medicinal preparation of (ii).

120. 118. The method of claim 117, wherein the composition or medical preparation of (i) and the composition or medical preparation of (ii) are administered together as a multivalent vaccine.

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