SEGMENT 1: Dr. Warren J. Leonard talks about his basic research and its potential applications in clinical settings.
NOTE: This transcript was edited for clarity.  

Dr. Gary H. Gibbons, Director, National Heart, Lung, and Blood Institute:  Hello, I’m Gary Gibbons and today my guest is Dr. Warren Leonard, Chief of the Laboratory Molecular Immunology and Director of the Immunology Center at the National Heart Lung and Blood Institute. Warren, welcome.

Dr. Warren J. Leonard, Division of Intramural Research, NHLBI:  Thank you.

Dr. Gibbons:  It’s our pleasure to have someone that’s part of our intramural program, and I want to thank you for taking the time to share with us some of your thoughts today. First off, could you tell us a little bit about where your interests in your current line of investigation really began?

Dr. Leonard:  Our work really began years ago with studies of the Interleukin-2 Receptor. IL-2 is a hormone of the immune system known as a cytokine, and we discovered – after early work on the IL-2 receptor that there were three components to the receptor –  that mutations in the gamma chain of the receptor that was cloned by Sugamura’s group in Japan resulted in X-linked severe combined immunodeficiency in humans, which many know as the Bubble Boy Disease. That discovery then led to a combination of both clinical and basic science advances, including that the gamma chain was shared by what now is known to be a total six cytokines [IL-2, IL-4, IL-7, IL-9, IL-15, and IL-21]. We work on that family of cytokines [known as the common cytokine receptor gamma chain family of cytokines], which broadly influence the development and the biology of the immune system. 

Dr. Gibbons:  Terrific. I’ve noticed that you, as a superlative basic scientist interested in immune systems, that throughout your work, there have been these intersections where some of your fundamental discovery science does have these links to clinical context, just like the one you just described as part of your early work. How did that come about, that interface between what you’re doing at the bench in discovery science and how it may relate to a clinical context. Is that serendipity or–

Dr. Leonard:  The original connection to X-linked SCID [severe combined immunodeficiency] was indeed serendipitous. We were working on basic aspects of interleukin-2, which already was appreciated to be clinically important, based on work of others.  But it really came from chromosomal localization of the gamma chain to the locus that had been defined as the defective locus in X-linked SCID that leapfrogged us to discovering the cause of a human-inherited immunodeficiency – so that we discovered the cause of the disease, probably a couple of years ahead of when people who were working on that problem in a directed fashion would have succeeded. This is one example that underscores the importance of pursuing basic research in biologically important areas, because that basic research, in addition to elucidating basic biological mechanisms, often leads to important clinical advances. We then, in what I would call a back-and-forth fashion between basic science ideas that we perceived from our original findings and clinical manifestations of SCID made predictions that led us to discover the basis for three other forms of inherited immunodeficiency. Coming out of those studies, in part, were other studies--  that we did together with John O’Shea—that resulted in our hypothesis that agents that inhibited JAK3 would be immunosuppressive. There are now, as you know, JAK inhibitors that are approved by the FDA, including for rheumatoid arthritis.

Dr. Gibbons:  Could you expand a little bit more on your work on the JAK/STAT sort of pathway and those relationships to disease if you could?

Dr. Leonard:  JAKs and STATS are signaling molecules. JAKs are kinases and STAT proteins are signal transducers and activators of transcription. They are activated by interferons and molecules that are type-1 cytokines, as well as certain other growth factors. They are very rapidly activated by tyrosine phosphorylation and translocate to the nucleus, where they act as transcription factors to modulate the expression of target genes. That pathway is extremely important for many growth factors, cytokines, and hormones-- not all, but many, and the JAK-STAT signaling pathways are important for mediating biological responses. For cytokines, such as those on which we work, these pathways have important ramifications within the immune system. When one interrupts the actions of important immunological cytokines, there are, for example, immunosuppressive effects, and indeed, ways of blocking cytokine actions are also being pursued for treatment of cancer and a range of other disease processes. 

Dr. Gibbons:  Could you tell me a little bit more about perhaps some of your more recent work related to interleukin-21 or other things that are active in your lab right now?

Dr. Leonard:  Sure. Interleukin-21 is an extremely important cytokine that was discovered in 2000. My own lab was involved in the first cloning of the IL-21 receptor. At the same time, another group cloned the receptor and also cloned the ligand. So a new field was basically created where, to be honest, it wasn’t clear that people thought that a system was missing, but it [discovering the IL-21 system] immediately nevertheless plugged holes and began to provide more information. IL-21, like many other cytokines, is pleiotropic, meaning it has a broad array of actions. It acts on multiple immunological lineages, it’s critical for terminal differentiation of B cells to plasma cells, it is extremely important as well for the expansion of CD8+ T cells, and has a range of key biological actions. 

I think the excitement related to IL-21 that interfaces at the clinical realm is in two areas. One, it is an anti-cancer agent, based on a number of animal studies, and is now in phase 2 clinical trials in humans. Additionally, based again on animal studies, but also on human correlative data, blocking IL-21 is important for blocking the development of certain autoimmune diseases. For example, type 1 diabetes, systemic lupus erythematosus, and autoimmune uveitis are three examples where when one looks at the development of disease in animal models, if one does so on an IL-21 receptor deficient background, disease will no longer develop. Therefore, there is the general idea that administering IL-21 may be beneficial in some forms of cancer, and blocking IL-21 may be beneficial in some forms of autoimmune disease. Obviously there has to be a lot of emphasis on the “may be”, because these are not yet established clinical treatments, but there is cautious optimism that maybe administering IL-21 or blocking IL-21 will be useful treatments in the future. 

