Research: Antigen-coated beads the New (Old) Cure of the Week


Aberrant T-cell activation underlies many autoimmune disorders, yet most attempts to induce T-cell tolerance have failed. Building on previous strategies for tolerance induction that exploited natural mechanisms for clearing apoptotic debris, we show that antigen-decorated microparticles (500-nm diameter) induce long-term T-cell tolerance in mice with relapsing experimental autoimmune encephalomyelitis. Specifically, intravenous infusion of either polystyrene or biodegradable poly(lactide-co-glycolide) microparticles bearing encephalitogenic peptides prevents the onset and modifies the course of the disease. These beneficial effects require microparticle uptake by marginal zone macrophages expressing the scavenger receptor MARCO and are mediated in part by the activity of regulatory T cells, abortive T-cell activation and T-cell anergy. Together these data highlight the potential for using microparticles to target natural apoptotic clearance pathways to inactivate pathogenic T cells and halt the disease process in autoimmunity.

 This is a picture of MD2’s urine after hearing about this article
(apologies for the poor taste, but that was his reaction as it made his p**s boil)
Many, many years ago the group of Stephen Miller showed that if you stuck myelin antigens onto cells with a chemical cross-linker that it could induce immunological unresponsiveness (immune tolerance) to disease induced with myelin antigens. As you may remember, I posted on using this approach as being half-way to a cure in humans when it was posted in ECTRIMS 2012. 

The group have realised that sticking peptides to the individuals own white blood cells is a problem, because it has to be individualized and cannot be standardized for every one. They thought that you could replace the cell with a polystrene bead or a microparticle and this NEW bit of biotechnology has turned out to be this weeks “Cure of the week”. 

They show that if you inject these microparticles into the blood they induce immunological tolerance in an antigen-specific way, via a mechanism that centres on T cell depletion and anergy (unresponsivenes of the T cell to stimulation) and not so much on T regulatory cells or interleukin-10 producing regulatory cells. Furthermre the peptide needs to be stuck on the surface. These microparticles are taken up in the spleen by macrophages expressing a receptor called MARCO, because in some of their past work they show that these cells took up dead (apoptotic) cells. 

As such tolerance was not present in MACRO-deficient mice when treated with antigen beads, but this surprisingly had no effect on tolerance induced by dead cells? They did not need polystyrene beads as this could also be done with biodegradable beads, approved for human use, so this could be an approach that could be easily translated into human use as both peptides and beads are available for human use. This is very good news let us hope they develop this, but the problem with this approach is that they are using peptides.

Peptides are easy to manufacture to the standards required for human use, but if you believe the data, it means that they need to know all the antigens that each individual will respond to, as it is very antigen-specific mechanism.  The solution to attach a mixture of whole proteins so that you do not need to know which epitopes the individuals will respond to. This is important because each individual will respond to a different set of proteins during disease. Furthermore to get optimal results in established disease they need to transiently deplete the T cells. How do I know this?  

Well if you care to read papers Pryce G, O’Neill JK, Croxford JL, Amor S, Hankey DJ, East E, Giovannoni G, Baker D.Autoimmune tolerance eliminates relapses but fails to halt progression in a model of multiple sclerosis. J Neuroimmunol. 2005;165:41-52 and notably Smith PA, Morris-Downes M, Heijmans N, Pryce G, Arter E, O’Neill JK, ‘t Hart B, Baker D, Amor S. Epitope spread is not critical for the relapse and progression of MOG 8-21 induced EAE in Biozzi ABH mice. J. Neuroimmunol. 2005 Jul;164(1-2):76-84 that the Nature journals of the time (Not sure there was Nature Biotechnology then) did not find interesting, then this may be clear.

There you will see that myelin antigens when chemically coupled to polystyrene beads (or microparticles) stop relapsing disease in EAE, not just given before disease ever develops as mainly occurs in this  current paper.

Reading these papers and you would know that you didn’t need cells to induce tolerance or apoptotic cells for that matter, that it works via depletion and anergy and a little by regulation and that the antigen ends up in macrophages in many different places and not only in the spleen but in the liver and lungs. 

