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MMRF » 2018 Vaccine Development Conference #14: Targeting Tissue-Resident Immune Cells for Enhanced Immune Protection [2018-06-27. Laura Kate Mackay. HVP/USC]

2018 Vaccine Development Conference Session #14: Targeting Tissue-Resident Immune Cells for Enhanced Immune Protection [2018-06-27. Laura Kate Mackay, Stanley Plotkin. HVP/USC]

2018 Vaccine Development Conference – Session #14: Targeting Tissue-Resident Immune Cells for Enhanced Immune Protection — Laura Kate Mackay, Stanley Plotkin.


Organized in conjunction with the Human Vaccines Project, the 1st Annual Conference on the Future of Vaccine Development was a one day event which took place at the USC Michelson for Convergent Bioscience on June 27, 2018.

By bringing together some of the world’s leading scientists in the fields of immunology, genomics, bioinformatics, and bioengineering, the Future of Vaccine Development annual conference aims to explore how the convergence of new technologies across disciplines is impacting the future of vaccine development. The conference will also honor the three inaugural winners of the Michelson Prizes for Human Immunology and Vaccine Research, both via their respective presentations and the remittance of their prizes during the Awards dinner ceremony following the conference itself.

About the Presenter: As laboratory head and senior lecturer at The Peter Doherty Institute for Infection and Immunity, Dr. Laura Kate Mackay is studying a recently described subset of immune cells called tissue resident memory T cells, which combat various viral infections and cancer. The research that will be funded by the Prize will examine immune responses by tissue resident memory T cells to harness their protective functions to improve vaccines and immunotherapies.


  • Laura Kate Mackay, PhD, Laboratory Head and Senior Lecturer at The Peter Doherty Institute for Infection and Immunity (Doherty Institute) at the University of Melbourne.
  • Stanley Plotkin, MD, Inventor of the Rubella vaccine, also worked extensively on vaccines for polio, rabies, cytomegalovirus, and rotavirus. Former medical and scientific director for Pasteur-Mérieux-Connaught Vaccines in Paris (1991-97), executive advisor to the CEO of Sanofi-Pasteur.

Stanley Plotkin: I said that the speakers are all my friends or friend of my friend, and Laura Mackay comes from the Doherty Institute. Peter Doherty was a colleague of mine at Wistar. Anyway, she is from the University of Melbourne as well as the Doherty Institute. She is senior lecturer at the University of Melbourne and also Howard Hughes Medical Institute, and she has an independent group at the Institute studying cellular immune responses and she is the recipient – this sounds very interesting. She is the recipient of the Victorian Young Tale Poppy Award. You’ll have to explain to us – oh, a tall poppy, excuse me, what a tall poppy is.

Laura Kate Mackay: It’s just an Australian science prize. Okay, thank you so much for the introduction. Tale poppy, yeah, tall poppy. So thank you so much. It’s such an honor to be here and be a recipient of this prize. I’m really, really, very, very greatly honored. So today, I’d like to talk to you about some of my research which centers around T-cells. My group and my career for a long time has really focused on two main questions. How do you generate good T-cell memory and also what are the flavors of memory T-cell subsets that give us the best type of immune protection?

And so we know when a T-cell first sees a pathogen or an antigen you’ll get this huge effector T-cell pool here, and then this pool will contract leaving memory T-cells that will protect us, and of course, this is the cardinal feature of vaccination. And really when we think about memory T-cells, for many years, many people have really just studied the T-cells that circulate throughout the body, our effector memory T-cells or our central memory T-cells.

But in the past almost decade now, this has just come into the textbooks in the last edition is that there’s a third subset of memory T-cells that we call tissue resident memory T-cells or TRMs, and these memory cells reside exclusively within the tissues and they don’t exchange with the circulation. A lot of groups are now working on these TRM cells and it’s really now relatively well accepted that if you want to get really efficient mucosal immunity, you want to boost T-cell responses in your tissues and not necessarily just boost the number of T-cells that you have circulating around the body, and a lot of our questions ask within this effector T-cell pool that we boost when we vaccinate or immunize, how can we gear these effector cells to becoming more of these tissue resident-like memory cells rather than these memory cells that circulate throughout the body.

