Melanie Cole, MS (Host): Welcome to Better Edge, a Northwestern Medicine podcast for physicians. I'm Melanie Cole. And today, we're highlighting increased autoreactivity and maturity of EBI2+ antibody-secreting cells from nasal polyps. Joining me is Dr. Bruce Tan. He's the Irene and Walter Wayne Templin Professor of Oral Pathology and Associate Professor of Otolaryngology, Head and Neck Surgery, and Medicine in Allergy and Immunology at Northwestern Medicine.

Dr. Tan, it's a pleasure to have you join us. Can you explain as we get into this really interesting topic why it's important that there are increased numbers of cells producing antibodies and anti-double-stranded DNA? I'd like you to explain a little bit about the antibodies in nasal polyp tissue and then the role of the EBI2+ antibody-secreting cells. Give us a bit of an overview so we could learn what we're talking about here.

Dr. Bruce Tan: Sure. Thank you very much for having me on this podcast, Melanie. A number of years ago, we had made this observation that unusually in nasal polyps, which is an inflammatory condition in the nose, we were seeing antibodies that were autoreactive, meaning that they targeted antigens that are typically found in yourself. And we had wondered where these antibodies might be coming from. The options were, were they coming in from the blood? Were they being produced locally within nasal tissue? We generally do not think of nasal polyps as an autoimmune disease, meaning we don't see systemic inflammation with nasal polyps. But we certainly see a lot of localized inflammation within the nose, especially within nasal polyp tissue itself.

And so, we started looking at the blood and we looked at nasal tissue, and we found that all these autoantibodies, which we use this anti-double-stranded DNA antibody as emblematic of, were found locally within nasal tissue and not seen systemically in the blood of patients with nasal polyps. We then started to look at which cells were the source of these anti-double-stranded DNA antibodies, and we had made an observation a number of years ago that there's this protein called EBI2 that's expressed on some B cells that are activated in an atypical manner called an extrafollicular activation and we have found that there were increased numbers of these EBI2 cells in nasal polyps and we looked at comparing whether or not the EBI2-positive cells compared to the EBI2-negative B cells, which was the more autoreactive pool of B cells, and we found that the EBI+ fraction of B cells within the nasal polyp tissue was the overwhelming source of autoreactive antibody production within nasal polyp tissue.

Melanie Cole, MS: So, how does this help to predict the recurrence of nasal polyps? How is knowing this about the autoreactivity help you to predict whether somebody is or isn't going to get these and have them recur?

Dr. Bruce Tan: One of the common treatments for nasal polyps is sinus surgery where we remove the nasal polyp tissue and we remove some of the surrounding inflamed tissue within the nasal cavity. It remains probably still one of the relatively first-line treatments for patients with severe chronic sinusitis with nasal polyps.

It is known that after surgery, some patients will have a recurrence of their nasal polyps, typically within a few years of having had that surgery. And that range of recurrence is variable, partially dependent on how far out you follow patients for. In our study, we found that at about an average of three years after surgery, the recurrence rate was slightly under 40%. We had Looked at predictors of this recurrence in a paper we published about two years ago. And we had looked at clinical predictors, things that a clinician could observe in biomarker predictors, things that you could measure in nasal tissue. And then, we use some machine learning algorithms to try and identify the best predictors for what predicted which patients eventually had recurrence of their nasal polyps.

And what we found was that amongst the biomarkers, there were three features that were predictive of polyp recurrence. Two of them related to this type of inflammation that we know drives nasal polyps called type 2 inflammation. But the third one, which was somewhat unexpected, was anti-double-stranded DNA, which is this autoreactive antibody that we had found previously and was also the topic of research in our current paper. And we found that these three biomarkers each had independent ability to predict recurrence after sinus surgery. We also have found some other predictors, such as presence of asthma, as well of highly severe radiographic score also added to the accuracy of the prediction.

