Hi, and welcome. Good evening, everyone. My name is Justin Roden. I'm a cardiologist in UT Southwestern, and welcome to our session entitled, where are we with cardiac amyloidosis? This is the agenda for this evening. We'll start with an introduction and then 3 great talks. That will be followed by a panel discussion and then eventually closing remarks. And we'll be done in just about an hour and a half. This is the target audience. These are the learning objectives for this session. The first is to review the current practice on the diagnosis and initiation of treatment for patients with ATTRCA. Then the next objective is to evaluate the current and emerging data on new therapies for the treatment of ATTRCA, followed by the objective to discuss the clinical trial data and role of RNAi therapies in the management of patients with ATTRCA. These are the accreditation and credit designation statements. These are the steps to claim credit. This is the quick off-label disclosure before we give our talks, and that is to say that we might be discussing off-label therapies within the context of our talks. The faculty for this session are as follows. So myself, my name is Justin Roden, as I mentioned before. I am a heart failure specialist, and I take care of patients with amyloidosis at UT Southwestern in Dallas, Texas. My other two colleagues are Dr. Kevin Alexander, who is an advanced heart failure specialist that also takes care of amyloid patients at Stanford, followed by Amy Brownell, who is a nurse practitioner and works at Ascension, Illinois. And, you know, I think it's very important. We'd like to thank our sponsor, Alnylam Pharmaceuticals, who have supported this activity from an educational grant. We're certainly grateful to all of their support. This is your pre-test before we get started. The first question is, which of the following leads to amyloid formation? A, valvular thickening, B, tetramer dissociation, C, cardiomyocyte formation, or D, amorphous oligomers? And I'll give you a few moments to answer that question. Our next question is which of the following statements is correct? A. A negative SPEP and UPEP may be used to rule out AL amyloidosis. B. Endomyocardial biopsy is not highly sensitive for cardiac amyloidosis. C. Technetium pyrophosphate bone scintigraphy plus a negative workup for an abnormal M protein has a very high specificity for ATTR and D. AL and ATTR are not macroscopically similar. The third question is currently in development CRISPR-Cas9 based in vivo gene editing therapy is targeting blank in adults with hereditary transthyretin amyloidosis and polyneuropathy with or without cardiomyopathy. A. Cardiac troponins T and I. B. N-terminal pro-B type natriuretic peptide. C. TTR produced by human hepatocytes. Or D. Urinary albumin to creatinine ratio. And our fourth question is data from the Apollo trial showed that patients in the blank group had improvement in multiple clinical manifestations of hereditary transthyretin amyloidosis including their neuropathy impairment score compared to those in the placebo group. A. Petitseran B. Inotericin C. Vutriceran and D. Eplantericin. And now with those questions I'd like to move on to the first talk which is done by me entitled ATTR Diagnosis Current Best Practices in Emerging Horizons Exploring Novel Techniques, Non-invasive Approaches and Biomarkers. So the outline for this discussion, you know, we'll cover these four bullet points. We'll discuss the ATTR pathophysiology and then this will resonate with the other talks through my colleagues. And then we will go over an overview of non-ATTR specific diagnostic strategies and I'm sure many of you who are clinicians in the audience use those in your routine practice. And then we're going to pivot. We're going to focus more so on ATTR specific diagnostic strategies. So these are going to be techniques that will actually secure a type specific diagnosis and I will detail how to use them. And then finally we're going to look ahead in that we'll discuss a few diagnostic strategies on the horizon. It will give you in the audience an idea of where this field is possibly headed. So the first section is we're going to discuss ATTR pathophysiology. So this is a ribbon diagram of the TTR tetramer. Now maybe you may or may not know that transthyretin is actually a portmanteau. That is to say is a combination of the three words transports thyroxin and retinol. Now ATTR amyloidosis really comes in two forms. So approximately 70% of all the ATTR that we see in North America and the United Kingdom is wild type. That is to say it is not hereditary and not caused by a pathogenic mutation. Or the remainder is hereditary or variant and it is caused by a mutation in TTR. Now what is key is the pathological formation of ATTR amyloid is caused by tetramer dissociation of the TTR molecule. So what does this look like? Well the TTR tetramer I should say 99% maybe more of the TTR in your body is made by the liver and it floats around in our bloodstream. There is a small amount that is made in the choroid plexus and the retinolipithelium that is in the cerebrospinal fluid and this does not cross the blood brain barrier. So TTR is secreted as a tetramer in the blood and it is really when this tetramer dimerizes we think it might dimerize it might break down into just the single monomers itself but this is really the rate limiting step in amyloidogenesis. And these folded dimers then dissociate into folded monomers which then go through this misfolding process and turn into amorphous oligomers and then this is really where mature amyloid fibrils start to form and deposit in different organs in the body. So now when we think about ATTR cardiac amyloidosis I think it is really important to at least have an idea of the natural history of the disease. Now this figure comes from an editorial I wrote with a colleague of ours Dr. Matt Maurer and really what it highlights if you look at this curve that highlights this rate of myocardial infiltration and notice it starts when individuals have no symptoms and it really isn't until later stages in the disease course that we start to see impairment in functional status, we see elevations in typical cardiac biomarkers, natriuretic peptides, troponins, then we see an increased risk of healthcare utilization manifested by hospitalizations and then ultimately death. Now what is unfortunate and you'll hear a lot about the treatments by Dr. Alexander is that the majority of the patients we see are really diagnosed in very advanced stages when they are symptomatic. We really don't see them when you know a lot of this amyloid formation is actually happening. So you know when we take a step back and we think you know we ask the question really what do these hearts look like, I think this picture highlights what a gross specimen, now this is a fresh specimen of a human being who was just undergoing a heart transplant for advanced cardiac amyloidosis and you can see in these sections that the muscle is really thick. Notice it's got kind of this glossy almost gelatinous texture that almost looks like plastic and if you can tell in your computer screens you'll see this kind of pale, palish ring on the inside of the heart muscle. So this is a gross example of what amyloid looks like. And when we take a step further and we look at what does amyloid look like in the myocardium, it is really what we call an infiltrative cardiomyopathy or an infiltrative heart disease. This is not a special stain, this is a very routine H&E stain of a patient with ATTR and what I'd like to point out is that these areas in light pink are actually amyloid infiltration into the cardiomyocytes which are shown here in dark pink and really what it highlights is how the amyloid infiltrates this network of cardiomyocytes. So it's almost like a, you know, and I usually give my patients this thought experiment, it's a heart with amyloidosis is almost like a brick wall with way too much mortar in between the bricks. So I think this figure or this image really encapsulates that concept. So now I want to pivot to an overview of some nonspecific or non-ATTR specific diagnostic strategies, many of them you use in your routine clinical practice. So I think the first and probably the most crucial step in trying to diagnose ATTR is really to develop a level of concern, a level of, you're trying to build a case that there maybe there are clues that this person might actually have the disease. To me this is really the rate limiting step because as you will see a lot of the diagnostic testing is actually kind of straightforward and can be algorithmic in a number of cases. So I think the first red flag or clue that might increase your index of suspicion for amyloidosis is unexplained LVH followed by unexplained restrictive cardiomyopathy. I think this one speaks for itself. Anybody who is over the age of 65 that has a history of carpal tunnel syndrome, especially when it's bilateral. Anybody who has an atraumatic biceps tendon rupture and on the next slide I'm going to show you an image of what this looks like. This tends to be quite ATTR specific. Anybody with an unexplained neuropathy. Now this might sound rather straightforward but we all know that diabetes is quite common in the United States and other parts of the world. But when you see that somebody's neuropathy is far more severe than what could be explained by what might be really well controlled in mild diabetes, that should really serve as a clue. Anybody with orthostatic hypotension, this could be a clue that