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T cells

TWiV 890: Looking into a booster crystal ball

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This episode of TWiV is focused on COVID-19 vaccines and antibodies: who should get boosters, whether a variant matched mRNA vaccine is superior to a historical vaccine, and how the interval between vaccination and infection influences the quality of the antibody response.

Hosts: Vincent Racaniello, Dickson Despommier, Rich Condit, and Brianne Barker

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Show notes at microbe.tv/twiv

An Intranasal SARS-CoV-2 Vaccine Candidate

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by Gertrud U. Rey

There are numerous advantages to administering a vaccine by the intranasal route: it is simple, painless, and does not require special training. The last time I discussed SARS-CoV-2 vaccine candidates for potential intranasal delivery, the idea was still fairly new. However, multiple new studies are now showing that delivery of a SARS-CoV-2 vaccine into the nose induces immune responses that are superior to those observed after delivery into the muscle.

The authors of one such recent study compared the efficacy of a single dose of two different adenovirus-based SARS-CoV-2 vaccine candidates after intramuscular injection to that after administration by the intranasal route. Both vaccines encoded three viral antigens: 1) the full-length S1 domain of the SARS-CoV-2 spike protein, which contains the region that binds to the host cell receptor ACE2; 2) the full-length nucleocapsid protein, which is essential for binding and packaging the viral genome; and 3) a truncated version of the RNA-dependent RNA polymerase (RdRp), the enzyme responsible for copying the viral genome during infection. The three antigens were inserted into either one of two adenovirus vectors: a virus of human origin to produce “Tri:HuAd” or a virus of chimpanzee origin to produce “Tri:ChAd.” Most currently approved SARS-CoV-2 vaccines only encode the full-length spike protein, but not nucleocapsid or RdRp. These two additional genes were likely included in the present study because nucleocapsid is the most abundant viral protein in infected cells, and because RdRp is well conserved among coronaviruses, thus potentially leading to broader immune responses.

To assess the antibody responses induced by either vaccine candidate, the authors immunized mice with a single dose of Tri:HuAd or Tri:ChAd into the nose or into the muscle and collected serum and lung wash samples 2, 4 and 8 weeks later. The collected samples were then tested for the presence of IgG antibodies, which provide the majority of antibody-based immunity. Analysis of sera collected at 4 weeks revealed that mice immunized intranasally with either vaccine had significantly higher levels of spike protein-specific IgG than did mice that were immunized intramuscularly, and these high levels were sustained through 8 weeks post-vaccination. However, only lung wash samples from mice immunized with either vaccine intranasally contained detectable levels of spike protein-specific IgG at the 4 week time point, suggesting that only intranasal immunization led to IgG production in the airways. Interestingly, the airways of Tri:ChAd recipients harbored almost twice as much spike-specific IgG as those of Tri:HuAd recipients, a difference that was not observed in the serum, suggesting that the vector of chimpanzee origin induced better local immune responses when administered via the nose. Additionally, only the lung wash samples from intranasal Tri:ChAd recipients contained any observable anti-spike IgA antibodies, which are prevalent in mucosal surfaces and represent a first line of defense against invasion by inhaled and ingested pathogens. These results suggested that both vaccines, but Tri:ChAd in particular, induced better immune responses when they were delivered intranasally.

Lung wash samples were also analyzed for vaccine antigen-specific T cells by detecting characteristic cell surface proteins on these T cells, and for specific cytokines produced by the T cells in response to their exposure to peptides encoding each of the three vaccine-encoded antigens: S1, nucleocapsid, or RdRp. This analysis revealed that when the vaccine was injected into the muscle, neither Tri:HuAd nor Tri:ChAd induced detectable airway antigen-specific cytotoxic T cells, which are capable of killing virus-infected cells. In contrast, both vaccines induced significant numbers of cytotoxic T cells in the airways when they were delivered intranasally. Although the detected T cells were specific for all three antigens, S1-specific cells were more abundant, and they were also multifunctional in that they produced more than one cytokine characteristic of cytotoxicity. However, in the context of intranasal vaccination, the Tri:ChAd vaccine induced higher total numbers of cytotoxic T cells than the Tri:HuAd vaccine, suggesting that a single dose of Tri:ChAd delivered intranasally is highly effective at inducing vaccine antigen-specific cytotoxic T cells in the respiratory tract.   

