Predicting COVID-19 booster immunogenicity against future SARS-CoV-2 variants and the benefits of vaccine updates (preprint)

The ongoing evolution of the SARS-CoV-2 virus has led to a move to update vaccine antigens in 2022 and 2023. These updated antigens were chosen and approved based on in vitro neutralisation titres against recent SARS-CoV-2 variants. However, unavoidable delays in viral manufacture and distribution meant that the updated booster vaccine was no longer well matched to the circulating SARS-CoV-2 variant by the time of its deployment. Understanding whether the updating of booster vaccine antigens improves immune responses to subsequent SARS-CoV-2 circulating variants is a major priority in justifying future vaccine updates. Here we analyse all available data on the immunogenicity of variant containing SARS-CoV-2 vaccines and their ability to neutralise later circulating SARS-CoV-2 variants. We find that updated booster antigens give a 1.4-fold [95%CI 1.07-1.82] greater increase in neutralising antibody levels when compared with a historical vaccine immunogen. We then use this to predict the relative protection that can be expected from an updated vaccine even when the circulating variant has evolved away from the updated vaccine immunogen. These findings help inform the roll out of future booster vaccination programs.

Mucosal and Systemic Immune Correlates of Viral Control following SARS-CoV-2 Infection Challenge in Seronegative Adults (preprint)

Human infection challenge permits characterisation of the associated immune response in unparalleled depth, enabling evaluation of early pre-symptomatic immune changes and the dynamic immune factors important for viral clearance. Here, 34 healthy young adult volunteers, seronegative to SARS-CoV-2, were inoculated with a D614G-containing pre-Alpha SARS-CoV-2 strain. Nasal and systemic soluble mediator and antibody responses, and peripheral blood T cell and B cell responses were measured by MesoScale Discovery and flow cytometry just before and up to 1 year after intra-nasal inoculation. In the 18 (53%) participants who became infected, both nasal and systemic mediator responses were dominated by interferons (IFN) but with divergent kinetics. T cell activation and proliferation in blood peaked at day 10 in CD4+ T cells and day 14 in CD8+ T cells, returning to baseline by day 28. Following infection, antigen-specific T cells were largely CD38+Ki67+ and displayed central and effector memory phenotypes. T cells contracted after viral clearance with expanded antigen-specific memory T cell populations persisting past day 28. Both mucosal and systemic antibodies became detectable around day 10 but nasal antibodies plateaued after day 14 while circulating antibodies continued to rise. Using piecewise linear regression modelling, viral load related closely to the induction of type I IFN responses, moreover, CD8+ T cell responses and early IgA responses were strongly associated with viral clearance. Detailed analysis of innate and adaptive immune responses to primary SARS-CoV-2 infection following human challenge thus revealed the relationship between immune kinetics and viral load as factors associated with resolution of infection.

Lasting first impression: Pre-existing immunity restricts mucosal antibody responses during Omicron breakthrough (preprint)

Understanding mucosal antibody responses from SARS-CoV-2 infection and/or vaccination is crucial to develop strategies for longer term immunity, especially against emerging viral variants. We profiled serial paired mucosal and plasma antibodies from: COVID-19 vaccinated only vaccinees (vaccinated, uninfected), COVID-19 recovered vaccinees (convalescent, vaccinated) and individuals with breakthrough Delta or Omicron BA.2 infections (vaccinated, infected). Saliva from COVID-19 recovered vaccinees displayed improved antibody neutralizing activity, Fc{gamma}R engagement and IgA compared to COVID-19 uninfected vaccinees. Furthermore, repeated mRNA vaccination boosted SARS-CoV-2-specific IgG2 and IgG4 responses in both mucosa biofluids (saliva and tears) and plasma. IgG, but not IgA, responses to breakthrough COVID-19 variants were dampened and narrowed by increased pre-existing vaccine-induced immunity to the ancestral strain. Salivary antibodies delayed initiation of boosting following breakthrough COVID-19 infection, especially Omicron BA.2, however, rose rapidly thereafter. Our data highlight how pre-existing immunity shapes mucosal SARS-CoV-2-specific antibody responses and has implications for long-term protection from COVID-19.

