Author Correction: Mosaic nanoparticle display of diverse influenza virus hemagglutinins elicits broad B cell responses.

In the version of this article initially published, the labels (50 Å) above the scale bars in Fig. 1b were incorrect. The correct size is 50 nm. The error has been corrected in the HTML and PDF versions of the article.

Mosaic nanoparticle display of diverse influenza virus hemagglutinins elicits broad B cell responses.

The present vaccine against influenza virus has the inevitable risk of antigenic discordance between the vaccine and the circulating strains, which diminishes vaccine efficacy. This necessitates new approaches that provide broader protection against influenza. Here we designed a vaccine using the hypervariable receptor-binding domain (RBD) of viral hemagglutinin displayed on a nanoparticle (np) able to elicit antibody responses that neutralize H1N1 influenza viruses spanning over 90 years. Co-display of RBDs from multiple strains across time, so that the adjacent RBDs are heterotypic, provides an avidity advantage to cross-reactive B cells. Immunization with the mosaic RBD-np elicited broader antibody responses than those induced by an admixture of nanoparticles encompassing the same set of RBDs as separate homotypic arrays. Furthermore, we identified a broadly neutralizing monoclonal antibody in a mouse immunized with mosaic RBD-np. The mosaic antigen array signifies a unique approach that subverts monotypic immunodominance and allows otherwise subdominant cross-reactive B cell responses to emerge.

Vaccine-Induced Antibodies that Neutralize Group 1 and Group 2 Influenza A Viruses.

Antibodies capable of neutralizing divergent influenza A viruses could form the basis of a universal vaccine. Here, from subjects enrolled in an H5N1 DNA/MIV-prime-boost influenza vaccine trial, we sorted hemagglutinin cross-reactive memory B cells and identified three antibody classes, each capable of neutralizing diverse subtypes of group 1 and group 2 influenza A viruses. Co-crystal structures with hemagglutinin revealed that each class utilized characteristic germline genes and convergent sequence motifs to recognize overlapping epitopes in the hemagglutinin stem. All six analyzed subjects had sequences from at least one multidonor class, and-in half the subjects-multidonor-class sequences were recovered from >40% of cross-reactive B cells. By contrast, these multidonor-class sequences were rare in published antibody datasets. Vaccination with a divergent hemagglutinin can thus increase the frequency of B cells encoding broad influenza A-neutralizing antibodies. We propose the sequence signature-quantified prevalence of these B cells as a metric to guide universal influenza A immunization strategies.

Induction of HIV Neutralizing Antibody Lineages in Mice with Diverse Precursor Repertoires.

The design of immunogens that elicit broadly reactive neutralizing antibodies (bnAbs) has been a major obstacle to HIV-1 vaccine development. One approach to assess potential immunogens is to use mice expressing precursors of human bnAbs as vaccination models. The bnAbs of the VRC01-class derive from the IGHV1-2 immunoglobulin heavy chain and neutralize a wide spectrum of HIV-1 strains via targeting the CD4 binding site of the envelope glycoprotein gp120. We now describe a mouse vaccination model that allows a germline human IGHV1-2(∗)02 segment to undergo normal V(D)J recombination and, thereby, leads to the generation of peripheral B cells that express a highly diverse repertoire of VRC01-related receptors. When sequentially immunized with modified gp120 glycoproteins designed to engage VRC01 germline and intermediate antibodies, IGHV1-2(∗)02-rearranging mice, which also express a VRC01-antibody precursor light chain, can support the affinity maturation of VRC01 precursor antibodies into HIV-neutralizing antibody lineages.

SARS-CoV-2 mRNA vaccine design enabled by prototype pathogen preparedness.

A vaccine for severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is needed to control the coronavirus disease 2019 (COVID-19) global pandemic. Structural studies have led to the development of mutations that stabilize Betacoronavirus spike proteins in the prefusion state, improving their expression and increasing immunogenicity1. This principle has been applied to design mRNA-1273, an mRNA vaccine that encodes a SARS-CoV-2 spike protein that is stabilized in the prefusion conformation. Here we show that mRNA-1273 induces potent neutralizing antibody responses to both wild-type (D614) and D614G mutant2 SARS-CoV-2 as well as CD8+ T cell responses, and protects against SARS-CoV-2 infection in the lungs and noses of mice without evidence of immunopathology. mRNA-1273 is currently in a phase III trial to evaluate its efficacy.