So just to give you a little bit of background on these T-cells, they’re a long-lived memory T-cell subset, they’re sequestered in tissues, and they were first found at really high densities at sites of previous infection. They’re distinct from memory T-cells that are found in the circulation. They have unique surface markers that allow us to identify these T-cells in the tissues, and over many years, we’ve been identifying the transcriptional identity of these T-cells saying what makes them different, what makes these T-cells survive in an autonomous fashion independently from those in the circulation.

A number of years ago now, we published that if you compare the transcriptomes of T-cell that circulate throughout the body both in the spleen and also in naïve T-cells that haven’t seen antigen, they cluster independently from these T-cells that are embedded within tissues, and the T-cells that are embedded in tissues, we isolate from the gut, the skin, and the lung, and they were embedded after various forms of infection. For example, to generate TRMs in the lung, we immunize with influenza. To generate them in the gut, immunize with LCMV. This is in the mouse, and in the skin, we immunize with herpes simplex virus.

We’ve also shown more recently that really these resident-like cells that are embedded within the tissues are actually more similar to other immune cell types such as AKT cells or tissue-resident AK cells as ILCs as compared to circulating memory T-cell subsets and we’ve identified a number of genes which we’re working on now which really identify these T-cells as being very unique.

So one of the cardinal features or hallmark features, if you like, of these T-cells is that they’re in disequilibrium with the circulating T-cell pool, and now these studies are really moving into the human realm and what we found is that if you look at facial transplants, for example, you can find these T-cells that are embedded within the skin tissue that have been found in patients carried across and they’ll be found several years later and they’ll be from original donor origin.

Also, many groups now have shown that these T-cells have a really important role in mediating local protective immunity against infections. In the last year or so, it’s been shown that these TRM cells are important in mediating immunity against cancer and there are also many studies now showing that having all these T-cells in the tissue is not necessarily always a good thing and they can mediate various tissue pathologies in the skin such as psoriasis or eczema, and they’re also linked to many autoimmune conditions.

So many studies in the mouse over the past say five to eight years have shown that if you immunize a mouse with various agents, you’ll get a high density of these TRM cells in the area in which the pathogen will replicate, for example. So again, he’s using influenza as an example. If you infect a mouse with influenza, you’ll get a high density of these TRM cells in the lung. In the liver, you’ll get a high density of these T-cells in the liver if you immunize with malaria.

Now the important point here is that if you embed these T-cells within specific organs of the body and then re-challenge the mouse with that specific pathogen, if the T-cells are with the right pathogen specificity, you can get enhanced local immunity specifically by embedding these T-cells in the tissue. Moving into the human space over the past few years, it’s now been shown that if you isolate pretty much all tissues now from all over the human body, you can identify these TRM cells as well. It’s a very similar surface phenotype to what we found in mouse and also we’ve done transcriptomes in TRMs isolated from human tissues and a lot of the genes that we found match up importantly within humans.

But of course, why does this miss for so many years? And this largely comes down to the fact that it’s what you can get access to, and of course, peripheral blood is largely what you can get access to, and to get a variety of tissues from a lot of human donors, Donna Farber at Columbia has been a real pioneer of this and she’s got a great cadaver cohort where she can isolate TRMs out of a range of human tissues.

So our group has shown that TRMs protect against local infection and if you infect with herpes once and you come back a second time, you can get site-specific infections, specifically at the region of skin where you’ve seen the pathogen before, but we started to move into saying how can we utilize these TRMs in a vaccine-like strategy, and so to do this, we’ll take naïve animals and we’ll embed pathogen-specific T-cells directly in the skin, and these memory T-cells, if they’re activated in addition then we introduce them into the skin, a fraction are able to enter the epidermis. They’re able to become these tissue resident memory T-cells and they’re able to persist pretty much for the lifespan of the mouse.

So one of the important facets here is that these T-cells, they don’t need to recognize antigen to persist within the skin. It’s a unique micro environment of the tissue that’s really critical in keeping these T-cells happy and it’s the micro environment that keeps these T-cells staying put.

So we embed these T-cells within the skin and they’ll look something like this. So these are two of the hallmark surface markers for TRM, so if we look in the skin, these are of the same antigen specificity. I’m just getting on transgenic T-cells here at TRMs express CD69 and the integrins CD103, the T-cells that we isolate from the circulation don’t. But we’ve got various tricks in the lab and here we’ve really been trying to get at what subset of T-cells are mediating the local protection that we can see against herpes so we can do various tricks whereby we’ll ablate all T-cells from the spleen. We’ll just have this population of resident memory T-cells in the skin, and we can also go the other way around where we can ablate T-cells out of the skin and introduce high numbers of circulating T-cells.

