Peter C. Doherty (2015) - The Killer Defence

Thank you. This is a very diverse audience, and my subject, immunology, is a very messy and complicated subject, half of which is usually wrong. I'm going to give you some personal reminiscences, some science, and some other stuff. I started in life as a ... I trained in veterinary science. I trained to be a veterinarian. I was going to save the world by producing more food by doing research. Of course, if I was doing that now, and I realize that cow's bells chime in there all the time, I probably would have become a plant scientist rather than an animal scientist. I did research on diseases of domestic animals for about the first ten years of my career. Worked on sheep, and cattle, and pigs, and chickens and became one of the world's leading sheep neuropathologists - a very small field, but still I was a neuropathologist. Then, I was supposed to go and work ... I was working in Scotland. I was supposed to go and work for the big government research organization in Australia, the CSRO, and took a bit of time out to learn a bit of immunology. Made a big discovery, never got back to the veterinary world, and became an MD, a mouse doctor. That's been my career, and now I'm mostly at the University of Melbourne but I also work at Saint Jude Children's Research Hospital in Memphis, which is a wonderful paediatric cancer place. If you're at all interested in that, they've got lots of money for post-docs and it's really ... You might think Memphis is a pretty weird place to work, but it's really actually quite nice because it's very nice socially for the post-docs. I work with two teams of bright young people in Memphis and in Melbourne. They allow me to get money for them, to help rewrite their papers in English and, occasionally, to talk about their work - though I may get it wrong. I think the time that you're most likely to make a real discovery is the time when you're very close to the data. And it's often only in those rather junior academic years that you're very, very close to the data. Of course, some of us as senior scientists side stay very close to the lab, and so we still have that opportunity, though most get off into more managerial roles and all the rest of it. It's a great privilege to work with smart young people. The killer defence. The immune system is all about protecting us from pathogens. That's why it's evolved. We're large, complex, multi-organ, multi-cellular systems. We replicate very slowly, and the various pathogens that want to live in and on us, the bacteria, the viruses, the protozoa, replicate very quickly. They can change very quickly, and we have to have a very complex immune defence mechanism to get rid of them and to control them. There's nothing predictable about it, because anything can be thrown at it. New things come at us all the time, and that's one of the things we encounter constantly, is that there's always a potential for new, major infections coming out of wildlife, in particular. Also out of domestic animal species, but the new ones tend to come out of wildlife. And with all environments being under increasing pressure, especially in countries like Africa, with forest clearance, increasing population numbers, we're seeing constant emergence of pathogens from nature that we hadn't seen before. And that's been going on and it will continue to go on. Years ago people were saying the year of infectious disease was over. That was before HIV/AIDS, before antibiotic resistance and all the rest of it, and we're constantly challenged. There are those totally new infections, but our biggest known pandemic threat is always influenza. And the reason for that is that the influenza A viruses change very quickly. They change by mutation of the surface glycoproteins, the hemagglutinin, and the neuraminidase, which are selected by antibody-mediated pressures, and all lab vaccines are basically antibody-directed. They bind to those surface proteins, the purple one and the peaky ones on that little diagram. And so that changes all the time. Influenza has a genome with eight different segments, and if a cell gets infected, say with an influenza virus from humans and an influenza virus from a bird, and they're basically diseases of aquatic birds, they're maintained in nature in aquatic birds, then you can get a totally new influenza virus out. And that can cause a pandemic. We get regular pandemics, the worst being the 1918-19 pandemic, where we think 50 million people died. It had a dramatic effect, helped bring the First World War to an end, probably led to a really lousy settlement of the Treaty of Versailles, and probably led to the Second World War as well. So flu causes everything. The reason flu is so dangerous is because when we get infected ... It's very infectious. When we get infected we don't necessarily feel ill, and we get on an aeroplane, and we cough and sneeze and we spread the virus. You're not at particular risk from the air handling system on the plane. The flu doesn't go through the whole plane, but if you're in two or three rows from someone with influenza, then