they have autonomic dysfunction and then any individual with hypertrophic cardiomyopathy that is diagnosed over the age of 60. So as I mentioned I was going to show you what an atraumatic biceps tendon rupture looks like. So this is a proximal biceps tendon rupture and what you see here is Popeye's sign. So you can see that this ball of muscle that is now rolled down to the bottom of the arm towards the antecubital fossa or the inside of the elbow. So be sure to look at that or be sure to ask your patients if you suspect they have amyloidosis whether or not this has occurred. Many will not volunteer it. You do have to ask. So now what are some tests that we use all the time that are great but really that are not ATTR specific? ECGs, echocardiography and of course cardiac magnetic resonance imaging with contrast. So ECGs are great. They're cheap. They're incredibly easy and what I'm showing you here is the classic ECG of an individual with amyloidosis. This is characterized by very, very low voltage. So I can tell you that this patient has LVH on echo. Notice that there is very low voltage in the limb leads and then we also see that there is this precordial kind of Q wave pattern where it looks like they had an anterior myocardial infarction. We call this a pseudo-infarct pattern. So this ECG is quite useful, however, it does have some limitations. There are a number of pitfalls. It's really not all that sensitive. It's great if you see it but if you don't, it doesn't necessarily mean the patient doesn't have amyloidosis. About 50% of individuals with AL amyloidosis meet this criteria, 30% with ATTR may meet this criteria and only about half have this pseudo-infarct pattern. Now what about echocardiography? This is a great tool. We use it all the time and I'm showing you two images here on this screen. The one on the left is a parasternal long axis view, an individual with no amyloid involved of his heart. Then what I'm showing you on the right is of a classic parasternal long axis view. So the same view and this is an individual that does have amyloidosis and you can see that it's really characterized by having markedly thickened left ventricular walls. We can't really see the RV free wall very well in this view. So now the next slides I'm going to show you how we, you know, this kind of goes over the pathophysiology of ATTR and in so doing, I will use different echocardiographic images to highlight certain clues that you might also find in somebody that may have amyloidosis. So I think the classic pattern that we see is that there is biventricular thickening. It's very important to recognize that this is not LV hypertrophy. The ventricles are stiff as far as we know because of all of these amyloid protein deposits that have infiltrated the myocardium. These ventricles are extremely stiff, they have poor compliance and so it's very common that we see abnormal markers of diastolic function and ejection fraction is typically very normal even in patients with advanced disease. It only drops in people with extremely advanced disease or when they're severe and they're getting to that point. The pericardium is frequently involved in effusions, are usually small. It's really rare that these patients go into tamponade. And historically, this has been a marker of risk. This is an image of a apical four-chamber view of the same patient with ATTR amyloidosis. And what it highlights here, if you look at these arrows, these AV valves, so the mitral valve on your right, the tricuspid valve on your left, are markedly thick. And this is likely due to amyloid infiltration on the surface of the valve. Now, it is very common to see mild to moderate regurgitation. It is also not uncommon that you can see severe leaks. Conduction delays are quite common in patients with amyloidosis, especially in ATTR. We see AV blocks, we see bundle branch blocks. It's unclear if this is due to amyloid infiltration of the conduction system, or is this just confounded because these individuals are typically older and older individuals are at greater risk to having some of these conduction delays. Now, what is important is the atria are universally involved. And it's not uncommon that you might notice that the atrial walls are mildly thick. Atrial function is very commonly poor in these individuals, and atrial fibrillation is very, very common. So, I could give you a whole talk on diastolic dysfunction in amyloidosis, I'm not gonna do that. Hopefully, none of you have eaten dinner, or maybe, you know, if you have, you're not easily grossed out, but this is another heart taken out of a human being. And what it highlights is just how stiff these hearts are. Now, I don't know if any of you have seen a human heart on a table, but just like a piece of meat on a cutting board they typically collapse under their own weight. And what the surgeon here is demonstrating to you is that this heart stays propped open much like a racquetball. So, this is how stiff these hearts are. So, what about cardiac MRI? Cardiac MRI is a great tool. We don't use radiation, it's a big magnet, and it's largely incredibly safe. So, these are some very characteristic images that we use from an MRI. On the left-hand side, you can see a heart with diffuse subendocardial LGE. Now, these are demarcated by kind of these white splotches on the inside of this horseshoe in the ventricle. And then on the right-hand side is a map of the extracellular volume. And you can see that all these areas in red, kind of in this donut shape of the left ventricle, indicate a markedly expanded ECV and is very consistent with amyloidosis. So, contrast-enhanced MRI is a great tool. It can distinguish carriers of a genotype who don't have any amyloid infiltration. You can see this on the left-hand panel. You can see this looks very similar to a non-carrier with heart failure with preserved ejection fraction. And then you can see how this differentiates from our patient who is genotype positive and does have the LGE phenotype and does have ATTR amyloidosis. Now, what about ECV? This, too, can distinguish healthy controls from genotype positive, phenotype negative carriers and non-carriers. And this is demonstrated here by if you look at that same donut hole, you can see that it looks yellow-green in the healthy controls, the phenotype negative carrier in the individual with HEP-PEP. And notice how it's all red, which marks an extremely expanded extracellular volume of this heart or the patient with this heart. So, you know, really one of the big issues with all of those tests is that they're wonderful, but AL and ATTR are macroscopically similar. These are two gross specimens from each of these types of cardiomyopathy. And I will tell you that I would challenge any of you all to tell the difference between any, between either of these hearts. They look overall similar. So this really calls for a need to use specific diagnostic strategies to diagnose ATTR. So what are those? So these are some amyloid-type specific tests that we use. This is one of the most important slides in my whole lecture to you all this evening. The first are free light chain assays with serum and urine immunofixation, bone scintigraphy of the heart, and then of course tissue biopsy. So one of the most important tests that we can do are measure free light chains in the blood. And you will see if we do nothing else, a free light chain ratio that is abnormal is 91% sensitive for the diagnosis of AL amyloidosis. And when we couple this with serum and urine immunofixation, you can see that the sensitivity reaches almost 99%. And I will remind you all, and this comes up on rounds all the time, that if you think you've ruled out AL amyloidosis with an SPEP or a UPEP, you have not. They are very insensitive. You need to check serum free light chains and you need to add the immunofixation or request it if your lab hasn't already done it. So a negative SPEP and UPEP, I will repeat again, does not rule out AL amyloidosis. You need those other tests. So what about bone scintigraphy? So there are a number of bone radio tracers that have been, or that we've observed over time that can identify ATTR amyloid infiltration of the heart. In the United States we use DPD, I'm sorry, we use HMDP and PYP. This is a very straightforward test to read. What I'm showing you here are some planar images of different individuals with different levels of uptake of the tracer in their myocardium. Now notice we do see the bones, much like a chest X-ray, but we see the panel that has grade one uptake, where there is a faint splotch over the heart that looks less than bone, grade two uptake, where the cardiac uptake is roughly equivalent to bone, and then of course grade three, where the cardiac uptake is greater than bone. In grade two and grade three are positive scans. Now we're not quite done there because you must pair this test with a workup for an abnormal M protein. And if you've ruled that out, the specificity and positive predictive value are excellent for ATTR. Now what is now standard of practice, and I think we can all agree is best practice, is we couple those planar images with more of a 3D imaging with SPECT. That way we can triangulate where the tracer is coming from. So both of these were positive scans by planar images. And you can see here on the panel on your left, all the orange or the uptake is in the blood pool and not in the myocardium. And we contrast that with the panel on your right, where all the uptake or all these hot counts are in the myocardium. So the panel on the right is