And then we take these naive animals that have pathogen-specific T-cells in different locations, infect these animals with herpes for the first time, and what we find is that it’s only when you’ve got resident memory T-cells in the skin you’re able to protect against skin disease. This is really a feature of if you want the T-cells at the right place at the right time. It’s nothing to do with numbers. You can keep on boosting circulating T-cells up to really high numbers but if you want to block a herpes virus infection which will go silent in the dorsal root ganglia incredibly quickly, you want to stop this virus in its tracks and you want the T-cells really there at the front line to be able to mediate this protection.

And the protection that we see is incredibly linked to the amount of T-cells that you can embed within the tissue, so this is just increasing the number of T-cells that we can embed within the skin, challenging here and looking at viral load within PFU, and the more T-cells you can get in the skin the better when it comes to protection against disease.

More recently, we’ve shown that TRMs can also protect against melanoma, so this is a really similar setup to what I’ve shown you before. We have B16 engineered with a herpes virus epitope so we can use it conjunction with transgenic T-cells here. We embed specific T-cells within the skin, deplete T-cells out from the circulation as shown here so you’ve just got your TRMs in the skin. We challenge with melanoma and what we find is that this is the takedown with tumors in naïve animals and here we’ve got both T-cells in the circulation and within the skin. We’ve depleted T-cells from the circulation and you can see that these T-cells embedded within the skin are able to protect against melanoma.

And what we find if we keep taking out these cohorts now and then we ablate our T-cells in the skin and we challenge with melanoma, 30 days later, we then ablate TRMs from the skin, what we find now is that these mice now start presenting with tumors, showing that it’s the TRMs that are contained within the skin which are mediating this cancer immune equilibrium.

So what about patients? There have been a lot of studies coming out in the past couple of years now. Now we’ve got the surface phenotype and the transcriptome of these TRMs showing that in various cancers, patients do better if they have T-cells that have this TRM-like phenotype, and we’ve shown this recently in breast cancer and in collaboration with clinician Sherene Loi at the Peter MacCallum Cancer Institute in Melbourne, we’ve done sequencing on various tumor infiltrating lymphocytes from breast cancers. This is getting on CD3+ T-cells and you can see there’s this unique cluster here that resembles the CD8 TRM cells that we see in mice. And again, we’ve developed a transcriptional signature here and what we find is that triple negative breast cancer patients that have a higher density of TRMs or have this TRM-like signature, this is indicative of an improved prognosis against breast cancer.

What we’ve found recently is that within mice and also within humans, if you gave on T-cells that are embedded within the tissues within circulation, and this is CD8 T-cells with exactly the same antigen specificity, and you look at all the checkpoints here, what we find is that all the checkpoint inhibitor markers actually screen much higher on these resident T-cells than on T-cells within the circulation, and if we isolate these T-cells out and we block with anti-PD-1, for example, it’s actually the resident T-cells that actually are really able to unleash that block and these T-cells are able to produce a lot of gamma in response.

And what we found again in humans, and this expression is both in humans and mice, but again, going back to our cohort of breast cancer patients, and again, it’s these patients that have a high amount of this TRM-like signature that actually are better responders against anti-PD-1 therapy.

So one of the major questions that drive us considering this proof of principle evidence that there are these T-cells that are embedded within the tissues and these T-cells are really linked to getting better immunity against infection and cancers is how can we regulate these TRMs specifically? How can we enhance their lodgment in tissues and how can we harness their protective function? And TRM differentiation is a really complicated process. We’ve been working on this for a number of years and it’s not just an effector T-cell or a memory T-cell that happens to wander into the tissue. These T-cells are really very distinct and they have really unique features and hallmark features that enable them to survive independently within the tissue.

