Viremic – Cases in HIV

Announcer:
Welcome to Viremic–Cases in HIV, hosted by Dr. Eileen Scully and Dr. Christopher Hoffmann, both HIV specialists at Johns Hopkins, who explore quandaries in adult HIV care. Each case discussion includes medical history and diagnoses, challenges in care and treatment, and key evidence and guidelines that inform clinical decision making.

Cases are presented as a composite from the hosts’ clinical practice, with all identifying details removed to protect the privacy of patients. Case discussions are for informational purposes only and not offered as medical or clinical practice advice for patients or clinicians. Any mention of specific medications or commercially available products is a description of use only, not an endorsement.

Dr. Eileen Scully:
Welcome to Viremic, a podcast where we discuss all things HIV. I’m Dr. Eileen Scully, coming to you from Johns Hopkins here in Baltimore. And before we begin, a reminder that if you like this episode, please share it with a colleague and follow us wherever you listen to podcasts. We’d also love to hear what you think, so please do leave a comment or send us an email at viremicpodcast@jh.edu. And feel free to leave suggestions for future topics that you’d like us to discuss on the podcast.

Today I am so thrilled to welcome Dr. Francesco Simonetti. Dr. Simonetti originally trained at the University of Milan and then completed a PhD here at Johns Hopkins, but that is just a tiny piece of what he means to the field of HIV. He’s done really incredibly important studies on the HIV reservoir, on how it expands and contracts over time, and I think will be one of the people who really brings us closer to achieving an HIV cure. Francesco, thank you so much for joining us today. Welcome to the podcast.

Dr. Francesco Simonetti:
Hi, Eileen. Thank you for having me. The feeling is mutual. I’m extremely thrilled to be here today. I’m a huge fan of the podcast, so it’s an honor to be here.

Eileen:
Well, great. I think that’s one of the things that makes your research so impactful, which is that you continue to engage in the clinical realities for people living with HIV, which I think really informs your work in a beautiful way. Before we start our discussion for today, I was wondering if you could tell us a little bit about your career path to this point. I hope that some of the people who listen to this podcast are people who are considering infectious disease, either clinically or for research, and always want to tell them the happy stories of how we landed in these careers.

Francesco:
Yes, absolutely. So, I think there are two reasons. A scientific one—early on in med school, when we learn about microbiology, I was really fascinated by pathogen evolution and how it can shape our immune system. And so what a better system to choose: a highly evolving pathogen such as HIV and a highly evolving system as our adaptive immunity. And the second reason that drew me towards HIV was the fact that it was affecting mostly disenfranchised groups. And, you know, 20 years ago in Italy there was still so much stigma against people living with HIV, LGTBQ+ people, migrants, and so it really made it even more important to choose that as my focus for my career.

Eileen:
I will just reflect back that I always think that HIV is the most wonderful thing to study because it has that combination of the most interesting science and also the most compelling social justice considerations. So, I completely share that motivation.

With that backdrop, I’ll start us out with a case. We’ll use this hypothetical patient to explore some of the research that is relevant in how we think about talking with our patients in the clinic about what’s going on with their HIV. Today we’ll consider a 45-year-old man. He has a history of HIV, which was acquired approximately 15 years ago. He has been engaged in care with full viral suppression achieved after initiating antiretroviral therapy, which at the time was with efavirenz/emtricitabine/tenofovir disoproxil [fumarate]. This was when he was diagnosed. His CD4 [cell] count is over 500, and his nadir was about 300 at the time of his diagnosis, so he did not reach advanced immune suppression, which is great. Approximately six years ago, he transitioned to bictegravir/emtricitabine/tenofovir alafenamide and has continued on that regimen through the current visit. He has no other significant comorbidities, and he actually doesn’t take any other medications.