Low-Dose Subcutaneous or Intravenous Monoclonal Antibody to Prevent Malaria.

BACKGROUND: New approaches for the prevention and elimination of malaria, a leading cause of illness and death among infants and young children globally, are needed. METHODS: We conducted a phase 1 clinical trial to assess the safety and pharmacokinetics of L9LS, a next-generation antimalarial monoclonal antibody, and its protective efficacy against controlled human malaria infection in healthy adults who had never had malaria or received a vaccine for malaria. The participants received L9LS either intravenously or subcutaneously at a dose of 1 mg, 5 mg, or 20 mg per kilogram of body weight. Within 2 to 6 weeks after the administration of L9LS, both the participants who received L9LS and the control participants underwent controlled human malaria infection in which they were exposed to mosquitoes carrying Plasmodium falciparum (3D7 strain). RESULTS: No safety concerns were identified. L9LS had an estimated half-life of 56 days, and it had dose linearity, with the highest mean (±SD) maximum serum concentration (Cmax) of 914.2±146.5 µg per milliliter observed in participants who had received 20 mg per kilogram intravenously and the lowest mean Cmax of 41.5±4.7 µg per milliliter observed in those who had received 1 mg per kilogram intravenously; the mean Cmax was 164.8±31.1 in the participants who had received 5 mg per kilogram intravenously and 68.9±22.3 in those who had received 5 mg per kilogram subcutaneously. A total of 17 L9LS recipients and 6 control participants underwent controlled human malaria infection. Of the 17 participants who received a single dose of L9LS, 15 (88%) were protected after controlled human malaria infection. Parasitemia did not develop in any of the participants who received 5 or 20 mg per kilogram of intravenous L9LS. Parasitemia developed in 1 of 5 participants who received 1 mg per kilogram intravenously, 1 of 5 participants who received 5 mg per kilogram subcutaneously, and all 6 control participants through 21 days after the controlled human malaria infection. Protection conferred by L9LS was seen at serum concentrations as low as 9.2 µg per milliliter. CONCLUSIONS: In this small trial, L9LS administered intravenously or subcutaneously protected recipients against malaria after controlled infection, without evident safety concerns. (Funded by the National Institute of Allergy and Infectious Diseases; VRC 614 ClinicalTrials.gov number, NCT05019729.).

Structure-based design of a single-chain triple-disulfide-stabilized fusion-glycoprotein trimer that elicits high-titer neutralizing responses against human metapneumovirus.

The Pneumoviridae family of viruses includes human metapneumovirus (HMPV) and respiratory syncytial virus (RSV). The closely related Paramyxoviridae family includes parainfluenza viruses (PIVs). These three viral pathogens cause acute respiratory tract infections with substantial disease burden in the young, the elderly, and the immune-compromised. While promising subunit vaccines are being developed with prefusion-stabilized forms of the fusion glycoproteins (Fs) of RSV and PIVs, for which neutralizing titers elicited by the prefusion (pre-F) conformation of F are much higher than for the postfusion (post-F) conformation, with HMPV, pre-F and post-F immunogens described thus far elicit similar neutralizing responses, and it has been unclear which conformation, pre-F or post-F, would be the most effective HMPV F-vaccine immunogen. Here, we investigate the impact of further stabilizing HMPV F in the pre-F state. We replaced the furin-cleavage site with a flexible linker, creating a single chain F that yielded increased amounts of pre-F stabilized trimers, enabling the generation and assessment of F trimers stabilized by multiple disulfide bonds. Introduced prolines could increase both expression yields and antigenic recognition by the pre-F specific antibody, MPE8. The cryo-EM structure of a triple disulfide-stabilized pre-F trimer with the variable region of antibody MPE8 at 3.25-Å resolution confirmed the formation of designed disulfides and provided structural details on the MPE8 interface. Immunogenicity assessments in naïve mice showed the triple disulfide-stabilized pre-F trimer could elicit high titer neutralization, >10-fold higher than elicited by post-F. Immunogenicity assessments in pre-exposed rhesus macaques showed the triple disulfide-stabilized pre-F could recall high neutralizing titers after a single immunization, with little discrimination in the recall response between pre-F and post-F immunogens. However, the triple disulfide-stabilized pre-F adsorbed HMPV-directed responses from commercially available pooled human immunoglobulin more fully than post-F. Collectively, these results suggest single-chain triple disulfide-stabilized pre-F trimers to be promising HMPV-vaccine antigens.