you're at increased risk on the plane. And if you're on an aisle seat you're at increased risk on the plane. Make of that what you will. Flu flies from city to city and people, and when people land, they go out, they spread the flu. We are the vectors of influenza, whereas mosquitoes are the vectors of the Chikungunya virus, which is a virus that's currently spreading all over the place, and we don't know why, and so on. Then, of course, we now know there are a whole lot of viruses carried by fruit bats. We didn't know that. If you had asked any of us back in the year 2000 how many viruses are carried by bats, we'd say rabies virus, vampire bats, South America, occasional lyssaviruses in fruit bats and insectivorous bats. An occasional case in England, we've had some in Australia. That's the same group of viruses. Now we know there's a whole spectrum of viruses carried by fruit bats. The first that emerged was the coronavirus. SARS in Southeast Asia. It caused a major epidemic, also spread to Toronto, so I suppose it can be called a pandemic. It comes out of fruit bats into a little animal called the Himalayan civet cats, and then gets into humans. If you saw the movie Contagion, which is about this type of virus, Gwyneth Paltrow catches it in Hong Kong. She shakes hands with a chef who has dressed a pig that's been infected by a fruit bat. Gwyneth gets kind of sick but she's not so sick that she doesn't have a bit of a liaison on the way back to America, and anyone who comes near this woman gets infected and dies horribly, and gives an enormous pandemic. Gwyneth has the top of her head taken off after she's dead. It's not real, it's a plastic one, a plastic Gwyneth. It's really quite a good movie. It didn't do well at the box office, it's too realistic. Now, of course, we have the Middle Eastern respiratory virus. Again, the same thing we think. Coming from fruit bats, they multiplied up in camels, and then going into humans, but also gong human-to-human. And of course now we've got an outbreak in Korea, an outbreak in Thailand, and it is getting around. It should be something we can handle reasonably easily because we understand the pathogenesis of it, how it works. We understand that people are infectious very late, which we didn't understand early on with SARS. Then, in Southeast Asia we've got henipaviruses, nipah virus in Thailand and Bangladesh. These are viruses that go from fruit bats to fruit, because fruit bats eat fruit. But then if that fruit is eaten by a pig, the partial remainder of the fruit is eaten by a pig, then they get infected and they can infect us. Also, if humans eat the sap that comes out of some of these trees where they cut it for sap and get sweet sap, the fruit bats feed on it. If we feed on it we can catch it directly, and the nipah virus can spread. Then, of course, the most famous bat-borne viruses that we've known about for a long time but didn't realize were carried by bats, are Marburg and Ebola. Of course, we also let the Ebola outbreak we had recently, a truly horrific outbreak, and basically because ... We should have been ready for it. We've always handled Ebola before, there have been regular outbreaks, simply by getting a lot of people in there quickly, practicing barrier nursing, and the local population understood what needed to be done. But then it got into West Africa, where people didn't know the disease. The international agencies didn't react quickly enough, WHO and so forth, CDC didn't get there quickly enough, and we had a really bad outbreak, which isn't totally over. We had an Ebola vaccine that was ready to go ten years ago. We had antivirals that were ready to go. But there's a real problem in that these, which were developed through the basic science mechanisms in places like NIH, they have no way of taking it back and saying, good for human use. That has to be taken up by a commercial company. Can't expect a commercial company to take up something for which there's no real market. So we've got a problem there, and we should be able to think about it. And we should be able to do this better by providing resources to develop vaccines and antivirals against potential threats, to the stage where they're ready to go into production and they're ready to be used. We should be doing that, but in these international agreements and international will, and we forget very quickly. We also need to keep our public health systems very good, and in these times of cost-cutting and so forth, that's a real issue. Now, immunity itself. Immunity, an extremely complex defence system. You've heard about part of it from Bruce Beutler and Jules Hoffmann. I'm not going to repeat any of that because they've discoursed on it so magnificently. The word immunity itself comes from the Latin, which comes from immunis, which means, "without tax". And it refers to the fact that soldiers who came back from the Rome wars were for a time exempt from tax. They're called the "Genio Immunium," just like rich Republicans in the United States, they pay no tax. They're there to prevent or to block the tax of infection, and of course