About a year prior to the current visit, he had a detectable viral load at 180 copies/mL. At that time, he denied any interruption in medications and reported stable and full adherence. He returned for an early follow-up about one month later, partially because he was as concerned about this viral load as his provider. And at that visit, he had a viral load of 210 copies. They did a very comprehensive review of medications—he wasn’t taking any still—and supplements, looking for vitamins or other things that could have affected his bictegravir. But those were all consistent with what he’d been reporting for years. His refill records were also reviewed, another way that clinicians can try to piece together whether or not someone’s missing medications, and he had regular refills throughout this time.

A subsequent viral load a few weeks later was again elevated, this time at 110 copies per mil [copies/mL]. And since that time, he’s continued with more frequent follow-up, with viral load checks occurring approximately every two to three months, and this is partially at his request. His viral load has remained between 80 copies at a low and 250 copies at a high with a stable CD4 [cell] count during this time. And, as previously, his refill records are appropriate, and he’s actually quite concerned about the detectable viremia. So, you have a relatively low suspicion about adherence being the issue. Francesco, can you tell me how you would characterize this type of a clinical scenario and what features of the history are most important to establish that assessment?

Francesco:
Yes. Well, the first thing I would say is that this is a persistent pattern rather than an isolated blip, which is an old term that we used to refer to these isolated detectable viral load measurements that are then followed by a return to suppression. Here we have a repeated detectable measurement for over a year. So clearly there’s something—whatever the mechanism underneath this positive viral load is—something that is sustained over time. And just as a reminder, the half-life of viral particles in blood is extremely short, and so for viremia to be detectable at multiple time points suggests that there’s something ongoing that also persists over time.

A few aspects that are very useful to look at in the history of a case like this that I would like to emphasize. Well, you mentioned some of them. We want to assess medication access and actual dosing. So, looking at refill records really help[s] to have a first initial assessment of adherence but of course cannot confirm 100% whether, you know, each dose has been taken. And as you well described, we also investigate whether there are other medications or supplements that may affect therapy. In this case, the patient is not taking mineral supplements, or you know, some of those other polyvalent cations that can reduce bictegravir absorption.

But again, the most important thing is the trajectory. A stable plateau of detectable viral load differs from a progressively, rapidly increasing viremia. This is also paralleled by stable CD4 counts. These are all a reassuring thing, even if they are not enough to establish the mechanism, which we’ll discuss in a second.

So overall, the relatively stable levels of HIV RNA in plasma over time, the long history of suppression before that, and the absence of an obvious issue in adherence raises the possibility for what we call nonsuppressible viremia [NSV]. Nonsuppressible viremia is this clinical scenario in which we have persistently detectable viral load that cannot be further decreased by optimization of treatment. And one important thing is that even though the pattern of viral load suggests NSV, the viral number alone cannot confirm whether this is due to release of virus, of replication, which is something that we should discuss in a second.

Eileen:
That’s incredibly helpful. And I just wanted to pause briefly on this idea of the viral load number itself. And so, with this concept of nonsuppressible viremia, are there limits around which you would say, “above that this no longer qualifies”? And where would you personally place that limit? Recognizing that this is not necessarily a clinical definition that we can put into action with all patients, but his viral loads have been 250 or less in our theoretical patient. And how do you think about the actual number?

Francesco:
Yes, so this is an extremely important aspect and, unfortunately, there are not standardized numbers across guidelines and clinical societies. For example, for the WHO, levels between 50 and 1,000 copies are those used to refer to low-level viremia, which includes also this scenario of nonsuppressible viremia. For the International AIDS Society, it’s everything that is above the limit of detection of the essay, which we know can vary based on the different clinic and geography, and it remains below 200 copies. So, our case for today falls exactly within this range: between 50 and 200 copies. Unfortunately, you know, viremia is a continuous variable, but we need to bin it into defined ranges so that we can provide operational definitions and guidelines.

Eileen:
I think that’s a really important point because for many of us who took care of patients when we actually didn’t have fully available ART to suppress everyone, we learned that if you put people on regimens that were partially effective, you could bring them down to a level, maybe get a half log or a log reduction in viral load, if not full suppression, with agents to which they were resistant. At some point, usually, that would break down, but it could be used as what we used to call a “holding regimen.”

