Transcript of ASCI Perspectives – Agbor-Enoh interview

Interview with Sean Agbor-Enoh, MD, PhD, NHLBI, NIH (elected 2023)

Interviewed by Patrick Nana-Sinkam, MD (elected 2019); member, ASCI Physician-Scientist Engagement Committee

Note: The text has been edited for readability by ASCI staff.

Patrick Nana-Sinkam: I’d like to welcome everyone to ASCI Perspectives. My name is Pat Nina- Sinkam. Today, I have the pleasure to have as our guest for ASCI Perspectives Dr. Sean Agbor. Dr. Agbor is Laboratory Chief, NIH Lasker Clinical Tenure-Track Investigator, NIH Distinguished Scholar, Lead of the Laboratory of Applied Precision Omics at the Division of Intramural Research at NHLBI. He’s a Lead Investigator and Director of the Genomic Research Alliance for Transplantation, Adjunct Associate Professor of Medicine, Lung Transplant Program, at the Johns Hopkins Hospital. He’s also the recipient of the 2021 James T. Willerson Award in Clinical Science. Dr. Agbor has dedicated his career to understanding genetics and genomics and how they are used to elucidate the pathogenesis and progression of diseases such as pulmonary arterial hypertension, COVID-19, and graft dysfunction post–lung transplant. In particular, his laboratory is really focused on cell-free DNA for the development of liquid biopsy biomarkers to inform clinical decision making in diagnosis, prognosis, and disease monitoring. Dr. Agbor, I’d like to welcome you to “ASCI Perspectives,” and I’d also like to congratulate you on your recent selection to ASCI. Thank you so much for joining us.

Sean Agbor-Enoh: It’s truly a pleasure. Thank you for having me.

PNS: So I would like to start out by maybe just asking you if you could share a bit with the audience your path or your journey to become a physician-scientist, and in particular, how did you decide to focus your line of investigation on transplant immunology?

SAE: So, I come from Cameroon; it’s in Central Africa. And this is relevant to why I became a physician-scientist. It’s in Central Africa — if you think about the map of Africa, where you come from Africa: Ghana, then Africa makes the turn to start going south. Right where it makes the turn, that’s Cameroon. Right where it makes that turn, that’s Limbé, where I grew up. Now, why is that relevant? On the other side of the Atlantic Ocean there is the highest volcanic mountain in western Central Africa, Mount Fako. I grew up there. Black volcanic sand. So part of the beaches are black. I tell you, it is . . . if you have time, go visit; it’s a very beautiful site in the world.

Now so when I was growing up, never in my mind thought of even becoming a doctor. We were surrounded by chemicals, and next to that area is SONARA [Societé Nationale de Raffinage], which is a petroleum company. So we grew up really poor. So you can imagine the rich kids in the area that you go to school with, their parents work for the petroleum company. So the dream for most young men growing up there, surrounded by rich volcanic soil, seeing money through petroleum, was to go towards chemical engineering or such fields. Medicine was the last thing in my agenda, truly the last thing. And so after high school I said, I’m going to find opportunities to go to the UK to study petroleum engineering, of course. And then my father of course didn’t have money to pay for me to go and said, well, if I give him about two–three years, he will raise that money. Now, he said, “In the meantime, don’t stay home. Why don’t you just apply to go to medical school while you’re waiting to go to the UK to start the petroleum engineering?” Most of my friends after high school went and did that. And so I went to medical school. I wrote an entrance exam. It’s a competitive — you write it, you get selected, you go. So I went there. But the first two years of my medical school was terrible. Grades? Oh, don’t even get me started. You need to see my transcript, because I was just . . . For me, it was just a place holder.

And then eventually I figured out my father: that was a trick, and I am going to just be in medical school. But I did . . . My thought was always petroleum chemicals until in my fourth year of medicine; it’s a seven-year program. We went to do an outreach activity for the WHO. When we went to this village, we walked about eight to twelve miles through the dense equatorial forest to get to these villages. We were giving Mectizan, which is a drug that WHO was giving at the time to treat filariasis. In one of the villages, we met the chief of the village, and he had a son. That son was about . . . They don’t have birth certificates there, but you could estimate somewhere between 16 and 20, and he was stuck in bed. When he sits up, he would get short of breath. Then when he lays down flat, he feels better. That just astounded me because I’m used to heart failure or other conditions that cause shortness of breath that you get better when you sit up. This child felt better when he lied down, Of course, I couldn’t figure it out until during my fellowship, many years later at Johns Hopkins. And I figured out what that patient had and indeed wrote a paper in one of the ATS (American Thoracic Society) journals about that experience: this idea of shortness of breath which gets better when you lie down. But that is the case that led me to, number one, be interested in medicine, that I could make a difference in a patient. Unfortunately, that young child died while we were there, and you can imagine, it hits you. I said, well, maybe if I knew a little bit more, I could help that child, that young man. And the young man died then. Then it just goes, the question is, what did this young man have?