the tax of infection can be death, and that's where we get "immunity". Immunology, I've been working in immunology now for forty-plus years. We've used immunology firstly to try and better understand the immune response to viruses, to develop better protective mechanisms, and to understand the pathology. Because the reason a virus kills is not just the effect of the virus, it's what happens in the tissue. Tissue damage, tissue repair, particularly in a very gentle and sensitive organ like the lung, the tissue repair process can be lethal. We don't necessarily know where to intervene in some of these mechanisms to get better clinical outcomes. But we're pulling it apart very quickly with work on the inflammazone, with knockout mice and all these sort of things. That pathogenesis, pathology thing, is to me enormously satisfying. We're actually starting to understand that now, that we haven't understood it for years. All specific immunity, and in fact pretty much all immunity ... well, not totally. All adaptive or specific immunity, which is the antibody response or T-cell response, is a property of white blood cells and their secreted products, that it's only there by white blood cells. The other thing that we've done over the years, is not to study the immune system itself, but use the immune response to viruses to actually study some of the basic biology of the system. And these are questions that are prominent in basic biology itself. Homeostasis, cell numbers, population size. The immune system, when you thing about it, we have two great systems for sensing the external environment. We have the brain, which obviously is in the skull and is a pretty stable organ, and we have the immune system, which is all over the body. We've got cells that are migrating everywhere. They increase in numbers, they increase in size, they invade into tissues. It's a totally fluid and dynamic system. And it also senses the external environment, not of the conscious level, of course, of the subconscious. They're the two systems we use to handle in a specific way, a very specific way, to handle new challenges and new stimuli. The other thing about the immune system, the T-cell system at least that I studied, I'll show you very, very briefly. It's great to study in differentiation, because we can get lineages of the cells with the same receptor that we can trace through from what we call the naïve state, before they've encountered any pathogen, where there are very, very low numbers, but they're there. To the effective state, where they're all armed, and angry, and ready to bump off any infected cell, and so forth. To the memory state, where they're ready to be recalled for protection more rapidly, which is of course the basis of memory. We can study those differentiation systems, we can study them in vitro. Of course, the perception of something external that's damaging in the immune system in the brain is a bit different. When you learn ... memory in the brain of course is a matter of connecting pathways. Memory in the immune system is a matter of cell division, increasing cell numbers, and also of differentiation and epigenetic change and all the rest of it. Now, if every time you had a thought you increased the numbers in your brain, you'd get terrible headaches, because the brain is so restricted by the skull. Just a little bit ... I'm kind of fascinated by history, and ... The red blood cells were actually seen by Leeuwenhoek way back when he first made those little microscopes. But it was another 140 years before we saw the white blood cells discovered, and that was because of better microscopes. It was a pathologist who discovered them, a couple of pathologists who discovered them. Not pathologists, the surgeons who discovered them. In 1840, surgeons, all they could do was cut legs off very quickly, and drain pus from abscesses and things. So they had to do something with their time in the meantime, when everyone's running all over, going around. And he actually described the colourless corpuscles. The white blood cells, which are rather infrequent, which is why Leeuwenhoek didn't describe them, though it does seem ... I've just gone back and read his paper in Philosophical Transactions of the Royal Society, the world's oldest journal in the English language, now available online. And it does seem that he actually did see the white blood cell but he didn't recognize it for what it was. They were called "colourless corpuscles" because there were no stains. And they could see these colourless corpuscles or globules, they called them, at very low frequency, in the blood and in the pus, and so the ones in the pus have come from the blood, which is absolutely correct. They could also see inside the colourless corpuscles other corpuscles or globules. That was the cell nucleus. Nobody had realized what the cell nucleus was. Then, a little bit later, with the German aniline dye industry developing dyes and Rudolph Virchow, the great pathologist, and Paul Ehrlich, the immunologist, they started to stain cells. And then we started to see all these cells and started to understand