Durable reprogramming of neutralising antibody responses following breakthrough Omicron infection (preprint)

SARS-CoV-2 breakthrough infection of vaccinated individuals is increasingly common with the circulation of highly immune evasive and transmissible Omicron variants. Here, we report the dynamics and durability of recalled spike-specific humoral immunity following BA.1 or BA.2 breakthrough infection, with longitudinal sampling up to 8 months post-infection. Both BA.1 and BA.2 infection robustly boosted neutralisation activity against the infecting strain while expanding breadth against other Omicron strains. Cross-reactive memory B cells against both ancestral and Omicron spike were predominantly expanded by infection, with limited recruitment of de novo Omicron-specific B cells or antibodies. Modelling of neutralisation titres predicts that protection from symptomatic reinfection against antigenically similar strains will be remarkably durable, but is undermined by novel emerging strains with further neutralisation escape.

Rapid recall and de novo T cell responses during SARS-CoV-2 breakthrough infection (preprint)

While the protective role of neutralising antibodies against COVID-19 is well-established, questions remain about the relative importance of cellular immunity. Using 6 pMHC-multimers in a cohort with early and frequent sampling we define the phenotype and kinetics of recalled and primary T cell responses following Delta or Omicron breakthrough infection. Recall of spike-specific CD4+ T cells was rapid, with cellular proliferation and extensive activation evident as early as 1 day post-symptom onset. Similarly, spike-specific CD8+ T cells were rapidly activated but showed variable levels of expansion. Strikingly, high levels of SARS-CoV-2-specific CD8+ T cell activation at baseline and peak were strongly correlated with reduced peak SARS-CoV-2 RNA levels in nasal swabs and accelerated clearance of virus. Our study demonstrates rapid and extensive recall of memory T cell populations occurs early after breakthrough infection and suggests that CD8+ T cells contribute to the control of viral replication in breakthrough SARS-CoV-2 infections.

Neutralising antibodies predict protection from severe COVID-19 (preprint)

Background: Vaccine protection from COVID-19 has been shown to decline with time-since-vaccination and against SARS-CoV-2 variants. Protection against severe COVID-19 is higher than against symptomatic infection, and also appears relatively preserved over time and against variants. Although Protection protection from symptomatic SARS-CoV-2 infection has been shown to be strongly correlated with neutralising antibody titres, however, this relationship has been is less well described for severe COVID-19. Protection against severe COVID-19 is higher than against symptomatic infection, and also appears relatively preserved over time and against variants. Here we analyse whether neutralising antibody titre remains predictive of protection against severe COVID-19 in the face of waning neutralising antibody levels and emerging variants. Methods: We extracted data from 15 studies reporting on protection against a range of SARS-CoV-2 clinical endpoints ("any infection", "symptomatic infection" and "severe COVID-19"). We then estimated the concurrent neutralising antibody titres using existing parameters on vaccine potency, neutralising antibody decay, and loss of recognition of variants and investigated the relationship between neutralising antibody titre and vaccine effectiveness against severe COVID-19. Findings: Predicted neutralising antibody titres are strongly correlated with vaccine effectiveness against symptomatic and severe COVID-19 (Spearman rho = .94 and 0.63 respectively, p

Efficient recall of Omicron-reactive B cell memory after a third dose of SARS-CoV-2 mRNA vaccine (preprint)

