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Tales from the front line

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  • Tales from the front line

    If you are sitting comfortably I am gong to tell you a story about a great battle in the war.

    We knew they were going to attack; it was only a matter of time. All our forces broadcast our friend or foe signal MHC and we had the signals scouts from B company & T company troops who knew what some of the enemy looked like - I wished we had more like them. When they came they were a new foe coming down blood road, no one initially recognised them, signals first suspected them, B cell changed over to Plasma Cell mode to broadcast their description in an antibody. Now the game was up with the antibody description plastered all over them they were easy to spot. The Killer T cells (CTLs) attacked lobbing granulocytes (chemical grenades, histamine and all kinds of toxic stuff), the NKs (Natural Killers) were already there because they had not detected an MHC so they just went in granulites blazing then there were the enormous Phagocytes, Monocytes gobbling up the enemy and Neutrophiles eating them and detonating granulocytes on them once confined. As if the battle was not dangerous enough the eosinphiles & basophiles were spraying toxic chemicals everywhere on us and them – very inflammatory! Meanwhile the signals section of the T cells sent out a stream lymphokine cytokines giving instruction to the whole army: whether new stem cells should be trained up into T (that’s an IL2), B, NK or phagocytes, calling more troops to hotspots (chemokines) and generally controlling the battle. Not to be outdone the Phagocytes had their macrophages & monocytes broadcasting monokine cytokines giving similar orders, the air was so thick with interlukine cytokines it was difficult to see. The battle was touch and go when B cells sent out an IgG antigen calling in the complement, about 25 proteins cascaded into the enemy forming an opening and it was guts all over blood street. After that it was mainly mopping up operations, some of the monocytes stopped patrolling blood street changed to macrophages and chased the enemy who had escaped into the tissues others phagocytes cleaned up the debris. Many of T & B company were disbanded but we always keep a few who can recognise them if they come back - and we have their antigen ready to broadcast – they will be sorry if they show there faces around here again. I feel it only fair to mention the T helper cells, while not in the thick of it they activated new T & B cells and called in macrophages but never get a mention in dispatches. I will try and find my old code book so I can show you some of those messages in the cytokines and immunoglobulin antibodies but that is a story for another time.

    Yes, it is confirmed, I am mad. I just have a visual memory and am awful with names, if I can thinks of it as a system it sinks in better.

  • #2
    Mab Libraries?

    I am not an immunologist so apologies for any errors.
    When H5N1 first came to my attention I started to try and find more information and used the internet to do so. I quickly discovered there was a community of others who thought this needed attention paid to it and who had amassed a lots of information already. As I got further into trying to evaluate the threat I realised there were a vast number of conflicting opinions and I was going to have to learn a fair bit of biology to be able to work out who was talking sense. Of all the areas I have tried to learn something about only once have I had to accept defeat and that was Immunology.
    Initially it is not too great a problem you can sort out active/passive and the main cell types (Bs, Ts, CTLs NKs etc), cytokines, chemokines and their various sub divisions. The problem comes with the up and down regulation. Knowing who the main players are and having some understanding of their roles is not enough; many of the players are either suppressing, or promoting, the activates of other players. As an illustration of the problem the cells listed above - B-cells, T-cells etc - all come from a common stem cell produced in the bone marrow. Which cell type a stem cell develops into is determined by the cytokines produced by the previous generation of T-cells, which is regulated by that T-cells chemical environment, this is being moderated by the output of other cells which are varying their output depending on any perceived immunological threat. There are positive and negative feedback loops everywhere.

    Having explained why you should not believe a word I say on Immunology I will now write some thoughts on exactly that subject. Shiloh posted a report on some research Scientists Create First Successful Libraries of Avian Flu Virus Antibodies which relates to some work to be published in PNAS which – regretfully – I have not got access to at the moment. This is along the lines of something I had advocated as a worth while avenue of research in a discussion with Mingus on the industrial scale use of Air lift fermentation production of Mabs as a longer term solution to pandemic vaccine production, some of which end up in a thread (this was a long wide ranging thread and the relevant sections are near the end - very sorry can not find link at the moment may have been archived if anyone recalls, started off being on 1918 recovered patients plasma being used to produce immunity). It would also be useful to look at Mingus’ thread on H5N1 vaccine matching to understand the ramifications of this work.