a true positive, the panel on the left is a false positive or a true negative test. Now what about endomyocardial biopsy? It's a great test. It's highly sensitive to identify cardiac amyloidosis. It's not without risks. Approximately 1%, again, it's in different hands and there are different risk factors per patient where it can cause a perforation in the right ventricle that could put you at risk for, or could put the patient at risk for developing tamponade. But what is crucial to know is that once these samples are stained, if they're Congo red, that tells you, it gives you one, it answers one question. Is there amyloid yes or no? If the answer is yes, that test must be followed with something that types the amyloid, either immunohistochemistry or laser capture microdissection mass spectrometry. Our lab utilizes both, but again, you must do that second step to type amyloidosis. What about FATPAD? So this is quite variable across the country. Centers have different experiences with this. I can tell you at our center, it's not all that useful, but data demonstrate that it might be, you can see here, it's got variable sensitivities in both AL and ATTR and might be very insensitive in individuals with ATTR wild type. It's highly dependent on the operator that's acquiring the sample, on the pathologist, and really how much tissue you've acquired. So at our center, and this is very similar to other centers, if there is a negative FATPAD biopsy, this really is not sufficient to rule out cardiac amyloidosis if the index of suspicion is high enough. So how do we put it together? The first thing to order is work up for an abnormal M protein with a free light chain and check serum and urine immunofixation. If they are abnormal, then we follow with an endomyocardial biopsy or a tissue biopsy. And the pathway we're on in this case is AL amyloidosis. Now, if the free light chains are normal and there's no evidence of an abnormal M protein, then we can get a PYP scan, and if it's positive, we then move on to genetic testing and sequence the ATTR gene and we've achieved a non-biopsy diagnosis of ATTR. Now, if they are all normal, then we either don't have amyloidosis or if the suspicion is high, you can always biopsy the patient's myocardium. So I think what's really important, and I think my colleagues would agree, that if really there's any ambiguity in any of this testing, that should really trigger consideration for an endomyocardial biopsy. So now I want to end in the last few moments with a few diagnostic strategies that are on the horizon. Now, some promising advances are highly sensitive PET and SPECT-based amyloid tracers. There are new, insightful, and very promising blood biomarkers that might give us a blood diagnosis of ATTR, and then of course, ECG-based AI tools. Some of you may be familiar with some of those. So I'm only gonna highlight the first two bullets, and the first is an agent called iodine-124-ebuzametide. You might also be familiar with it as ATO1. This is an investigational agent that is a pan-amyloid peptide that binds to amyloid. And what you can see here are two successive specimens of the myocardium, and you can see here in the areas that are conga-red positive on the right-hand side directly correspond to the biotinyl-labeled ebuzametide, or the biotinyl-labeled ebuzametide, where that tracer has bound in this successive site. So what you're looking at with this tracer is amyloid. Now, these are some very early patients that were given this tracer, and I think what it demonstrates is the capacity of this tracer with PET imaging to identify individuals that have clinical symptoms or not, but still do have perhaps subclinical amyloid involvement. And then I wanna pivot to a really unique and novel peptide probe developed by Lorena Solis Gomez called TAD1 that is highly sensitive and highly specific for ATTR in the blood. You can see here on the left-hand side that it differentiates ATTR from a number of different controls, including other molecules of TTR or forms of amyloid. And then you can see here that it might vary, if you look at the panels to the right, in patients that are allele carriers that don't have symptoms, those that might be pretreatment, and those that might be post-treatment. We kinda see the predicted concentrations in those individuals. So that is my last slide. So we discussed ATTR pathophysiology. We gave an overview of nine ATTR-specific diagnostic strategies. We highlighted ECG, MR, and echocardiography. And then we talked about some very useful ATTR-specific diagnostic strategies, light chains, PYP scan, et cetera, and then we discussed a few diagnostic strategies on the horizon. So the take-home points of my talk are as follows, is that non-ATTR-specific diagnostic strategies can enhance suspicion for cardiac ATTR. Centigraphy with bone tracers can lead to a non-biopsy diagnosis of ATTR if an abnormal M protein has been ruled out. Now remember, when the diagnostic data are uncertain, always consider tissue biopsy. And lastly, promising strategies are on the horizon. They're really exciting, and they might be able to facilitate a more rapid ATTR diagnosis. Thank you all for your attention. These are the post-test questions. Which of the following leads to amyloid formation? I can't see your answers, but if you answered B, tetramer dissociation, you were correct. And for question 2, which of the following statements is correct? Again, I can't see your answers, but if you answered C, PYP bone scintigraphy plus a negative workup for an abnormal M-protein has a very high specificity for ATTR is correct. And now I'd like to pivot to my colleague, Dr. Kevin Alexander, who will give the following talk entitled Breaking Ground, the Latest Data and Game-Changing Therapies in ATTR. Great. Thanks, Dr. Grodin. And I think Dr. Grodin's talk really set the stage for what I'm going to talk about now, which are some of the advances in ATTR therapies. And this has been a very dynamic space. And one of the fun things about it is developing slides for talks like this, you'd look six to 12 months into the future and you have to change a lot of stuff around. And I think that it's a really good sign that there's a lot of innovation in the field. So in a relatively short period of time, I'm going to go over some of those advances in therapies and look forward to the discussion later, panel discussion later. So, Dr. Grodin touched on this, but I'm just going to review the pathogenesis of ATTR because this is really important to understand the targets for ATTR therapies. So the majority of the transthyretin is produced by the liver as a homotetramer, and its main job is to transport thyroid hormone and retinal binding protein throughout the body. And people with ATTR, they develop dissociation of these tetramers for a variety of reasons. They may have a point mutation that predisposes the tetramer to being unstable, and that's the hereditary form of ATTR. And there's also probably other mechanisms that lead to protein instability that are less well understood, age-related factors, sex-related factors, and this is an area of ongoing investigation to understand some of these molecular determinants for tetramer dissociation. Regardless of the reasons that lead to tetramer dissociation, this is really the rate-limiting step in forming amyloidosis. So when the tetramers dissociate into monomers, these monomers are thermodynamically unstable and have an increased propensity for misfolding, and these misfolded monomers are really the backbone of what later become mature amyloid fibrils. They start to aggregate, and these aggregates form small oligomers and amorphous aggregates, and then these are really what go on to develop into the mature fibrils that deposit in the heart, peripheral nervous system, and other places. So for the first class of ATTR therapies that I'm going to highlight, I want to go back to a clinical observation that was made a few decades ago and really was the foundation for development of one of the classes of ATTR therapies. So there's a TTR variant called V30M or V50M, depending on the nomenclature that you're using, and this is a type of ATTR, hereditary ATTR, that leads to aggressive neuropathy, and it can manifest as early as 30 years of age and can be quite devastating. A lot of families in Portugal were studied with this variant, and what the investigators there observed was that some of these family members were not affected by polyneuropathy, in fact, had no evidence of neuropathy and normal lifespans, etc., and at the time when they did further genetic testing, it turned out that some of these individuals had a second mutation in the transloretin protein, the T119M variant, and further investigation showed that these individuals are resistant to developing ATTR amyloidosis, and the reason is the T119M variant, in contrast to other mutations which have a destabilizing effect on the tetramer, this variant actually has a hyperstabilizing effect and protects the tetramer from dissociating into monomers, and so that led to the hypothesis by Jeff Kelly and his colleagues that if you can develop a small molecule that mimics the effects of this stabilizing mutation, a stabilizing variant, then that could potentially stabilize the tetramer and prevent further dissociation of the tetramer, thereby leading to improved clinical outcomes, and so after many years of hard work, there's been a number of small molecules that have been developed, and we call these transloretin stabilizers. These are small molecules that