The rest, I’ll tell you, it’s practical, it’s really history. After medical school, I got an opportunity, through the Fogarty International [Center], to travel to Georgetown and did a one-year research internship and just loved it; and so I decided to do a PhD. At the time, I did a PhD in biochemistry, molecular biology, and trying to clone a gene for malaria. And that got me interested in genomics. And so a PhD comes, residency at Hopkins, and everything. But then I started asking the question, how do I then apply what I have learned to address a major problem? During my last year of residency and my Chief year, I had an encounter with a lung transplant patient, and I started learning about the condition: the idea that lung transplant definitely saved lives. These patients would otherwise die if you do not get a lung transplant. So it’s a beautiful story. I loved it. However, unfortunately, within only five, six, seven years of transplantation, half — 50% — of these patients unfortunately die because of complications of rejection.

That’s number one. Number two, it amazed me that this transplantation is one of the only medical conditions where you take an organ from a different person, i.e., a different genome, and put it in another person, a different environment, and that genome needs to make a decision. The genome could adapt in that new environment and survive, or if the genome fails to adapt, it unfortunately would be rejected. So it marries, then, some principles of rejection, some principles of genomics that I learned in graduate school. And this idea of trying to ask why to address a major problem. There is the path to lung transplantation and to lung transplant research.

After my fellowship, I was literally barbecuing behind my house. I can tell I have many kids. We were barbecuing one Sunday, and I got a phone call, “Hey, my name is Hannah Valentine. I’m from Stanford. I want to set up a genomic research lab.” Guess what? In transplantation. I’m there, wait a minute, where is this coming from? And so Hannah Valentine called me and said, “Can you come from Hopkins to the NIH, join me at the NIH, and set up this?” And a wonderful mentor, just a wonderful person. And that interest just married together. And that’s where the pathway, the path of this is. The path has been traced way early in Cameroon. But that’s where I finally ended up, and that research has been ongoing since then.

PNS: That’s an amazing and inspiring story. And I’d like to use that to kind of delve into the question about your actual . . . the research that you’ve been doing. As you know over the last probably one to two decades, liquid biopsies have been a real focus for many, many investigators. And in many diseases, the liquid biopsy is considered almost the holy grail for diagnosis, for treatment decisions, prognostication, and so forth. Yet when we look at liquid biopsies, every group has their compartment of interest that they like to focus on, whether it’s microRNAs, proteins, extracellular vesicles, circulating tumor DNA. Yet your group has focused on cell-free DNA. Can you share with the audience a little bit about why cell-free DNA? What have you learned from the work that you’ve done with cell-free DNA, and how do you see cell-free DNA perhaps being the holy grail for liquid biopsy in understanding human disease?

SAE: Thank you very much. So cell-free DNA is one of the measures of liquid biopsy, but overall the concept truly amazed me while I was thinking about where in genomics I should approach transplantation. Because a lot of these models, you can approach them using genomics, microRNAs. Vesicles have a big content of nucleic acid. You could approach them in that. But why cell-free DNA? And I would give a plug about that and then talk a little bit more, in a perspective about all the other biomarkers, of my thoughts about how they would come together. So cell-free DNA are short DNA fragments that are released when cells die or they’re undergoing regeneration, and that ends up in body fluids. It could be blood, urine, CSF, and all that. In plasma, for example, in the healthy patient, there’s a 100 billion — this is billion with a B — 100 billion fragments of cell-free DNA, all coming from all over your body. That intrigued me. And this is DNA: DNA drives everything that each of your cells does. And it seems like this DNA, not only is your DNA sequence coming, but the epigenetic landscape of that cell where the DNA is coming from, that epigenetic fingerprints are preserved on cell-free DNA. Therefore, it felt to me that with just a little bit of plasma from a patient, could I sample that patient and trace what is happening in different organs in your body, almost providing a whole-body molecular scan of what’s happening?

So transplantation felt like a good place to start, because in transplant, again, you have genomic admixture. You have the genome of the donor, and you have the genome of the recipient. So in addition to these epigenetic fingerprints that I talked about — and I’ll come to that in a little bit — you have also single nucleotide polymorphisms. The transplant organ has DNA sequences or single nucleotide, but that are distinct from that of the recipient. So with plasma, therefore, you could use those SNPs, or single nucleotide polymorphisms, to track cell-free DNA coming from that allograft. And if DNA is released when an allograft cell dies, therefore, the amount of allograft-derived or donor-derived cell-free DNA could guide you into what is happening to that allograft.

