Three SARS-CoV-2 spike protein variants delivered intranasally by measles and mumps vaccines are broadly protective.

As the new SARS-CoV-2 Omicron variants and subvariants emerge, there is an urgency to develop intranasal, broadly protective vaccines. Here, we developed highly efficacious, intranasal trivalent SARS-CoV-2 vaccine candidates (TVC) based on three components of the MMR vaccine: measles virus (MeV), mumps virus (MuV) Jeryl Lynn (JL1) strain, and MuV JL2 strain. Specifically, MeV, MuV-JL1, and MuV-JL2 vaccine strains, each expressing prefusion spike (preS-6P) from a different variant of concern (VoC), were combined to generate TVCs.

A next-generation intranasal trivalent MMS vaccine induces durable and broad protection against SARS-CoV-2 variants of concern.

As SARS-CoV-2 variants of concern (VoCs) that evade immunity continue to emerge, next-generation adaptable COVID-19 vaccines which protect the respiratory tract and provide broader, more effective, and durable protection are urgently needed. Here, we have developed one such approach, a highly efficacious, intranasally delivered, trivalent measles-mumps-SARS-CoV-2 spike (S) protein (MMS) vaccine candidate that induces robust systemic and mucosal immunity with broad protection. This vaccine candidate is based on three components of the MMR vaccine, a measles virus Edmonston and the two mumps virus strains [Jeryl Lynn 1 (JL1) and JL2] that are known to provide safe, effective, and long-lasting protective immunity.

Recombinant measles virus expressing prefusion spike protein stabilized by six rather than two prolines is more efficacious against SARS-CoV-2 infection.

Measles virus (MeV) has been an excellent vector platform for delivering vaccines against many pathogens because of its high safety and efficacy, and induction of long-lived immunity. Early in the COVID-19 pandemic, a recombinant MeV (rMeV) expressing the prefusion full-length spike protein stabilized by two prolines (TMV-083) was developed and tested in phase 1 and 1/2 clinical trials but was discontinued because of insufficient immunogenicity and a low seroconversion rate in adults. Here, we compared the immunogenicity of rMeV expressing a soluble prefusion spike (preS) protein stabilized by two prolines (rMeV-preS-2P) with a rMeV expressing a soluble preS protein stabilized by six prolines (rMeV-preS-6P).

Prime-Pull Immunization of Mice with a BcfA-Adjuvanted Vaccine Elicits Sustained Mucosal Immunity That Prevents SARS-CoV-2 Infection and Pathology.

Vaccines against SARS-CoV-2 that induce mucosal immunity capable of preventing infection and disease remain urgently needed. In this study, we demonstrate the efficacy of Bordetella colonization factor A (BcfA), a novel bacteria-derived protein adjuvant, in SARS-CoV-2 spike-based prime-pull immunizations. We show that i.

Interferon-induced transmembrane protein 3 (IFITM3) limits lethality of SARS-CoV-2 in mice.

Interferon-induced transmembrane protein 3 (IFITM3) is an antiviral protein that alters cell membranes to block fusion of viruses. Conflicting reports identified opposing effects of IFITM3 on SARS-CoV-2 infection of cells, and its impact on viral pathogenesis in vivo remains unclear. Here, we show that IFITM3 knockout (KO) mice infected with SARS-CoV-2 experience extreme weight loss and lethality compared to mild infection in wild-type (WT) mice.

5-methylcytosine (mC) RNA modification controls the innate immune response to virus infection by regulating type I interferons.

5-methylcytosine (mC) is one of the most prevalent modifications of RNA, playing important roles in RNA metabolism, nuclear export, and translation. However, the potential role of RNA mC methylation in innate immunity remains elusive. Here, we show that depletion of NSUN2, an mC methyltransferase, significantly inhibits the replication and gene expression of a wide range of RNA and DNA viruses.

SARS-CoV-2 prefusion spike protein stabilized by six rather than two prolines is more potent for inducing antibodies that neutralize viral variants of concern.

The spike (S) protein of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is the main target for neutralizing antibodies (NAbs). The S protein trimer is anchored in the virion membrane in its prefusion (preS) but metastable form. The preS protein has been stabilized by introducing two or six proline substitutions, to generate stabilized, soluble 2P or HexaPro (6P) preS proteins.

A highly efficacious live attenuated mumps virus-based SARS-CoV-2 vaccine candidate expressing a six-proline stabilized prefusion spike.

With the rapid increase in SARS-CoV-2 cases in children, a safe and effective vaccine for this population is urgently needed. The MMR (measles/mumps/rubella) vaccine has been one of the safest and most effective human vaccines used in infants and children since the 1960s. Here, we developed live attenuated recombinant mumps virus (rMuV)-based SARS-CoV-2 vaccine candidates using the MuV Jeryl Lynn (JL2) vaccine strain backbone.