Despite a clear role in protective immunity, the durability and quality of antibody and memory B cell responses induced by mRNA vaccination, particularly by a 3rd dose of vaccine, remains unclear. Here, we examined antibody and memory B cell responses in a cohort of individuals sampled longitudinally for ~9-10 months after the primary 2-dose mRNA vaccine series, as well as for ~3 months after a 3rd mRNA vaccine dose. Notably, antibody decay slowed significantly between 6- and 9-months post-primary vaccination, essentially stabilizing at the time of the 3rd dose. Antibody quality also continued to improve for at least 9 months after primary 2-dose vaccination. Spike- and RBD-specific memory B cells were stable through 9 months post-vaccination with no evidence of decline over time, and ~40-50% of RBD-specific memory B cells were capable of simultaneously recognizing the Alpha, Beta, Delta, and Omicron variants. Omicron-binding memory B cells induced by the first 2 doses of mRNA vaccine were boosted significantly by a 3rd dose and the magnitude of this boosting was similar to memory B cells specific for other variants. Pre-3rd dose memory B cell frequencies correlated with the increase in neutralizing antibody titers after the 3rd dose. In contrast, pre-3rd dose antibody titers inversely correlated with the fold-change of antibody boosting, suggesting that high levels of circulating antibodies may limit reactivation of immunological memory and constrain further antibody boosting by mRNA vaccines. These data provide a deeper understanding of how the quantity and quality of antibody and memory B cell responses change over time and number of antigen exposures. These data also provide insight into potential immune dynamics following recall responses to additional vaccine doses or post-vaccination infections.

Dynamics of immune recall following SARS-CoV-2 vaccination or breakthrough infection (preprint)

Vaccination against SARS-CoV-2 results in protection from acquisition of infection as well as improved clinical outcomes even if infection occurs, likely reflecting a combination of residual vaccine-elicited immunity and the recall of immunological memory. Here, we define the early kinetics of spike-specific humoral and T cell immunity after vaccination of seropositive individuals, and after breakthrough infection in vaccinated individuals. Intensive and early longitudinal sampling reveals the timing and magnitude of recall, with the phenotypic activation of B cells preceding an increase in neutralizing antibody titres. In breakthrough infections, the delayed kinetics of humoral immune recall provides a mechanism for the lack of early control of viral replication but likely underpins accelerated viral clearance and the protective effects of vaccination against severe COVID-19.

Tracking the kinetics and phenotype of spike epitope-specific CD4 T cell immunity in the context of SARS-CoV-2 infection and vaccination (preprint)

CD4+ T cells play a critical role in the immune response to viral infection. SARS-CoV-2 infection and vaccination elicit strong CD4+ T cell responses to the viral spike protein, including circulating T follicular helper (cTFH) cells that correlate with the development of neutralising antibodies. Here we use a novel HLA-DRB1*15:01/S751 tetramer to precisely track spike-specific CD4+ T cells following recovery from mild/moderate COVID-19, or after vaccination with spike-encoding vaccines. SARS-CoV-2 infection induces robust S751-specific responses with both CXCR5- and cTFH phenotypes that are maintained for at least 12 months in a stable, CXCR3-biased, central memory pool. Vaccination of immunologically naïve subjects similarly drives expansion of S751-specific T cells with a highly restricted TCR repertoire comprised of both public and private clonotypes. Vaccination of convalescent individuals drives recall of CD4+ T cell clones established during infection, which are shared between the CXCR5- and cTFH compartments. This recall response is evident 5 days after antigen exposure and includes a population of spike-specific cTFH that persist in the periphery after losing expression of PD-1. Overall this study demonstrates the generation of a stable pool of cTFH and memory CD4+ T cells that can be recalled upon spike antigen re-exposure, which may play an important role in long-term protection against SARS-CoV-2 infection.

mRNA Vaccination Induces Durable Immune Memory to SARS-CoV-2 with Continued Evolution to Variants of Concern (preprint)