So cell-free DNA then offered that opportunity. It is also indeed quite stable. They’re wrapped around nucleosomes. As you know, nucleosomes are these eight-mer proteins that are dense in positively charged amino acids like lysine and all that. So their positive charge on everything almost spells it — they’re so resistant to endonucleases and everything. As I tell my students in the lab, you can take cell-free DNA, literally put it on a stone, get a hammer, and hit it. It does not break down. So, a very stable molecule, very abundant in plasma. You can use plasma to almost derive what is happening in several tissue all across the body. And we started the story in transplant, and we showed some really key principles: that number one, it is sensitive. It can track injury coming from that organ, indeed. Indeed. It can pick up that a patient is getting rejection two to four months before the patient shows any clinical symptoms or before the biopsy becomes positive. That, I’m telling you, that just wowed me.

And so early detection — and we have studies now trying to figure out if early detection and early treatment of rejection guided by cell-free DNA would improve lung transplant outcome. As I mentioned earlier, these patients die within five to six years, half of them from complications of rejection. So those studies are ongoing. But then we backed off and said, “Wait a minute, could we test and try to validate some of these principles in other conditions, such as pulmonary hypertension, COVID-19?” And that story seems to be the same: that with these conditions, you can not only use this to detect complications early, but you can also use this to re-stratify patients for poor outcome. Like in pulmonary hypertension, you can re-stratify patients for transplant-free survival. Patients who will need a transplant or will die from their disease — you can identify them years in advance. With COVID-19, at the time of patient admission, you can re-stratify the patient to see which patient will need ICU care and therefore maybe do interventions a little bit early. So that is what cell-free DNA can do. This idea that it’s a stable marker in plasma, that you just noninvasively get a blood collection and you can sample different things happening in that organ. Now you can define tissue injury in pulmonary hypertension and COVID-19, for example. These are nontransplant conditions.

However, how do I know that there’s injury from the heart, injury from the kidney, injury from the liver? This is simply because while the DNA sequence in your entire body is identical: your heart pumps, your lung breathes and it’s an immune organ, your liver does all kinds of metabolism, your kidney does excretion. So how can the same DNA with the same sequence orchestrate such distinct functions in different organs? The reason is because each organ has epigenetic fingerprints that are specific to that organ. Those epigenetic fingerprints — DNA methylation, histone, chromatin footprinting — they are preserved on cell-free DNA. Therefore, if I get plasma from a patient, I scan the epigenetic fingerprints on cell- free DNA. Could I therefore assign the organ or tissue which is producing that cell-free DNA? So that’s the technology that we then transfer from, to test the principles that we learned in transplantation to study pulmonary hypertension and COVID-19 and a few other diseases that would be coming up. So in summary, that’s been the love for cell-free DNA.

Now, however, the other biomarkers in the field, the non-liquid biopsies, such as the microRNAs, the extracellular vesicles, I think these carry similar information, but I think each of them have some small added advantages. Now we have people specializing in different areas. My hope is that coming very soon, as we have learned about these molecular approaches, that at some point we’re having now the technology, huge computing, people doing machine learning, big data — that it is time for marriage, where, say, a cell-free DNA person gets, collaborate[s] with a microRNA or extracellular vesicles, and we see how these two approaches combine, better informed how to triage our patients and how to select appropriate treatment or re-stratify them. Or even better, further understand the mechanism of what disease is happening. And again, the long game would be that these approaches would help us identify what is happening in disease in a way that we do not need to go get biopsy samples from an organ. But could we identify what is happening looking at just blood and looking at these different approaches?

PNS: So I’d like to kind of just extend on that and ask you to perhaps reflect on and maybe even predict what you see as the next series of important scientific questions to ask and to hopefully answer. So for those early-career investigators who are thinking about perhaps a career in transplant immunology and maybe even studying lung transplant — recognizing that we’ve made tremendous progress, but as you said earlier, the outcomes in lung transplants still tend to lag behind some of the other solid-organ transplantation. So what do you see really as those important scientific questions that we need to be asking and answering to start to narrow some of those gaps when it comes to lung transplantation outcomes?

SAE: So lung transplant truly has a need. I’m telling you, if there’s a branch of medicine that is in real need of a physician-scientist, lung transplant is. For most other diseases, you have to wait years before you see patients with poor outcomes. While it’s unfortunate for the patients, the idea that you can get some of these outcomes early and you can track patients over time to get these outcomes, is that an opportunity to understand what is going on in these patients? Let me be a little bit more specific. In all solid organ transplantation, the lung and the small bowel have the poorest outcomes. It’s not exactly clear why, but there’s several hypotheses, and that’s what we are all working on.