There are a number of different molecules that are involved which we’ve shown over the years, but in short, an effector T-cell, you have to be within a short window of activation to become one of these TRM cells, so an effector T-cell that is circulating throughout the body, that infiltrates throughout the skin, the first thing that this T-cell needs to do is it needs to block tissue retention, so you need to be retained within the tissue first. We’ve tracked many genes over the years as T-cells come into the tissue and then get up into the epidermis, for example. Some genes that are shut down such as S1PR1, this makes sense. This molecule needs to go down for T-cells to be embedded within the tissue and also to shut down tissue egress. There are also genes that need to be induced once you get into the tissue to enable their adhesion and survival specifically within this compartment.

Now of course it’s not a one size fits all situation. T-cells are embedded within different tissues. There are similarities. There are also differences. At the moment, we’re really concentrating on the similarities between TRMs in different sites, but the microenvironment, for instance, there are different features between skin TRMs or gut TRMs, for example, but there are different tissue-specific requirements for the recognition of local antigen in the environment and also different cytokines that keep these T-cells happy.

So going back to transcriptional signature, we’ve been working through a number of genes over the years and we’ve been honing in on certain genes that are uniquely expressed in resident T-cells and these are resident T-cells at a wide variety of tissues and also in innate lymphocytes that also establish residence as well, and here’s just one example here to show proof of principle. So there’s a transcription factor called Hobit, which is exclusively expressed in resident immune cells, and what we find if we take CD8 T-cells and we enforce them to express Hobit using a retrovirus, and this transcription factor is not expressed by T-cells that circulate.

It’s not expressed in effector T-cells or those that are circulating throughout the body, but if we make CD8 T-cells express it and then we look at the lodgment in different tissues, compared to T-cells that circulate either in the blood or that are found in the spleen, when you overexpress Hobit and then look at your recovery of CD8 T-cells, this is 14 or 30 days later, no difference on the number of CD8 T-cells that you’ll find throughout the blood or the spleen. You’ll get increased TRM lodgment in the skin, the liver, and in other sites.

And just to show you an example going the other way, so here this is a gene S1PR5, which is important for tissue egress, and we take CD8 T-cells in a similar fashion. We enforce these T-cells to permanently express S1PR5 and what we can find is this exclusively affects the lodgment of our TRM cells. You can see again, no effect on the recovery of cells that we recover from the spleen or the blood. We’re able to dislodge our TRM cells from different tissues and now we’re working with different antagonists against this molecule to try to enhance TRM cell lodgment.

So this schematic really kind of sums up what we’re trying to do in the lab at the moment, so when a DC first primes or pool the effector T-cells, we’re looking within this T-cell pool. We’re looking for precursors of resident memory T-cells and the sort of genes that we can identify here. One thing that we have found is that for a T-cell to go on and form these resident T-cells rather than T-cells that circulate throughout the body, it’s actually T-cells that undergo fewer divisions when they first see DCs for the first time. It’s also T-cells that don’t express molecules such as KLRG1 and CX3CL1, so we’re able to exclusively gate on T-cells that are going to go on to form this fate and then T-cells will go up into the skin tissue, for example, and switch on other molecules, and CD103 is a prime example of that.

In some of the genes that we’re finding they’re expressed early which can identify T-cells that will go on to a resident memory T-cell fate as shown here, and we’re doing lots of single cell analysis tracking genes from the precursor stage up into commitment when these T-cells are long lived within the tissue, and then there’ll be a second wave of gene imprinting within the tissue which exclusively comes on to T-cells once they’re embedded within the tissue and we’re tracking these gene changes.

And one of the things that we’re tracking in here, this is just GP fate analysis of single cell within the lymph node, is we’re tracking T-cells within the lymph node at the earliest time after these T-cells see antigen and we’re seeing a commitment divide here where T-cells will go on to form effector T-cells, or effector memory T-cells, or resident memory T-cells, and we’re trying to identify these genes now and try to boost these genes to try and get more T-cells to commit to this TRM cell fate.

So just to wrap up what I’ve shown you today, I’ve shown you that these TRM cells, they mediate local immune surveillance and provide immunity against a wide range of infections and various malignancies. They’re also associated with a better prognosis against various cancers. Importantly, they display a distinct developmental program. They have a distinct identity as compared to circulating T-cells, and so this really needs to be factored into the sorts of immunization strategies or adjuvants that you might use because if you just use seeing T-cells in the blood is your readout, you actually might be boosting the wrong sort of T-cells if you want, for example, really great mucosal immunity in the female reproductive tract.