the immune system. Macrophages, basophils, all these spectacular-looking cells. The actual white blood cells that are important to us, the lymphocyte, are the least interesting to a morphologist because they're tiny. They just have a nucleus, therefore almost no cytoplasm, and the lymphocytes weren't worked out until the 19... We didn't start to work them out till the 1960s with the work of Jim Gowans and then Jacques Miller identifying the T-cell, Henry Claman and Bruce Glick identifying the B-cell and B-lymphocyte. Or Max Cooper and the T-helper cell and the killer T-cell and all the rest of it. This is the basis of immunity. We start with a very few naïve precursor cells with a specific receptor, they multiply under the stimulus of the infection. We get cell division, we get differentiation, we get the effector cells which clear out the infection. All of those cells die off and then we get it in memory, which goes on for ages. For the life of a laboratory mouse or for up to fifty years in humans. That all goes in those lymph nodes in our neck, which swell up when we get sick. We're very interested in making better vaccines. We don't have a vaccine against HIV. Influenza, we'd like not to have to make a new vaccine every time an infection came along. We despaired of making a vaccine against hep-C, but now we've got a very good therapeutic and we can clear it with a drug. Of course, if you can handle a persistent infection with a small molecule, that's really fantastic. We handle HIV, of course, with drugs but because the thing hides back in the genome, you've got to keep giving the drug, and you've got to give triple therapy. It's really problematic from the cost point of view, especially in developing countries, though it's a tremendous advance. The CD8+ "killer" T-cell that I study is the kind of legionnaire of the immune system. It's the guy that comes up to the cell that's infected with a virus, because viruses only grow within cells, and they have to be bumped off in short range. So it comes up to it and bumps it off. It's a legionnaire. The legionnaire carried a very short, stabbing sword, and they had big shields. They'd come up close and they'd go like that. The dog is an immunological joke because there's nothing funny about it. There's nothing funny about jokes told by immunologists. Here's the killer cell. The green cell is going to kill that big cell, and when it does it also kills the virus inside it. You can see it permeablises the cell membrane in this red dye in the fluid, and that's kind of dead. That's what these T-cells do. These CD8 or "killer" T-cells, they invade the tissues, they go where viruses are growing, bump off the cells, cure the infection. They're highly specific. They don't actually kill, they actually turn on the cell's death path way in the cell. They cause apoptosis, so they're actually inducers of suicide cells rather than killer cells. And their recognition is very specific. And it goes through the fact that a small piece of the bar as a peptide is bound into the cleft of our transplantation molecules, the ones that are recognized in graft rejection. And that's carried to the cell surface and these cells recognize that. I don't have time to go into all that detail, but that's the way it works. We made a very superficial discovery way back in the 1970s. We made this discovery before there was any recombinant DNA technology, so you couldn't sequence small amounts of protein from cell surface. You could sequence bucket-loads of, like, myoglobin or something. We made it before molecular antibodies, before PCR, and it was a pretty primitive time. We made some good guesses, and we drew this hokey diagram that's on the far side there of the circles and things. And we wrote two 2-page letters in The Lancet, and a 5-page theoretical article in ... two 2-page letters in Nature, a 5-page letter in The Lancet in 1974-75. And we got the Nobel Prize years later after many, many people had worked on it and we got all the credit. That was a very good resolution. And that's me with my colleague Zinkernagel. That's after the Nobel Prize in 1996. We waited 23 years for the Nobel Prize, a fairly normal time to wait for a Nobel Prize. That photograph illustrates which surface ... much later. This was an honorary degree ceremony. It illustrates two things for you, and that is, one, that if all men aren't fools they can be made to look like them. And the other thing is men have absolutely no idea what to do with flowers. We're both standing there, there was no hand-off. There's usually a hand-off. You go to Asia, you give a lecture, they give you flowers, there's a nice lady, you hand them to her. But there was no handoff and we're stuck with them. We'd like to throw them away, but instead we're standing with them in front of our crotch. I don't know what actual thing was. The fact that we could do this work very quickly and we got a real resolution, was a consequence of the fact that we had just been able to tap into mouse transplantation and genetics. And that was thirty years of work particularly started by George Snell, who