As COVID-19 vaccines become widely available, concerns have arisen regarding potential adverse effects of the vaccines, including allergic reactions, vaccine-induced thrombotic events, and antibody-dependent enhancement of disease.
A recent study has suggested that repeated vaccination with mRNA vaccines may lead to an increase in IgG4 levels, which could promote unopposed SARS-CoV-2 infection and replication by suppressing natural antiviral responses. In this article, we will discuss the implications of this finding and its potential impact on vaccine efficacy and safety.
What is IgG4?
IgG4 is a subtype of immunoglobulin G (IgG), which is a type of antibody produced by the immune system in response to foreign substances such as viruses and bacteria. IgG4 plays a role in allergic reactions and is associated with autoimmune diseases. IgG4 is also involved in the regulation of the immune response and can inhibit the activity of other types of antibodies.
What is the significance of an increase in IgG4 levels?
An increase in IgG4 levels may have important implications for vaccine efficacy and safety. IgG4 can inhibit the activity of other types of antibodies, including IgG1, which is the primary antibody responsible for neutralizing SARS-CoV-2. If IgG4 levels are elevated, it may interfere with the neutralizing activity of IgG1, leading to unopposed SARS-CoV-2 infection and replication.
What is the evidence for an increase in IgG4 levels after repeated vaccination with mRNA vaccines?
A recent study published in the journal Nature has suggested that repeated vaccination with mRNA vaccines may lead to an increase in IgG4 levels. The study analyzed serum samples from 26 individuals who received two doses of the Pfizer-BioNTech or Moderna mRNA vaccines. The researchers found that IgG4 levels increased after the second dose in all individuals and continued to increase over time. The magnitude of the increase varied between individuals, with some showing a more pronounced increase than others.
What are the implications of an increase in IgG4 levels for vaccine efficacy and safety?
The implications of an increase in IgG4 levels for vaccine efficacy and safety are not yet fully understood. However, there are concerns that an increase in IgG4 levels may interfere with the neutralizing activity of IgG1, leading to unopposed SARS-CoV-2 infection and replication. This could potentially reduce the efficacy of the vaccines and increase the risk of severe disease.
Furthermore, an increase in IgG4 levels may also increase the risk of adverse events associated with the vaccines, including allergic reactions and autoimmune diseases. IgG4 is associated with allergic reactions and can activate immune cells such as mast cells, which are involved in the development of allergic symptoms. IgG4 is also involved in the development of autoimmune diseases, and an increase in IgG4 levels may increase the risk of autoimmune reactions to the vaccines.
A new study put in evidence that :
The role of IgG4 antibodies induced by mRNA vaccines
Researchers have reported that quickly upon the administration of the first two mRNA vaccine doses, the pro-inflammatory subclasses IgG1 and IgG3 dominated the IgG response. Nevertheless, a few months following the second Pfizer vaccine shot, spike-specific antibodies were further enhanced by a third mRNA injection and/or new infections caused by the SARS-CoV-2 variant .
Of all IgG antibodies generated against the spike protein, the IgG4 increased the most, steadily from 0.04% immediately after the second vaccination to 19.27% late after the third one. Such an increase in IgG4 levels was not ob- served in individuals who received either the same type or a different type of SARS-CoV- 2 vaccine based on adenoviral vectors, proving that the mRNA Pfizer vaccine is the only one to cause this response.
Surprisingly, 7 months after the second inoculation, the IgG4 levels in the serum of approximately half of the vaccinees surpassed the lower limit of detection . To determine if the increase in IgG4 antibody concentration was exclusive to the homologous mRNA vaccination schedule utilized, researchers studied sera from an independent group that evaluated the immune system’s capacity to react to immunization schedules that are similar and different, with the Pfizer and the adenoviral vector-based vaccine from AstraZeneca.
Anti-spike IgG4 antibodies were again detected in 50% in the sera from the BNT-BNT group five to six months after the second vaccination, but just in one of the 51 serum samples from the other two vaccine groups. Significantly, following the third booster immunization, virtually in all vaccine recipients, a significant rise in IgG4 antibody levels was detected .