bind to different areas in the TTR tetramer to decrease the amount that dissociates into those toxic monomers, so the first landmark trial to study a transloretin stabilizer is the ATTR-ACT study, and this looked at a drug called tefamidase. The study design briefly is shown here, so patients were randomized to either placebo or 20 milligram or 80 milligram dose of tefamidase. There's over 441 patients in the trial, which particularly at the time when this was thought to be a rare disease, took a heroic effort among all the investigators to recruit this large number of ATTR patients. They had either wild-type or variant ATTR, and they had NYH class 1 to 3 heart failure, and the primary endpoint in this trial is a composite of cardiovascular hospitalization, mortality, and a few other surrogate endpoints that were analyzed in a hierarchical fashion, and what you can see here from the survival curves is that there's a statistically significant benefit in terms of mortality, as well as the other secondary endpoints for the tefamidase group, and those curves separated around 18 months. The number needed to treat to reach for the primary endpoint was about seven and a half, so very strong data for any kind of cardiovascular trial, and certainly for a disease where there were no treatments beforehand, this was really exciting when these data came out. Looking a little bit further into the subgroup analyses, there's benefit across various subgroups, but I think one thing to highlight is that when you look at the class 3 heart failure patients, they did not derive the same benefit from tefamidase as the other groups, and I think it highlights a point that Dr. Grodin mentioned during his talk about the pathophysiology and the natural history of this disease. Later stages of the disease, patients often have much more amyloid burden and probably much more structural remodeling of their hearts, and if we think about the path, if you think about the mechanism of action of tefamidase, it really works by stabilizing the tetramer and preventing or reducing the amount of new amyloid that forms, and we don't necessarily think it directly intervenes on amyloid that's already in the myocardium, so it's really further emphasizing the importance of diagnosing patients earlier. You know, we have effective treatments, but they clearly work much better in the earlier stages of disease. We recently had approval of a second transthlaretin stabilizer called aciramidase, so this is another small molecule that stabilizes transthlaretin. It was studied in the TRIBUTE-CM trial. This had a similar study design as well as primary endpoint shown here, and this trial, aciramidase, also reached its primary endpoint, which is the composite of mortality, cardiovascular hospitalization, six-minute walk, and KCCQ. There is a 50% relative risk reduction in cardiovascular hospitalization and a benefit across a number of subgroups shown here. I think one thing that we learned from this trial as well, just looking at the event rates and survival rates, this population clearly is different than the ATTRAC population, so you fast-forward several years into the future, we're starting to diagnose patients a little bit earlier with the emergence of technetium-based scanning, which is a non-invasive alternative to endocardial biopsy for many patients, and so with that, we see that the natural history of the patients presenting to us and also that are enrolling in trials have changed over time, and so that's one of the things I think, you know, perhaps in the discussion we talk about these various treatments, it's important to keep in mind that the cross-trial comparisons are extremely challenging, and I would shy away from those things because the groups of patients studied in each of these trials is a little bit different. So, the second class of medications that I'd like to talk about are therapies that focus on knocking down the protein expression of transthyretin in the liver, so if we know that transthyretin is a substrate by which amyloid, this type of amyloids form, can we decrease that production to decrease the amount that goes on to form amyloid, and so there's been a number of therapies that have been developed in this space. The three kind of mechanisms of action that have been used or approaches that have been used have been siRNA approaches, antisense aglonucleotides, so these are things that target the messenger RNA, and then there's also been next-generation gene editing to potentially provide a one-time treatment through permanent genome editing. So, shown here is one of the first phase three trials for looking at these TTR knockdown therapies. This is the Apollo trial that looked at patisserin, and so much of the early data for the knockdown or silencer agents for ATTR were studied in a polyneuropathy population, and so Apollo looked at 225 patients with variant ATTR polyneuropathy, and they were randomized to either patisserin or placebo and followed over 18 months. These neuropathy trials use a primary endpoint of the MNIST plus seven, which is a composite of neurologic deficits, sensory, motor, and autonomic. The more deficits that you have, the higher your score, so the score increasing is bad, and what you can see here for the placebo group that there's a 28-point worsening over an 18-month period, and just to put that in context, even a few, if you look at a poorly controlled diabetic neuropathy population, you might anticipate that there's a worsening of several points over that same period of time, so 28 really highlights how devastating and progressive ATTR polyneuropathy is. If we look at the treatment group with patisserin, not only does their MNIST plus seven score not worsen, but there's actually some improvement of six points by the 18 months, and so this was a very striking result and a clear benefit for patisserin in terms of not only slowing down, but improving neuropathy symptoms in ATTR patients. The next generation of patisserin is an sRNA called mutrisserin, and this was studied in the HELIOS-A trial. Some comparative differences in terms of the route and frequency administration are shown here, so patisserin is an intravenous medication given every three weeks. Mutrisserin is given subcutaneously and only every three months, so some advantages for patients. The HELIOS-A trial used the historical control from the Apollo group, so shown here in the red line, and you can see with mutrisserin that the MNIST plus seven score is stabilized to somewhat improved by the end of the trial at 18 months. In terms of the degree of knockdown that we get with these sRNAs, for both patisserin and mutrisserin, that's around 80 to 85 percent, and you can see that transliterated knockdown happens pretty quickly in the first few weeks of treatment initiation. Another group of transliterated silencers that have been studied are antisense-augment nucleotides, and the first generation ASO is onotericin, which was studied in the NeuroTTR trial. Similar design to the Apollo study, looking at variant ATTR patients with polyneuropathy, 172 patients, and what you can see here is that there's a worsening in the MNIST plus seven in the placebo group, similar to what we saw for Apollo, and with inotericin, there is a slowing of that MNIST, of that worsening in neuropathy, so not quite a reversal in their symptoms, but clearly a difference in terms of how patients progress in the treatment group. There's a couple important side effects to note that were observed in that trial. Patients could have rarely thrombocytopenia, as well as glomerulonephritis, so those are things on the label in terms of monitoring patients when they're on this therapy. The next generation of antisense-augment nucleotides is eplantericin, and so shown here is the difference in administration, so inotericin is subcutaneous, given every week, and eplantericin is subcutaneous, given every month. This was studied in the neurotransform trial, and you can see that eplantericin stabilizes that worsening in the MNIST plus seven compared to the historical control. And the knockdown that we see with eplantericin is in that 80 to 85 percent range, and you can see kind of how quickly translucent knockdown happens. So, what about cardiomyopathy? So, we're just starting to get some data for transdiuretin knockdown in cardiomyopathy, and the first phase three, large phase three trial to look at this is the Helios B study with mutriceran. So, this was, this looked at wild type and hereditary patients, 665, 55 patients. Important to note that about 40 percent were on tefamidus, and about 40 percent were on tefamidus, and what you see here is the primary endpoint in the overall population, and so there's a statistically significant benefit for the mutriceran group over placebo at 36 months, and not shown here are some extension data that go out to 42 months that show similar trends in the treatment group, and so with these data, mutriceran is currently under FDA review for extension of its label into cardiomyopathy patients. And finally, in the knockdown space, our therapy is looking at permanent lowering of transdiuretin, and so NEX-Z is one of those, is that therapy that's currently in development for this, and this is a CRISPR-Cas9-based therapy that targets the hepatocytes in the liver to try and knock down transdiuretin with a one-time treatment. This is one of the initial dose escalating studies that was published a few years ago, and you can see at the higher doses that you get greater than 90 percent knockdown with NEX-Z. So, if