Now, typically those would be much, much higher than in these low 100 ranges, and that’s a very different clinical scenario, where we do think there’s active replication that’s ongoing, we’re just sort of reducing how much. And I think you were alluding to this a bit in terms of thinking about what these particles are. Just to bring us back to our case, if we classify this gentleman as having nonsuppressible viremia, we could do some additional tests to feel valid in making that determination. One is to confirm that there’s no known or suspected resistance to the regimen that he’s on. And that would basically put us in a position of saying we don’t think there’s any added value of additional agents because the agents that he’s on are capable of interrupting the life cycle [of HIV].

In his case, also, it’s a departure from his prior clinical history. So previously he was durably suppressed and there was no immediate tie to a change in medications. It wasn’t like you just changed and this started happening, which can also occur, but in this case that’s not true. And we know that, as you said, by very definition, the nonsuppressible viremia actually can’t be suppressed with additional clinical interventions. We don’t have great evidence for anything working in this situation. And I was wondering if you could try to discuss a little bit about how we sort out what, about having this low-level detectable viral load, might be concerning to a patient and to their provider?

Francesco:
Yes, of course. You know, for someone who has been undetectable for years, seeing a number above the limit of detection again can feel like losing something that they work so hard to achieve. And since our first visit with a patient, we instruct them that achieving undetectable viral load is the most important goal of their treatment. And so, NSV brings a lot of concerns and anxiety. Patients will worry about their health, the risk of transmission for their partners, and whether that ongoing regimen that they are taking is going to fail. So, it’s very important, I think, to acknowledge all of these concerns even before we try to explain the biology underneath it.

Regarding the concern of virologic failure or development of resistance, in the past, larger cohort studies of low-level viremia or of levels compatible with nonsuppressive viremia have shown a signal of a higher risk for virological failure. However, these larger cohort studies lump together cases in which both viremia caused by the natural reservoir biology and ongoing replication are both present. Indeed, when instead we look at studies that were just on highly selected patients, at all the features of nonsuppressive viremia, usually we don’t see selection for resistance, we don’t see virological failures, and very stable CD4 counts as in the case that you described earlier.

Eileen:
That’s a really critical point. And I’ll just highlight here that if an individual takes their medications before they come in for appointments—which is not that infrequent; people are sometimes motivated by an upcoming visit and they do much better with their medications—they may have detectable viremia. And that situation, as you’re describing, is actually very different from someone who takes their medicines every day and develops this type of detectable viremia.

And so, the case we’re talking about today has been sort of very clearly proven to fall in this other category because that’s what we mostly wanted to talk about today. But I think you’re making a really important distinction here, which is that if you just look at the population level, these detectable viral loads might be associated with an increased risk of failure, but they also likely include people who have less than 100% adherence to their regimens. Is that what you were outlining for us there?

Francesco:
Yes, absolutely. And indeed, we like the term “nonsuppressible viremia” because it shifts the focus from the copy numbers, from the magnitude of viremia, to the fact that it’s a matter of mechanism, and changing treatment or adding additional antiretroviral drugs will not help. For example, we have studied patients with NSV even above 1,000 copies that after some sequencing and more in-depth studies [have] shown that those were not caused by replication, just by release of viral particles. But those will fall well above the 200 cutoff. So that’s why we like to talk about the fact that it’s nonsuppressible rather than focusing solely on numbers.

Eileen:
That’s perfect, it really highlights that understanding the biology does require, as I said at the top, a really thoughtful understanding of the clinical realities, and that, as any provider knows, there’s about 400 reasons why someone might have detectable viremia. And identifying that group who has no clinical reason based on their adherence or other features to have viremia has helped us to learn more about the reservoir. So, in our clinical community here at Hopkins, we have this advantage of trying to learn more about what’s happening with these rare individuals. And that is through collaboration with scientists like you.