Interactions between lung macrophage cells and SARS-CoV-2 can increase the expression of the host-derived peptide MHCII on macrophage surfaces, allowing these cells to become a critical component of “trained innate immunity.” This altered innate immunity results from a functional modification of innate immune cells that leads to an antibody-independent temporary immune memory that is characterized by an accelerated response to a second challenge with the same pathogen. Neither Tri:HuAd nor Tri:ChAd induced detectable MHCII expression on macrophages in the lungs of mice when delivered intramuscularly. In contrast, both vaccines induced increased levels of MHCII-expressing lung macrophages 8 weeks after immunization when delivered intranasally, with a higher number of these cells in the lungs of Tri:ChAd recipients, suggesting that intranasal immunization with Tri:ChAd induces strong trained innate immunity.

The authors next tested the ability of the vaccines to protect mice from SARS-CoV-2 infection by intentionally infecting immunized mice with a lethal dose of SARS-CoV-2 at 4 weeks after immunization, and monitoring them for clinical signs of illness in terms of weight loss. Unvaccinated control animals and 80% of animals immunized intramuscularly with either vaccine were not protected from infection or disease and died by 5 days post-infection. In contrast, animals immunized with Tri:HuAd showed a temporary and slight weight loss, but rebounded over the course of two weeks, while animals immunized with Tri:ChAd did not lose weight at all for the duration of the two-week time course. This result correlated with reduced viral load and reduced lung pathology at 14 days post-infection in mice immunized intranasally compared to unvaccinated controls and intramuscular vaccine recipients, with Tri:ChAd recipients exhibiting the most reduced viral burden and lung pathology.   

Having determined that intranasal administration of Tri:ChAd induced the best immune responses, all subsequent experiments were carried out using this vaccination strategy. To assess the individual contributions of B cells and T cells in the vaccine-induced immune response, the authors immunized two groups of mice: one group that was genetically deficient in B cells and a second group of genetically wild type mice that were depleted of T cells by receiving injections of antibodies against T cell-characteristic cell surface molecules. Because T cells are needed for B cell stimulation and production of antibodies, the antibody injections were done on day 25 after immunization to ensure that vaccine-induced B cell functions and antibody production remained intact in the T cell-depleted mice. At 28 days after vaccination, both groups of animals were infected with SARS-CoV-2 and monitored for weight loss. Unvaccinated mice that were either B cell-deficient or T cell-depleted and control unvaccinated mice with normal B and T cell functions were subject to drastic weight loss. In stark contrast, neither B cell-deficient nor T cell-depleted mice that had been intranasally immunized with Tri:ChAd showed any weight loss throughout the 2-week experimental time course, despite the fact that both groups of animals had significantly elevated viral burden in the lung. This result suggested that Tri:ChAd was immunogenic enough to compensate for the lack of B or T cells and protect the mice from clinical disease after SARS-CoV-2 infection.

The authors next wanted to analyze the role of trained innate immunity in the absence of B and T cell immunity, so they immunized two groups of mice. The first group consisted of B cell-deficient animals that were depleted of T cells shortly after vaccination. Because T cells induce vaccine-mediated trained innate immunity early after vaccination, this strategy produced mice that lacked B cells, T cells, and trained innate immunity. The second group consisted of B cell-deficient animals that were depleted of T cells starting at day 25 post-vaccination to allow for induction of trained innate immunity and thus produce mice that lacked B cells and T cells, but possessed intact trained innate immunity. At 28 days after vaccination, both groups of animals were infected with SARS-CoV-2 and monitored for weight loss and lung pathology. Unvaccinated control mice with functional B cells, T cells, and trained innate immunity and unvaccinated mice lacking B cells, T cells, and trained innate immunity showed severe weight loss and severe lung pathology. In contrast, vaccinated mice lacking B cells and T cells but possessing trained innate immunity appeared healthy with no weight loss and no pathology in the lungs, suggesting that the immune protection observed in animals lacking B cells, T cells, or both, was likely mediated by trained innate immunity.

