Broadly neutralizing antibodies against sarbecoviruses generated by immunization of macaques with an AS03-adjuvanted COVID-19 vaccine.

The rapid emergence of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) variants that evade immunity elicited by vaccination has placed an imperative on the development of countermeasures that provide broad protection against SARS-CoV-2 and related sarbecoviruses. Here, we identified extremely potent monoclonal antibodies (mAbs) that neutralized multiple sarbecoviruses from macaques vaccinated with AS03-adjuvanted monovalent subunit vaccines. Longitudinal analysis revealed progressive accumulation of somatic mutation in the immunoglobulin genes of antigen-specific memory B cells (MBCs) for at least 1 year after primary vaccination. Antibodies generated from these antigen-specific MBCs at 5 to 12 months after vaccination displayed greater potency and breadth relative to those identified at 1.4 months. Fifteen of the 338 (about 4.4%) antibodies isolated at 1.4 to 6 months after the primary vaccination showed potency against SARS-CoV-2 BA.1, despite the absence of serum BA.1 neutralization. 25F9 and 20A7 neutralized authentic clade 1 sarbecoviruses (SARS-CoV, WIV-1, SHC014, SARS-CoV-2 D614G, BA.1, and Pangolin-GD) and vesicular stomatitis virus-pseudotyped clade 3 sarbecoviruses (BtKY72 and PRD-0038). 20A7 and 27A12 showed potent neutralization against all SARS-CoV-2 variants and multiple Omicron sublineages, including BA.1, BA.2, BA.3, BA.4/5, BQ.1, BQ.1.1, and XBB. Crystallography studies revealed the molecular basis of broad and potent neutralization through targeting conserved sites within the RBD. Prophylactic protection of 25F9, 20A7, and 27A12 was confirmed in mice, and administration of 25F9 particularly provided complete protection against SARS-CoV-2, BA.1, SARS-CoV, and SHC014 challenge. These data underscore the extremely potent and broad activity of these mAbs against sarbecoviruses.

A ferritin-based COVID-19 nanoparticle vaccine that elicits robust, durable, broad-spectrum neutralizing antisera in non-human primates.

While the rapid development of COVID-19 vaccines has been a scientific triumph, the need remains for a globally available vaccine that provides longer-lasting immunity against present and future SARS-CoV-2 variants of concern (VOCs). Here, we describe DCFHP, a ferritin-based, protein-nanoparticle vaccine candidate that, when formulated with aluminum hydroxide as the sole adjuvant (DCFHP-alum), elicits potent and durable neutralizing antisera in non-human primates against known VOCs, including Omicron BQ.1, as well as against SARS-CoV-1. Following a booster ~one year after the initial immunization, DCFHP-alum elicits a robust anamnestic response. To enable global accessibility, we generated a cell line that can enable production of thousands of vaccine doses per liter of cell culture and show that DCFHP-alum maintains potency for at least 14 days at temperatures exceeding standard room temperature. DCFHP-alum has potential as a once-yearly (or less frequent) booster vaccine, and as a primary vaccine for pediatric use including in infants.

Robust T cell responses to Pfizer/BioNTech vaccine compared to infection and evidence of attenuated peripheral CD8+ T cell responses due to COVID-19

T cells are a critical component of the response to SARS-CoV-2, but their kinetics after infection and vaccination are insufficiently understood. Using "spheromer” peptide-MHC multimer reagents, we analyzed healthy subjects receiving two doses of the Pfizer/BioNTech BNT162b2 vaccine. Vaccination resulted in robust Spike-specific T cell responses for the dominant CD4+ (HLA-DRB1∗15:01/S191) and CD8+ (HLA-A∗02/S691) T cell epitopes. Antigen-specific CD4+ and CD8+ T cell responses were asynchronous, with the peak CD4+ T cell responses occurring one week post the second vaccination (boost), whereas CD8+ T cells peaked two weeks later. These peripheral T cell responses were elevated compared to COVID-19 patients. We also found that prior SARS-CoV-2 infection resulted in decreased CD8+ T cell activation and expansion, suggesting that prior infection can influence the T cell response to vaccination. Graphical Our understanding of T cell responses in COVID-19 and vaccination is incomplete. Gao et al. examine SARS-CoV-2-specific T cell responses to infection and vaccination, revealing disparate kinetics between CD4+ and CD8+ T cells. Furthermore, compared to vaccination alone, circulating CD8+ T cells are attenuated during infection and in subsequent vaccination.