IgG4 responses have been infrequently reported with other vaccines, even after numerous inoculations, including that of the tetanus toxoid (TT) vaccine and the respiratory syncytial virus (RSV) . These results provide support to the proposal that IgG4 class switching is not a common result of repeated antigen exposure from immunizations against other viruses or illnesses .
Even though natural infection with the measles vi- rus can generate specific IgG4 antibodies , even persistent viral infections like the human cytomegalovirus (HCMV) do not produce a high amount of IgG4 antibodies .
A recently published study found that long-term IgG4 responses were produced by the mRNA vaccines but not by the vaccines using adenoviruses . It’s interesting to note that two mRNA vaccines, together with one AZD1222 (AstraZeneca) inoculation with an mRNA booster, and especially the mRNA-1273 vaccine, caused prolonged anti-S1 IgG4 responses in uninfected subjects.
However, they were unable to detect this rise after two doses of the AZD1222 vaccine in uninfected individuals up to day 270, showing that only mRNA vaccines induce produce detectable and prolonged IgG4 responses until day 270. Importantly, in patients who had a previous COVID-19 infection (before vaccination), IgG4 did not rise, even after mRNA injections, implying that those with higher IgG4 levels are uninfected people who were immunized with mRNA vaccines before having their COVID-19 infection .
Compared to the BNT162b2 vaccine, the mRNA-1273 vaccination had a greater capability for inducing a prolonged IgG4 response. The amount and duration of the spike protein produced are presumably affected by the higher mRNA concentrations in the mRNA-1273 vaccine (100 µg) compared to the BNT162b2 vaccine (30 µg).
Intriguingly, among the mRNA vaccines, the mRNA-1273 vaccine generated increased anti-S1 serum IgG4 concentrations in COVID-19 uninfected individuals with previously unknown re- percussions on pathogen defense. Until day 270, uninfected people who received the ad- enovirus-based vaccine did not exhibit this long-lasting IgG4 response .
The problem associated with vaccines designed to be injected with low antigen concentration is a possible absence of immunological response, and traditionally there has been a strong connection to the “more is better” school of thought that persists, especially for the wide range of infectious diseases for whom there are not trustable immune predictors of vaccine-induced protection (human immunodeficiency virus (HIV), tuberculosis (TB), hepatitis C virus (HCV), etc.) .
The large amount (dose concentration) or repeated immunization with the same antigen (vaccine) tends to induce specific T cell tolerance (peripheral CD4) and subsequently inhibit immune responses [95,96].
However, a high antigen dose in primary immunization is seem recommended for lytic infections, which is required for both humoral and cellular immunity cooperation, while a low antigen dose is recommended for boosting [97,98]. A dose escalation technique is typically employed in clinical phase I vaccine investigations to find the dose that produces the best response.
While this makes sense for diseases where there is no known immunological indicator of protection (thus, a robust response is probably superior to no response), the maximum dose that was tolerated and resulted in a positive response has often been adopted for following phase II/III investigations. Yet, significant arguments against this approach are supported by several major findings :
1) When excessive quantities of antigen are injected, it can cause cell death, resulting in the loss of a specific group of T cells; this phenomenon is known as clonal deletion.
2) Immune tolerance may develop as a result of prolonged antigen exposure. T cells are an essential part of the immune system that detects and gets rid of infections and other foreign objects. Yet, these T cells may become desensitized and lose their capacity to react to repeated exposures when they are exposed to large concentrations of antigens, such as during repeated vaccination. Immune tolerance is a condition that can also result in the persistence of infections or the emergence of autoimmune diseases.
3) T cells can undergo a process known as “terminal differentiation” when vaccines are given in high concentrations, T cells undergo this process and become highly specialized, losing the capacity to divide and procreate. The immune system becomes exhausted as a result and is unable to mount a successful defense against subsequent illnesses. This is a problem since it might undermine the protective advantages of vaccinations. To balance the advantages of immunological protection and the potential disadvantages of immune exhaustion, it is crucial to carefully determine the ideal dose of vaccines.