These TRM cells, they have distinct requirements for their formation or survival and also we’re able to manipulate some of these TRM-specific genes to enhance their lodgment, which will likely lead to improved immune protection, and this is something that we’re working on at the moment.

So just to thank everyone involved in the work. This is my team. We’re relatively new. The melanoma studies were all carried out by my Ph.D. student, Simone Park, in collaboration with Tom Gebhardt, my mentor, and Frank Carbone, who’s given me absolutely terrific support. Sherene Loi at the PeterMac was my collaborator on the breast cancer project, and thanks so much for your attention.


Stanley Plotkin: That was indeed a tall poppy, but so perhaps with your work on resident memory cells, you could clarify for us the difference between the inactivated influenza vaccine and the LAIV. There really is no established correlative protection for the LAIV, although one assumes that mucosal responses have some role, but perhaps this could be done in ferrets, actually, that is using the inactivated vaccine, the LAIV, to try to show whether resident T-cells are the difference. Well, without going into a lot of detail about LAIV, it certainly is problematic at the moment.

Laura Kate Mackay: Yeah, you make a really good point and something to bear in mind of course is that this field and the description of these T-cells is still incredibly new, and so really kind of looking at these T-cells in any sorts of trials is only just starting to be done. Only the proof of principle in the mouse system, if you have enhanced T-cells in the parenchyma of the lung or in the nasal tissue, for example, will give you better protection against influenza. That only came out say 18 months ago, so this is incredibly new and I think that moving forward, I think maybe ferrets might be a really good way to go because of course you need access to the tissue and you need to identify whether it is these T-cells that are making the difference, and I think we’re going to be seeing that in the next few years that people actually now we can identify them. We can say this is a resident cell and this is an effector T-cell which wane over time. I think we’re going to see those studies coming out relatively soon.

Stanley Plotkin: Other questions for Dr. Mackay? Yes.

Audience: This is really exciting stuff. The switch from effector to TRM, I have a detailed question, which is let’s say it’s malaria programmed TRM from the liver. Is it happening because it’s surrounded by liver cells or is it happening because of the microbiome of the liver and cross communication from the microbiome in these fibers?

Laura Kate Mackay: Yeah, so I know a couple of groups that are starting to work on the microbiome in the gut and the skin. In the liver, I’m not sure, but the way I see it is there’s a two wave of programming. There’s a commitment which is made when you see antigen for the first time and this kind of preprograms whether you’re going to become a short-lived effector or an effector memory T-cell which also doesn’t live very long. It’s interchangeably used for the resident memory T-cells sometimes but they’re completely different. They wane. And then there’s the second step of commitment once you’re in the tissue, and I think that there are interactions with different cell types within the tissue which are really important on imprinting the second part of the program in keeping these T-cells happy within that tissue.

Now the communication between different cells within a tissue for being important and maintaining these resident memory T-cells, really there aren’t many good examples that are being published yet, but a lot of groups are working on it at the moment of what cell types are important within a tissue, which are important for keeping say a malaria T-cell happy within the liver, and the studies on malaria came out last year as well showing that say within a mouse, embedding more T-cells in their liver gives you better protection against malaria, and that was only last year as well. So now kind of dissecting it down to who makes the decisions in keeping these T-cells happy in the tissue hasn’t been done yet, as far as I know.

Stanley Plotkin: Marcello.

Audience: Very nice stuff. The question that I have based on everything that was said, what would be the relative proportion of activity of a given pathogen that has been circulating in memory cells and the T resident cells, because you should expect that whatever circulating cells are in the tissue at any particular time, that that specifically also have a host.

Laura Kate Mackay: Absolutely. Yeah, that’s a really great question and I absolutely don’t want to oversell resident memory in any way because you absolutely want both, and the optimal protection is having both. Of course now there’s a problem with circulating memory T-cells will wane over time, at least the CD8s that have the ability to keep trafficing into the tissue. If you don’t have repeated antigen stimulation, that’s where the TRMs can come into play, but one of the major protective functions of TRMs is actually secreting a lot of gamma which will recruit more circulating T-cells more rapidly into the site of infection, so that’s one of their key mechanisms is bringing in a lot more circulating T-cells, so you absolutely want both.

Stanley Plotkin: All right, well, thank you very much.