shared the 1980 Nobel Prize for transplantation, and there's a whole history there. I make that point, we would have gotten nowhere without having those mouse strains available, mutant in the H2 system compatibility area, and in recombinance in the transplantation area. Another point from it is, Zinkernagel and I did this work together as very junior people, and we were complementary. Rolf did the in vitro cytotoxicity experiments, I did the mouse stuff because I was a vet, and I also wrote the papers. If I had done the in vitro experiments and Rolf had written the papers, we would have remained totally unknown and completely obscure. Of course, with regards to George Snell and his colleagues, we reflect Isaac Newton's statement, That was in a letter to Robert Hooke. Actually, he was having a go at Hooke, he hated him. Hooke was very short, that's why he was saying "giants." And not only that. Towards the end of his life, when Newton was president of the Royal Society, he outlived Hooke and he had the one portrait of Hooke destroyed. That's real malevolence. Not all senior scientists are totally nice, so be careful of some of these Nobel Prize winners. They seem nice, but you're never quite sure. That's a recreation of what Hooke looked like. Nobody knows what the guy looked like. Not a problem there. We all get painted after you win the Nobel Prize. If you don't win all the Nobel Prize you don't get all the prizes usually after the Nobel Prize, unless you're incredibly smart, which some of these people are. I'm not, but you do get painted. The other thing is, you get things named after you. This is a street in my hometown, they've named the street in my honour, and the building in the background is Boggo Road Jail. Now, the killer T-cell response is very interesting, because a lot of the peptides that it recognizes come from internal proteins, and those proteins are shared between a lot of viruses. So it can get cross-protection. I'm running out of time, so I'm not going to be able to tell you what most of my slides are about because I talk too much. But we can make these tetrameric complexes of MHC like a protein, and peptide, and we can use them to quantitate the T-cell response, which we can only do from about 1997. And we can also use them to sort T-cells and actually look at their molecular profiles by using RTPCR, seeing what message is expressed in them. And even if we don't have an antibody to sign the cells of its protein, we can do single-cell PCR straight out of a mouse or straight out of a human. We've got an enormous amount of information very quickly. We've been able to study clonal lineages, clonal expansion, and recall responses. We've been able to study how clonal lineages survive by tracking through the T-cell receptors in these responses. We've developed that technology. We've been able to look at when the various effective molecules that did the killing are turned on. That's totally tied to cell division. Every time the cells divide they turn on more of these effective molecules. They turn most of them off actually afterwards, and we've been able to work all those things through. Now we're studying the actual epigenetic signatures that define these different cell lineages. The kind of question then in an intellectual sense is, what are these cells and what characterizes them, and can we really tell them apart and are there subdivisions of them? And that we're rapidly working through. The other important question, of course, is, can we optimize T-cell memory so that it would be very good for recall responses, because there is a capacity for cross-reactive responses between, say, different flu viruses. They can share the same internal proteins, though they may have totally different cell surface proteins, and so we can get some cross-protection. And here exactly is ... we can show that in a mouse, and we've shown it for years. Here is an experiment with the H7N9 influenza outbreak we've had recently in China, where an H7N9 chicken virus, which is an infection that's completely inapparent in birds, which is really dangerous, can kill humans. Particularly it kills older people, particularly it kills older men. Because when they retire the women send them out to buy the live chickens from the chicken market to get them out of the apartment, and they get infected and die. What we've seen, and the wonderful collaboration that Katherine Conzias, my colleague, has had with a group in Shanghai, is that the people who turn on the CD8-T-cells most quickly, that is the red cells in that diagram, they're the ones who survive. These are all people who were admitted to the hospital and they were quite sick, but if they turn on that response quickly they survive. If they don't turn it on fast, it takes longer to emerge, then you see other components in the immune system coming up. And that's a theme across immunity. If one part of it doesn't handle it, then other parts will get emphasized. The most would be, everything turns on very strongly and the people that die, they're just lacking the whole thing so that it doesn't work. We think there may