With that as a frame, we will say that we had the clinical discussion in the office with this individual about the implications of this low-level virus. We can’t comment on whether it increases your cardiovascular risk per se. We don’t love that it’s there, but we actually don’t have a clinical intervention to reduce it. From the perspective of U=U—for this person in particular, it falls under the threshold of viral suppression for undetectable equals untransmittable, so we can give some reassurance about the risk of ongoing transmission.

But in terms of the other questions, this individual is someone who wanted to know more and was really ready and willing to be engaged in research. So, Francesco, as you mentioned, you’ve studied a number of individuals with nonsuppressible viremia who’ve been confirmed through history, through drug levels, through a bunch of different modalities to be taking their medications and still having detectable viremia. And I was wondering if you could take us through some of the different stories for how this arises that your research has uncovered.

Francesco:
Yes, that would be my pleasure. We have been sequencing virus in plasma from people with NSV for more than a decade now, and we were able to learn so much just from looking at the sequence of the virus. You know, HIV, it’s highly diverse, and so each viral sequence becomes almost like a barcode that allows us to infer whether there is evolution over time, whether instead HIV persists because the cells that carry that virus are persisting during treatment. And so, one important thing that we found out is that when you sequence virus in plasma, this is very often highly clonal, so meaning that every sequence is 100% identical.

Which reflects one of the most important features of HIV persistence, which is the clonal expansion, the proliferation of the infected T cells. This is what I call a T-cell day job is: to proliferate, to function, and survive and carry our immunological memory. So, the first take-home message is that virus causing NSV is extremely clonal. There’s very few infected clones that drive this virus in plasma, because every day a handful of cells will have transcriptional activity and express the viral gene and hence viral particles.

Another important observation that we made a few years ago is that not all the time the virus that is produced and causing viremia is infectious. We found a handful of cases in which the clones driving viremia were indeed defective, and they all have defects, all narrowly focused in the same region and—without going too much into details, but—it’s a noncoding region that orchestrates the HIV replication and the life cycle. So, even small defects in this region can completely abrogate infectiousness, but they still allow for virus expression. And then, recently, we tried to follow up on this and ask the question, “how often are cases of NSV driven by defective HIV genomes?” And so, we put together a large cohort, we studied almost 50 cases of NSV, and we found that, on average, 95% of the virus causing NSV was defective, all because [of] these defects in this region that we call the 5′ leader. So, this is very common in people with NSV.

Eileen:
Francesco, there’s just a few points I’d love to pull out there just for the sake of our listeners who are maybe thinking about this for the first time. So, when we look at the HIV in a person who’s not on antiretroviral therapy and has 1,000 just because that’s how many copies they have, we’ll see a very high level of diversity. Most sequences will be different from each other. And that first finding that the HIV is nearly the same all the time, says kind of two things. One is that they’re coming from the same place, and the second is that it’s not actually able to go into a new cell because every time you go into a new cell, you get the opportunity for the mutation magic to happen and to have a slight variation. And so it creates this situation where the virus comes out, but it doesn’t have anywhere to go. So, I’m just going to check back that the way I’ve explained that fits with what you’re describing.

Francesco:
Yes, that was beautifully said. I think the simplest way to put this is to distinguish virus replication, which is the full cycle, the one that leads to the accumulation of mutations over time, versus virus expression, which is sort of like half of the cycle in which an infected cell just releases viral particles and this happens in all people living with HIV on therapy, there’s always some extent of daily dribbling of viral particles, even if therapy is fully effective.

Eileen:
Perfect. So then I think we can address some of the concerns, right? So, if we think that the virus is there in the blood or plasma but not able to cause new rounds of infection, we can feel good [that} we’re not making new reservoir cells, but we still have all these same concerns about number one, onward transmission, and number two, the inflammation of being exposed to HIV.

And so now I’ll just pull out the next thing that you highlighted, which is that another finding from your work has been that in many of these cases, this virus has this defect, or not necessarily one particular defect but a couple in a family of types of defects, so that you can make the particle, but then the particle, which is measured by the viral load, actually is not good at causing a new infection. And that’s where one of those classifications is those 5′-leader defects. So, you have viremia, it’s almost all the same, and that virus actually is not capable of infecting new cells efficiently.

