MYOCARDITIS IN YOUNG MEN AFTER mRNA-BASED COVID19 VACCINES: COULD PEGYLATION BE THE CULPRIT?
A testable hypothesis with practical consequences.
Acute myocarditis (inflammation of the heart muscle) has emerged as a possible but rare complication of COVID-19 vaccines that predominantly affects young male adults and typically has a mild clinical course requiring only supportive therapy. Importantly, myocarditis appears to be specifically associated with the mRNA-type vaccines (both from Moderna and Pfizer/BioNtech) but not with other types of vaccines, such as adenovirus-based vaccines (e.g. from AstraZeneca or J&J) although all current COVID19 vaccines use the same antigen: the full-length Spike-protein(or in some cases key domains of it) of the SARS-Cov2 virus that causes COVID19.
With almost 300 million doses of mRNA vaccines administered in the U.S. (as of June 11, 2021), 1226 cases of myocarditis have been documented (ACIP June 23, meeting). Case reports have been published here, here, here; and three recent reports were discussed in JAMA Cardiology’s June 29 editorial). The crudely estimated risk for myocarditis would thus be roughly 1 in (all) 250,000 mRNA-vaccine recipients — an exceedingly small number. This frequency is too small for this complication to have been detected in the Phase 3 clinical trials of these vaccines with a cohort size of 30,000–43,000 participants, and also too small to question the favorable evaluation of risk/benefit ratio of mRNA vaccines by the CDC with regard to myocarditis risk. If we take only young men, the frequency increases, in men of age 16–24, to 1 in 3000–6000, as an Israeli study showed. In any case, such anecdotal, small and retrospective case reports do not permit a comparison with background population incidence to support an etiological association of myocarditis with the vaccine. (But just for illustration, a ballpark number: a 2013 Finnish study found ~ 3000 cases of myocarditis in 1.7 million hospital admissions, also predominantly in male adolescents).
So let’s say the above numbers are “suggestive” of a causal link.
However, timing of onset of symptoms (within a week after mRNA vaccine), clinical similarity of the reported cases, the concentration of cases among young male adults, the absence of other known etiology (viral infections, autoimmunity) and notably, the selectivity for mRNA-based vaccines warrant the assumption of a causal association –if only to motivate the academic discussion on a possible biological mechanism.
Circumstances and demographics of patients exhibit an interesting pattern that should stimulate rigorous reasoning, allowing for logically and factually consistent and plausible speculation given all existing knowledge about the biological mechanisms. Such an academic exercise for the sake of getting to the bottom of a problem, independent of any explicitly declared practical utility but in the name of our scientific curiosity and inquisitiveness has great pedagogical value for biomedical researchers. But biological thinking in medicine has been sidelined by editors and educators in an era of data-driven descriptive research. So let’s think a bit.
We start with the pattern of the cases of mRNA-vaccine-associated myocarditis: they were observed predominantly in young (median age < 26), male (66%) adults and symptom onset was typically within 1–5 days after receiving the vaccination. While 75% of the myocarditis occurred after the second dose of the mRNA vaccine, a substantial number occurred after the first dose (see ACIP report).
A pathogenesis hypothesis on mRNA-vaccine-induced myocarditis must be consistent with these three characteristics. They are the “explanandum” that constrain the hypothesis, which in turn must be constrained by established biomedical knowledge.
To make a long story short, here is the hypothesis, which is explained step-by-step further below: The myocarditis following administration of mRNA vaccines may be an acute auto-immune reaction involving COMPLEMENT activation, which in turn is triggered by preexisting circulating anti-PEG antibodies (PEG=polyethylene-glycol). These antibodies bind to the PEG-conjugated (“PEGylated”) lipids on the lipid-nanoparticles (LNP) used in both mRNA vaccines (Fig 1). LNPs are necessary in mRNA vaccines to protect the mRNA and carry it into the antigen-presenting cells (APC) upon injection without causing inflammation. COMPLEMENT activation and anti-PEG antibody engagement further stimulate production by B-cells of these very antibodies that bind to PEG.