Interprotomer disulfide-stabilized variants of the human metapneumovirus fusion glycoprotein induce high titer-neutralizing responses.

Human metapneumovirus (HMPV) is a major cause of respiratory disease worldwide, particularly among children and the elderly. Although there is no licensed HMPV vaccine, promising candidates have been identified for related pneumoviruses based on the structure-based stabilization of the fusion (F) glycoprotein trimer, with prefusion-stabilized F glycoprotein trimers eliciting significantly higher neutralizing responses than their postfusion F counterparts. However, immunization with HMPV F trimers in either prefusion or postfusion conformations has been reported to elicit equivalent neutralization responses. Here we investigate the impact of stabilizing disulfides, especially interprotomer disulfides (IP-DSs) linking protomers of the F trimer, on the elicitation of HMPV-neutralizing responses. We designed F trimer disulfides, screened for their expression, and used electron microscopy (EM) to confirm their formation, including that of an unexpected postfusion variant. In mice, IP-DS-stabilized prefusion and postfusion HMPV F elicited significantly higher neutralizing responses than non-IP-DS-stabilized HMPV Fs. In macaques, the impact of IP-DS stabilization was more measured, although IP-DS-stabilized variants of either prefusion or postfusion HMPV F induced neutralizing responses many times the average titers observed in a healthy human cohort. Serological and absorption-based analyses of macaque responses revealed elicited HMPV-neutralizing responses to be absorbed differently by IP-DS-containing and by non-IP-DS-containing postfusion Fs, suggesting IP-DS stabilization to alter not only the immunogenicity of select epitopes but their antigenicity as well. We speculate the observed increase in immunogenicity by IP-DS trimers to be related to reduced interprotomer flexibility within the HMPV F trimer.

SARS-CoV-2 S2P spike ages through distinct states with altered immunogenicity.

The SARS-CoV-2 spike is the primary target of virus-neutralizing antibodies and critical to the development of effective vaccines against COVID-19. Here, we demonstrate that the prefusion-stabilized two-proline "S2P" spike-widely employed for laboratory work and clinical studies-unfolds when stored at 4 °C, physiological pH, as observed by electron microscopy (EM) and differential scanning calorimetry, but that its trimeric, native-like conformation can be reacquired by low pH treatment. When stored for approximately 1 week, this unfolding does not significantly alter antigenic characteristics; however, longer storage diminishes antibody binding, and month-old spike elicits virtually no neutralization in mice despite inducing high ELISA-binding titers. Cryo-EM structures reveal the folded fraction of spike to decrease with aging; however, its structure remains largely similar, although with varying mobility of the receptor-binding domain. Thus, the SARS-CoV-2 spike is susceptible to unfolding, which affects immunogenicity, highlighting the need to monitor its integrity.

Structure-based design of a quadrivalent fusion glycoprotein vaccine for human parainfluenza virus types 1-4.