    If we assume an H5N1 viron has entered the body but not infected a cell but has made first contact with the immune system. Step one is the immune system needs to see it as ‘not self’. Only the external surfaces of the virus are ‘visible’ and in our case that principally means parts of the H & N proteins. Ignoring all the detail of what happens next the salient points are that certain surface features of these proteins’ tertiary structure (antigenic sites) are ‘learnt’ and communicated (antibodies are produced by the B-cells) to the rest of the immune system which then gears up to fight the infection. This is an important step change; the immune system has changed from looking for any thing that is ‘not self’ (generalised xenophobia) to identifying a specific threat and targeting it. A second viron – with identical surface feature – will now have a much lower chance of successfully reaching its target cell type as it is being actively targeted. If the surface features are similar – but not identical - then protection will fall somewhere between the two states. This learnt protection is not based on the whole overall viral surface shape but on lots of small features with an antibody produced for each. Monoclonal antibodies (Mabs) are artificially produced antibodies and these researchers will have produced as many as they can based on those found circulating in the survivors of Turkish clusters. They, of course, only have antibodies to the virus they encountered (a strain of Clade 2.2 HPAI H5N1) so this is an incomplete library and will not include antibodies specific to other clades or serotypes. The flu virus is famous for its ability to mutate and selective pressure will favour variants that are not easily recognised by the immune systems of its hosts; so much of the surface is constantly reinventing itself to avoid detection. Certain areas are less free to change their appearance than others, particularly if the surface shape has an important biological function (like the receptor binding site on the Hemagglutinin protein which is involved in ‘docking’ with the host cell) a change in this area may help avoid immune system attack but will probably also prevent cell surface binding and penetration. The long term aim would be to broaden the library to include key antibodies to a wide range of flu (or any other disease’s) surface antigens and then be able to quickly identify the surface antigens of a given flu – or flus – and select a number of Mabs from the library to form a vaccine cocktail targeted at that specific strain. As there may be many antigenic sites we could prime a response for, it gives us the opportunity to produce different cocktails for the same virus which has the added benefit of making it difficult for the virus to adapt against. That last bit might need a little clarification.
    An analogy: Assume there is a spy in our midst and I broadcast that he is bald and has a birth mark on the top of his head - if he puts on a hat he is safe. If to some of you I describe his birth mark, to others his finger prints, to others eye colour, his height, weight, age etc. He now has a much bigger problem. Each Mabs is like one of these bits of data all I need to do is give out enough to make his detection likely, if I keep varying which bits it is difficult for him to counter.

    With apologies for the anthromorphising, gross over simplification (including completely ignoring the non H & N antigenic sites that get posted by MHC cells etc) and many other crimes.
    Last edited by JJackson; December 15th, 2015, 05:16 PM.

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    • #3
      Re: Tales from the front line

      "I just have a visual memory and am awful with names, if I can thinks of it as a system it sinks in better."

      Hi JJ.,
      imagin you DON'T have a visual memory - it would be much worst than.

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      • #4
        Re: Mab Libraries?

        Originally posted by JJackson View Post
        (this was a long wide ranging thread and the relevant sections are near the end - very sorry can not find link at the moment may have been archived if anyone recalls, started off being on 1918 recovered patients plasma being used to produce immunity)..
        Found the thread http://www.flutrackers.com/forum/showthread.php?t=9783 and Dutchy posted this one Researchers find quick way to make human monoclonal antibodies against flu yesterday.

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        • #5
          Re: Tales from the front line

          I am posting this here as it is both a reply to gsgs's post in Dutchy's thread (linked to in the previous post) and relevant to this thread. Post #2 (and the other link in the last post) were offering industrial Mab production as a pandemic vaccine alternative but did not explain why the existing system was unsuitable, this tries to redress that omission. As gsgs points out partial immunity - and better booster priming - are achieved if you can get the homology of the strain and vaccine as close as possible, again see the Mingus vaccine matching link in Post #2.

          Originally posted by gsgs View Post
          1) prepandemic vaccine as recently ordered by HHS, Qinghai strain, $5 per dose
          for all US-citizens
          2) advance purchase agreement (APA) with vaccine producers for strain-specific vaccine in case of
          a pandemic to be delivered after 4-5 months in a pandemic.
          the prepandemic vaccine can be given as first dose ("mock up") immediately.
          It will produce some cross-protection depending on the strain.
          In connection with the 2nd strain-specific dose it should probably work almost as well
          as two specific doses.