FIRST, SOME BACKGROUND FOR THE NON-TECHNICAL READER: The human body has hundreds of millions of distinct antibodies that bind antigens because of a snug fit (due to the complementary shapes of antigen and antibody). ‘Distinct’ means: each antibody recognizes and binds a different antigen molecule. And each one is made by a distinct B-cell — the antibody-producing cells of the immune system. Antigens are molecule on microbes and viruses, or even artificial products, recognized by the immune system as foreign in part because antibodies exist that bind to them.
Some antibodies are present in higher amounts in circulation because of previous exposure to an antigen (from an earlier infection) and can neutralize new infections by microbes and viruses that carry these same antigen molecules. These antibodies can be rapidly produced by armies of so called B-cells that have been “primed” (immunized) by the past first antigen contact. Once primed, these B cells are ready to mass-produce these antibodies quickly when reactivated by a recurring infection that has the same antigen. (This mechanism also accounts for the fact that allergic reactions typically require “sensitization” and are much stronger on the second exposure with the antigen).
On the other hand the body has a vast library of other antibodies present in minute amounts. These low-level antibodies have never encountered an antigen. The B-cells that make them are rare and not primed. When a new antigen enters the body, one of these rare “virgin antibodies” may detect them (by binding them). This engagement will cause exactly those B-cell that makes them to become “primed” and mass produce these antibodies. This priming is thus antigen-specific and requires amplification of the tiny signal sent by the rare virgin but now engaged antibodies to convey to the immune system that a new antigen has been detected. Such signal amplification that primes the B cells upon first antigen encounter is achieved by a clever system in the body consisting of a powerful protein cascade called the COMPLEMENT.
The complement system is a biochemical mechanism through which a cascade of protein activation can signal to the immune system that an antibody has encountered an antigen; thereby it launches the immune response which results in the “priming” (=activation) of the B cells or awakening the already primed B cells (officially termed memory B cells). But the complement system not only acts as a signal amplifier. It also acts as an executor in first line defense: the complement protein cascade ends with the activation of a potent cell-destructing protein complex.
In fact, this cell-destroying activity runs continuously in the background in the entire body! It would destroy any cell that is not protected by a security system. The latter is a protective protein machinery on every cell in the body and ensures that the activated complement system is continuously being inactivated. This strategy of an active balance between continuous background activation and a continuously operating control system is akin to the system of armed fighter jets waiting with running engines around the clock in peace times that are safely secured by a command chain system to prevent undue launch. Such stand-by but secured system is widely used in nature because it ensures a fast launch of a defense system in case of an invading microbe. The price we pay for this “preloaded system” of the complement is that its accidental undue activation can cause diseases, such as auto-immune disease. Injured tissue not caused by infectious agents, such as a heart attack, can mimic foreign invasion antigens, and may thus, also activate the complement system which contributes to the expansion of the tissue damage. END OF BACKGROUND.]
With this background information we can now proceed with explaining the hypothesis for the pathogenesis of post-mRNA-vaccine myocarditis. The following premises (known biomedical facts or observations) jointly support the above hypothesis that preexisting antibodies against the antigen “PEGylated molecule” plays a role and explain the particular pattern of this complication.
1. First, both mRNA vaccines, but not adenovirus-based vaccines, use PEG-conjugated (“PEGylated”) lipids in their LNP. PEGylation is a standard biocompatible chemical modification that stabilizes therapeutic macromolecules. Thus, the mRNA-vaccine particles expose PEG on their surface. PEGylated molecules have recently been suspected to be the antigen that could trigger allergic reactions, especially if the patient has pre-existing antibodies against PEG.
2. Second, there is indeed a high prevalence for presence of circulating anti-PEG antibodies (IgG and IgM), ranging from 25–72% of the healthy population, depending on the study. Importantly, a systematic survey in 2016 revealed that among adults in Taiwan, prevalence was highest in 20-year old (60%) and decreases with age to 20% for 80-year old, who also have less than half the amount of anti-PEG-IgG than the 20 year old (Fig 2). There was no gender differences. Thus, while epidemiology of preexistence of anti-PEG levels coincides with the predominance of myocarditis in the young adolescents, it does not explain why myocarditis primarily affects men.