And so, overall, we think that type 2 inflammation, overall severity of disease, but also this presence of autoreactive antibodies, as represented by this double-stranded DNA antibody are all predictors of how well a patient does after sinus surgery; thus, where these double-stranded DNA antibodies might be coming from was of relevance.

Melanie Cole, MS: This is very advanced technology and pretty exciting in your field, Dr. Tan. Now, speak about the characteristics of the cells themselves that are producing antibodies in nasal polyps and how are they different from cells in other tissues? What have you noted?

Dr. Bruce Tan: So, the cells that make antibodies in the human are obviously B cells and B lineage cells. But B cells are kind of complex cells to study because they kind of evolve into different phenotypes. So, they acquire features that make them unique as they evolve from being what they call a naïve B cell to a mature B cell or an antibody-producing cell.

And one of the features we had noticed about nasal polyps in the past was that a lot of the cells in nasal polyps are what we call antibody-secreting cells, which have historically been thought to be plasma cells, which are these antibody factories that just churn out a ton of antibodies. And so, we had seen that there were more plasma cells found within nasal polyps in the past. And we were then interested to know whether these were obviously the source of the majority of autoantibodies that we have found in these nasal polyp tissues.

Melanie Cole, MS: So, how did the composition of the B cells you were discussing differ between tonsils and nasal polyps?

Dr. Bruce Tan: We had studied this question in relation to tonsils because tonsils are lymphoid structures, and it is known that B cells typically undergo an activation process, that is in our lymphoid structures like your tonsils. So when you encounter an antigen or virus, for example, one of the places that the B cells respond to this new antigen and make cells that make antibodies to target this antigen are in structures like your tonsils.

One of the unique things about tonsils is that, in that environment, B cells get activated in very close coordination with another cell type called your T cell. And the B cells and T cells interact with each other to allow the B cells to mature and become antibody-secreting cells. But in that process, it is well known that the T cells really try very hard to censor autoreactivity or autoimmunity, meaning that they will kill off B cells that acquire ability to bind to self-antigens. And whereas in a nasal polyp, we do not see these lymphoid structures. We do not see that there's this B cell and T cell-coordinated dance that goes on. And so, what we found was that certainly comparing the B cells that you find in abundance in tonsils to the B cells you find in a nasal polyp. We see that the B cells in the nasal polyp don't have some of this phenotype of cell that interacts with the T cell called the germinal center B cell. We don't see those present in nasal polyp tissue. We see a lot more antibody-secreting cells in nasal polyp tissue compared to a tonsil, almost hundred fold higher numbers of antibody-secreting cells.

And so, our interpretation from this is that the environment in the nasal polyp allows antibody-secreting cells to be generated in abundance. And therefore, you've got these little factories of antibodies that are locally present within tissue. But they have features that don't suggest this tight interaction with T cells in their activation. So, perhaps that might be a mechanism by which these B cells are allowed to make antibodies that react to self; and therefore, these self-reactive antibodies are present in abundance within nasal polyp tissue.

Melanie Cole, MS: Then, what about the changes in certain cellular pathways in the cells producing antibodies for these nasal polyps? Are there changes in these pathways?

Dr. Bruce Tan: So, we compared the antibody-secreting cells using this technology called single-cell RNA seq. We compared the antibody-secreting cells you found in a nasal polyp, compared to the antibody-secreting cells found in a tonsil, and we found a few relatively unique features about them. One was that the antibody-secreting cells in a polyp were much more mature, which was interesting because these mature plasma cells in our immunity are known to live forever. It's the reason why when you got vaccinated as a child with the MMR vaccine, as an adult, you still have antibodies in most cases against these childhood vaccines because these plasma cells can live essentially for decades. And what we found was that the antibody-secreting cells in the polyp have features of maturity, meaning that they up-regulated some of the machinery needed for these mature plasma cells to persist for a long time.

Correspondingly, compared to a tonsil, they down-regulated some of the features that made them B cells, which is a known feature of these long-lived plasma cells. They basically lose their cell surface features that make them look like B cells. And so, that overall gave us this impression that these antibody-secreting cells within nasal polyp tissue were long lived, and mature type plasma cells.