Transcript curation: Alison Deshong

Dr. Laura Kate Mackay, winner of the 2018 Michelson Prize for her work in vaccine research and human immunologyDr. Laura Kate Mackay – 2018 Michelson Prize Winner for Human Immunology and Vaccine Research.

Dr. Laura Kate Mackay; Targeting Tissue-Resident Immune Cells for Enhanced Immune Protection. 1st Annual Conference on the Future of Vaccine Development. [2018-06-27] {#401} (Credit: Marv Steindler / Steve Cohn Photography)Dr. Laura Kate Mackay addresses a question from the audience while demoing her presentation ‘Targeting Tissue-Resident Immune Cells for Enhanced Immune Protection’ during the 1st Annual Conference on the Future of Vaccine Development. [2018-06-27] {#0401} (Credit: Marv Steindler / Steve Cohn Photography)

Dr. Laura Kate Mackay, winner of the 2018 Michelson Prize, attends The Future of Vaccine Development Symposium DinnerWayne Koff, PhD, President / CEO of the Human Vaccines Project presents the 2018 Michelson Prize for Human Immunology and Vaccine Research to Laura Kate Mackay, Laboratory Head and Senior Lecturer at the University of Melbourne for her studies of a subset of immune cells called Tissue-resident memory T (Trm) cells which reside in the skin and combat various viral infections and cancer. [2018-06-27] {#0700} (Credit: Marv Steindler / Steve Cohn Photography)

Francis Carbone, PhD, Dr. Laura Kate Mackay, Michelson Prizes 2018; The Future of Vaccine Development Symposium Dinner. {#0719} [2018-06-27. Marv Steindler / Steve Cohn Photography]Francis Carbone, PhD, Professor in the Department of Microbiology and Immunology (DMI) at the University of Melbourne and Mentor of Laura Kate Mackay, 2018 Michelson Prize for Human Immunology and Vaccine Research winner, pose together to celebrate the win of Dr. Mackay during the dinner awards ceremony. [2018-06-27] {#0719} (Credit: Marv Steindler / Steve Cohn Photography)

Dr. Patricia Therese Illing, Dr. Ansuman Satpathy, Dr. Laura Kate Mackay, Michelson Prizes 2018, Dr. Gary K. Michelson, Wayne Koff, PhD, Ian Gust AO, Dr. Steve A. Kay; The Future of Vaccine Development Conference. {#0005} [2018-06-27. Marv Steindler / Steve Cohn Photography](Left to Right) The 3 winners of the 2018 Michelson Prize for Human Immunology and Vaccine Research: Dr. Patricia Therese Illing, Dr. Ansuman Satpathy and Dr. Laura Kate Mackay pose with Dr. Gary K. Michelson, Founder of the Michelson Medical Research Foundation, Wayne Koff PhD, Ian Gust AO of the Human Vaccines Project and Steve A. Kay, Director of the USC Michelson Center for Convergent Bioscience, for a group picture ahead of the 1st Annual Conference on the Future of Vaccine Development held at the USC Michelson Center for Convergent Bioscience. [2018-06-27] {#0005} (Credit: Marv Steindler / Steve Cohn Photography)

Ian Gust AO, Dr. Ansuman Satpathy, Dr. Laura Kate Mackay, Dr. Gary K. Michelson, Wayne Koff, PhD, Steve A. Kay, PhD, Dr. Patricia Therese Illing, Michelson Prizes 2018; The Future of Vaccine Development Conference. {#0011} [2018-06-27. Marv Steindler / Steve Cohn Photography]Ian Gust, AO (Human Vaccines Project), Dr. Ansuman Satpathy, Dr. Laura Kate Mackay (2018 Michelson Prize winners of the Human Immunology and Vaccine Research Prize), Dr. Gary K. Michelson (Michelson Medical Research Foundation), Wayne Koff, PhD (Human Vaccines Project), Steve A. Kay, PhD (USC Michelson Center for Convergent Bioscience) and Dr. Patricia Therese Illing (2018 Michelson Prize winner of the Human Immunology and Vaccine Research Prize) pose outside the USC Michelson Center for Convergent Bioscience ahead of the 1st Annual Conference on the ‘Future of Vaccine Development’. [2018-06-27] {#0011} (Credit: Marv Steindler / Steve Cohn Photography)

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