SARS-CoV-2 mRNA vaccines have shown remarkable efficacy, especially in preventing severe illness and hospitalization. However, the emergence of several variants of concern and reports of declining antibody levels have raised uncertainty about the durability of immune memory following vaccination. In this study, we longitudinally profiled both antibody and cellular immune responses in SARS-CoV-2 naive and recovered individuals from pre-vaccine baseline to 6 months post-mRNA vaccination. Antibody and neutralizing titers decayed from peak levels but remained detectable in all subjects at 6 months post-vaccination. Functional memory B cell responses, including those specific for the receptor binding domain (RBD) of the Alpha (B.1.1.7), Beta (B.1.351), and Delta (B.1.617.2) variants, were also efficiently generated by mRNA vaccination and continued to increase in frequency between 3 and 6 months post-vaccination. Notably, most memory B cells induced by mRNA vaccines were capable of cross-binding variants of concern, and B cell receptor sequencing revealed significantly more hypermutation in these RBD variant-binding clones compared to clones that exclusively bound wild-type RBD. Moreover, the percent of variant cross-binding memory B cells was higher in vaccinees than individuals who recovered from mild COVID-19. mRNA vaccination also generated antigen-specific CD8+ T cells and durable memory CD4+ T cells in most individuals, with early CD4+ T cell responses correlating with humoral immunity at later timepoints. These findings demonstrate robust, multi-component humoral and cellular immune memory to SARS-CoV-2 and current variants of concern for at least 6 months after mRNA vaccination. Finally, we observed that boosting of pre-existing immunity with mRNA vaccination in SARS-CoV-2 recovered individuals primarily increased antibody responses in the short-term without significantly altering antibody decay rates or long-term B and T cell memory. Together, this study provides insights into the generation and evolution of vaccine-induced immunity to SARS-CoV-2, including variants of concern, and has implications for future booster strategies. GRAPHICAL ABSTRACT O_FIG O_LINKSMALLFIG WIDTH=146 HEIGHT=200 SRC="FIGDIR/small/457229v1_ufig1.gif" ALT="Figure 1"> View larger version (32K): org.highwire.dtl.DTLVardef@16c64b1org.highwire.dtl.DTLVardef@146ca3aorg.highwire.dtl.DTLVardef@86b7edorg.highwire.dtl.DTLVardef@956879_HPS_FORMAT_FIGEXP M_FIG C_FIG

SARS-CoV-2 variants: levels of neutralisation required for protective immunity (preprint)

A number of SARS-CoV-2 variants of concern (VOC) have been identified that partially escape serum neutralisation activity elicited by current vaccines. Recent studies have also shown that vaccines demonstrate reduced protection against symptomatic infection with SARS-CoV-2 variants. Here we integrate published data on in vitro neutralisation and clinical protection to understand and predict vaccine efficacy against existing SARS-CoV-2 variants. We find that neutralising activity against the ancestral SARS-CoV-2 is highly predictive of neutralisation of the VOC, with all vaccines showing a similar drop in neutralisation to the variants. Neutralisation levels remain strongly correlated with protection from infection with SARS-CoV-2 VOC (r=0.81, p=0.0005). We apply an existing model relating in vitro neutralisation to protection (parameterised on data from ancestral virus infection) and find this remains predictive of vaccine efficacy against VOC once drops in neutralisation to the VOC are taken into account. Modelling of predicted vaccine efficacy against variants over time suggests that protection against symptomatic infection may drop below 50% within the first year after vaccination for some current vaccines. Boosting of previously infected individuals with existing vaccines (which target ancestral virus) has been shown to significantly increase neutralising antibodies. Our modelling suggests that booster vaccination should enable high levels of immunity that prevent severe infection outcomes with the current SARS-CoV-2 VOC, at least in the medium term.

Tear antibodies to SARS-CoV-2: implications for transmission (preprint)