Dr. Gibbons:  Have there been some insights gained from again, either rare mendelian or more common? Do you watch those studies that relate to that axis in autoimmune disease?

Dr. Leonard:  In autoimmune disease yes. There are a number of autoimmune diseases where the disease locus [by GWAS studies] has been related to either IL-21 receptor or IL-21, but very often it really is IL-2 or IL-21 because they’re adjacent genes and those studies normally do not distinguish which of those would be directly involved.

Dr. Gibbons:  Could you tell us – given this line of investigation that you’ve been systematically engaged in – any recent findings that either intrigued or surprised you recently in some of your work?

Dr. Leonard:  One of the studies that I think is particularly interesting that we engaged in, as did three other groups, was a study where we came from the angle of interleukin-21 and others came from somewhat different approaches. Our studies began from studies of IL-21-mediated gene regulation, where we had shown that a transcription factor, IR4, and STAT3 were involved in the regulation of a range of genes and that many of those genes had composite elements such that both factors [IRF4 and STAT3] were required in concert. However, a puzzling thing was that IR4 was known to act in B-cells in concert with a factor called PU.1, which generally is not expressed within the T lineage. But yet we knew that IRF4 was acting within T cells, and so we needed another partner, and that partner turned out to be BATF, which is an AP-1 family FOS-like protein that acted with JUN family proteins. We discovered, as did the other groups – Dan Littman, Harinder Singh, and Ken Murphy – that these factors [i.e., BATF/JUN family proteins and IRF4] act together in concert to mediate the actions of IR4 within the T-lineage, which is different from how IR4 is acting within the B lineage. What I think is very exciting about that from a basic perspective are the fundamentally different mechanisms of how transcription factors work within different lineages, and from a more clinically applicable perspective, there may be different ways of targeting the actions of IR4 and thereby, for example, IL-21, in a lineage-specific fashion.

Dr. Gibbons:  That’s very intriguing. Are there other sort of accessory co-factors or scaffolding proteins or interactions that further cement that lineage specificity to the signaling or any other players that you’re–

Dr. Leonard:  It’s still an area of active investigation, but we’re far beyond the ideas of early studies that single factors did everything. I think it’s universally appreciated that factors are generally acting in a setting where other factors are also acting together, of course with epigenetic considerations as well, whether chromatin is open or closed at particular loci, and in part that will depend on the lineage. Some genes are expressed, even if certain factors are present, only in certain lineages. 

Dr. Gibbons:  That’s fascinating. If we step back a little bit, Warren, you’ve again been exploring and really setting the leading edge in your field. As you sort of look forward, maybe even beyond your particular findings right at the current time, what do you see as particularly exciting horizons in the space that you’re in in terms of immunology or some of the areas that you’re exploring? What’s sort of exciting that you see on the horizon?

Dr. Leonard:  I think that we are really just scratching the surface of new knowledge in many areas, and the “we” there is a “general field we” rather than limited to my individual lab. What I mean by that is we are appreciating how multiple things connect together along the lines of, let’s say, what we just discussed about multiple transcription factors. We’re understanding a lot more about the players that are involved. Obviously next generation sequencing has provided a wealth of data regarding the range of sites to which transcription factors bind in an in vivo setting, and RNA seq[uencing], even better than microarray expression, has been teaching us more about the range of expression of genes in response to appropriate stimuli and in lineage-specific fashion. Collectively, those data, both in primary cells that are studied in the lab as well as from patients with a range of diseases, are providing a tremendous amount of information, both at the basic science level and that may – and hopefully will – be translated into better understanding of disease and better treatment of disease.

Dr. Gibbons:  Perfect. As you know, the NHLBI portfolio includes a number of chronic disorders – heart disease, lung disease, in particular in which increasingly it’s appreciated that there's a major immune component that we often talk about – proinflammatory disorders – in sort of loose way. Could you say something related to the emerging sense of immunology and how it might translate to a number of disorders that are part of the NHLBI portfolio?

Dr. Leonard:  Certainly. I think immunology probably relates to diseases of every organ system. In particular, related to pulmonary, there is a wealth of data related to asthma, which obviously is a major increasing problem, where immunological-related modulations and immunological basis for the disease are certainly components. For atherosclerosis, there certainly is an inflammatory component related to the development of atherosclerotic plaques and foam cells/macrophages, as well as immunological components to other inflammatory disorders of the cardiac system. I think that there is evolving a greater appreciation all the time of the connection of the immune system to cardiovascular and pulmonary disease and, of course, regarding blood, there are blood diseases that are autoimmune such as ITP, and moreover, malignancies of the hematologic system include of course lymphomas and leukemias, where immunological considerations are important not only in the therapy but also potentially in the evolution.

Dr. Gibbons:  Could you say a word about what I believe is some of your work that links pneumonia and the work that you’ve been doing in the interleukins?

Dr. Leonard:  We did a study with a virus called pneumonia virus of mice or PVM, which causes a disease in mice that is similar to some severe aspects of disease of human respiratory syncytial virus. The viruses are highly related to each other, and what we demonstrated in mice in the PVM system is that interleukin-21 mediates the pathological response to the virus, so that disease is diminished in animals lacking IL-21 receptor. Moreover, if one pretreats animals with an IL-21 receptor Fc fusion protein, one can block the development of disease. Hopefully some of those lessons learned in PVM will have applicability to the human disease as well.

Dr. Gibbons:  Very intriguing.