Precision-Guided (MALT-Targeting) Mucosal Vaccines
This page contains a list, with brief overviews, of topics that are addressed in more detail on the pages that follow. If you click on any red header, below, it will take you to a page with more info on that topic.
1. BRIEF INTRODUCTION TO MUCOSAL MEMBRANES, AND MUCOSAL PATHOGENS
This page describes how mucous membranes are very different from dry skin. “Epidermal cells” actually are empty-bag pseudo-cells, which makes them ideal for absorbing and entrapping infective microbes, and preventing them from reproducing. Since mucosal membranes are covered by a different type and class of complete and functional cells (“epithelial” cells), they are major targets for pathogenic microbes, and “mucosal pathogens” are – by far – the largest and most important class of pathogens on this planet.
2. MALT PATCHES, AND HOW “M CELLS” SAMPLE AND PULL IN APPARENT PATHOGENS
MALT (“mucosal associated lymphoid tissue”) patches are specialized lymph nodes, mounted on the outer exposed surfaces of mucous membranes. They are a crucial “first line of defense” against pathogens that try to infect mucous membrane cells.
M cells are “sampling cells” on the surfaces of MALT patches. If they detect a particle carrying a “pathogen pattern”, they will grab it and pull it in; however, rather than processing it, they will push it through the cell, as rapidly as possible, and eject it into a “docking site” where a different type of immune cell – an “immature” (or “pre-committed”) dendritic cell – is waiting for that type of “pathogen delivery”.
Therefore, “MALT-targeting” peptides are “pathogen pattern” peptides that can be attached to the surfaces of vaccine particles, so that they will “trick” the M cells into pulling them in, and passing them on to dendritic cells that are waiting for pathogen deliveries.
Dendritic cells play the absolutely crucial role, in determining: (i) which foreign particles are NOT really important and dangerous, and should be gobbled up and digested on the spot, without further ado; versus (ii) which foreign particles appear to be dangerous and important pathogens, which need to be taken to a lymph node (while being broken apart and semi-digested along the way), so that T and B cells in a lymph node can make antibodies that will bind to those alien/invading particles. With the help of “chemo-attractant” signals, newly-formed dendritic cells find and settle into the “docking sites" on the undersides of M cells, to await a pathogen delivery that will cause them to transform from an “immature” dendritic cell, into an “antigen-presenting” cell. Therefore, “MALT-targeting peptides” – when attached to the surfaces of mucosal vaccine particles – offer an ideal way to get those vaccine particles rapidly delivered to dendritic cells, and to get the dendritic cells which receive those particles to do exactly what is needed, to launch an antibody-forming response to any antigens on the surfaces of those vaccine particles.
The Y-shaped internal IgG antibodies are not large or powerful enough to hinder a virus, let alone a bacterial cell, so they fight pathogens by using a shape-changing “tag and flag” process. When either “sticky arm” latches on to a particle, the antibody “stem” changes from a “Leave me alone, I’m an antibody” shape, into a different shape that signals, “I’ve latched onto something important, so somebody go find an immune cell, and tell it to come here and help me.” Guided to that location by “complement proteins” which “amplify” that signal, immune cells do the actual work of fighting and killing pathogens.
However, in secreted mucosal fluids that are outside of any cells or tissues, there are not enough immune cells (or complement proteins) to respond to any signals from shape-shifted antibodies. So, “secreted mucosal antibodies”, needed a totally different structure, and function, to be able to function effectively, and the mucosal immune system developed a remarkably clever way to strap two antibodies to each other, via their stem components, in a way that creates an “antibody dimer”, with FOUR different “sticky arms” (with two, at each end of an elongated molecular complex). Secreted IgA antibody dimers perform a “grab-and-drag” function, which prevents pathogens from penetrating into any mucous membrane cells. If a pathogen is grabbed in the mouth or nasal cavity, it will be dragged down into the stomach acids, which will kill nearly all pathogens; if grabbed in the intestines, it will be kept suspended in the food that is being digested, until it is “pooped out” and eliminated.
“Adjuvants” are compounds added to injected vaccines, to make them more effective and potent. However, in practical terms, they are harsh, irritating, inflammatory, distress-causing agents. They are added to injected vaccines, to cause the muscle cells at an injection site to send out distress signals (“cytokine” molecules), to recruit any nearby immune cells to come to the injection site, before the vaccine particles can be diluted, diffused, or degraded. Because of how they work, MALT-targeting vaccines have the potential to completely eliminate any need for harsh and irritating adjuvants. If that turns out to be the case, they will offer a major benefit, for all vaccines.