So this new technology is really not so new. We are pretty sure that at least some of the authors of the current paper are aware of the original study, so I wonder where the idea really came from?

However, no citation of the original idea by MD2 (…..again). It would show that much of this is not really innovative, which would limit the chances of publication. They have however made some biodegradable beads. These were available 7 years ago but not with the side chains to link proteins to them…we looked. I will not get on the high horse of reporting and refereeing standards, otherwise my wee may boil too.

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  • Sad that the original research is not quoted, though not particularly surprising.
    Good to see though that the original research holds up. Wonder how Steve Miller explains this with regard to his previous work on tolerance?

  • If you believe in the epitope spread hypothesis presented here, that one relapse conditions the next relapse by a response to a different epitope, then how many epitopes are needed and which ones do you use if it is an orderly progression you may have missed the boat, now put into the mix that for every relapse there are at least ten or more lesional events then the number of epitopes need to grow and grow.

    Epitope spread does occur such that the autoimmune response expands with time, but barring a very few labs, it is not sequential and orderly and can be reactivation of responses focused one epitope how do we get relapsing disease in T cell receptor transgenic mice, where the spread in that…maybe a load of lard:-)

  • The cited papers are poor articles to use to call this an old idea. Indeed, the use of polymers and the role of MARCO are the most impacting components of this work. While the MS model used certainly has some pitfalls, the EAE model remains the most reliable autoimmunity model in the lab setting. Clinically, celiac disease and/or allergy based diseases with well defined epitopes will need to be tested to truly test the technology.

    • The core of this article is stick antigen to beads. The rest is topping it is a paper called biotechnology.Had the authors cited our paper of seven years ago it would be clear sticking antigen to beads is not an original idea and i would bet the paper would of had trouble getting published in Nature as. lack of originality is a killer
      A literature review is part ofgood science

      I grant you there is some original work the biodegradsble beads but this should have been the starting point of the paper not the finishing point

    • Read. Smith et al 2005 and you will see antigen stuck to bead inhibits eae. Before that antigen was stuck to white blood cells. At that time the miller group was saying that antigen stuck to cells was working via anigen the cell onto which the cell was stuck. We showed that this was unlikely to be the case the bead experiment has one part of this

  • The papers cited above are not the ones you are referring to, which articles did you publish- i would certainly be interesting in reading those as well.


    • Are you talking about papers questioning a sequential epitope spread? There are lots do a search on eae and do not read those with sd miller who has dug himself into a hole with this hypothesis. Hard to back track out except just ignore it

  • Actually i went back and read your article above– any idea why when you infused peptides in animals with ongoing disease you did not see any anaphylaxis. The premise of the NBT article is that this causes issues (published by miller as well– Differential induction of IgE-mediated anaphylaxis after soluble vs. cell-bound tolerogenic peptide therapy of autoimmune encephalomyelitis. Smith CE, Eagar TN, Strominger JL, Miller SD. Proc Natl Acad Sci U S A. 2005 Jul 5;102(27):9595-600. Epub 2005 Jun 27)

    • The peptides were coupled either to white blood cells or in one case carboxlated polystyrene beads. This means the peptides were not soluble and therefore did not cause anaphylaxis.

    • Why did we not see anaphylaxis….we must not of had a relevant specific IgE stuck to mast cells.

      PART 1 Now the history to this question.

      It was reported by the Steinman (Stanford Group) group that intravenous myelin peptide in sensitized EAE animals can cause IgE mediated anaphylaxis (allergic reaction). The Miller (Chicago) group then later showed that intravenous peptide injected into sensitized animals could also cause anaphylactoid reactions, but according to the Miller group this was caused by IgG rather than IgE. These reports in mice were in response to an even earlier study by the Steinman (Group) that myelin peptide injected subcutaneous caused anaphylaxis in MSers.

      A Nature paper saying the trial in MSers taking years to organise did not work and then another a bit later again in Nature in an experiment taking 5-6 weeks of work in mice saying why trial would not work and cause anaphylaxis that killed mice was gobsmacking.