          Switzerland and others(Singapore,Austria,Denmark?,Ireland?,.. )did it for 100% of their population
          UK has planned/ordered it for 50% of the population other EU-countries have similar
          solutions. USA mainly relies on their capacities to be completed 2010,2011

          I don't know, what 2) costs. The agreement alone without production, maybe $10 per dose.
          1) may have to be renewed after 3? years
          in 2008 other solutions become available, capacity is bigger, prices lower.


          Some expensive antibody-doses for health care personnell etc. could then quickly be produced
          when/if panflu starts.
          But I assume the bulk will still be vaccine. I.e. for poorer countries.
          Provided the vaccine works well, of course - better have several options

          I do not disagree. Although I think we may be answering different question.

          What I want is a solution to: What can we realistically do to control the potentially catastrophic consequences of a highly virulent flu pandemic. H5N1 is showing us that 1 or 2 % are not an upper limit on CFR.

          What you have suggested is what we can do now and I agree it is worth doing but it is not a solution to my problem.

          It is not a solution to my problem because
          There is inadequate production capacity.
          Egg production is not fast enough and not scaleable.
          The efficacy of the primer dose is dependent on correctly predicting the serotype and getting close to the strain – and there is not the capacity
          The strain specific booster can not be produced in time – and there isn’t the capacity.

          In a little more detail
          Production capacity
          Seasonal flu is nasty but can generally be dispatched by a fit adult immune systems so it is not cost effective to protect this group, which makes up the bulk of the population. H1N1 and H5N1 have showed that some flus have no problem killing even the fit and healthy – and in unacceptable numbers. The situation is actually less rosy than this because capacity is further limited to match the subset of those who can afford it, mainly in the rich industrial nations. If the richer nations were fortunate enough to correctly predict the pandemic strain and save every citizen from an H1N1 or – god forbid H5N1 – type pandemic then they would still have lost their world: A gated community in a smouldering city. That is assuming we were so bereft of compassion we were willing to be spectators. Vaccine campaigns can have positive feedback. The goal is heard immunity in which a disease dies out, or spreads very slowly, because it has difficulty finding a susceptible target – most already being immune. Enlightened self-interest: This benefit is greatly diluted if you have a pool of high immunity in a sea of lower immunity. This sea acts as the disease’s reservoir from where it can keep picking of the vulnerable in the pool. Tamiflu epi-modelling predicted a better outcome for Americans – given a fixed amount of Tamiflu – if they did not keep it all for themselves but distributed part of it to other parts of the world (from memory I think the optimum split was keep ~90%) the principle is the same. One last point: Non animal vax plants produce significant quantities (especially in China) and I have never understood why these could not be built/upgraded-to dual use, I know this was to be looked at a couple of years ago but would be grateful if anyone can supply a link to any developments or reasons why this would not work.


          Speed of production
          While egg based vaccine production suffices for seasonal flu it is a hand tool technology in a CAD/CAM world. It takes months to produce 1 dose and – given our current manufacturing capacity – you could never produce enough to get effective heard immunity. The first vials would be long past their use-by-date before the last came of the line - that is assuming you could persuade the flu strains to stop mutating for a few years while you built up you stocks. You could build more vax plants but it would not cut down time to first dose and they may not be needed for 30 years. For pandemic production you need a flexible process which, like CAD/CAM, can build tractor parts for 30 years on a 40 hr week and switch over night to tank parts running 24/7 if you are attacked. It needs to be dual use with fast switching and scalable.

          Primer dose
          The next pandemic may or may not be caused by H5N1. H7s are in the news as well and are just as good at infecting humans and h2h – also available in both LP & HP. The 1968 pandemic was a reassortment virus. Furthermore there is little evidence that any of the 20th century pandemics gave us any warning prior to emergence so building a strategy based on knowing the serotype and strain in advanced is very dodgy. This year the Northern Hemisphere seasonal flu jab was largely ineffective despite knowing in advance exactly what the serotypes were and having a triple strain vaccine to improve the odds of getting the right one.

          Booster dose
          Most of this is already covered. If the pandemic is due to a zoonotics emergence the virus should undergo accelerated genetic change under the increased selection pressures of a new host. While we know little about the inter-strand interactions we do know of various sites that are typically avian or mammalian markers (PB2 E627 => K627 etc) and we can expect these to change. Artificial pressures like anti-virals will also fix changes on NA & M. If it takes 6 months plus to begin to get meaningful amounts of booster vaccine (assuming you scored a hit with your primer) the virus may have evolved significantly away from the case zero you based the booster on.