Caspase-4/11 exacerbates disease severity in SARS-CoV-2 infection by promoting inflammation and immunothrombosis.

Severe acute respiratory syndrome coronavirus 2 (SARS–CoV-2) is a worldwide health concern, and new treatment strategies are needed. Targeting inflammatory innate immunity pathways holds therapeutic promise, but effective molecular targets remain elusive. Here, we show that human caspase-4 (CASP4) and its mouse homolog, caspase-11 (CASP11), are up-regulated in SARS–CoV-2 infections and that CASP4 expression correlates with severity of SARS–CoV-2 infection in humans.

Heparanase Blockade as a Novel Dual-Targeting Therapy for COVID-19.

The coronavirus disease 2019 (COVID-19) pandemic, caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), has caused over 5 million deaths worldwide. Pneumonia and systemic inflammation contribute to its high mortality. Many viruses use heparan sulfate proteoglycans as coreceptors for viral entry, and heparanase (HPSE) is a known regulator of both viral entry and inflammatory cytokines.

Interferon-induced transmembrane protein 3 (IFITM3) limits lethality of SARS-CoV-2 in mice.

Interferon-induced transmembrane protein 3 (IFITM3) is a host antiviral protein that alters cell membranes to block fusion of viruses. Published reports have identified conflicting pro- and antiviral effects of IFITM3 on SARS-CoV-2 in cultured cells, and its impact on viral pathogenesis remains unclear. Here, we show that IFITM3 knockout (KO) mice infected with mouse-adapted SARS-CoV-2 experienced extreme weight loss and lethality, while wild type (WT) mice lost minimal weight and recovered.

Viral RNA N6-methyladenosine modification modulates both innate and adaptive immune responses of human respiratory syncytial virus.

Human respiratory syncytial virus (RSV) is the leading cause of respiratory tract infections in humans. A well-known challenge in the development of a live attenuated RSV vaccine is that interferon (IFN)-mediated antiviral responses are strongly suppressed by RSV nonstructural proteins which, in turn, dampens the subsequent adaptive immune responses. Here, we discovered a novel strategy to enhance innate and adaptive immunity to RSV infection.

A Methyltransferase-Defective Vesicular Stomatitis Virus-Based SARS-CoV-2 Vaccine Candidate Provides Complete Protection against SARS-CoV-2 Infection in Hamsters.

The current pandemic of coronavirus disease 2019 (COVID-19) caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has led to dramatic economic and health burdens. Although the worldwide SARS-CoV-2 vaccination campaign has begun, exploration of other vaccine candidates is needed due to uncertainties with the current approved vaccines, such as durability of protection, cross-protection against variant strains, and costs of long-term production and storage. In this study, we developed a methyltransferase-defective recombinant vesicular stomatitis virus (mtdVSV)-based SARS-CoV-2 vaccine candidate.

N6-methyladenosine promotes induction of ADAR1-mediated A-to-I RNA editing to suppress aberrant antiviral innate immune responses.

Among over 150 distinct RNA modifications, N6-methyladenosine (m6A) and adenosine-to-inosine (A-to-I) RNA editing represent 2 of the most studied modifications on mammalian mRNAs. Although both modifications occur on adenosine residues, knowledge on potential functional crosstalk between these 2 modifications is still limited. Here, we show that the m6A modification promotes expression levels of the ADAR1, which encodes an A-to-I RNA editing enzyme, in response to interferon (IFN) stimulation.

A safe and highly efficacious measles virus-based vaccine expressing SARS-CoV-2 stabilized prefusion spike.

The current pandemic of COVID-19 caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) highlights an urgent need to develop a safe, efficacious, and durable vaccine. Using a measles virus (rMeV) vaccine strain as the backbone, we developed a series of recombinant attenuated vaccine candidates expressing various forms of the SARS-CoV-2 spike (S) protein and its receptor binding domain (RBD) and evaluated their efficacy in cotton rat, IFNARmice, IFNAR-hCD46 mice, and golden Syrian hamsters. We found that rMeV expressing stabilized prefusion S protein (rMeV-preS) was more potent in inducing SARS-CoV-2-specific neutralizing antibodies than rMeV expressing full-length S protein (rMeV-S), while the rMeVs expressing different lengths of RBD (rMeV-RBD) were the least potent.

Nonsegmented Negative-Sense RNA Viruses Utilize -Methyladenosine (mA) as a Common Strategy To Evade Host Innate Immunity.

-Methyladenosine (mA) is the most abundant internal RNA modification catalyzed by host RNA methyltransferases. As obligate intracellular parasites, many viruses acquire mA methylation in their RNAs. However, the biological functions of viral mA methylation are poorly understood.