Interestingly, the aforementioned ACIP analysis of demographics revealed a dramatic age demarcation. Not only are predominantly young male adolescents affected, but the incidence of post-mRNA vaccine myocarditis (reported rates) was 20x higher per second dose vaccine administered to males aged younger than 29 year than to those older than 30 years. This sharp drop coincides with the exponential decrease of prevalence of circulating anti-PEG antibodies during the first three decades of life. Such a sudden drop is also consistent with the added effect from the decrease with age in the plasma concentrations of anti-PEG antibodies in individuals that carry the antibodies, as reported in the same study.
3. Third, once the anti-PEG antibodies bind the PEGylated LNP injected by vaccination, they can activate the COMPLEMENT system. This is particularly likely because with their size (optimized by Moderna scientists to maximize immune activation) of approximately ~100nm, the antibody-coated lipid nanoballs are sufficiently large to act like a typical opsonized pathogen. (Opsonization is what renders a pathogen able to be recognized and eaten up by immune cells). Therefore, they are likely to be effective in activating the complement system. The ensuing protein activation cascade not only can destroy the antigen-bearing target but also stimulates the B cell clone that produces the anti-PEG antibody — thus starting the mechanism that physiologically connects the initial innate immune defense to the activation of the adaptive immune system that primes the respective (anti-PEG antibody producing) B-cells for future encounter. Interestingly, small pox vaccination with attenuated vaccinia virus, a giant virus which at 200 nm diameter is in the size range of the LNPs, is also a strong activator of the complement system, and has been reported to cause myocarditis in a cohort of military personnel. Thus, perhaps the LNP of the mRNA vaccine mimic the big vaccinia virus?
4. Fourth, complement activation has been shown to be critical in triggering immune myocarditis in several mouse models (e.g. here and here). More generally, heart muscle tissue is, for poorly understood reasons, particularly sensitive to complement activation and vice versa, injured heart tissues itself can activate complement. The role of complement in myocardial pathology is most prosaically manifest in coxsackie virus-induced auto-immune myocarditis and in complement activation during myocardial infarct. Such mutualism can start a pathophysiological vicious cycle capable of overcoming homeostatic safeguard systems.
5. Finally, in contrary to anti-PEG levels where no gender difference has been observed, in the Caucasian population woman have significantly lower level of the terminal (effector) components of the complement cascade. This previous finding may explain the observation that men are two times more frequently affected by post-mRNA vaccine myocarditis than women. Moreover, studies in mice have shown that testosterone increased, and estrogen decreased, complement activity. Thus, while the predominance of males in post-mRNA vaccine myocarditis is not be explained by bias in the prevalence for circulating in anti-PEG, it could be result of the higher proclivity of complement activation in men.
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In summary, the above five known facts or prior observations connect the dots between the LNP in the mRNA vaccine (with its PEGylation), with preexisting anti-PEG antibodies in the vaccine recipients, with the complement system and with myocarditis as complication of vaccination with mRNA/LNP. The patterns of timing and epidemiology of myocarditis, and possibility (though less frequent) of occurrence after the first vaccine dose, is consistent with a complement activation-mediated process triggered by preexisting anti-PEG antibodies that bind to the PEGylated LNP of the mRNA vaccine.
This hypothesis can be tested by measuring the levels of anti-PEG antibodies and complement proteins (notably C3, C5) as well as (technically more challenging) complement activation in the serum of the affected patients at time of myocarditis diagnosis. If the postulated pathogenetic mechanism finds evidential support, for instance, if post-vaccine myocarditis patients have exceedingly high anti-PEG antibodies, then anti-PEG titer could serve as a biomarker to identify individuals who would have to avoid mRNA vaccines that contain PEGylated LNP and instead, should seek alternatives, such as the adenovirus-based vaccines. This is particularly important as we now begin to vaccinate teenagers. A screen for anti-PEG antibodies can be as simple and inexpensive as any point-of care COVID19 cartridge test.