The study demonstrated similar results after infection of mice with alpha and beta variants of SARS-CoV-2; however, it would be interesting to see if these vaccines have similar efficacy against the later variants, including delta and omicron. Although these results are promising, it is important to remember that mice can be poor predictors of human disease outcomes. Nevertheless, I look forward to seeing the results of the clinical trial that is currently underway to compare the relative efficacies of Tri:HuAd and Tri:ChAd following intranasal delivery in humans.

[For a more detailed discussion of this paper, please check out TWiV 867.]

TWiV 867: I love the smell of vaccines in the morning

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TWiV reviews an experimental, intranasally administered SARS-CoV-2 vaccine that utilizes adenovirus vectors to deliver three viral proteins and induces immunity to ancestral and variant isolates.

Hosts: Vincent Racaniello, Alan Dove, Rich Condit, and Kathy Spindler

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Show notes at microbe.tv/twiv

Heterologous Vaccine Regimens Might be Better

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by Gertrud U. Rey

Have you ever wondered if you can “mix and match” SARS-CoV-2 vaccines? For example, would it be ok to boost a first dose of the vaccine produced by AstraZeneca with a dose of the vaccine produced by Pfizer/BioNTech? The latest science shows that such a vaccine regimen actually induces a stronger immune response than a regimen consisting of two doses of the same vaccine.

The occasional incidence of thrombosis in people under the age of 60 after receiving an adenovirus-vectored vaccine like the ones made by AstraZeneca and Johnson & Johnson has prompted several European governments to recommend the use of these vaccines only in people over 60. Because many people under 60 had already been vaccinated with a first dose of the AstraZeneca vaccine and still needed a second dose, they had to decide whether they should continue their regimen with another dose of the same vaccine (i.e., a homologous regimen), or receive an mRNA vaccine instead (i.e., a heterologous regimen).

In an effort to evaluate the efficacy of a heterologous SARS-CoV-2 vaccine regimen, the authors of this Brief Communication engaged the participants of an ongoing clinical trial, which aims to monitor the immune responses to SARS-CoV-2 in health care professionals and individuals with potential exposure to SARS-CoV-2. All study participants had already received a first dose of the AstraZeneca vaccine (referred to as “ChAd” in the study) and were given the option of receiving a second dose of the same vaccine or a dose of Pfizer/BioNTech’s mRNA vaccine (referred to as “BNT”). Although both of these vaccines encode the gene for the full-length SARS-CoV-2 spike protein, it is housed slightly differently. In the ChAd vaccine, the spike protein is encoded in an adenovirus vector of chimpanzee origin, while in the BNT vaccine it is surrounded by a lipid nanoparticle.

Out of the 87 individuals who participated in the study, 32 chose a second dose of ChAd and 55 chose to be vaccinated with BNT instead. Participants who chose a homologous ChAd/ChAd regimen received their second dose of ChAd on day 73 after the initial dose and donated a blood sample for analysis 16 days later. Participants who chose a heterologous ChAd/BNT regimen received their dose of BNT 74 days after their initial ChAd dose and donated a blood sample 17 days later. All results obtained from the analyses of the blood samples from these two groups were compared to results obtained from a group of 46 health care professionals who had been vaccinated with two doses of BNT (i.e., a homologous BNT/BNT regimen).

Briefly, the findings were as follows. Relative to the antibody levels induced by the first dose, homologous boosting with ChAd led to a 2.9-fold increase in IgG antibodies against the SARS-CoV-2 spike protein. IgG antibodies are mostly present in the blood and provide the majority of antibody-based immunity against invading pathogens. In contrast, heterologous boosting with BNT led to an 11.5-fold increase in anti-spike IgG, within the range observed in BNT/BNT-vaccinated individuals. A similar pattern was observed with anti-spike IgA antibodies, which are predominantly found in mucus membranes and their fluids. Boosting with BNT induced significantly higher increases in anti-spike IgA than did boosting with ChAd, suggesting that a heterologous boosting regimen induces better antibody responses. Interestingly, although the booster immunization induced an increase in neutralizing (i.e., virus-inactivating) antibodies in both vaccination groups, only heterologous ChAd/BNT vaccination induced antibodies that neutralized all three tested variants (Alpha, Beta, and Gamma), similar to what was observed in BNT/BNT vaccine recipients.