Spheromers reveal robust T cell responses to the Pfizer/BioNTech vaccine and attenuated peripheral CD8+ T cell responses post SARS-CoV-2 infection.

T cells are a critical component of the response to SARS-CoV-2, but their kinetics after infection and vaccination are insufficiently understood. Using "spheromer" peptide-MHC multimer reagents, we analyzed healthy subjects receiving two doses of the Pfizer/BioNTech BNT162b2 vaccine. Vaccination resulted in robust spike-specific T cell responses for the dominant CD4+ (HLA-DRB1∗15:01/S191) and CD8+ (HLA-A∗02/S691) T cell epitopes. Antigen-specific CD4+ and CD8+ T cell responses were asynchronous, with the peak CD4+ T cell responses occurring 1 week post the second vaccination (boost), whereas CD8+ T cells peaked 2 weeks later. These peripheral T cell responses were elevated compared with COVID-19 patients. We also found that previous SARS-CoV-2 infection resulted in decreased CD8+ T cell activation and expansion, suggesting that previous infection can influence the T cell response to vaccination.

Progress in vaccine development for infectious diseases-a Keystone Symposia report.

The COVID-19 pandemic has taught us many things, among the most important of which is that vaccines are one of the cornerstones of public health that help make modern longevity possible. While several different vaccines have been successful at stemming the morbidity and mortality associated with various infectious diseases, many pathogens/diseases remain recalcitrant to the development of effective vaccination. Recent advances in vaccine technology, immunology, structural biology, and other fields may yet yield insight that will address these diseases; they may also help improve societies' preparedness for future pandemics. On June 1-4, 2022, experts in vaccinology from academia, industry, and government convened for the Keystone symposium "Progress in Vaccine Development for Infectious Diseases" to discuss state-of-the-art technologies, recent advancements in understanding vaccine-mediated immunity, and new aspects of antigen design to aid vaccine effectiveness.

Durability of immune responses to mRNA booster vaccination against COVID-19.

BackgroundMaintaining durable immunity following vaccination represents a major challenge, but whether mRNA booster vaccination improves durability is unknown.MethodsWe measured antibody responses in 55 healthy adults, who received a booster dose of the Pfizer-BioNTech or Moderna vaccine against SARS-CoV-2 and calculated the half-life of the antibody titers. We also measured memory B and T cell responses in a subset of 28 participants. In 13 volunteers who received a second booster vaccine, we measured serum antibody titers and memory B and T cell responses.ResultsThe booster (third immunization) dose at 6 to 10 months increased the half-life of the serum-neutralizing antibody (nAb) titers to 76 days from 56 to 66 days after the primary 2-dose vaccination. A second booster dose (fourth immunization) a year after the primary vaccination further increased the half-life to 88 days. However, despite this modestly improved durability in nAb responses against the ancestral (WA.1) strain, there was a loss of neutralization capacity against the Omicron subvariants BA.2.75.2, BQ.1.1, and XBB.1.5 (48-, 71-, and 66-fold drop in titers, respectively, relative to the WA.1 strain). Although only 45% to 65% of participants demonstrated a detectable nAb titer against the newer variants after the booster (third dose), the response declined to below the detection limit in almost all individuals by 6 months. In contrast, booster vaccination induced antigen-specific memory B and T cells that persisted for at least 6 months.ConclusionThe durability of serum antibody responses improves only marginally following booster immunizations with the Pfizer-BioNTech or Moderna mRNA vaccines.

A TLR7-nanoparticle adjuvant promotes a broad immune response against heterologous strains of influenza and SARS-CoV-2.