be some possibility for some cross-reactive immunity, and possibly some sort of vaccination strategy that's working around that in combination with other effectors, other mechanisms. There's a lot of interest on that. That's about where we are. Much of what we're finding with the "killer" T-cells and the virus infections is probably applicable to some forms of cancer. Renal cell carcinoma, melanoma, we've long known, are under a degree of immune control, and of course, if you follow the recent experiments with anti-PD-1, anti-CLT-4, releasing those cells from apparent suppression of some sort, allows them to work. We're getting for the first time over the last few years real progress in the cancer immunotherapy area, which is really fantastic. As for me, I am still involved with the science, I still talk to the scientists. I've stopped running a lab, I'm now 75, and I've stepped back from that but I'm still involved in the research programme. I've been writing books. I think Steve Chu showed a baby picture. The one on the tricycle is me at age three. I've never looked better in my whole life. It was just downhill from then, and I've written other books, which increasingly I'm like many people at this meeting. I'm extremely perturbed about the situation with climate change, and I've been reading into that. I've been writing books about it, writing from an outsider point of view, but I'm not insider. Largely I can read the biology papers, but I can't really handle the mathematical modelling. And the latest book is a bit on writing about medical science from an insider perspective and about climate science from an outsider perspective. And try to talk to people about, "Hey, you can look at these sorts of issues," and that's going to be out later this year. I would like to say to you that I think science communication is enormously important. Not in the sense of telling people stuff. People are sick of being told stuff. There's too much stuff that comes top-down, and that's a bit of a problem with books. But you young guys that use social media, and YouTube, and you're generating great video material and all the rest of it, start a conversation out there if you can. Get things going as a sort of discourse, and put great video material up on YouTube. And just use that medium to try to have some connection, also try to connect with school kids. The other thing is, in Australia we just started this thing called theconversation.com. Take a look at The Conversation. You can sign up for it, get it free to your mailbox. It's in the UK, it's in the USA, Australia, and in Africa. You just go to theconversation.com, you put in your email address, it will deliver the content every day, like an online newspaper, to your blog. What it is, it's a thousand-word articles written by people who have an academic affiliation. You can sign up and write for it. You have to declare a conflict of interest. You can write really good stuff about what you know about, the things of general interest. It will then go through a professional newsroom, and they'll go backwards and forwards with you on that line. It's a very sophisticated platform. They'll add some nice illustrations, and your article that you publish in this can be picked up by any newspaper that wants to republish it. It's all free. They can pick it up for free, and a lot of it gets republished. And you can get metrics on how many times it's been republished, how many references have been to it, and all the rest of it. It's really good. You don't have to be in one of those countries to sign up. All you have to do is be affiliated with an academic institution. You can sign up for it anyway. And it's a way to start honing your skills in science communication with the broader community, and starting conversations, because there are responses and all the rest of it, with a broader spectrum of people. And getting also a view of how the general public is receiving and reacting to science. I'll leave you with that. Thank you very much.

Peter C. Doherty (2015)

The Killer Defence

Peter C. Doherty (2015)

The Killer Defence

Abstract

Immune surveillance by virus-specific CD8+ cytotoxic T lymphocytes (CTLs), or killer T cells, has long been known to be central to the control of acute infections and some cancers, though the role of CTL memory in the rapid recall of immune protection has been less clear. Focused onto the surface of virus-infected cells by the T cell receptor interaction with self MHC class I glycoproteins presenting non-self (viral) peptides (p), the availability of pMHCI tetramers from the latter part of the 1990’s has allowed us to both quantitate naïve, effector and memory CTLs and to explore their qualitative status by probing profiles of gene and protein expression. Apart from suggesting possibilities for novel immunotherapies and vaccines, the CTLs thus provide a fantastic system for probing two major issues, the nature of differentiation in immunity and the characteristics of homeostasis and clonal longevity in this extremely fluid and dynamic system.

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