It so happens that these are organs that have just truly big lymph nodes. If you think about the small bowel, they have pouches underneath the gut epithelium. The lung has all kinds of lymphoid organs as well in there. Second, they’re exposed to the environment. They’re exposed to all the particulate pollutants and everything that is available in the environment. You have that. They’re available, they’re all microbial particles and all that. You have that. And then third, these patients are immunosuppressed. What a cocktail to trigger inflammation! In the lung, we’re looking at things like cell-free DNA. Lung transplant patients at baseline, whatever you can define as baseline: when they’re not having rejection, their cell-free DNA is about tenfold higher — tenfold higher — than in heart transplant patients. Cell-free DNA is a measure of baseline injury. Just think about how much lung injury these patients have compared to patients who have heart transplant. So there is true opportunity there to understand this interaction, what the environment contributes, and what inflammation contributes and the underlying autoimmunity or now allograft immunity — because you have an allograft, so you call it that — it’s indeed autoimmunity.

But this interaction between these three components that a lot of focus is on. Lung transplant provides that basic field. It feels to me that understanding those basic principles that drive these baseline injury patterns in these patients will help to better understand what is going on and how we can stop it. Not only that, not only that, we have also seen that by studying things like environmental particulate matters in lung transplantation: Would that help patients with asthma? COPD? Think about the gut, inflammatory bowel disease. The principles that you would learn from these rare diseases such as lung transplantation, these principles are potentially broadly applicable in other conditions beyond transplantation.

So this is a call to the young physician-scientists that are coming. That, in my mind — pick a problem that is passionate to you. Something that you wake up and you think about; something that in your mind, you believe is a problem. And then that devotion towards that mission to improve outcomes, at least that’s mine, is to put a dent to improve outcomes in those patients for whom lung transplantation is the only cure. That’s my mission. And now rejection and early detection is an approach that I’m taking now. My hope is we can make a dent. But do that, pick a problem that is passionate about, and then harvest that passion. As a physician-scientist — I’m telling you this idea of taking a problem that you see in clinic, bringing it to the lab, diving deep into it, looking at mechanisms, identifying interventions, going back to see how you can improve these patients: this is the holy grail of medical practice.

This passion I hope I can transmit to my colleagues who are coming up and thinking about this. It is just something that, I will tell you, if you develop that and pick the right thing that’s good for you, that you feel so strongly about, it is hard to not love it. And so it’s a field — we do have challenges: trying to find grants, trying to find dedicated time for research, trying to find so many components that we need to do this work. It’s hard. It’s really hard. There are lots of mentors that have done it. My people say — I’m from Cameroon, as I said earlier — If you walk on the shoulders of giants, you can see further. So mentors are just amazing. I would advise that they look for mentors.

Someone that truly, you can tell, that they just want you to succeed. They don’t have to be in the same field as you, but they can guide. And then my hope is that you can find those components, a mentor, a problem that you’re passionate about, and then the two together, bringing those two together, it would be likely that you’ll find the resources and the time to do it. Then call me back and tell me if you do not find joy just doing what you’re passionate about. So that is what I would leave as advice to the junior physicians coming up.

To big groups like ours or the group that I’m so excited and I’m so honored to belong and to come join: ASCI . . . I think we have started advocacy already. We are right at the forefront of advocating what we think is going help promote this track of physician-scientists. My hope is that joining ASCI, that will be something that I could also lend a hand. And as physician-scientists, we have to all continue that advocacy to try to see how we can make the bed easier for these colleagues of ours that are coming up with so much passion to do this. If we can give them the tools and help them to integrate their life and their work such that they can both live and do this so passionately, then I think we would be moving in a straightforward path.

So let me summarize that. For my colleagues coming up: passion, passion, find the problem with passion. Find a mentor or mentors. And truly a big shout out to ASCI, for an association like this, for advocacy. My hope is that we all can join in this advocacy to try to provide the microenvironment that will just make this feel richer. And it ends up benefiting everyone: the patients; the government, because we help save money and we do all these things. So why not?

PNS: I think that’s a fantastic note to end on. I certainly speak for all of ASCI in saying how much we appreciate and how fortunate we are to have you as a member of this organization. Your commitment, your passion, your scientific excellence are beyond reproach. And we wish you the very, very best. We’ll be looking for great things from your group. You’re certainly a role model, and we hope that there’ll be many more who follow in your footsteps. So I really want to thank you on behalf of all of ASCI for just taking the time to speak with me today.

SAE: No, thank you. I’m truly honored for the opportunity to give my perspective as well. Thank you.

PNS: Thank you so much.