The ideal vaccine against viruses such as influenza and SARS-CoV-2 must provide a robust, durable and broad immune protection against multiple viral variants. However, antibody responses to current vaccines often lack robust cross-reactivity. Here we describe a polymeric Toll-like receptor 7 agonist nanoparticle (TLR7-NP) adjuvant, which enhances lymph node targeting, and leads to persistent activation of immune cells and broad immune responses. When mixed with alum-adsorbed antigens, this TLR7-NP adjuvant elicits cross-reactive antibodies for both dominant and subdominant epitopes and antigen-specific CD8+ T-cell responses in mice. This TLR7-NP-adjuvanted influenza subunit vaccine successfully protects mice against viral challenge of a different strain. This strategy also enhances the antibody response to a SARS-CoV-2 subunit vaccine against multiple viral variants that have emerged. Moreover, this TLR7-NP augments antigen-specific responses in human tonsil organoids. Overall, we describe a nanoparticle adjuvant to improve immune responses to viral antigens, with promising implications for developing broadly protective vaccines.

Ablation of Adar1 in myeloid cells imprints a global antiviral state in the lung and heightens early immunity against SARS-CoV-2.

Under normal homeostatic conditions, self-double-stranded RNA (self-dsRNA) is modified by adenosine deaminase acting on RNA 1 (ADAR1) to prevent the induction of a type I interferon-mediated inflammatory cascade. Antigen-presenting cells (APCs) sense pathogen-associated molecular patterns, such as dsRNA, to activate the immune response. The impact of ADAR1 on the function of APCs and the consequences to immunity are poorly understood. Here, we show that ADAR1 deletion in CD11c+ APCs leads to (1) a skewed myeloid cell compartment enriched in inflammatory cDC2-like cells, (2) enhanced numbers of activated tissue resident memory T cells in the lung, and (3) the imprinting of a broad antiviral transcriptional signature across both immune and non-immune cells. The resulting changes can be partially reversed by blocking IFNAR1 signaling and promote early resistance against severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection. Our study provides insight into the consequences of self-dsRNA sensing in APCs on the immune system.

Epigenetic adjuvants: durable reprogramming of the innate immune system with adjuvants.

Development of effective vaccines is a critical global health priority. Stimulating antigen-specific B and T cells to elicit long-lasting protection remains the central paradigm of vaccinology. Adjuvants are components that enhance vaccine immunogenicity by targeting specific innate immune receptors and pathways. Recent data highlight the capacity of adjuvants to induce durable epigenetic reprogramming of the innate immune system to engender heightened resistance against pathogens. This raises the prospect of developing epigenetic adjuvants that, in addition to stimulating robust T and B cell responses, convey broad protection against diverse pathogens by training the innate immune system. In this review, we discuss our emerging understanding of the various vaccines and adjuvants and their effects on durable reprogramming of the innate immune response, their putative mechanisms of action, and the promise and challenges of developing epigenetic adjuvants as a universal vaccine strategy.

Early immune markers of clinical, virological, and immunological outcomes in patients with COVID-19: a multi-omics study.

Background: The great majority of severe acute respiratory syndrome-related coronavirus 2 (SARS-CoV-2) infections are mild and uncomplicated, but some individuals with initially mild COVID-19 progressively develop more severe symptoms. Furthermore, there is substantial heterogeneity in SARS-CoV-2-specific memory immune responses following infection. There remains a critical need to identify host immune biomarkers predictive of clinical and immunological outcomes in SARS-CoV-2-infected patients. Methods: Leveraging longitudinal samples and data from a clinical trial (N=108) in SARS-CoV-2-infected outpatients, we used host proteomics and transcriptomics to characterize the trajectory of the immune response in COVID-19 patients. We characterized the association between early immune markers and subsequent disease progression, control of viral shedding, and SARS-CoV-2-specific T cell and antibody responses measured up to 7 months after enrollment. We further compared associations between early immune markers and subsequent T cell and antibody responses following natural infection with those following mRNA vaccination. We developed machine-learning models to predict patient outcomes and validated the predictive model using data from 54 individuals enrolled in an independent clinical trial. Results: We identify early immune signatures, including plasma RIG-I levels, early IFN signaling, and related cytokines (CXCL10, MCP1, MCP-2, and MCP-3) associated with subsequent disease progression, control of viral shedding, and the SARS-CoV-2-specific T cell and antibody response measured up to 7 months after enrollment. We found that several biomarkers for immunological outcomes are shared between individuals receiving BNT162b2 (Pfizer-BioNTech) vaccine and COVID-19 patients. Finally, we demonstrate that machine-learning models using 2-7 plasma protein markers measured early within the course of infection are able to accurately predict disease progression, T cell memory, and the antibody response post-infection in a second, independent dataset. Conclusions: Early immune signatures following infection can accurately predict clinical and immunological outcomes in outpatients with COVID-19 using validated machine-learning models. Funding: Support for the study was provided from National Institute of Health/National Institute of Allergy and Infectious Diseases (NIH/NIAID) (U01 AI150741-01S1 and T32-AI052073), the Stanford's Innovative Medicines Accelerator, National Institutes of Health/National Institute on Drug Abuse (NIH/NIDA) DP1DA046089, and anonymous donors to Stanford University. Peginterferon lambda provided by Eiger BioPharmaceuticals.