The authors also analyzed the effect of the two different boosting regimens on spike-specific memory B cells, which circulate quiescently in the blood stream and can quickly produce spike-specific antibodies upon a subsequent exposure to SARS-CoV-2. All vaccinees from both the ChAd/ChAd and ChAd/BNT groups produced increased levels of spike-specific memory B cells after receiving their booster shot, with no significant differences observed between the two groups. These results emphasize the importance of booster vaccination for full protection from SARS-CoV-2 infection.

ChAd/BNT recipients also had significantly higher levels of spike-specific CD4+– and CD8+ T cells compared to ChAd/ChAd recipients. CD4+ T cells are a central element of the adaptive immune response, because they activate both antibody-secreting B cells and CD8+ T cells that kill infected target cells. Compared to the ChAd/ChAd regimen, ChAd/BNT vaccination also induced significantly increased levels of T cells that produce spike-specific interferon gamma – a cytokine that inhibits viral replication and activates macrophages, which engulf and digest pathogens. Overall, these results suggest that a heterologous ChAd/BNT regimen induces significantly stronger immune responses than a homologous ChAd/ChAd regimen.

The study did have several limitations. First, it was not a randomized, placebo-controlled trial where each participant was randomly assigned to an experimental group or a control group. Randomizing trial participants eliminates unwanted effects that have nothing to do with the variables being analyzed, so that the only expected differences between the experimental and control groups are the outcome variable studied (efficacy in this case). Subjects in a control group receive a placebo – a substance that has no therapeutic effect, so that one can be sure that the effects observed in the experimental group are real and result from the experimental drug. Second, the authors were unable to test the antibodies for their ability to neutralize the Delta variant, which has recently become the predominantly circulating variant in many parts of the world. Third, the study mostly included young and healthy people, making it difficult to extrapolate the data to other specific patient groups outside of this category. Fourth, the data only show the results of assays performed in vitro, which may not necessarily manifest a clinical significance. Extended studies aimed at determining the practical importance of the observed immune responses are needed to validate the relevance of these responses. 

Despite its limitations, the study provides information that could have some valuable practical implications. Most of the currently approved SARS-CoV-2 vaccine regimens involve two doses of the same vaccine. However, access to two doses of the same vaccine may be limited or absent under some circumstances, thus necessitating the use of a different type of vaccine to boost the first dose.

[For a more in-depth discussion of this study, I recommend TWiV 782.]

TWiV 684: Persistence of SARS-CoV-2 immune memory

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Daniel Griffin provides a clinical report on COVID-19, and Shane Crotty explains a study of antibodies, B cells and T cells in patients which suggests that immunological memory to SARS-CoV-2 might be long-lived.

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T Cell Responses to Coronavirus Infection are Complicated

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by Gertrud U. Rey

Throughout the current pandemic, there has been a lot of talk about T cells and their role in protecting against SARS-CoV-2 infection and disease. Some data suggest that 20-50% of people with no prior exposure to SARS-CoV-2 have T cells that recognize SARS-CoV-2 peptides, and that these T cells may be a result of recent infections with one or more of the seasonal human coronaviruses. However, it is unclear whether these “cross-reactive” T cells actually protect from SARS-CoV-2 infection and disease.  

T cells are an important part of the adaptive immune response, which initiates during a first exposure to a pathogen and protects from re-infection and disease upon a second exposure to the same pathogen. During that first exposure, T helper cells sense the presence of one or more proteins (i.e., antigens) on the surface of the invading pathogen and release a variety of signals that ultimately stimulate B cells to secrete antibodies to those antigens, white blood cells to destroy ingested microbes, and cytotoxic T cells to directly kill infected target cells (see schematic). Before T cells encounter their first antigen, they are considered to be “naïve.” Upon their first contact with an antigen, they begin to mature and differentiate into either cytotoxic T cells or memory T cells. As we age and encounter more and more pathogens, the ratio of our memory T cells to our naïve T cells increases – a phenomenon sometimes referred to as “immunological age.” 

Based on evidence from several labs, some have suggested that pre-existing cross-reactive memory T cells in people with no prior exposure to SARS-CoV-2 may have a protective effect. However, recent findings indicate that this may not be the case.

