We will introduce it orally

Why create vaccines that can be sprayed in the nose, eat and drink

Daria Spasskaya, N+1

For almost a year now, RNA vaccines and adenoviral vectors have been protecting us from covid. But dramatically accelerated progress in the development of vaccines may affect not only platforms, but also means of delivery. After a while, we may have vaccines in the form of pills, nasal spray, chewing gum or kefir in general. We tell you how close these methods are to implementation and how they can complement traditional injections.

The most common method of vaccination is intramuscular or intravenous injection. All current coronavirus vaccines are administered intramuscularly. This method of delivering an antigen (that is, a molecule or even a whole virus against which it is necessary to develop immunity) stimulates systemic immunity: immune cells and protein molecules that float in the blood and from there penetrate into tissues.

But if, for example, you eat an antigen instead of an injection, it will interact with the mucous membranes of the body. And there the mucosal immunity functions: its own, partially autonomous defense, which is represented by slightly different types of immune cells and antibodies.

Systemic immunity mainly uses IgG immunoglobulins floating in the blood to protect the body from the pathogen (their number is measured by tests for antibodies to coronavirus after illness or vaccination). Mucosal immunity relies more on IgA-type molecules. They have a secretory component attached to them — a protein that allows antibodies to work on the surface of the mucous membranes and protects them from splitting. Secretory molecules of IgA (more precisely, already sIgA) prevent the attachment of microorganisms and toxins to the epithelium of the mucous membranes and thereby prevent them from penetrating inside. IgG can also be found in the mucous membranes, but for this their concentration in the blood must be high enough. In addition, these molecules are much less stable and poorly preserved, for example, in the digestive tract.

The structural basis of the local immunity of the mucous membranes is the lymphoid tissue MALT (mucosal—associated lymphoid tissue). It is divided into several departments: nasopharynx, lungs, intestines, and so on, but this division is quite conditional, since all departments are connected to each other due to the migration of lymphocytes. For example, lymphocytes trained in the intestine are sent first to the nearest lymph nodes, and from there through the lymphatic vessels and circulatory system to the mucous membranes of the nasopharynx and lungs, where they continue to produce IgA against the target antigen. In the lymphoid tissue of the mucous membranes, inductive sites can be distinguished, where the antigen is presented to the immune system and effector sections, where immune cells already familiar with the antigen move through the bloodstream from the induction site.

Subtypes of mucosal-associated lymphoid tissue in the pharynx, lungs and intestinesGallorini S et al.
/ Mucosal Delivery of Biopharmaceuticals. Springer, Boston, MA, 2014

So the stimulation of immunity in the digestive tract can lead to its strengthening in the nose and vice versa — and, apparently, not only from a specific pathogen, but also non-specific protection. All this explains, for example, the hypothesis that taking probiotics helps to get colds less. He also explains how a capsule with bacteria can protect a person from cholera or coronavirus infection.

The most studied site of induction of local immunity of the mucosa are Peyer's plaques in the intestine, which were discovered in the XVII century by the Swiss anatomist Johann Peyer. If we imagine that the control center of all mucosal immunity is located in the intestine, it becomes clear where the legs of the concept of edible vaccines grow from.

Peyer's plaques in the intestine of a transgenic mouse, highlighted with green fluorescent proteinVojtech.dostal / Wikimedia Commons

Drink me up

Like other pathogens of respiratory infections, coronavirus enters the human body, as a rule, through the mucous membrane. In theory, if the human mucosal immunity is trained to recognize and destroy SARS-CoV-2 virions, it will be able to prevent further penetration of the virus into tissues — and, consequently, the development of systemic symptoms, ranging from pneumonia to blood clotting disorders, nerve damage, and so on. In addition, the binding of the virus on the mucous membranes will protect not only the vaccinated person, but also prevent him from infecting others (that is, it will lead to the formation of sterilizing immunity — at least in mice, this property of mucosal vaccines has been shown).

Such an "edible" vaccine against covid is currently being developed at the Institute of Experimental Medicine of the Russian Academy of Sciences. As a delivery system, Russian scientists use a recombinant (genetically modified) strain of enterococcus, on the surface of which fragments of the S-protein of the SARS-CoV-2 virus are expressed. The virus uses the S-protein to attach to human cells, is recognized by antibodies in the first place, and therefore has become a target for most of the vaccines available today.