Parainfluenza virus types 1-4 (PIV1-4) are highly infectious human pathogens, of which PIV3 is most commonly responsible for severe respiratory illness in newborns, elderly, and immunocompromised individuals. To obtain a vaccine effective against all four PIV types, we engineered mutations in each of the four PIV fusion (F) glycoproteins to stabilize their metastable prefusion states, as such stabilization had previously enabled the elicitation of high-titer neutralizing antibodies against the related respiratory syncytial virus. A cryoelectron microscopy structure of an engineered PIV3 F prefusion-stabilized trimer, bound to the prefusion-specific antibody PIA174, revealed atomic-level details for how introduced mutations improved stability as well as how a single PIA174 antibody recognized the trimeric apex of prefusion PIV3 F. Nine combinations of six newly identified disulfides and two cavity-filling mutations stabilized the prefusion PIV3 F immunogens and induced 200- to 500-fold higher neutralizing titers in mice than were elicited by PIV3 F in the postfusion conformation. For PIV1, PIV2, and PIV4, we also obtained stabilized prefusion Fs, for which prefusion versus postfusion titers were 2- to 20-fold higher. Elicited murine responses were PIV type-specific, with little cross-neutralization of other PIVs. In nonhuman primates (NHPs), quadrivalent immunization with prefusion-stabilized Fs from PIV1-4 consistently induced potent neutralizing responses against all four PIVs. For PIV3, the average elicited NHP titer from the quadrivalent immunization was more than fivefold higher than any titer observed in a cohort of over 100 human adults, highlighting the ability of a prefusion-stabilized immunogen to elicit especially potent neutralization.

Immunogenicity and structures of a rationally designed prefusion MERS-CoV spike antigen.

Middle East respiratory syndrome coronavirus (MERS-CoV) is a lineage C betacoronavirus that since its emergence in 2012 has caused outbreaks in human populations with case-fatality rates of ∼36%. As in other coronaviruses, the spike (S) glycoprotein of MERS-CoV mediates receptor recognition and membrane fusion and is the primary target of the humoral immune response during infection. Here we use structure-based design to develop a generalizable strategy for retaining coronavirus S proteins in the antigenically optimal prefusion conformation and demonstrate that our engineered immunogen is able to elicit high neutralizing antibody titers against MERS-CoV. We also determined high-resolution structures of the trimeric MERS-CoV S ectodomain in complex with G4, a stem-directed neutralizing antibody. The structures reveal that G4 recognizes a glycosylated loop that is variable among coronaviruses and they define four conformational states of the trimer wherein each receptor-binding domain is either tightly packed at the membrane-distal apex or rotated into a receptor-accessible conformation. Our studies suggest a potential mechanism for fusion initiation through sequential receptor-binding events and provide a foundation for the structure-based design of coronavirus vaccines.

Protective Efficacy of Nucleic Acid Vaccines Against Transmission of Zika Virus During Pregnancy in Mice.

Zika virus (ZIKV) caused an epidemic of congenital malformations in 2015-2016. Although many vaccine candidates have been generated, few have demonstrated efficacy against congenital ZIKV infection. Here, we evaluated lipid-encapsulated messenger RNA (mRNA) vaccines and a DNA plasmid vaccine encoding the prM-E genes of ZIKV in mouse models of congenital infection. Although the DNA vaccine provided comparable efficacy against vertical transmission of ZIKV, the mRNA vaccines, including one that minimizes antibody-dependent enhancement of infection, elicited higher levels of antigen-specific long-lived plasma cells and memory B cells. Despite the induction of robust neutralizing antibody titers by all vaccines, breakthrough seeding of the placenta and fetal head was observed in a small subset of type I interferon signaling-deficient immunocompromised dams. In comparison, evaluation of one of the mRNA vaccines in a human STAT2-knockin transgenic immunocompetent mouse showed complete protection against congenital ZIKV transmission. These data will inform ongoing human ZIKV vaccine development efforts and enhance our understanding of the correlates of vaccine-induced protection.

Importance of Neutralizing Monoclonal Antibodies Targeting Multiple Antigenic Sites on the Middle East Respiratory Syndrome Coronavirus Spike Glycoprotein To Avoid Neutralization Escape.