we look at the current FDA indications for the therapies I mentioned with the phase three trial results, currently tefaminis and aciramis are both approved, FDA approved for ATTR cardiomyopathy, whether you have variant or wild-type ATTR, and for the knockdown therapies, batuceran, inotericin, vitriceran, and eplantericin, those are all FDA approved for hereditary ATTR with polyneuropathy, and vitriceran is currently under review for expansion of its label into ATTR cardiomyopathy, and we expect a decision for that next month. So, in earlier stages of development for therapies, so I think that for both the silencers and stabilizers, these treatments work the best in earlier stage disease, and we think that they work predominantly on the production end, so decreasing the amount of new amyloid that's forming, but what about amyloid that's already in the heart? There's an unmet need there to try to remove that amyloid and potentially reverse remodel the heart, and so an approach that's in kind of earlier stages of development are antibodies to see if we can remove ATTR from the heart. This is a letter that was published in the New England Journal a little under two years ago, where the amyloidosis group in the UK demonstrated that there are a few patients in their cohort that had naturally occurring ATTR antibodies, and that led to clearance of amyloid over time, which could be seen through a variety of imaging modalities, and I think this helped. This is a good proof of concept that perhaps we can develop targeted antibodies to ATTR amyloid epitopes by promoting clearance of amyloid fibrils, and so there's a number of antibodies in development to try to remove amyloid. This is just one of the studies shown here. This is a phase one study looking at one of these antibodies, and what you can see in the 40 patients that were studied in this trial is that some of them had evidence of clearance of their amyloid by MR and technetium based imaging. There was no significant safety signal, and I think it's encouraging data for the phase three trials that are ongoing. So, this is just to highlight some of the trials that are ongoing in later stages of development. We have eflantericin, which is one of the antisense aglonucleotides. It's already approved for neuropathy. There's a cardiomyopathy trial that's been ongoing for a few years now, and so we wait the results for that. Next, we have a few antibody therapies, karamatog, which is in phase two trial ATO2, another monoclonal antibody being studied in ATTR. Next, Z is the CRISPR-Cas9 based therapy, which is an ongoing phase three study called Magnitude, and then finally, we have another monoclonal antibody being studied in the depleter CEM trial, and so that was kind of an overview of a lot of the therapies that are in development, and there's even more things that are in the preclinical space, so definitely a really exciting time for ATTR in terms of providing a variety of therapies for our patients, and I think one of the things kind of in years to come will be to figure out what the right sequencing or combination of therapies are for patients. So, now I'm going to go through the post-test questions. Currently in development, CRISPR-Cas9-based in vivo gene editing therapy is targeting A, cardiac troponins, B, intubrobian P, C, TTR, and human hepatocytes, or D, urinary albumin to creatinine ratio in adults with hereditary transthyretin amyloidosis and polyneuropathy. So, for those of you that answered C, that's the correct answer. The next question is, data from the Apollo trial showed us that patients in the choices are patisserin, A, B, inotericin, C, botryserin, or D, eplantericin, had improvement in multiple clinical manifestations of hereditary amyloidosis including their neuropathy impairment compared to those in the placebo group. And the answer for this question is A, butyricerin was studied in the Apollo trial and showed these benefits. So at this time I'd like to hand over to my colleague Amy Brunel to go over a few ATTR cases. Thank you so much Dr. Alexander and Dr. Grodin. Now that we're all experts in our diagnosis process, pathophysiology, and treatment, we'll talk a little bit about some engaging case vignettes and real-world insights. And throughout this presentation I'd like to engage my colleagues with some questions because these are really patients that we've all taken care of in the amyloid space. All right, so case number one, it's not just the valve when aortic stenosis brings a plus one. We're going to keep this really light-hearted here. So Mr. K is a 79-year-old Caucasian gentleman with a history of severe aortic stenosis and hypertension. He's referred for a TAVR evaluation for symptomatic severe aortic stenosis. Ongoing symptoms are he endorses fatigue, dyspnea, exertional dyspnea, mild bilateral lower extremity edema over the last year. Some of his pre-TAVR workup, he had an echo done which did show severe aortic and thickened ventricular walls, septal thickness of 2.1 centimeters, concerning for an infiltrated cardiomyopathy. He also had global longitudinal strain with apical sparing patterns. These are all those red flag symptoms that we kind of talked about earlier. Cardiac MRI demonstrated elevated ECV and diffused late gadolidium enhancement with sub-adicardial enhancement, abnormal myocardial nulling, and by atrial enlargement and mild pericardial attenuation. He had some nuclear syntagraphy imaging done which showed to be positive for amyloidosis. And of course, his negative serum-free light chains, serum and urine IFE, and he had genetic testing done which was deemed negative. So what does Mr. K have? So he has our wild-type cardiomyopathy, amyloidosis. So some of our diagnostic challenges that we incur are related to concurrent pathophysiologies. The presence of both severe aortic stenosis specifically with wild-type ATTR cardiomyopathy does complicate the clinical picture. Symptoms of dyspnea and fatigue could initially be attributed solely to his aortic stenosis, especially as severe as it is, and may also be driven by an underlying amyloidosis. So this overlap can lead to an unrecognition of our ATTR wild-type cardiomyopathy. So undergoing our TAVR decision-making process, the discovery of ATTR wild-type cardiomyopathy introduces the complexity in our decision-making for this population. There is limited data really on outcomes of TAVR patients with co-existing amyloidosis, and this can make it challenging for predicting procedural risks and benefits. So we have a lot of unmet needs with this patient population. Enhanced screening, there is a need for a more rigorous screening regarding this population. Our elderly patients undergoing TAVR, we estimate about 16% of these patients undergoing TAVR evaluation. It could be attributed to amyloidosis, and especially those with unexplained ventricular thickening or heart failure symptoms that are disproportionate to the degree of aortic stenosis. And we really do need some more research. It is needed specifically in this patient population to help guide treatment decisions to improve that patient counseling portion. This disease requires multidisciplinary care. Increased collaboration specifically between cardiologists, heart failure specialists, amyloid experts are needed to manage this patient population that have concurrent aortic stenosis and amyloidosis effectively. So I'm going to open up the floor to our resident experts, Dr. Alexander and Dr. Grogan. So regarding this diagnostic overlap in this case vignette, how do you differentiate between symptoms that are caused by severe aortic stenosis and those that are potentially caused by the amyloidosis in this elderly patient? Dr. Grogan, do you want to answer first? Sure. Well, Amy, I'm delighted that you presented this case because it is a challenge that's not that uncommon that we do see from time to time. And I think it's not without consequence, right? Because the implication, as you so astutely put, is whether or not we should address the valve and fix it. I can tell you that, you know, based on my experience in our evaluation, you know, we tend to do a pretty thorough hemodynamic evaluation where we'll leverage even invasive hemodynamics to truly understand is the valve severe. We also rely on CT and other non-invasive techniques. But, you know, we do intervene on these individuals. And I can tell you at least empirically, and of course, maybe this is a selection bias, but a lot of patients do feel better and do get improvement post TAVR. Now, would they gain the same amount of benefit that somebody who does not have amyloidosis achieve? You know, I think that's harder to know. I would assume probably not. But I'd also be interested to hear Dr. Alexander's opinion on this as well. Yeah, I think, you know, we have a similar approach, you know, because there's going to be a spectrum of patients where perhaps the valve is causing more of the symptoms in pathophysiology, or it's more the ATTR. And it can be really challenging to parse out. But I think hemodynamic assessments and imaging can help to understand how much of this is the aortic stenosis. In many cases, we pursue TAVR in them. And I think that the fact that they, like Dr. Grodin pointed out, feel immediately better after the TAVR suggests that the aortic stenosis is a big driver for many of these patients. But I think there's great opportunity to identify ATTR at this stage because of the therapies that we have for this. And perhaps that will help patients to feel better in the long run than if they