The S-protein expression system on the surface of the vaccine strain by embedding a fragment of the S-gene into the ebpS gene frame.Alexander Suvorov

According to scientists, the bacteria should be taken orally — in the form of a solution, a fermented milk drink or in a capsule — and, once in the intestine, they will multiply for some time and produce an antigen, stimulating mucosal immunity.

The ability to pack the vaccine into a tablet or kefir is a huge plus of such technology. It will be much more convenient to store and use, and it is psychologically easier for many people to drink medicine (not to mention kefir) than to inject it intramuscularly.

So far, the research team has tested their recombinant strains on mice and confirmed that they form the necessary IgA class immunoglobulins. In animals that were given the vaccine orally, the antibody titer significantly exceeded the value for the control group. The next step is to test the protective effect on hamsters: first they will be injected with a vaccine in the form of a solution, and then they will be infected with a coronavirus.

Formation of IgA class antibodies in mice that were given the vaccine strain orally (per os) or injected subcutaneously (sub cut), compared with the control group. Alexander Suvorov

In conversation with N+1 Alexander Suvorov, the head of the development team, claims that they plan to conduct tests according to all the rules: first finish the pre—clinic on animals, and only then conduct full-fledged clinical trials on humans - despite the fact that there is no shortage of volunteers right now.

Breathe me in

Stimulation of mucosal immunity is possible not only through the intestine, but also through the respiratory tract. Therefore, instead of feeding people a vaccine, you can give them a nasal spray. This is how at least one market-based flu vaccine, FluMist, works.

Discussing the prospects of intranasal covid vaccines on the pages of the journal Science, scientists Francis Land and Troy Randell suggest that in the case of respiratory infections, local stimulation of the IgA pathway in the mucous membranes will have a more pronounced protective effect — since IgG antibodies, which are formed during intramuscular administration of the antigen, enter the mucous membranes only at a sufficiently high titer.

They stipulate, however, that mucosal immunity is most effectively formed in "naive" people who have not previously encountered this antigen, and synthesized IgA probably will not live long. Nevertheless, cellular immunity and memory cells are also formed during mucosal stimulation. Perhaps the best strategy would be to use nasal vaccines in combination with traditional injections, for the joint work of systemic and local immunity.

However, so far all such vaccines are only undergoing the first phases of clinical trials — in July, Science counted seven such vaccines. Since the summer, at least one more has joined these developments — the nasal variation of Sputnik V, which this October received approval from the Ministry of Health for clinical trials. According to the entry in the Register of Permits for Clinical Trials, the second phase of testing of the nasal "Satellite" on 500 people will end in December 2023.

 It is also known that even before they began, at least one person experienced its effect on himself — in June, the chairman of the Federation Council, Valentina Matvienko, became the chairman of the Federation Council, who already in October said (however, without reference to any scientific research or the opinion of specific experts) that nasal vaccination "does not cancel the need to get a regular vaccination." There is no other information about the effect and limitations of the "satellite zilch" in the public domain yet.

In addition, the nasal form of Sputnik V is going to be tested by another Russian manufacturer, Generium Pharmaceutical Company, which created it "based on the development of the Gamalei Institute" — however, there is no corresponding entry in the Register of Permits for Clinical Trials (RCTs) yet.

Therefore, we will not find out in the very near future how much the rather obvious idea of stimulating local immunity in the nose to protect against a virus that penetrates through the nose actually works.

Is it even effective?

There is still quite little data on how important the role of mucosal immunity is in protecting against infection with the SARS-CoV-2 virus. But studies in 2009 showed that when infected with another coronavirus, the causative agent of the common cold, IgA antibodies appear in patients, and their presence correlates with the duration of infection. For covid, the formation of antibodies of this type has also already been shown — and, apparently, unlike IgG, they are formed with any severity of the course of the disease. However, it is not clear whether the protection of IgA trained to fight the viral particles of the coronavirus is enough to prevent the disease.