Middle East respiratory syndrome coronavirus (MERS-CoV) causes a highly lethal pulmonary infection with ∼35% mortality. The potential for a future pandemic originating from animal reservoirs or health care-associated events is a major public health concern. There are no vaccines or therapeutic agents currently available for MERS-CoV. Using a probe-based single B cell cloning strategy, we have identified and characterized multiple neutralizing monoclonal antibodies (MAbs) specifically binding to the receptor-binding domain (RBD) or S1 (non-RBD) regions from a convalescent MERS-CoV-infected patient and from immunized rhesus macaques. RBD-specific MAbs tended to have greater neutralizing potency than non-RBD S1-specific MAbs. Six RBD-specific and five S1-specific MAbs could be sorted into four RBD and three non-RBD distinct binding patterns, based on competition assays, mapping neutralization escape variants, and structural analysis. We determined cocrystal structures for two MAbs targeting the RBD from different angles and show they can bind the RBD only in the "out" position. We then showed that selected RBD-specific, non-RBD S1-specific, and S2-specific MAbs given prophylactically prevented MERS-CoV replication in lungs and protected mice from lethal challenge. Importantly, combining RBD- and non-RBD MAbs delayed the emergence of escape mutations in a cell-based virus escape assay. These studies identify MAbs targeting different antigenic sites on S that will be useful for defining mechanisms of MERS-CoV neutralization and for developing more effective interventions to prevent or treat MERS-CoV infections.IMPORTANCE MERS-CoV causes a highly lethal respiratory infection for which no vaccines or antiviral therapeutic options are currently available. Based on continuing exposure from established reservoirs in dromedary camels and bats, transmission of MERS-CoV into humans and future outbreaks are expected. Using structurally defined probes for the MERS-CoV spike glycoprotein (S), the target for neutralizing antibodies, single B cells were sorted from a convalescent human and immunized nonhuman primates (NHPs). MAbs produced from paired immunoglobulin gene sequences were mapped to multiple epitopes within and outside the receptor-binding domain (RBD) and protected against lethal MERS infection in a murine model following passive immunization. Importantly, combining MAbs targeting distinct epitopes prevented viral neutralization escape from RBD-directed MAbs. These data suggest that antibody responses to multiple domains on CoV spike protein may improve immunity and will guide future vaccine and therapeutic development efforts.

Self-assembling influenza nanoparticle vaccines elicit broadly neutralizing H1N1 antibodies.

Influenza viruses pose a significant threat to the public and are a burden on global health systems. Each year, influenza vaccines must be rapidly produced to match circulating viruses, a process constrained by dated technology and vulnerable to unexpected strains emerging from humans and animal reservoirs. Here we use knowledge of protein structure to design self-assembling nanoparticles that elicit broader and more potent immunity than traditional influenza vaccines. The viral haemagglutinin was genetically fused to ferritin, a protein that naturally forms nanoparticles composed of 24 identical polypeptides. Haemagglutinin was inserted at the interface of adjacent subunits so that it spontaneously assembled and generated eight trimeric viral spikes on its surface. Immunization with this influenza nanoparticle vaccine elicited haemagglutination inhibition antibody titres more than tenfold higher than those from the licensed inactivated vaccine. Furthermore, it elicited neutralizing antibodies to two highly conserved vulnerable haemagglutinin structures that are targets of universal vaccines: the stem and the receptor binding site on the head. Antibodies elicited by a 1999 haemagglutinin-nanoparticle vaccine neutralized H1N1 viruses from 1934 to 2007 and protected ferrets from an unmatched 2007 H1N1 virus challenge. This structure-based, self-assembling synthetic nanoparticle vaccine improves the potency and breadth of influenza virus immunity, and it provides a foundation for building broader vaccine protection against emerging influenza viruses and other pathogens.

Broadly Neutralizing Human Immunodeficiency Virus Type 1 Antibody Gene Transfer Protects Nonhuman Primates from Mucosal Simian-Human Immunodeficiency Virus Infection.