          On the question of cost this was discussed in the thread linked to above but the short answer is I could not find out why Mabs are always thought of as expensive apart from they are not usually produced in an industrial manner and have not therefore benefited from the economies of scale. Where is Henry Ford when you need him.
          Last edited by JJackson; May 3rd, 2008, 08:06 AM.

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          • #6
            Re: Tales from the front line

            this capacity problem - we had discussed it at fluwiki
            with no result.
            http://www.newfluwiki2.com/showDiary.do?diaryId=2334
            http://www.newfluwiki2.com/showDiary.do?diaryId=2419
            and other threads=diaries

            We do already have cell-based vaccine. Licenced and producing.
            This can be scaled up in a pandemic with the limit only being
            payment for the vaccine - or not ?

            We often see these capacity calculations and statements.
            Why ? Why isn't this question addressed and answered ?
            I wrote to Novartis, they wanted to reply ... but then refused
            when I said I were no reporter, just posting to birdflu forums.

            You just need more bioreactors, more personnell etc.
            for additional production. I think, this can be quickly
            achieved when/if panflu starts.
            Since years they had told us that we need cell based production
            because eggs might go out in a pandemic.
            Now we have it, but we still hear capacity concerns.

            Maybe new capacities currently have to be approved by some strange laws
            which are supposed to change in a severe pandemic ?
            I'm interested in expert panflu damage estimates
            my current links: [url]http://bit.ly/hFI7H[/url] ILI-charts: [url]http://bit.ly/CcRgT[/url]

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            • #7
              Re: Tales from the front line

              mutating away from the vaccine during production
              ------------------------------------------------

              Apparantly this had not happened in the previous 2 pandemics.
              When one strain suddenly appears with dramatically increased
              transmission - this is supposed to be a unique event,
              a reassortment probably. From that point we have a unique
              new strain which will mutate at typical rate from there on.
              Not as with seasonal flu where we already have much diversity
              and are just guessing which strain will dominate next season.
              I'm interested in expert panflu damage estimates
              my current links: [url]http://bit.ly/hFI7H[/url] ILI-charts: [url]http://bit.ly/CcRgT[/url]

              Comment


              • #8
                Re: Tales from the front line

                Gsgs thanks for the links. The first one in particular was excellent particularly as I had not seen the extract from Dr.Woodson’s forthcoming book which tied in very well with the three inter-related threads in this workshop.
                This one - which starts with the immune system and moves on to ways of boosting it.
                The WHO thread which starts by looking at the need for global oversight and guidance from the WHO and goes on to argue why it is structurally unable to fulfil the role we need from it. The second post then looks at the effects of past catastrophic events as instruments of societal change – sadly concluding, like Woodson, that our petty self interest will prevent us from taking the necessary steps until after the horse has bolted. The third post tries to relate all this to the Indonesia sample sharing debate with some proposed solutions in post #7.
                The third thread in this series Pandemic Plans & Hubris is an argument relating to the dangers of over interpreting the data on the 20th century pandemics echoed by Woodson’s concerns re pandemic waves.
                The longer term aim is – time permitting – to try and pull all these together with some practical suggestions on where we go from here and a continued discussion of the impediments to progress. (don’t hold your breath some of these post are already 2 years old)

                You suggested, and Dr.Woodson advocated, cell based vaccine production which I view as a major step forward but a generation behind Mabs. Both technologies are currently available I only favour Mabs because it gives more flexibility and has greater potential for speed and varying formulation to prevent strain drift.

                On the subject of strain drift you correctly point out that the seasonal vaccine matching problem is not due to the drift of one strain but because we do not know which of the circulating strains will be dominant in any given season. My comment on evolving away from case zero homology was really a continuation of the argument in Pandemic Plans & Hubris about underestimating the virus and lack of data on its rate of change. There was significant discussion a while back about the difference between the sequences emanating from different labs which were growing on samples from the same area. After a while it became apparent that at least part of the confusion was being caused by the rapid evolution of the virus in the labs dependent on that labs choice of substrate. The sequenced sample was not taken from the sample supplied to the lab (vivo) it was from virus grown in the lab from the original sample (vitro) and the labs choice of substrate avian or mammalian (MDCK) was effecting the result. While they were growing virus for sampling it was rapidly adapting to the host cells – hence the different sequences. I would also argue that the emergence of the Fujian strain in China is likely to have been as a result of its occupying the niche left by the suppression of those strains more closely matched to their poultry vaccine. This is obviously harder to prove but would be a logical conclusion based on basic evolutionary theory.
                Last edited by JJackson; December 15th, 2015, 05:31 PM.

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