      It was unbelievable, because the inference was the anaphylaxis was a surprise. This was because their past work supporting the trial was doing treating before animals got disease. That they did not do the study in relapsing disease in the mice until after the trial is the shocking bit.

      An experiment that costs a few bob compared risking MSers lives.

      However, once clinical trials are put in place I suspect that companies do not want to do further experiments that could throw up something they do not want to see or hear.

      These types of experiments are doable but so often do not get done (70% of EAE treatments are done before disease is induce). This is because the interest of the scientist may be in a mechanism not because they are concerned with MS. The person doing the trial is often not a person who came up with the idea and does not do experimental work, so they (professional trialists) would not go back to do the right experiments) and is one of the reason that stuff in EAE fails to translate into humans.

    • PART II

      Furthermore they (Smith CE et al. 2005 in Paper above from the Miller Lab) showed that if you stuck the peptide to cells that the anaphylactoid response did not occur. This makes sense because if you have a destructive antibody response targeting the peptide stuck onto a cell/bead it does not matter if the destructive response destroys the cell because the cell is already dead anyway, because it was killed during the cross-linking (Fixation) process when you stick the antigen on.

      If the animals were not sensitized when the treatment was used prophylactically (before disease induction) there was no anaphylaxis because no antibody response had yet developed.

      Why did we not get anaphylaxis in our studies (Smith PA et al J Neuroimmunol 164:76) as we did virtually all our experiments in sensitized animals. Well when we used proteins we either stuck them on cells/beads and the peptides we used did not drive an antibody response as they were T cell epitopes.

      It was well known at the time that T cells see short linear bits of protein whereas B cells see longer conformational bits of a protein

      We did do some later experiments with a long (twenty amino acid) myelin oligodendrocyte peptide and this could cause anaphylactoid response whereas a shorter (thirteen amino acid) peptide myelin oligodendrocyte peptide did not.

      However we did not get this using whole protein either, but maybe because the dominant immune in the animals are not against the antibody inducing elements. But getting antibody responses after intravenous antigen is to be an expected response. This property has been used in thousands of mice when they are given the final boost to drive antibody production before you take the spleen for monoclonal antibody production.

      So in most cases anaphylaxis which would kill the mouse does not occur. This approach has been used in humans with myelin basic protein without a major problem being reported.When we did something similar we were ready with the adrenaline in case of this predictable activity. Likewise this is a risk with any delivery of protein such as antibodies like Tysabri, even CAMPATH if you have a re-existing antibody response.

    • PART III
      The story starts long before this. This boils done to many, many studies showing that if you deliver soluble antigens by the intravenous route it causes T cell immunological tolerance, which is unresponsiveness of T cells.

      The history also told me that injecting peptides under the skin is usually a sensitizing route.

      Now it was shown that if you have a peptide that has some substitutions in the sequence that they can be even more potent action than the normal sequence at inducing T cell tolerance when the peptide was inhaled (via a Th2 response).

      This gave rise to the term altered peptide ligand. This had the amino acid sequence of myelin basic protein with a few amino acids changed. They appeared to convince themselves that all altered peptide ligand were a better tolerogens than the normal native protein and forgot that the route of delivery was all important for T cell tolerance induction.

      A myelin basic protein altered peptide ligand when presented by HLA-DR2 was made. Maybe because of fear of adverse reactions when it came to the human trials they decided to inject it under the skin rather than intravenously and disaster and some MSers suffered the consequence.

      In a different study to the one inducing anaphylaxis, MSers suffered more attacks. They seemed not to tissue type the MSers in the trial and not all were HLA-DR2 positive and in fact some were HLA-DR4+ were the altered peptide ligand did not enhance a tolerogenic response but acted just like the natural peptide and stimulated a pathogenic response.

      They put this peptide in via a sensitizing route and surprise, surprise some attacks were triggered. This is the only bit of real evidence that suggests that MS is an autoimmune disease. However some of the MSers got peripheral nerve disease (MBP is present in both peripheral and central nerve) which suggests to me that it is not the most important antigen in MS

      Could history repeat itself, possibly.

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