Broadly neutralizing antibodies (bnAbs) can prevent lentiviral infection in nonhuman primates and may slow the spread of human immunodeficiency virus type 1 (HIV-1). Although protection by passive transfer of human bnAbs has been demonstrated in monkeys, durable expression is essential for its broader use in humans. Gene-based expression of bnAbs provides a potential solution to this problem, although immune responses to the viral vector or to the antibody may limit its durability and efficacy. Here, we delivered an adeno-associated viral vector encoding a simianized form of a CD4bs bnAb, VRC07, and evaluated its immunogenicity and protective efficacy. The expressed antibody circulated in macaques for 16 weeks at levels up to 66 g/ml, although immune suppression with cyclosporine (CsA) was needed to sustain expression. Gene-delivered simian VRC07 protected against simian-human immunodeficiency virus (SHIV) infection in monkeys 5.5 weeks after treatment. Gene transfer of an anti-HIV antibody can therefore protect against infection by viruses that cause AIDS in primates when the host immune responses are controlled.

Comparative analysis of the magnitude, quality, phenotype, and protective capacity of simian immunodeficiency virus gag-specific CD8+ T cells following human-, simian-, and chimpanzee-derived recombinant adenoviral vector immunization.

Recombinant adenoviral vectors (rAds) are the most potent recombinant vaccines for eliciting CD8(+) T cell-mediated immunity in humans; however, prior exposure from natural adenoviral infection can decrease such responses. In this study we show low seroreactivity in humans against simian- (sAd11, sAd16) or chimpanzee-derived (chAd3, chAd63) compared with human-derived (rAd5, rAd28, rAd35) vectors across multiple geographic regions. We then compared the magnitude, quality, phenotype, and protective capacity of CD8(+) T cell responses in mice vaccinated with rAds encoding SIV Gag. Using a dose range (1 × 10(7)-10(9) particle units), we defined a hierarchy among rAd vectors based on the magnitude and protective capacity of CD8(+) T cell responses, from most to least, as: rAd5 and chAd3, rAd28 and sAd11, chAd63, sAd16, and rAd35. Selection of rAd vector or dose could modulate the proportion and/or frequency of IFN-γ(+)TNF-α(+)IL-2(+) and KLRG1(+)CD127(-)CD8(+) T cells, but strikingly ∼30-80% of memory CD8(+) T cells coexpressed CD127 and KLRG1. To further optimize CD8(+) T cell responses, we assessed rAds as part of prime-boost regimens. Mice primed with rAds and boosted with NYVAC generated Gag-specific responses that approached ∼60% of total CD8(+) T cells at peak. Alternatively, priming with DNA or rAd28 and boosting with rAd5 or chAd3 induced robust and equivalent CD8(+) T cell responses compared with prime or boost alone. Collectively, these data provide the immunologic basis for using specific rAd vectors alone or as part of prime-boost regimens to induce CD8(+) T cells for rapid effector function or robust long-term memory, respectively.

Cholesterol reduction by immunization with a PCSK9 mimic.

Proprotein convertase subtilisin/kexin type 9 (PCSK9) is a plasma protein that controls cholesterol homeostasis. Here, we design a human PCSK9 mimic, named HIT01, with no consecutive 9-residue stretch in common with any human protein as a potential heart attack vaccine. Murine immunizations with HIT01 reduce low-density lipoprotein (LDL) and cholesterol levels by 40% and 30%, respectively. Immunization of cynomolgus macaques with HIT01-K21Q-R218E, a cleavage-resistant variant, elicits high-titer PCSK9-directed antibody responses and significantly reduces serum levels of cholesterol 2 weeks after each immunization. However, HIT01-K21Q-R218E immunizations also increase serum PCSK9 levels by up to 5-fold, likely due to PCSK9-binding antibodies altering the half-life of PCSK9. While vaccination with a PCSK9 mimic can induce antibodies that block interactions of PCSK9 with the LDL receptor, PCSK9-binding antibodies appear to alter homeostatic levels of PCSK9, thereby confounding its vaccine impact. Our results nevertheless suggest a mechanism for increasing the half-life of soluble regulatory factors by vaccination.

Gene-based vaccination with a mismatched envelope protects against simian immunodeficiency virus infection in nonhuman primates.