Phenotypes of disease severity in a cohort of hospitalized COVID-19 patients: Results from the IMPACC study.

BACKGROUND: Better understanding of the association between characteristics of patients hospitalized with coronavirus disease 2019 (COVID-19) and outcome is needed to further improve upon patient management. METHODS: Immunophenotyping Assessment in a COVID-19 Cohort (IMPACC) is a prospective, observational study of 1164 patients from 20 hospitals across the United States. Disease severity was assessed using a 7-point ordinal scale based on degree of respiratory illness. Patients were prospectively surveyed for 1 year after discharge for post-acute sequalae of COVID-19 (PASC) through quarterly surveys. Demographics, comorbidities, radiographic findings, clinical laboratory values, SARS-CoV-2 PCR and serology were captured over a 28-day period. Multivariable logistic regression was performed. FINDINGS: The median age was 59 years (interquartile range [IQR] 20); 711 (61%) were men; overall mortality was 14%, and 228 (20%) required invasive mechanical ventilation. Unsupervised clustering of ordinal score over time revealed distinct disease course trajectories. Risk factors associated with prolonged hospitalization or death by day 28 included age ≥ 65 years (odds ratio [OR], 2.01; 95% CI 1.28-3.17), Hispanic ethnicity (OR, 1.71; 95% CI 1.13-2.57), elevated baseline creatinine (OR 2.80; 95% CI 1.63- 4.80) or troponin (OR 1.89; 95% 1.03-3.47), baseline lymphopenia (OR 2.19; 95% CI 1.61-2.97), presence of infiltrate by chest imaging (OR 3.16; 95% CI 1.96-5.10), and high SARS-CoV2 viral load (OR 1.53; 95% CI 1.17-2.00). Fatal cases had the lowest ratio of SARS-CoV-2 antibody to viral load levels compared to other trajectories over time (p=0.001). 589 survivors (51%) completed at least one survey at follow-up with 305 (52%) having at least one symptom consistent with PASC, most commonly dyspnea (56% among symptomatic patients). Female sex was the only associated risk factor for PASC. INTERPRETATION: Integration of PCR cycle threshold, and antibody values with demographics, comorbidities, and laboratory/radiographic findings identified risk factors for 28-day outcome severity, though only female sex was associated with PASC. Longitudinal clinical phenotyping offers important insights, and provides a framework for immunophenotyping for acute and long COVID-19. FUNDING: NIH.

Durable protection against the SARS-CoV-2 Omicron variant is induced by an adjuvanted subunit vaccine.