ObjectivesSARS-CoV-2 can be transmitted by aerosols and the ocular surface may be an important route of transmission. Little is known about protective antibody responses to SARS-CoV-2 in tears after infection or vaccination. We analysed SARS-CoV-2 specific IgG and IgA responses in human tears after either COVID-19 infection or vaccination. MethodsWe recruited 16 subjects with COVID-19 infection an average of 7 months previously and 15 subjects before and 2 weeks after Comirnaty (Pfizer-BioNtech) vaccination. Plasma, saliva and basal tears were collected. Pre-pandemic plasma, saliva and basal tears from 11 individuals were included as healthy controls. Antibody responses to 5 SARS-CoV-2 antigens were measured via multiplex. ResultsIgG antibodies to Spike and Nucleoprotein were detected in tears, saliva and plasma from subjects with prior SARS-CoV-2 infection in comparison to uninfected controls. While RBD-specific antibodies were detected in plasma, minimal RBD-specific antibodies were detected in tears and saliva. In contrast, high levels of IgG antibodies to Spike and RBD, but not Nucleoprotein, were induced in tears, saliva and plasma of subjects receiving 2 doses of the Comirnaty vaccine. Increased levels of IgA1 and IgA2 antibodies to SARS-CoV-2 antigens were detected in plasma following infection or vaccination, but were unchanged in tears and saliva. ConclusionBoth infection and vaccination induce SARS-CoV-2-specific IgG antibodies in tears. RBD-specific IgG antibodies in tears were induced by vaccination but were not present 7 months post-infection. This suggests neutralising antibodies may be low in the tears late following infection.

Relating in vitro neutralisation level and protection in the CVnCoV (CUREVAC) trial. (preprint)

A recent study analysed the relationship between neutralising antibody response and protection from SARS-CoV-2 infection across eight vaccines platforms. The efficacy results from a phase 2b/3 trial of a ninth vaccine candidate, CVnCoV (CUREVAC), was announced on 16 June 2021. The low efficacy of this new mRNA vaccine, which showed only 47% protection from symptomatic SARS-CoV-2 infection, was surprising, given the high efficacy of two previous mRNA-based vaccines. A number of factors have been suggested to play a role in the low efficacy in the CVnCoV study, particularly around the dose and immunogenicity of the vaccine (which uses a slightly different mRNA construct) and the potential role of infection with SARS-CoV-2 variants (which were the dominant strains observed in the CVnCoV trial).

What level of neutralising antibody protects from COVID-19? . (preprint)

Both previous infection and vaccination have been shown to provide potent protection from COVID-19. However, there are concerns that waning immunity and viral variation may lead to a loss of protection over time. Predictive models of immune protection are urgently needed to identify immune correlates of protection to assist in the future deployment of vaccines. To address this, we modelled the relationship between in vitro neutralisation levels and observed protection from SARS-CoV-2 infection using data from seven current vaccines as well as convalescent cohorts. Here we show that neutralisation level is highly predictive of immune protection. The 50% protective neutralisation level was estimated to be approximately 20% of the average convalescent level (95% CI = 14-28%). The estimated neutralisation level required for 50% protection from severe infection was significantly lower (3% of the mean convalescent level (CI = 0.7-13%, p = 0.0004). Given the relationship between in vitro neutralization titer and protection, we then used this to investigate how waning immunity and antigenic variation might affect vaccine efficacy. We found that the decay of neutralising titre in vaccinated subjects over the first 3-4 months after vaccination was at least as rapid as the decay observed in convalescent subjects. Modelling the decay of neutralisation titre over the first 250 days after immunisation predicts a significant loss in protection from SARS-CoV-2 infection will occur, although protection from severe disease should be largely retained. Neutralisation titres against some SARS-CoV-2 variants of concern are reduced compared to the vaccine strain and our model predicts the relationship between neutralisation and efficacy against viral variants. Our analyses provide an evidence-based prediction of SARS-CoV-2 immune protection that will assist in developing vaccine strategies to control the future trajectory of the pandemic.

Decay of Fc-dependent antibody functions after mild to moderate COVID-19 (preprint)