They also had a few other interesting molecular mechanisms that made them tick. One was that we saw expression of some of the antibody genes that were known to bind to double-stranded DNA. We found these at higher levels of transcripts in the nasal polyp antibody-secreting cells compared to the tonsil-derived ones, suggesting that perhaps there was some favorable environment for selecting these autoreactive antibodies. And then, we also noticed that there were some other molecular mechanisms, such as increase in this NF-kappa B signaling, which perhaps may help these antibody-secreting cells become more mature and long lived. They also expressed a receptor for a type 2 cytokine called IL-5, which was unusual because human B cells and plasma cells were not known to express IL-5 until recently. There have been a few papers talking about these IL-5 positive plasma cells that are found in specific environments like the nasal polyp. In mice where IL-5 receptor had been first discovered, it is known that IL-5 does drive maturity of plasma cells. But it's been demonstrated that human plasma cells could express the IL-5 receptor.

Melanie Cole, MS: Dr. Tan, with this lesson on cellular physiology that we're getting here on this podcast today, and it's really interesting, take us from bench to bedside. How might these findings influence current approaches to diagnosing or treating nasal polyps and how do they contribute overarchingly to our understanding of the immune response, antibodies, and inflammation in respiratory conditions? Because it seems that it has even bigger implications.

Dr. Bruce Tan: The clinical implications are something that I think of much as a clinician. I think one aspect of this is certainly recognition that the B cells and plasma cells that live inside the nose and the airway may have relatively unique activation mechanisms that are independent of the ones that we are most studied in lymphoid organs. Thus, you may have antibodies in the nose that react to antigens, including autoantigens, that you'd have to really look inside the nose to find and measure and discover their presence. Thus, a blood test may not always accurately reflect what kind of antibody responses go on inside the nose. So, that's one aspect.

The second aspect is this study suggests this concept of tissue resident memory that is, I think, gaining importance in a number of inflammatory conditions where they see that immune cells in inflamed tissue may have its own ability to remember antigens, in our case, perhaps through the B cell and plasma cell compartments, such that because these cells can live for such a long time, that there may be mechanisms by which inflammation can persist because you have these local cells that can make autoantibodies. They can persist for a long time. And so, these tissue resident cells that have memory in their antigen recognition may play an important role in what causes these chronic inflammatory conditions of the airway to persist.

From a therapeutic perspective, there also are implications. There are a new class of drugs coming up in nasal polyps, some of which target this IL-5 pathway. In clinical trials, interestingly, they have been shown to be effective in reducing polyp size. But they aren't as effective as some of the other drugs that target the IL-4 receptor, but this study may implicate a different effect of these IL-5-targeting drugs in that, if these plasma cells that express IL-5 receptor need IL-5 in order to survive, then it may suggest that using these IL-5 drugs may potentially reduce or even eliminate some of these tissue resident memory cells that might cause disease to persist. And so, we need to do some further investigation around this question, but I would say that that is something that this study raises some potential therapeutic implications for.

Melanie Cole, MS: Interesting implications, as you say. Final thoughts, Dr. Tan, for other providers listening, what you would like the key takeaways to be from these studies.

Dr. Bruce Tan: That the antibody responses found within inflamed respiratory tissue can be very different from the antibody responses found systemically. That nasal polyps, although not a classic autoimmune disease, may have some manifestations of autoimmunity locally within the airway. And that understanding in the mechanisms that allow these local autoreactive pathology to exist within the airway might help us better treat patients with very severe disease such as nasal polyps or asthma better in the future. And I would say that is sort of the direction that I think this research takes us.

Melanie Cole, MS: Thank you so much, Dr. Tan, for joining us today and sharing your incredible expertise. To refer your patient or for more information, please visit our website at breakthroughsforphysicians.nm.org/ent to get connected with one of our providers. That concludes this episode of Better Edge, a Northwestern Medicine podcast for physicians. I'm Melanie Cole. Thanks so much for joining us today.