would if they weren't started on ATTR therapies, you know, at the time around TAVR. Thank you so much, Dr. Grodin and Dr. Alexander. Now, Dr. Alexander, to kind of spin off of that, are there specific risks or benefits that are associated with undergoing TAVR in this patient population if they do have amyloidosis? Or do we just not have enough information and research at this time? Yeah, that's a good question. I mean, I think that if someone has underlying amyloidosis, that they're probably at higher risk for things like conduction abnormalities or perhaps atrial arrhythmias post-procedure. I think it's just kind of an added risk factor for some of the things that we typically watch for post-TAVR. But I think most of the managements are being pretty similar. Great. Dr. Grodin, do you have anything else to add regarding that? Well, I mean, I also think, you know, we still have to put our internist hat on and assess frailty and really make sure that what we're doing we think will, you know, give some global benefit much in the same way we would do with anyone with aortic stenosis. And I think to dovetail with what Dr. Alexander was saying is we really do take a heart team approach to these individuals and make sure that we're being as thoughtful as possible and really, you know, identifying all the domains that might influence their outcome. That's great. Dr. Alexander, you so beautifully covered all of the new technologies that we're utilizing as well as therapies. Is there a specific role for amyloidosis specific therapies to improve these outcomes in patients that have wild type ATTR that are undergoing TAVR? Yeah. So, you know, this hasn't been studied in any randomized controlled trial, but there's a number of groups that have been looking at their respective cohorts. There's a group in France as well as some groups in the U.S. that have looked at outcomes for ATTR therapies. Meaning to families predominantly. And, you know, I think that in observational cohorts, there's some signal that there is a benefit to identifying ATTR earlier and starting treatments. But, you know, I think the big caveat is, you know, this hasn't been studied in a randomized fashion. I think that as long as ATTR is a significant component of their pathogenesis, kind of going back to how Dr. Groh and I were talking about parsing out kind of how much of this is the valve, how much of this is cardiomyopathy. But I think, you know, to me, it would make sense if someone has, you know, some degree of clinical ATTR that they benefit from being on therapy in the long run. Yeah. Who knows if we do this talk six months from now, things may change. I mean, just in the last few months, since we were supposed to have HFSA until now, we've had a newer therapy that's been FDA approved. And there's just a lot of exciting things that are on the market for this disease process. So diving into our second case vignette, when your DNA throws a party, but your heart doesn't RSVP. So this is a very common case now that we're doing familial testing and following up. So Ms. J is a 42-year-old African-American female. And she comes to you reportedly with a family history of amyloidosis. She's currently asymptomatic, but she's undergoing genetic testing and counseling after her older brother was diagnosed with the hereditary variant of B.1.22i associated. She didn't have genetic testing completed, which demonstrated that she was positive for the B.1.22i variant, which we know is in about three to 4% of the African-American Afro-Caribbean population here. Our cardiac workup that we did upon our initial clinical evaluation, her EKG demonstrated normal sinus rhythm, no evidence of arrhythmia, no conduction abnormalities, so pretty normal EKG. Echocardiogram demonstrated normal left and right ventricular size and function, no evidence of hypertrophy or any structural abnormalities, normal wall thickness, and any sort of valvular issues. We also drew an antepropion P intraponin, which were all within normal limits. So this is case vignette really clearly covers our genotype positive, phenotype negative patients. And some of our challenges that we see diagnostic, our screening recommendations, you know, these challenges lie with determining that optimal timing and method for screening in this specific population of genotype positive, phenotype negative individuals. There's really not clear enough guidelines on how frequently these patients should be monitored or specific tests such as echocardiogram, cardiac MRI, or serial biomarkers being utilized just to detect early signs of involvement. Now this may change in the future as our technology develops. Patient management, there is that uncertainty of disease progression with it being autosomal dominant with that variable penetrance and creating that challenging counseling those patients about the risks of developing cardiac manifestations, the need specifically for lifestyle modifications to maintain a healthy heart and positive benefits for early therapeutic interventions. Additionally, I'd say there's that whole psychological impact that's associated with the genetically inherited disease. And that can really have a cascade effect on family and the patients themselves. Regarding our unmet needs, we really need to develop standardized screening protocols specifically for genotype positive, phenotype negative individuals and improve biomarkers or imaging techniques that can help to predict that clinical onset of disease, helping us to better understand that natural history progression of our ATTR cardiomyopathy in this population of genotype positive, phenotype negative to help guide early intervention and strategies. And I know there's some pipeline clinical trials that are ongoing for acting early. So we're, you know, excited about what this brings. So I'm going to kind of open it up to some questions regarding this patient because this is a common patient that we'll see in our clinic. And regarding screening and early detection. So what, Dr. Brodin, what are current evidence supporting on the optimal timing and method for screening of genotype positive, phenotype negative individuals for early cardiomyopathy? This is like, we could go down a rabbit hole with this, I guess. Yeah, welcome to the rabbit hole, everyone. So I mean, this is an area of research interest of mine. So I could probably have a whole other session on this. But I think you highlight a really, really important point, especially as genetic testing becomes more common as to, you know, what do we do with these individuals that find out their carriers? I think everybody's aware that 23andMe reports three variants, B.1.22i, T.60a and B.30m. So there are people that are finding this out in the community. And then you asked a great question, what is the evidence? In short, it's not great. So much of the recommendations are based on expert opinion. And, you know, that expert opinion identifies a crucial time, which is called the predicted age of disease onset for a carrier with a specific variant. Now, one of the nuances about this predicted age of disease onset is that this can vary for the different mutations. So it's really incumbent to understand what is the natural history of the mutation? And then what are the types of, or what are the, what is the clinical syndrome or maybe what types of early signs should we be looking for? And that does vary. So when we talk about what is the data, much of what we know from the natural history of carriers of pathogenic variants comes from individuals that carry the B.1.22i or B.1.42i pathogenic variant. And that we really don't see that it's all that penetrant until individuals are later in life. Most frequently from the seventh or the sixth, maybe, but the seventh decade of life and beyond, we start to see a greater risk of heart failure in comparison with non-carrier controls. So I'd be very interested to know what, you know, Dr. Alexander's experiences in this field. Of course, I'm sure we're going to get into, you know, how do you test these people? How frequently do you see them? But, you know, I know Kevin's given some thought about this as well. So I'd be interested to hear his comments. Yeah, another fun and exciting thing in the amyloid space is thinking through how to manage these patients. And, you know, like Dr. Groten is saying, we're really at the expert opinion phase of all this. So I guess just some thoughts to add to it would be that the, I think our approach will largely be kind of age dependent. So meaning, I think the buckets of people that are very young, decades before their mutation, people that are kind of approaching that predicted age of disease onset, and then people that are kind of well into that. And, you know, I think just kind of thinking of those three big buckets, you know, our approach is going to be tailored to those. And so I think for the very young people, and this patient's kind of in that range at age of 42, you know, I think we have to be cautious not to over test and over, you know, stress people in terms of what we do for them. You know, these things aren't trivial in terms of cost necessarily, if you're doing technetium pyrophosphate scans annually, or, you know, I had a lady the other day, that was 35 and gotten biopsied somewhere to screen for ATTR. I mean, I think that while we want to have detect disease early, there's kind of a, there's always kind of a threshold where it probably makes sense to do that. And then, you know, also the psychological impact. I mean, I think that if you're 30, and now