Francesco:
Exactly, that’s correct. We have tested this extensively. Every time that we introduce one of the defects found in patients into one of our infectious lab strains, we completely lose, by, you know, multiple orders of magnitude, infectiousness. And so, the cell can make viral particles—these will contain HIV RNA, which is what we detect with the clinical assays—but the viral particle doesn’t have the machinery required to infect a new cell.

Eileen:
That really effectively addresses the concerns about onward transmission and also goes back to that same issue about reservoir expansion, which should not occur. And if you can screen using some of the assays that you developed to identify this, that would all be very reassuring for those first two points: reservoir expansion and onward transmission. I wonder though, if you could comment on whether or not these defective particles have any immune consequences, because as we all know, one of the problems with HIV is ongoing immune stimulation that can lead to inflammation and may enhance the risk of comorbidities.

Francesco:
Yes, that’s a very important aspect and I think it’s something that remains a research priority. I think a clear linkage has been made between HIV expression and higher inflammation. And you know, since the majority of proviruses within an individual on ART are defective and can still express some level of RNA, some proteins, and as we were discussing, even viral particles, they are likely a potential driver of this inflammation. However, we still miss a direct link with poorer clinical outcome, because I think, oftentimes, it’s difficult to disentangle what was the immunological damage that happened before therapy that remains as an immunological scar, as a sequela, even after someone has been on therapy for many years, with the impact of this daily expression of virus, even when they are defective. So, I think in the future we need to find ways to understand to which extent daily virus expression and NSV really are a source of concern regarding inflammation, immune activation, and immune aging.

Eileen:
I think those are really important points and it is a really important area for research. I find that when I am talking with people living with HIV about this, I usually end up saying, “We certainly can’t exclude that there is an ongoing contribution of this amount of virus, whether it’s defective or not, to some immune stimulation.” But that said, we all, not all of us, but the vast majority of us have Epstein Barr virus, have CMV, and other sources of little tickles of the immune system over time. And if there is no further clinical intervention, I certainly don’t want people adding chronic stress about their low detectable viremia to the list of factors that will increase their risk for comorbidities and just try to focus on the other interventions that we can do to reduce risk, including things like exercise, smoking cessation, managing your other cardiovascular risk profile or neuroaging profile. But it is an important question and one that I hope that research will address in the future.

Francesco:
Yeah. Until we have an intervention that really decreases HIV expression, such as, potentially, a new class of antiretrovirals, it’s impossible to really assess what’s the impact of residual HIV expression on ART.

Eileen:
I think that’s a great point and also highlights that the ART that we have currently work on the sort of entry replication integration primarily side of the viral life cycle, which is the part of the viral life cycle that’s being blocked. But the other part, where you go from integrated provirus to viral particle, largely relies on cellular machinery that is harder to target with drugs and so therefore is not a major antiretroviral therapy target.

But maybe there is something where we can get some viral specificity into that back half of virus production that could help with this nonsuppressible viremia.

Francesco:
Yes, exactly. And a frequent misconception that I ran into talking with other clinicians is, for example, that protease inhibitor[s] would instead affect HIV expression. While instead protease inhibitor[s] affect the quality, the maturation, of the viral particles, rendering them noninfectious, but they don’t prevent them from being released. And regarding an outlook of future new classes of medications, there could be future generations of capsid inhibitors like the ones that are in use right now, such as lenacapavir, that not only block new infection events, but they are so powerful at preventing virus assembly that you even block virus expression.

Eileen:
That’s a really hopeful thing for the future, for sure. Francesco, the one other thing I wanted to talk about a little bit is how you think about why things like nonsuppressible viremia can come up spontaneously and then regress, and what that says about the composition of the reservoir and what cells in the body are holding the HIV provirus.