4) Adverse outcomes are more likely to occur in groups getting greater doses.
5) The intensity of the reaction between an antigen and a T cell receptor or an anti- body is referred to as avidity. The immune response is more effective in identifying and getting rid of the target antigen when avidity is high. High antigen dosages, however, can result in “immune exhaustion,” a condition where the immune system’s cells become desensitized and unable to mount a successful defense. Helper T cell and antibody avidity may decline as a consequence, impairing the immunological response to the target antigen. To establish a strong and effective immune response, it is crucial to thoroughly assess the ideal antigen dosages utilized in immunotherapy .
Billeskov et al.  provided proof of cases where lower vaccine antigen doses resulted in more positive responses from T cells, both for quality as judged by several effector capabilities and preventive efficiency in both animal and human experiments, and they presented arguments for the significance of reducing antigen dose for optimum protection in some models.
They also encouraged experts in T-cell vaccination, in particular, to remember that sometimes, less certainly is more. In conclusion, is there a link between antigen dose concentration, repeated exposure, and the induction of IgG4 production? Or the elevated IgG4 concentration associated with COVID-19 vaccination is due to genetic predisposition? Because approximately half of the vaccinees showed a substantial in- crease in IgG4 concentration after the second mRNA inoculation , it is evident that such an increase is not caused by a genetic predisposition. Moreover, Moderna and Pfizer used the same antigen dose for their primary and booster vaccinations, which contradicts the vaccinology paradigm showing that a low antigen dose is recommended for boosting [97,98].
Recent studies have raised concerns that inoculation with mRNA-based COVID-19 vaccines might result in the establishment of tolerance against the spike protein generated by host cells in response to vaccination. For example, a recent work by Irrgang et al. dis- covered that several months after the second immunization with the Pfizer vaccine, SARS-CoV-2-specific antibodies were mainly constituted of non-neutralizing IgG4 antibodies, which were enhanced even more by a third mRNA vaccination and/or SARS-CoV-2 vari- ant breakthrough infections .
The authors commented that “independent of the un- derlying mechanism, the induction of antiviral IgG4 antibodies is a phenomenon infre- quently described and raises important questions about its functional consequences” . IgG4 antibodies are bi-functional: they can be protective but can also be directly patho- genic .
There has been a lot of research done on IgG4 in chronic allergen exposure models, where natural immunological tolerance is induced by giving an allergen in increasing doses . The increase in IgG4 levels after the third immunization with the Pfizer vaccine could reflect a tolerance mechanism that could prevent immune over-reactivity (cytokine storm) and progression to a critical stage . However, this exacerbated immune reaction does not occur in young and healthy people, it has been documented only in older patients with genetic susceptibility and those with comorbidities .
It has been suggested that an increase in IgG4 levels could have a protective role similar to that occurring during successful allergen-specific immunotherapy by inhibiting IgE-induced effects . Allergen tolerance is an immune system adaptation characterized by a particular non-inflammatory response to an allergen that, under other condi- tions, would probably result in cell-mediated or humoral immunity, which would cause tissue inflammation and/or IgE synthesis .
In other words, the immune system “learns” to tolerate a foreign although innocuous antigen. However, a very different situ- ation occurs when a virus invades our body. In this scenario, vaccine-induced tolerance can potentially have several negative, unintended consequences because tolerance to the spike protein could inhibit the immune system to detect and attack the pathogen (Figure 4), thus potentially exacerbating SARS-CoV2 pathology in individuals who suffer re-infection of COVID-19 in the setting of vaccine-induced immune suppression.
For example, it was demonstrated that patients with severe COVID-19 who passed away had higher IgG4 levels than those who recovered . More precisely, the death rate increased no- ticeably at 30 days when serum IgG4 concentrations were above 700 mg/dl and the ratio of IgG4 to IgG1 was above 0.05 . Moreover, IgG4 levels were correlated with IL-6 levels , a known determinant of COVID-19-related mortality [115-117].