you have this variant, and potentially you and others attribute symptoms to that variant, it's very unlikely to be due to that. And again, I've seen kind of other cases where other cardiac conditions were present. And they, that person just happened to have the D122I. So I think that the education messaging around there has to be important. And also how we go about cascade testing in those younger, younger patients. I think genetic counseling is extremely important to kind of even decide when to test. If someone's 20, I don't think they necessarily need to be tested unless they've been educated and counseled and really, really, you know, want to do it. I don't think we should, as a medical community, push that on someone that young. And, you know, as people get into that predicted age of disease onset, I think the testing approach is going to, you know, require study. But some of the things, I think some of the biomarkers that are being developed, like Dr. Grodin was saying, cardiac MRI, I think are tests that can really catch patients in those, that phase where there's TTR aggregates or early myocardial infiltration before there's any kind of arrhythmia or heart failure consequence. And, you know, I firmly believe that treatment initiation at that phase is going to be highly effective at preventing progression or slowing progression dramatically. So kind of what tests and the timing of that I think still needs to be worked out. But cost and invasiveness and radiation, I think, are some of the factors to think through. And then, you know, for those patients that are in that age where they could develop disease, so say a sibling that gets detected during cascade testing, then I think we should be more aggressive about how we follow those patients longitudinally. And if I can add, you know, Amy, I think one thing to what Kevin said, I mean, he's right on the money, but I think this really emphasizes a number of the key points he just made. But we do know from epidemiological cohorts, at least of individuals that are carriers of the B.1.22i variants, and then even other cohorts of patients that might, or of individuals that might have other variants, that there are some subtle clues, or at least there's some signals that something might be going on. So for example, we know that individuals such as this, that, you know, the case that Amy presented that are in their 40s that carry a B.1.22i variant, that even individuals on average in their mid 40s up to the mid 50s may have subtle differences, maybe in their heart's wall thickness, or maybe their heart's stiffness, that could suggest there could be amyloid involvement. But I think of one clue, or one key point to recognize is that nobody tested for amyloid in a lot of these studies, but it is compelling. Now it does suggest that something might be there. So I think, you know, so this really supports the importance of what Dr. Alexander was saying, and you really have to be thoughtful about what you order and when you order it, and then all of the downstream implications. Yeah, those are really great points. And I think, you know, especially the Dr. Alexander's point that familial testing and also counseling piece is just so incredibly important. You know, we have laws that protect against preexisting conditions and coverage for health insurance, but people forget about life insurance all the time. So I think that's an important part. You know, if you're six years old, I mean, you're not loading up on life insurance, so to say. So making those informed decisions are very important. And like you mentioned, Dr. Groban, that downstream effects of cost of screening, you know, we don't have clear guidance, you know, how routine, is routine cardiac imaging being done? But what I find is really fascinating, especially with, you know, certain variants, like the V122I, is that you look back retrospectively at these patients, and the Penn Medicine study showed, you know, 48% of them had a history of like bilateral carpal tunnel with at least 10 to 15 years before they ever developed any cardiomyopathy symptoms. So doing a better job, I think, at recognizing those red flag symptoms, and then initiation of therapy, whatever that therapy is, as sooner as possible, we'll have better outcomes long-term. So regarding the natural progression of disease, what do we, I mean, we talked a little bit about, you know, V122I, are there certain variants that you may, you know, see neurologic features sooner than others, and kind of where do we start? If I'm working even in cardiology or primary care, what does that natural progression look like for a patient that's a genotype positive, phenotype negative? And then how, as a clinician, do you kind of guide your clinical decision-making based off of that genotype? I don't know, Dr. Alexander, if you want to start. Sure. So, I mean, I'll focus on the three variants that we see more commonly in the US. It'd be V122I, T60A, and V30M. So those latter two, many of those patients have some component of neuropathy, or it might be the primary driver of their amyloid manifestations. And so you can either have a scenario where the cardiomyopathy and neuropathy develop in parallel or mostly neuropathy symptoms that develop and the cardiac symptoms become, you know, manifest later in the disease. And so I think that that's one of the challenges, is that what patients develop might not necessarily just be kind of the pure cardiomyopathy and people could present to other providers with symptoms. And so if you have someone that's gene positive, I think it's important to have a multidisciplinary assessment and have that patient on your radar. And they don't necessarily have to see six specialists longitudinally, but I think that getting some baseline assessments and at least having someone that's quarterbacking and recognizes when to refer to the appropriate person's important. And I think that's what a lot of amyloidosis centers, what we provide is that guidance. And also from the patient's perspective, if a new symptom develops, having a low threshold to alert, you know, your amyloid team to do further investigation, certainly if you're going for some procedure, say GI procedure, and you're getting endoscopy, you know, getting biopsies and that sort of thing and standing for amyloid or things to consider. But so I think to answer your question, you know, it's important. I think you highlight an important concept, which is that you have to think outside of the heart for some of the, for these variants, all the variants, but particularly the ones that can have a mixed phenotype or primary neuropathy phenotype. And that's going to take a multidisciplinary approach. That's great. Dr. Groden? Dr. Groden thinks so. Yeah. Yeah, I couldn't agree more. I think really what, I mean, this is a nice example of taking a genotype first approach in medicine. So, you know, if we see a carrier, then as Dr. Alexander highlighted, I think it's really incumbent, you know, to understand what is the expected phenotype. And I think you see this cartoon here on your, or this figure here on this slide. And I think it really conceptualizes this a lot. I think it's very important to have an understanding of when you would expect the disease to show up in an individual with that variant, and then what might be some of the features. And then that can certainly inform how you would follow this person, how you would do so thoughtfully, and what testing and what signs you would look for. That's great. Let's open it up to both of you. Are there, this is a million dollar question. Are there any current or emerging therapies that we can possibly use prophylactically at this point in time, in this case scenario, genotype positive, phenotype negative patients, to prevent the onset of cardiomyopathy from occurring? Anyone want to, Dr. Alexander, you want to go first? Sure, either way. We're going to give the same answer. I know, I know. Which one? Drink your green tea, I don't know. Yeah, I mean, I think that, so we don't have trial level data yet for prevention, but as you both know, there's the ACT-Early study, which is going to look at, or is looking at aciramidus, so one of the stabilizers approved for cardiomyopathy, to see if that can prevent the onset or delay the onset of ATTR, polyneuropathy or neuropathy, in that gene positive asymptomatic population. And so patients will kind of get ruled out for amyloid at screening, and then we follow it longitudinally with cardiac and neuropathy assessments for development of amyloid. And I think the important thing in that trial, and it comes back to what Dr. Grodin's comments, is you want to study a population that's in that predicted age of disease onset range. You know, if you look at a bunch of 20 year olds, then you'd have to run a trial, you know, probably long after we've all retired from medicine. So, you know, I think that that'll be interesting to see, kind of, you know, do we hit the sweet spot of identifying that at risk group of patients, and then see in that group, does TTR stabilization improve outcomes? Dr. Grodin sounds about right. Yeah, I was going to say the same thing. I mean, I think we'll see. I think it's very, very compelling. And, you know, again, this is, you know, we're talking about treatments that are developmental, but, you know, some of the questions that come along are, you know, I think Dr. Alexander in his talk highlighted the potential for gene therapy with CRISPR-Cas9 based technologies. You know, I mean, do we take it to that extreme? And somebody