Despite the remarkable efficacy of COVID-19 vaccines, waning immunity and the emergence of SARS-CoV-2 variants such as Omicron represents a global health challenge. Here, we present data from a study in nonhuman primates demonstrating durable protection against the Omicron BA.1 variant induced by a subunit SARS-CoV-2 vaccine comprising the receptor binding domain of the ancestral strain (RBD-Wu) on the I53-50 nanoparticle adjuvanted with AS03, which was recently authorized for use in individuals 18 years or older. Vaccination induced neutralizing antibody (nAb) titers that were maintained at high concentrations for at least 1 year after two doses, with a pseudovirus nAb geometric mean titer (GMT) of 1978 and a live virus nAb GMT of 1331 against the ancestral strain but not against the Omicron BA.1 variant. However, a booster dose at 6 to 12 months with RBD-Wu or RBD-ß (RBD from the Beta variant) displayed on I53-50 elicited high neutralizing titers against the ancestral and Omicron variants. In addition, we observed persistent neutralization titers against a panel of sarbecoviruses, including SARS-CoV. Furthermore, there were substantial and persistent memory T and B cell responses reactive to Beta and Omicron variants. Vaccination resulted in protection against Omicron infection in the lung and suppression of viral burden in the nares at 6 weeks after the final booster immunization. Even at 6 months after vaccination, we observed protection in the lung and rapid control of virus in the nares. These results highlight the durable and cross-protective immunity elicited by the AS03-adjuvanted RBD-I53-50 nanoparticle vaccine.

Distinct sensitivities to SARS-CoV-2 variants in vaccinated humans and mice.

The emergence of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) in 2019 has led to the development of a large number of vaccines, several of which are now approved for use in humans. Understanding vaccine-elicited antibody responses against emerging SARS-CoV-2 variants of concern (VOCs) in real time is key to inform public health policies. Serum neutralizing antibody titers are the current best correlate of protection from SARS-CoV-2 challenge in non-human primates and a key metric to understand immune evasion of VOCs. We report that vaccinated BALB/c mice do not recapitulate faithfully the breadth and potency of neutralizing antibody responses elicited by various vaccine platforms against VOCs, compared with non-human primates or humans, suggesting caution should be exercised when interpreting data obtained with this animal model.

Adjuvanting a subunit SARS-CoV-2 vaccine with clinically relevant adjuvants induces durable protection in mice.

Adjuvants enhance the magnitude and the durability of the immune response to vaccines. However, there is a paucity of comparative studies on the nature of the immune responses stimulated by leading adjuvant candidates. In this study, we compared five clinically relevant adjuvants in mice-alum, AS03 (a squalene-based adjuvant supplemented with α-tocopherol), AS37 (a TLR7 ligand emulsified in alum), CpG1018 (a TLR9 ligand emulsified in alum), O/W 1849101 (a squalene-based adjuvant)-for their capacity to stimulate immune responses when combined with a subunit vaccine under clinical development. We found that all four of the adjuvant candidates surpassed alum with respect to their capacity to induce enhanced and durable antigen-specific antibody responses. The TLR-agonist-based adjuvants CpG1018 (TLR9) and AS37 (TLR7) induced Th1-skewed CD4+ T cell responses, while alum, O/W, and AS03 induced a balanced Th1/Th2 response. Consistent with this, adjuvants induced distinct patterns of early innate responses. Finally, vaccines adjuvanted with AS03, AS37, and CpG1018/alum-induced durable neutralizing-antibody responses and significant protection against the B.1.351 variant 7 months following immunization. These results, together with our recent results from an identical study in non-human primates (NHPs), provide a comparative benchmarking of five clinically relevant vaccine adjuvants for their capacity to stimulate immunity to a subunit vaccine, demonstrating the capacity of adjuvanted SARS-CoV-2 subunit vaccines to provide durable protection against the B.1.351 variant. Furthermore, these results reveal differences between the widely-used C57BL/6 mouse strain and NHP animal models, highlighting the importance of species selection for future vaccine and adjuvant studies.

Mechanisms of innate and adaptive immunity to the Pfizer-BioNTech BNT162b2 vaccine.