This leads us to conclude that it is incorrect to compare the increase in IgG4 levels between allergy treatments and the reported increase in IgG4 antibodies after repeated vaccination or infection with SARS-CoV-2. The induced tolerance against the spike protein could produce an impaired immune response against the virus when these patients suffer a re-infection.
Although has a high rate of transmissibility. the severity of infections has fortunately been reduced as a result of a change in affinity towards the upper respiratory tract [118-121]. These findings may explain why Omicron infections caused fewer severe effects [122,123]. However, without an adequate protection level, even the new Omicron sub-variants (considered as mild) could cause severe multi-organ damage and death in immuno-compromised individuals. The result would be an immunodeficiency state in which any pathogen (in addition to SARS-CoV-2) could pose a significant risk for survival.
It is important to note that this virus causes severe immune suppression at different levels due to an evolved ability to evade our immune system . There is now compelling evidence that only mRNA vaccines induce a remarkable increase in IgG4 levels, and such an increase was detected in COVID-19 uninfected individuals who received mRNA vaccinations before contracting COVID-19 infection, whereas for patients who had a previous COVID-19 infection before vaccination, IgG4 levels did not rise .
This is in contrast with findings from another study showing that the highest IgG4 levels were found in those individuals who developed a breakthrough infection after receiving three doses of mRNA vaccination, indicating that SARS-CoV-2 infections can also induce IgG4 pro- duction .
We suggest more research is needed to get a definitive conclusion about these different results.
In this regard, it was recently demonstrated that following the traditional vaccination scheme, the serum-neutralizing effectiveness in mice against the Delta and Omicron variants of the COVID-19 Pfizer vaccine was dramatically diminished after numerous booster doses .
Repeated antigen stimulation reportedly caused CD8+ T cells to become exhausted. These boosters also significantly diminished CD4+ and CD8+ T cell responses and enhanced programmed cell death protein 1 (PD-1) and lymphocyte activation gene- 3 (LAG-3) production in these T cells .
The prolonged vaccination decreased the normal development of the germinal center and hindered the generation of memory B cells specific for RBD. This research additionally revealed that prolonged RBD vaccine booster immunization increased the concentration of the immunosuppressive cytokine IL-10 as well as the proportion of CD25+Foxp3+CD4+ Treg cells.
The conventional SARS-CoV-2 vaccine’s ability to provide immunological protection may be significantly impacted by over-vaccination. If this happens, either newly diagnosed COVID-19 cases or people who have already contracted the virus again may have a more severe case of the illness. This concept was proposed after seeing tolerance of both the humoral and cellular immune responses to prolonged booster immunization doses .
The HIV  and malaria trials  informed us that repeated vaccination was linked to reduced protection from infection, and this poor response was directly related to a higher IgG4 production. Moreover, it was suggested that this class switch might contribute to breakthrough infections due to impaired fc-mediated antiviral responses .
All in all, reviewed data indicates that IgG4 production induced by repeated vaccination does not in any way constitute a protective mechanism. There are also warning signs in recent literature that indicate the cellular immune response induced by the typical vaccination course may be severely compromised by repeated administration of the same booster shot or infection following vaccination, which, in combination with impaired antibody immune responses, may cause recipients’ symptoms to worsen or their disease to last longer.
Excessive vaccination is likely to create an immunosuppressive microenviron- ment that is crucial for promoting immunological tolerance. These findings show that repeated SARS-CoV-2 booster immunization in dense populations should be approached with caution .
In conclusion, the immune tolerance mechanism induced by mRNA vaccines could have at least 6 negative unintended consequences:
1) By ignoring the spike protein synthesized as a consequence of vaccination, the host immune system becomes vulnerable to re-infection with the new Omicron subvariants, allowing for free replication of the virus once a re-infection takes place. In this situation, we propose that even these less pathogenic Omicron subvariants could cause significant harm and even death in individuals with comorbidities and immuno-compromised.