that is harboring such a pathogenic variant that has afflicted their whole family, at what point does that prevention, you know, when do we, when would we implement something like that? So there are a number of really interesting hypotheses that could be tested, you know, but right now, I mean, I think until we, until we have the results from ACT early, and, you know, it's really going to be, it's really going to be in question, which I think makes it, you know, really key just to underscore everything we've just discussed in the last few minutes that, you know, we really want to find the disease early, and then that's really a key point to implement therapies where, you know, we can kind of halt that progression, that curve that I showed you earlier in this session. Yeah, and to your point, Dr. Grodin, is there, you know, if I'm an ancient patient coming with genotype positive, phenotype negative presentation, is there any advice that you would give me as a patient regarding lifestyle modifications or any non-pharmacological interventions that I could incorporate to potentially delay the disease, or we just don't have that information at that time? Well, I can tell you that there are anecdotes, and there are off-label therapies with, you know, that really don't have a strong evidence backing to support their use. You know, some of these include green tea extracts, doxycycline with something called turo-urso-deoxycholic acid, maybe turmeric, but it's really unclear if those do anything, number one, in a human being, and it's also unclear if they have any efficacy beyond, you know, their own pharmacological uses. So I don't necessarily routinely recommend those, and I think what is key is, just like Dr. Alexander said, you know, laying out expectations, reassuring about a timeline, and I think one thing that's very important to emphasize is that this disease does not necessarily move very fast. It's very common that it is very slowly progressive when it's in its kind of preclinical or subclinical phases, and so I think, you know, reassuring them that, look, if we see anything, we're gonna be catching it early, and then we'll implement real therapies at that point when we detect the disease. Absolutely. So we have a couple questions from the audience, and feel free, we have about 10 minutes left of this webinar. We'll open it up. If you guys wanna ask us some live questions right now, we're able to answer them on the air. So the first question I have is, do you recommend MRI screening as a test for genotype-positive, phenotype-negative patients, like the one we just discussed? Dr. Alexander? Sure, yeah. Dr. Grunin, this is one of his primary areas of research, so we'll give a, I think, a little more nuanced answer, but I think that, I like it as a screening tool for gene-positive, phenotype-negative for falling patients, because I think it's quite sensitive for early amyloid, and it kind of comes back to our discussion earlier about the age at which you do that, and so I think when people are getting to that predicted age of disease onset range, it's a nice test because you avoid the radiation from serial nuclear imaging, but still have a lot of sensitivity there. So I think it's reasonable. I mean, in the US, I think MR is a little bit more expensive and difficult to get, and the interpretation, there's some variability about where, depending on where you go, compared to the UK, but I think that, you know, in amyloid centers and places that have, you know, good MR readers, it's a great test, I think, for this group of patients. Yeah, and I would agree. I would say if we have to take our own temperature, we really don't know what an early amyloid heart looks like. I mean, we have some ideas, and I think a lot of people, and I would agree with Dr. Alexander, we speculate that maybe MR is more sensitive than some other diagnostic techniques. I think what we don't want to do is, you know, is encounter a patient like Dr. Alexander suggested earlier, where they were a carrier, and we biopsied a young person way before their predicted age of onset. So I agree, MRI's got a number of advantages. It's really the gold standard for assessing, you know, the structure and function of the heart, and complete with contrast-enhanced images. You know, but an MRI is really kind of like a, you know, a filet mignon. You know, you can get one at a restaurant around the corner that's not that fancy, but you can also go to the fancy steakhouse downtown, and even though they're technically filets, they're very, very different. So again, it's not something that's widespread, or the expertise can be a little bit variable. I tend to use them a lot for many of the same reasons that Dr. Alexander highlighted. You know, we are currently actively answering, or trying to answer this question, where we're enrolling a nested case-control study of individuals that carry pathogenic TTR variants in age, sex, and race-matched non-carrier controls, and they're going through this really interesting battery of exercise and resting images, complete with strain everywhere, on a 3-Tesla MRI, and so we'll see what we can see, and that will hopefully get us a better answer, you know, to your question. But yes, I use it, but I don't repeat it on an annual basis. You know, I might, depending on their age, I might pepper in just ECGs or echocardiography, and I try to be thoughtful with the cadence with which, you know, we order these tests. Absolutely. Have you had any issues with prior authorization or insurance coverages because of a patient like this? You know, type positive, you know, type negative. No, not for an MRI or PYP. We've actually been pretty easy. Great. So, Dr. Grodin, you did a fabulous job covering the pathophysiology of the disease. One of the questions that just came through was, are there any changes that are noted in the thyroid function with TTR cardiomyopathy being used for treatment? That's a great question, because, and so, TTR is really not the major carrier of thyroxin in the blood, and this is a safety endpoint that's assessed in all studies, and to my knowledge, I don't believe that there has been any signal for impairments in thyroid function in any of the studies, and this would obviously be far more important for the silencers, which deplete circulating TTR. Regarding, you know, Dr. Alexander, you covered the medications that are available, some that were experimental. It's kind of a brave new world. We've only really had FDA-approved therapies since 2018, so it'll be something we'll talk about 20 years from now and say, remember when we had nothing for amyloid and all we could do was a liver transplant or a heart transplant? But what about diplonasol? Are you still using that at all in any of your patients and individuals who are at high risk, like homozygous, and this is a question that came through. Yeah, I mean, I think backstory on diplonasol, so that's a non-steroidal anti-inflammatory drug that was later found to have TTR-stabilizing properties, and I think many of us used it more before tefamines was approved. I mean, when there are no therapies, we know that there's stabilization, you know. I think some patients are, you know, we try that and there's a benefit. One of the challenges is a lot of those patients have cardiac conditions where usually the brain says they're contraindicated and so really it's kind of a niche group of patients where we might think about it. And, you know, some people do use it in those pre-symptomatic carriers. To me, you know, and this is probably, there's probably variations of opinions, but I mean, I think now that we have multiple approved therapies with better side effect profiles and, you know, we're developing more nuance about how to manage these gene-positive, gene-positive patients, I think we can monitor many of them closely and then start, you know, an FDA-approved therapy if they were to develop disease. And for homozygous patients, you just move their predicted age of disease onset earlier and their penetrance is higher than, you know, those heterozygous patients. So probably whatever screening protocol you're using, you're going to look at, you know, you're going to monitor them at an earlier age and you're going to probably monitor them a little bit more closely. But I think a similar, you know, I would probably do a similar approach to how I would follow heterozygous, gene-positive phenotype negative patients. Thank you for that. Okay, that concludes an excellent panel discussion. I'd like to thank both of you for outstanding talks, outstanding sessions. I think the panel discussion was really thought-provoking and then I especially want to thank every one of you taking time out of your evenings to share it with us and listen to this session. Again, reminders to claim credits. These are the steps. You've seen those quite a bit in this talk and before the session. So again, I'd like to thank you all. I'd also like to thank our sponsor, Alnylam, the HFSA for organizing and supporting this very important session. And then I'd really like to thank my colleagues, Dr. Alexander and Amy Brindel for really fabulous talks and discussions and really great remarks. So I hope you all have a good night. I would like to thank you again for tuning into this session entitled, Where Are We With Cardiac Amyloidosis?

cardiac amyloidosis

ATTRCA

diagnostic practices

emerging therapies

transthyretin stabilizers

silencer therapies

CRISPR-Cas9

multidisciplinary care

early intervention

MRI

genotype-positive

aortic stenosis