There is exactly one approved mucosal vaccine for respiratory disease — this is the above-mentioned nasal spray for influenza. The FluMist vaccine is based on a weakened vector virus, in which antigens of influenza strains are embedded. If we talk about other, non-respiratory viral diseases, there are also approved mucosal vaccines against polio and rotavirus infection, which are used in the form of a solution.

As for the bacterial mucosal vaccines available on the market (namely, St. Petersburg scientists are going to make one), they are all designed to provide protection not from respiratory diseases, but from "dirty hands diseases" — in particular, cholera and typhus. They are based on killed bacteria, and the composition of the Dukoral cholera vaccine, in addition to the Vibrio cholerae strain, also includes cholera toxin as a component that enhances the immune response (adjuvant).

Instructions for vaccination with Dukoral European Medicines Agency

Thus, the development of scientists from the IEM is really innovative — and therefore risky. It is completely unknown whether a combo of bacteria and S-protein will work in the body, and how strong the formed immunity will be, how long the modified bacteria will multiply in the intestine and whether they will be at all.

The results of the tests of the "kefir vaccine" from covid will have to wait more than a year — and it is unknown how much the virus will mutate during this time. Preliminary test results of another recombinant vaccine speak in defense of St. Petersburg enterococcus. However, this is a vaccine against diarrhea, which contains killed strains of E. coli with toxin genes inserted into them. She successfully completed the second phase of clinical trials in Bangladesh and led to the appearance of the necessary antibodies in the subjects, while not showing serious side effects, including on children.

In general, vaccines in capsules, yoghurts or sprays are not limited to. For example, more extravagant vaccination options have been discussed for quite a long time: with the help of transgenic tomatoes, bananas and potatoes.

Eat me

The idea of making vegetable vaccines has been floating in the scientific community since the discovery of genetic modification of plants. This form of vaccine release seems ideal: it is convenient to produce, store and use.

In the XX century, quite a lot of attempts were made to turn vegetables into vaccinations. For example, they tried to produce E. coli toxins, hepatitis B virus proteins and gastroenteritis pathogen in potatoes. These developments have reached clinical trials in humans. However, since protein antigens tend to denature during cooking, the volunteers had to chew raw potatoes.

In 2000, in an American trial of transgenic potatoes producing proteins of norovirus, which causes intestinal infection, two groups of 10 participants ate 150 grams of transgenic tubers twice or three times (the control group was given ordinary potatoes). Antibodies of the IgA class against the norovirus capsid protein then appeared in 19 out of 20 participants, but the concentration spread turned out to be huge: from 6 to 280 conventional units. The authors attributed this to the unstable expression of the antigen in the tubers themselves, and recognized that oral immunization with pure recombinant capsid protein, without any potatoes, gives a more stable result.

The same research team then tested transgenic corn that synthesized E. coli toxin. Despite the fact that seven of the nine volunteers who were fed cornmeal had antibodies against the desired antigen, the tests did not continue — perhaps because secretory antibodies in the intestine were found in only four. A similar story happened with salad loaded against hepatitis B, and with spinach against rabies. And attempts to make carrots against measles and tomatoes expressing HIV proteins did not go further than experiments on mice.

Formation of antibodies of two different classes in volunteers vaccinated with transgenic corn with E. coli toxin. The arrows indicate the days of use of the corn vaccine
Tackle With et al / Vaccine, 2004

Why is it so difficult? Firstly, it is difficult to calculate the necessary doses for the development of immunity: different fruits can express antigens at different levels, and in addition, they are destroyed both during cooking and in the stomach. Secondly, banal legislative restrictions. After all, we are talking about genetically modified plants that are not allowed to be grown in all countries. And for full-fledged tests, large samples are needed and, consequently, a lot of product.

Scientists, however, do not abandon their attempts. In September 2021, scientists from the University of Tokyo reported on the results of the first phase of clinical trials of the MucoRice-CTB anti—cholera vaccine - transgenic rice against cholera.

In the meantime, plants are more often used not as a method of delivering vaccines to the human body, but as platforms for developing the proteins and antibodies needed by scientists. Tobacco is most often used as such a platform: in its plants, they are trying to grow, among other things, the S-protein of the coronavirus for use in subunit vaccines.

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