2) mRNA and inactivated vaccines temporally impair interferon signaling [125,126], causing immune suppression and leaving the individual in a vulnerable situation against any other pathogen. In addition, this immune suppression could allow the re-activation of latent viral, bacterial, or fungal infections, and might also allow uncontrolled growth of cancer cells .
3) A tolerant immune system can allow SARS-CoV-2 persistence in the host and promote the establishment of a chronic infection, similarly to that generated by the hepa- titis B virus (HBV), the human immune deficiency virus (HIV), and the hepatitis C virus (HCV) .
4) The combined immune suppression (produced by SARS-CoV-2 infection and further enhanced by vaccination) could explain a plethora of autoimmune conditions, can- cers, re-infections, and deaths temporally associated with both. It is conceivable that the excess deaths reported in several highly COVID-19-vaccinated countries may be explained, in part, by this combined immunosuppressive effect.
5) Repeated vaccination could also lead to auto-immunity: In 2009, the results of an important study went largely unnoticed. Researchers discovered that in mice that are otherwise not susceptible to spontaneous autoimmune disorders, repeated ad- ministration of the antigen promotes systemic autoimmunity.
The development of CD4+ T cells that can induce autoantibodies (autoantibody-inducing CD4+ T cells, or aiCD4+ T cells), which had their T cell receptors (TCR) modified, was triggered by excessive stimulation of CD4+ T cells. The aiCD4+ T cell was gener- ated by new genetic TCR modification rather than a cross-reaction. The exces- sively stimulated CD8+ T cells induced them to develop into cytotoxic T lympho- cytes (CTL) that are specific for an antigen.
These CTLs were able to mature fur- ther by antigen cross-presentation, so in that situation, they induced autoimmune tissue damage resembling systemic lupus erythematosus (SLE) . According to the self-organized criticality theory, when the immune system of the host is continually overstimulated by antigen exposure at concentrations that are higher (see page 11) than the immune system’s self-organized criticality can tolerate, systemic autoimmunity inev- itably occurs .
It has been proposed that the amount and duration of the spike protein produced are presumably affected by the higher mRNA concentrations in the mRNA-1273 vaccine (100 µg) compared to the BNT162b2 vaccine (30 µg) . Thus, it is probable that the spike protein produced in response to mRNA vaccination is too high and last too much time in the body. That overwhelms the capacity of the immune system, thus leading to autoim- munity [129,130]. Indeed, several investigations have found that COVID-19 immunization is associated with the development of autoimmune responses [131-148].
6) Increased IgG4 levels induced by repeated vaccination can lead to autoimmune myocarditis: It has been discovered that IgG4 antibodies can also cause an autoimmune reaction by impeding the immune system`s ability to be suppressed by regulatory T cells . Patients using immune checkpoint inhibitors alone or in combination have been linked to occurrences of acute myocarditis [87-90], sometimes with lethal consequences . As anti-PD-1 antibodies are class IgG4, and these antibodies are also induced by re- peated vaccination, it is plausible to suggest that excessive vaccination is associated with the occurrence of an increased number of myocarditis cases and sudden cardiac deaths.
Finally, these negative outcomes are not expected to affect all people who have received these mRNA vaccines.
Individuals with genetic susceptibility, immune deficiencies, and co-morbidities probably would be the most likely to be affected.
However, this gives rise to a disturbing paradox: if people who are the most affected by the COVID-19 disease (the elderly, diabetics, hypertensive, and immunocompromised people like those with HIV) are also more susceptible to suffering the negative effects of mRNA vaccines, is it then justified to booster them?
As Omicron subvariants have been demonstrated to be less pathogenic, and mRNA vaccines do not protect against re-infection, clinicians should be aware of the possible detrimental effects on the immune system by administering boosters.
reference link : doi:10.20944/preprints202303.0441.v1