The capacity of antibodies to engage with innate and adaptive immune cells via the Fc region is important in preventing and controlling many infectious diseases, and is likely critical in SARS-CoV-2 infection. The evolution of such antibodies during convalescence from COVID-19 is largely unknown. We developed novel assays to measure Fc-dependent antibody functions against SARS-CoV-2 spike (S)-expressing cells in serial samples from a cohort of 53 subjects primarily with mild-moderate COVID-19, out to a maximum of 149 days post-infection. We found that S-specific antibodies capable of engaging dimeric FcγRIIa and FcγRIIIa decayed linearly over time. S-specific antibody-dependent cellular cytotoxicity (ADCC) and antibody-dependent phagocytosis (ADP) activity within plasma declined linearly as well, in line with the decay of S-specific IgG. Although there was significant decay in S-specific plasma ADCC and ADP activity, they remained readily detectable by all assays in 94% of our cohort at the last timepoint studied, in contrast with neutralisation activity which was only detectable in 70% of our cohort by the last timepoint. Our results suggest that Fc effector functions such as ADCC and ADP could contribute to the durability of SARS-CoV-2 immunity, particularly late in convalescence when neutralising antibodies have waned. Understanding the protective potential of antibody Fc effector functions is critical for defining the durability of immunity generated by infection or vaccination.

Evolution of immunity to SARS-CoV-2 (preprint)

The durability of infection-induced SARS-CoV-2 immunity has major implications for public health mitigation and vaccine development. Animal studies and the scarcity of confirmed re-infection suggests immune protection is likely, although the durability of this protection is debated. Lasting immunity following acute viral infection requires maintenance of both serum antibody and antigen-specific memory B and T lymphocytes and is notoriously pathogen specific, ranging from life-long for smallpox or measles4, to highly transient for common cold coronaviruses (CCC). Neutralising antibody responses are a likely correlate of protective immunity and exclusively recognise the viral spike (S) protein, predominantly targeting the receptor binding domain (RBD) within the S1 sub-domain. Multiple reports describe waning of S-specific antibodies in the first 2-3 months following infection. However, extrapolation of early linear trends in decay might be overly pessimistic, with several groups reporting that serum neutralisation is stable over time in a proportion of convalescent subjects. While SARS-CoV-2 specific B and T cell responses are readily induced by infection, the longitudinal dynamics of these key memory populations remains poorly resolved. Here we comprehensively profiled antibody, B and T cell dynamics over time in a cohort recovered from mild-moderate COVID-19. We find that binding and neutralising antibody responses, together with individual serum clonotypes, decay over the first 4 months post-infection, as expected, with a similar decline in S-specific CD4+ and circulating T follicular helper (cTFH) frequencies. In contrast, S-specific IgG+ memory B cells (MBC) consistently accumulate over time, eventually comprising a significant fraction of circulating MBC. Modelling of the concomitant immune kinetics predicts maintenance of serological neutralising activity above a titre of 1:40 in 50% of convalescent subjects to 74 days, with probable additive protection from B and T cells. Overall, our study suggests SARS-CoV-2 immunity after infection is likely t 66 o be transiently protective at a population level. SARS-CoV-2 vaccines may require greater immunogenicity and durability than natural infection to drive long-term protection.

Immunogenic profile of SARS-CoV-2 spike in individuals recovered from COVID-19 (preprint)

The rapid global spread of SARS-CoV-2 and resultant mortality and social disruption have highlighted the need to better understand coronavirus immunity to expedite vaccine development efforts. Multiple candidate vaccines, designed to elicit protective neutralising antibodies targeting the viral spike glycoprotein, are rapidly advancing to clinical trial. However, the immunogenic properties of the spike protein in humans are unresolved. To address this, we undertook an in-depth characterisation of humoral and cellular immunity against SARS-CoV-2 spike in humans following mild to moderate SARS-CoV-2 infection. We find serological antibody responses against spike are routinely elicited by infection and correlate with plasma neutralising activity and capacity to block ACE2/RBD interaction. Expanded populations of spike-specific memory B cells and circulating T follicular helper cells (cTFH) were detected within convalescent donors, while responses to the receptor binding domain (RBD) constitute a minor fraction. Using regression analysis, we find high plasma neutralisation activity was associated with increased spike-specific antibody, but notably also with the relative distribution of spike-specific cTFH subsets. Thus both qualitative and quantitative features of B and T cell immunity to spike constitute informative biomarkers of the protective potential of novel SARS-CoV-2 vaccines.