Most people have never even heard of “secreted antibody dimers”, but there are more than twice as many of those – in just the few pounds of saliva, nasal mucus, lung fluids, and digestive juices, in a healthy human – as all of the internal antibodies in the entire remaining weight and bulk of that person's body. Our bodies would not devote so many resources to creating huge numbers of mucosal antibodies, unless they were truly important, and immunologists know, full well, that if vaccines could be created which could safely and reliably create mucosal AND internal antibodies, those “balanced, bi-functional vaccines” would be better and more effective (especially against “upper respiratory tract” infections) than vaccines which can only create internal antibodies. However, scientists and vaccine companies have never previously been able to make vaccines which can trigger BOTH internal AND mucosal antibodies, without using truly nasty chemicals which are not acceptable for livestock or pets, let alone humans. MALT-targeting mucosal vaccines have the potential to change that, dramatically.
7. STARTING-POINT FACTS ABOUT PHAGES, “PHAGE LIBRARIES”, AND "SCREENING TESTS" THAT CAN BE USED TO ISOLATE THOSE PHAGES WHICH CAN PERFORM SOME SPECIAL KIND OF TRICK OR FEAT.
“Phages” (originally called “bacteriophages”) are the smallest viruses ever discovered. Each type can infect only a small set of bacteria, and none can infect plants or animals, so they are treated as harmless and non-pathogenic.
Phage “libraries” (aka phage display libraries) contain billions of different phage particles, and each particle carries a different, randomly-created “foreign peptide insert" on an exposed “coat protein”. They took decades to develop, but now, a top-quality library with a trillion different phage particles can be purchased for less than $800 from companies like New England Biolabs.
“Screening tests” are clever ways, thought up by scientists, to subject millions of phage particles to a fair competition, usually involving something like, “Uptake and processing, by a specific and unusual type of cell”, so that they can isolate and then analyze those few phages which happened to be carrying an inserted peptide sequence which caused those cells to perform that activity. Rather than trying to predict, design, or control which protein sequences will be able to perform "the XYZ trick", it often is easier to design a way to let millions of phages give it a try, and figure out some way to isolate those phages which were able to do that particular trick.
So, this page provides background information, on what phages, phage libraries, and "screening tests" are, to enable non-experts to understand what we did, why we did it that way, and what we got, as the results.
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Building on all the information and factors described above, Precision-Guided Vaccines LLC thought up and used a new type of screening test, which isolated about 100 phage particles (out of a billion candidates/contestants) which happened to be carrying peptide inserts which made those particles appear to be so dangerous, and so important, to M cells and dendritic cells (in mice), that those specific particles were taken in and processed – as quickly as possible – by those M cells and dendritic cells, in ways that would have led to antibody formation, if we had allowed the cells to continue. Instead, we extracted a mixed batch of mucosal surface cells, and used a clever screening method, to isolate only those dendritic cells which had become transformed and activated, by their contact with the specific phages they had taken in. We then broke those activated dendritic cells open, and analyzed the foreign inserts in the phages they had taken in.
Once we knew the DNA and amino acid sequences for the phage inserts that functioned as “MALT-targeting” peptides, we hired a phage assembly lab to create “the first testable constructs”, carrying 15 copies of the best-performing MALT-targeting sequences; and hundreds of copies of a well-known antigen that is easy to test for. In both mice and pigs, a single nasal infusion of those particles (with no adjuvants) caused "robust” formation of BOTH: (i) internal IgG antibodies, in blood, AND, (ii) secreted IgA antibody dimers, in saliva, as shown by both ELISA and SDS-PAGE/Western assays. Those results were so good that we shifted over to a different type of phage that is better suited for pathogen challenge tests.
We do not want to compete against any vaccine companies; we do not want to become a manufacturing company; and, we do not want to assemble, and then learn how to use, biosafety equipment, to be able to do pathogen testing on animals. Other people are already experts in all those things, and we truly and genuinely respect their expertise, and hope to work with them, in ways that will let them use their talents and skills to make the best vaccines that can possibly be made, both for humans (some day), and for non-human animals (as soon as possible).
Therefore, our goals are: (i) to become a licensing company; and (ii) to promote more research into MALT-targeting mucosal vaccines, as quickly as possible, by as many animal vaccine companies as possible, by offering to provide them, at the lowest possible cost, ready-for-testing T7 phages carrying both: (a) about 40 copies/particle of the three best-performing MALT-targeting sequences we have identified, so far, and (b) about 400 copies/particle of whatever antigen/epitope sequence a requestor wants to test, in one or more specific types of animal(s). And, to help encourage, motivate, and incentivize experts in animal testing to begin doing that research, we hereby commit to offering worldwide exclusive licenses, for any use of MALT-targeting peptides, to the first companies that are able to generate enough solid data to qualify for governmental approvals to sell the vaccines that they have shown to work.
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