Several research groups from the U.S. and Australia analyzed blood samples from individuals with no prior SARS-CoV-2 exposure with the intent of better defining the range of T helper cells that can recognize antigenic portions known as epitopes in the SARS-CoV-2 genome. To ensure that the blood donors had never been infected with SARS-CoV-2, the researchers used stored samples that had been collected between 2015 and 2018. The authors found that the blood samples contained T cells that can recognize SARS-CoV-2 sequences that have at least 67% similarity to seasonal coronavirus sequences. However, the authors also found that people who had experienced a previous infection with SARS-CoV-2 had stronger and more specific (i.e., higher avidity) memory T cell responses to SARS-CoV-2 peptides than people with no prior exposure, and more than half of these responses were directed to epitopes in the spike protein. This suggests that SARS-CoV-2 memory T helper cells preferentially target viral proteins that are made in abundance during infection. 

The authors conclude that an infection with a seasonal coronavirus may induce a range of memory T cells that have substantial cross-reactivity to SARS-CoV-2. However, they are careful to note that the clinical relevance of these data remains unclear and that there is no evidence that these memory T cells have any functional role in protecting from infection or disease.  

Based on these findings, several groups of investigators in Germany wanted to determine whether the observed pre-existing T cell memory response in people with no prior SARS-CoV-2 exposure is protective against COVID-19. They first tested T cell activity in the blood of donors with and without prior exposure to SARS-CoV-2 by exposing their blood to highly immunogenic SARS-CoV-2 peptides. Blood from donors with a prior exposure contained memory T cells that recognized SARS-CoV-2 peptides very well, especially peptides derived from the spike, membrane, and nucleocapsid proteins. Blood from donors with no prior exposure also contained low levels of SARS-CoV-2-reactive memory T cells, but this reaction was more scattered and directed against multiple viral proteins. 

To further characterize the pre-existing cross-reactive T cells in blood samples from people with no prior SARS-CoV-2 exposure, the authors compared the numbers of memory T cells and naïve T cells in the blood samples by analyzing them for the presence of protein markers that are characteristic for each type of cell. A substantial portion of SARS-CoV-2-reactive T cells from donors with no prior exposure were naïve T cells, whereas from COVID-19 patients most were mature memory T cells. Because memory T cells are more easily activated following an infection than naïve T cells, the authors speculate that deficient, low avidity memory T cells in people with no prior SARS-CoV-2 exposure may compete with naïve T cells and prevent their activation and maturation into highly specific memory T cells upon infection with SARS-CoV-2. This could potentially lead to an inferior immune response in people with no prior SARS-CoV-2 exposure. 

Some have suggested that young patients and children may be particularly well protected from SARS-CoV-2 infection and/or disease because they are frequently infected with seasonal coronaviruses and thus presumably have high levels of pre-existing memory T cells. However, the authors found that compared to young people, older people with no prior SARS-CoV-2 exposure actually had higher numbers of SARS-CoV-2 cross-reactive memory T cells, but these T cells had a decreased avidity to SARS-CoV-2 epitopes compared to memory T cells from people with prior SARS-CoV-2 exposure.   

A further comparison of T cell responses in patients with mild or severe COVID-19 revealed that although the latter had high numbers of SARS-CoV-2 specific T cells, these T cells had reduced target specificity and avidity compared to T cells from patients with moderate disease. The authors conclude that this unfocused response may result from recruitment of a broad range of pre-existing memory T cells in people with increased immunological age and may contribute to development of severe COVID-19 in the elderly. 

The initial discovery of SARS-CoV-2-specific memory T cells in individuals with no prior exposure to SARS-CoV-2 had inspired the hypothesis that these T cells could possibly protect these individuals from disease and might partially explain why children are less susceptible to COVID-19. However, in light of these new findings it is likely that pre-existing SARS-CoV-2-specific memory T cells may contribute to the wide spectrum of disease severity among the general population and may actually be partially responsible for severe COVID-19 in the elderly. 

[For an in-depth discussion of these two papers, I recommend TWiV 657 and Christian Drosten’s “Das Coronavirus Update,” episodes 58 and 60.]