The RV144 trial demonstrated that an experimental AIDS vaccine can prevent human immunodeficiency virus type 1 (HIV-1) infection in humans. Because of its limited efficacy, further understanding of the mechanisms of preventive AIDS vaccines remains a priority, and nonhuman primate (NHP) models of lentiviral infection provide an opportunity to define immunogens, vectors, and correlates of immunity. In this study, we show that prime-boost vaccination with a mismatched SIV envelope (Env) gene, derived from simian immunodeficiency virus SIVmac239, prevents infection by SIVsmE660 intrarectally. Analysis of different gene-based prime-boost immunization regimens revealed that recombinant adenovirus type 5 (rAd5) prime followed by replication-defective lymphocytic choriomeningitis virus (rLCMV) boost elicited robust CD4 and CD8 T-cell and humoral immune responses. This vaccine protected against infection after repetitive mucosal challenge with efficacies of 82% per exposure and 62% cumulatively. No effect was seen on viremia in infected vaccinated monkeys compared to controls. Protection correlated with the presence of neutralizing antibodies to the challenge viruses tested in peripheral blood mononuclear cells. These data indicate that a vaccine expressing a mismatched Env gene alone can prevent SIV infection in NHPs and identifies an immune correlate that may guide immunogen selection and immune monitoring for clinical efficacy trials.

A multivalent polyomavirus vaccine elicits durable neutralizing antibody responses in macaques.

In 2019, there were about 100,000 kidney transplants globally, with more than a quarter of them performed in the United States. Unfortunately, some engrafted organs are lost to polyomavirus-associated nephropathy (PyVAN) caused by BK and JC viruses (BKPyV and JCPyV). Both viruses cause brain disease and possibly bladder cancer in immunosuppressed individuals. Transplant patients are routinely monitored for BKPyV viremia, which is an accepted hallmark of nascent nephropathy. If viremia is detected, a reduction in immunosuppressive therapy is standard care, but the intervention comes with increased risk of immune rejection of the engrafted organ. Recent reports have suggested that transplant recipients with high levels of polyomavirus-neutralizing antibodies are protected against PyVAN. Virus-like particle (VLP) vaccines, similar to approved human papillomavirus vaccines, have an excellent safety record and are known to induce high levels of neutralizing antibodies and long-lasting protection from infection. In this study, we demonstrate that VLPs representing BKPyV genotypes I, II, and IV, as well as JCPyV genotype 2 produced in insect cells elicit robust antibody titers. In rhesus macaques, all monkeys developed neutralizing antibody titers above a previously proposed protective threshold of 10,000. A second inoculation, administered 19 weeks after priming, boosted titers to a plateau of ≥ 25,000 that was maintained for almost two years. No vaccine-related adverse events were observed in any macaques. A multivalent BK/JC VLP immunogen did not show inferiority compared to the single-genotype VLP immunogens. Considering these encouraging results, we believe a clinical trial administering the multivalent VLP vaccine in patients waiting to receive a kidney transplant is warranted to evaluate its ability to reduce or eliminate PyVAN.

EV-D68 virus-like particle vaccines elicit cross-clade neutralizing antibodies that inhibit infection and block dissemination.

Enterovirus D68 (EV-D68) causes severe respiratory illness in children and can result in a debilitating paralytic disease known as acute flaccid myelitis. No treatment or vaccine for EV-D68 infection is available. Here, we demonstrate that virus-like particle (VLP) vaccines elicit a protective neutralizing antibody against homologous and heterologous EV-D68 subclades. VLP based on a B1 subclade 2014 outbreak strain elicited comparable B1 EV-D68 neutralizing activity as an inactivated viral particle vaccine in mice. Both immunogens elicited weaker cross-neutralization against heterologous viruses. A B3 VLP vaccine elicited more robust neutralization of B3 subclade viruses with improved cross-neutralization. A balanced CD4+ T helper response was achieved using a carbomer-based adjuvant, Adjuplex. Nonhuman primates immunized with this B3 VLP Adjuplex formulation generated robust neutralizing antibodies against homologous and heterologous subclade viruses. Our results suggest that both vaccine strain and adjuvant selection are critical elements for improving the breadth of protective immunity against EV-D68.