Despite the success of the BNT162b2 mRNA vaccine, the immunological mechanisms that underlie its efficacy are poorly understood. Here we analyzed the innate and adaptive responses to BNT162b2 in mice, and show that immunization stimulated potent antibody and antigen-specific T cell responses, as well as strikingly enhanced innate responses after secondary immunization, which was concurrent with enhanced serum interferon (IFN)-γ levels 1 d following secondary immunization. Notably, we found that natural killer cells and CD8+ T cells in the draining lymph nodes are the major producers of this circulating IFN-γ. Analysis of knockout mice revealed that induction of antibody and T cell responses to BNT162b2 was not dependent on signaling via Toll-like receptors 2, 3, 4, 5 and 7 nor inflammasome activation, nor the necroptosis or pyroptosis cell death pathways. Rather, the CD8+ T cell response induced by BNT162b2 was dependent on type I interferon-dependent MDA5 signaling. These results provide insights into the molecular mechanisms by which the BNT162b2 vaccine stimulates immune responses.

Antibodies elicited by SARS-CoV-2 infection or mRNA vaccines have reduced neutralizing activity against Beta and Omicron pseudoviruses.

Multiple severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) variants that have mutations associated with increased transmission and antibody escape have arisen over the course of the current pandemic. Although the current vaccines have largely been effective against past variants, the number of mutations found on the Omicron (B.1.1.529) spike protein appear to diminish the protection conferred by preexisting immunity. Using vesicular stomatitis virus (VSV) pseudoparticles expressing the spike protein of several SARS-CoV-2 variants, we evaluated the magnitude and breadth of the neutralizing antibody response over time in individuals after infection and in mRNA-vaccinated individuals. We observed that boosting increases the magnitude of the antibody response to wild-type (D614), Beta, Delta, and Omicron variants; however, the Omicron variant was the most resistant to neutralization. We further observed that vaccinated healthy adults had robust and broad antibody responses, whereas responses may have been reduced in vaccinated pregnant women, underscoring the importance of learning how to maximize mRNA vaccine responses in pregnant populations. Findings from this study show substantial heterogeneity in the magnitude and breadth of responses after infection and mRNA vaccination and may support the addition of more conserved viral antigens to existing SARS-CoV-2 vaccines.

Immune imprinting, breadth of variant recognition, and germinal center response in human SARS-CoV-2 infection and vaccination.

During the SARS-CoV-2 pandemic, novel and traditional vaccine strategies have been deployed globally. We investigated whether antibodies stimulated by mRNA vaccination (BNT162b2), including third-dose boosting, differ from those generated by infection or adenoviral (ChAdOx1-S and Gam-COVID-Vac) or inactivated viral (BBIBP-CorV) vaccines. We analyzed human lymph nodes after infection or mRNA vaccination for correlates of serological differences. Antibody breadth against viral variants is lower after infection compared with all vaccines evaluated but improves over several months. Viral variant infection elicits variant-specific antibodies, but prior mRNA vaccination imprints serological responses toward Wuhan-Hu-1 rather than variant antigens. In contrast to disrupted germinal centers (GCs) in lymph nodes during infection, mRNA vaccination stimulates robust GCs containing vaccine mRNA and spike antigen up to 8 weeks postvaccination in some cases. SARS-CoV-2 antibody specificity, breadth, and maturation are affected by imprinting from exposure history and distinct histological and antigenic contexts in infection compared with vaccination.

Early non-neutralizing, afucosylated antibody responses are associated with COVID-19 severity.

A damaging inflammatory response is implicated in the pathogenesis of severe coronavirus disease 2019 (COVID-19), but mechanisms contributing to this response are unclear. In two prospective cohorts, early non-neutralizing, afucosylated immunoglobulin G (IgG) antibodies specific to severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) were associated with progression from mild to more severe COVID-19. To study the biology of afucosylated IgG immune complexes, we developed an in vivo model that revealed that human IgG-Fc-gamma receptor (FcγR) interactions could regulate inflammation in the lung. Afucosylated IgG immune complexes isolated from patients with COVID-19 induced inflammatory cytokine production and robust infiltration of the lung by immune cells. In contrast to the antibody structures that were associated with disease progression, antibodies that were elicited by messenger RNA SARS-CoV-2 vaccines were highly fucosylated and enriched in sialylation, both modifications that reduce the inflammatory potential of IgG. Vaccine-elicited IgG did not promote an inflammatory lung response. These results show that human IgG-FcγR interactions regulate inflammation in the lung and define distinct lung activities mediated by the IgG that are associated with protection against, or progression to, severe COVID-19.