Francesco:
NSV is another reflection of a major mechanism of HIV persistence, like we said before, which is the proliferation of infected cells. And so our T cells, our immune repertoire, is constantly changing, is characterized by memory. So, some T-cell clone will persist our entire lifetime while others will be lost and new ones will be formed, but it’s never fixed. It’s a highly dynamic system. And so you can imagine if that one infected T-cell clone might be at a very low frequency for many years and then stimulated by a specific antigen or by other immunological stimuli might increase in size to the point that that little virus that was made daily now is higher and hence it falls within the detectability range of clinical assay. Likewise, we lose clones, we lose immune memory against some pathogens over time, and so we have seen that many people with NSV resolve completely the detectable viremia, because that clone contracts or is completely extinguished just because of the natural variable fate of our T cells.

Eileen:
And this is one of those things that can be challenging to explain, and one of those situations where when we look longitudinally at patients, we can be tempted to assign causality to interventions that happen: regimen changes, weight gain, other diagnoses. But we have to have the humility to say that we don’t actually know in many cases what’s happening to the underlying T-cell repertoire and which T cells happen to be holding the HIV reservoir. And so, some of those things that may appear to be causally linked may actually be related to a whole different set of dynamics that we don’t measure clinically.

Francesco:
Yeah, absolutely. And if we think about some recent data from longitudinal studies from the ACTG [AIDS Clinical Trials Group], where they followed for many years people with NSV, about a third of them resolved NSV without any clear clinical event or change in therapy or change in habit or environmental exposure. It’s part of that randomness of our immune system dynamics.

Eileen:
Thanks so much for this discussion. And we touched on this a little bit, but I just am wondering how you’re thinking forward to how what you’ve learned in your group and in other groups about nonsuppressible viremia is going to impact what we do for people living with HIV in the future. What is your sort of dream scenario?

Francesco:
Well, first of all, one take-home message is not to immediately stop at adherence, and sort of like challenge what patients tell us about adherence, because there might be biological reasons for having low-level viremia. And approach this with compassion and try to understand really “what are the mechanisms.”

And the other thing that we learned from our work is that there’s really a technological gap. We need to be, get better at sequencing the virus, even when there’s little of it in plasma, so that we can offer tools, we can offer assays to improve the initial assessment to understand whether the virus is clonal, it’s intact, it’s defective, it has drug resistance. But, unfortunately, as in the case of our patient today, sometimes it’s challenging to sequence the virus from plasma when it’s below 500 copies. So, my dream is to be able to develop additional tools to empower both clinicians and people living with HIV. They might want to better understand why after so many years they suddenly have detectable viral load.

Eileen:
Well, I think you’re well on the way to developing those tools, and I personally look forward to being able to deploy them clinically. So, thank you so much. And just to summarize, today we talked about an individual who had nonsuppressible viremia, so someone whose viral load remained detectable despite high adherence and an effective therapy. We talked about how this can be identified using assays that are not currently clinically available but that can really define that the virus is coming from one place and in many cases is actually defective, and how this changes our assessment of the risk in terms of treatment failure or reservoir expansion, which is functionally eliminated, and also the potential risk of transmission, which is also substantially mitigated, but that it doesn’t answer all of the questions about what having HIV particles does to the immune system with respect to inflammation.

And I think we’re all left hoping that we can get the Simonetti assay to determine what’s going on with our patients in the future. Thank you so much for joining me today.

Francesco:
And thank you for having me. It’s been a blast.

Eileen:
So, to our listeners, thank you for joining in on another episode of Viremic. Please send any comments or questions you have to viremicpodcast@jh.edu, and we’ll be back in two weeks to talk about immune reconstitution inflammatory syndrome.

Announcer:
Viremic is sponsored by the Clinical Guidelines Program, a collaborative effort of the New York State Department of Health AIDS Institute and the Johns Hopkins University Division of Infectious Diseases.

Visit the program’s website at: HIVguidelines.org.

Viremic’s case discussions are for informational purposes only and not offered as medical or clinical practice advice for patients or clinicians. Any mention of specific medications or commercially available products is a description of use only, not an endorsement

The podcast is produced and edited by Jen Ham and Brian Hatcher with assistance from Jesse Ciekot and Laura LeBrun Hatcher.