Stable Attenuation of Human Respiratory Syncytial Virus for Live Vaccines by Deletion and Insertion of Amino Acids in the Hinge Region between the mRNA Capping and Methyltransferase Domains of the Large Polymerase Protein.

Human respiratory syncytial virus (RSV) is the leading viral cause of lower respiratory tract disease in infants and children worldwide. Currently, there are no FDA-approved vaccines to combat this virus. The large (L) polymerase protein of RSV replicates the viral genome and transcribes viral mRNAs.

Vesicular Stomatitis Virus and DNA Vaccines Expressing Zika Virus Nonstructural Protein 1 Induce Substantial but Not Sterilizing Protection against Zika Virus Infection.

The nonstructural protein 1 (NS1) of several flaviviruses, including West Nile, dengue, and yellow fever viruses, is capable of inducing variable degrees of protection against flavivirus infection in animal models. However, the immunogenicity of NS1 protein of Zika virus (ZIKV) is less understood. Here, we determined the efficacy of ZIKV NS1-based vaccine candidates using two delivery platforms, methyltransferase-defective recombinant vesicular stomatitis virus (mtdVSV) and a DNA vaccine.

Porcine Epidemic Diarrhea Virus Deficient in RNA Cap Guanine-N-7 Methylation Is Attenuated and Induces Higher Type I and III Interferon Responses.

The 5' cap methylation of viral RNA plays important roles in RNA stability, efficient translation, and immune evasion. Thus, RNA cap methylation is an attractive target for antiviral discovery and development of new live attenuated vaccines. For coronaviruses, RNA cap structure is first methylated at the guanine-N-7 (G-N-7) position by nonstructural protein 14 (nsp14), which facilitates and precedes the subsequent ribose 2'-O methylation by the nsp16-nsp10 complex.

N-methyladenosine modification enables viral RNA to escape recognition by RNA sensor RIG-I.

Internal N-methyladenosine (mA) modification is one of the most common and abundant modifications of RNA. However, the biological roles of viral RNA mA remain elusive. Here, using human metapneumovirus (HMPV) as a model, we demonstrate that mA serves as a molecular marker for innate immune discrimination of self from non-self RNAs.

Viral N-methyladenosine upregulates replication and pathogenesis of human respiratory syncytial virus.

N-methyladenosine (mA) is the most prevalent internal modification of mRNAs in most eukaryotes. Here we show that RNAs of human respiratory syncytial virus (RSV) are modified by mA within discreet regions and that these modifications enhance viral replication and pathogenesis. Knockdown of mA methyltransferases decreases RSV replication and gene expression whereas knockdown of mA demethylases has the opposite effect.

Efficient Production of Human Norovirus-Specific IgY in Egg Yolks by Vaccination of Hens with a Recombinant Vesicular Stomatitis Virus Expressing VP1 Protein.

Human norovirus (HuNoV) is responsible for more than 95% of outbreaks of acute nonbacterial gastroenteritis worldwide. Despite major efforts, there are no vaccines or effective therapeutic interventions against this virus. Chicken immunoglobulin Y (IgY)-based passive immunization has been shown to be an effective strategy to prevent and treat many enteric viral diseases.

A Lactic Acid Bacteria (LAB)-Based Vaccine Candidate for Human Norovirus.

Human noroviruses (HuNoVs) are responsible for more than 95% of the non-bacterial acute gastroenteritis epidemics in the world. The CDC estimates that every year 21 million individuals suffer from HuNoV-induced gastroenteritis in the United States. Currently, there is no FDA-approved vaccine for HuNoVs.

A Zika virus vaccine expressing premembrane-envelope-NS1 polyprotein.

Current efforts to develop Zika virus (ZIKV) subunit vaccines have been focused on pre-membrane (prM) and envelope (E) proteins, but the role of NS1 in ZIKV-specific immune response and protection is poorly understood. Here, we develop an attenuated recombinant vesicular stomatitis virus (rVSV)-based vaccine expressing ZIKV prM-E-NS1 as a polyprotein. This vectored vaccine candidate is attenuated in mice, where a single immunization induces ZIKV-specific antibody and T cell immune responses that provide protection against ZIKV challenge.

IFITM3 Restricts Human Metapneumovirus Infection.

Human metapneumovirus (hMPV) utilizes a bifurcated cellular entry strategy, fusing either with the plasma membrane or, after endocytosis, with the endosome membrane. Whether cellular factors restrict or enhance either entry pathway is largely unknown. We found that the interferon-induced transmembrane protein 3 (IFITM3) inhibits hMPV infection to an extent similar to endocytosis-inhibiting drugs, and an IFITM3 variant that accumulates at the plasma membrane in addition to its endosome localization provided increased virus restriction.