Welcome, everyone. My name is Dr. Clay Hayes, and I'm welcome to, I'm proud to welcome everyone to our program tonight, sponsored by the American Society of Nuclear Cardiology. We'll be talking about suspecting and diagnosing transveritin amyloid cardiomyopathy in underdiagnosed populations at risk for heart failure. Our program faculty, as you can see here, we want to talk about our learning objectives. You can see the different ones, and they should have been provided in your packet. Here are the disclosures that we'd like to let everybody know about. And then tonight is our agenda that we want to try to discuss when we talk about cardiac amyloidosis. Some housekeeping requests. Please try to mute your personal cell phone as well as muting your computer audio. And if you have questions at the end of the program, we'll be taking those via Zoom, the Q&A button at the bottom of your screen. If you want to obtain your CME or certificate, you can claim that at the links listed below. Obviously, if you're a technologist, you also would like to claim credit, and we'll be displaying a six-digit code at the end of the program. And also, social media. It's important to do that and let people know what kind of experience you had at today's program. We will start next with a case presentation. I will briefly switch over to this in just a minute. Nothing like the power of Zoom. Okay, today's case presentation is about a gentleman that I first saw when I first started practice. He was an 83-year-old man who I started treating for coronary artery disease, and I had a chance to put a stent in his circumflex artery, and he did well for a number of years. He got the usual risk factors, or developed the usual problems that men do in America, including hypertension, hyperlipidemia, carotid artery disease, paroxysmal atrial fibrillation, prostate cancer, and GERD. As part of a preoperative workup for prostate surgery, but he had prostate cancer, he had a normal exercise MyoView study in 2020, and also an echocardiogram. This was his preoperative EKG during that time period. You can see he was in sinus rhythm and had a right bundle branch block. This was a preoperative echocardiogram that he had. You can see his LV function is normal. He does have a thickened left ventricle, but no major abnormalities, and I'll just show some of the brief images here. That was a parasternal long axis. Here's a four-chamber apical view. You can see that his LV function was fine. Just some of the diastolic inflow you can see. Also the tissue Doppler on the lateral wall, and also the medial segment. He had his surgery, had a prostatectomy for his prostate cancer that was uneventful and did well. He came back to see me for his regular checkups, and in July, he started having chest pain and underwent a nuclear stress test, which I thought showed inferolateral ischemia, or we did. Here are his nuclear images. You can see his LV function still was good, but he did have a reversible defect in the inferolateral wall. He went on to have a heart catheterization and was found to have multivessal disease and had a three-vessel bypass. His ejection fraction following surgery was normal, and he did well. In the spring of, or the late winter of February, he did have an inguinal hernia repair. He had no problem, but over the summer of 2023, he noted there was a decrease in his stamina, and he had more dyspnea with exertion. In August of 2023, he developed chest tightness, which was concerning. We did another cardiac PET scan this time to look and see, and we were concerned about the abnormality, particularly on the top row that you see there. There's definitely a difference. So we thought his ejection fraction had dropped down to 25%. We did another heart catheterization, which showed that his lema was patent, but the vein grafts up to his marginal branch and diagonal branch were occluded, and again, we were concerned about his poor LV function. We adjusted his medicines by putting up titrating his beta blockers. We talked with the surgeons. They didn't think he was a good candidate for redo bypass surgery because of his lema was so good. Again, at the end of September, he had some improvement of his ejection fraction, a lot of improvement in his ejection fraction, but his right heart pressures were elevated, and then he still complained of dyspnea, particularly in January of 2024. Again, he's an 83-year-old man, and we're thinking coronary artery disease. We got a follow-up echo in January of 2024, and we noticed some things had changed about this particular echo. We noticed that the thickness of his wall seemed to be a little different, and this is the apical four chamber. Again, we had some concerns and said, well, maybe we're not thinking about this the right way. Again, you see his diastolic inflow, tissue doppler from the lateral wall and also from the medial wall, and then we said, well, we check in his lab work. We said his pro BNP was quite elevated, 12,000, and his creatinine was 1.6, and we said, hmm, what should we do? We had some concerns. Obviously, his right heart pressures are still quite high. He had mild to moderate aortic insufficiency and moderate mitral regurgitation. So our next step was to think about amyloid. We did a PYP scan. We have this particular modality in our hospital, and this is the scan results here. So I'm gonna stop this particular presentation, my portion of this, and we're gonna move on to our next speaker, Dr. Frederick Rueberg, who's gonna discuss what is cardiac amyloidosis. Dr. Rueberg. Thank you very much, Dr. Hayes. I hope everybody can hear me okay. I'll begin my screen share. Okay, hopefully we can see this. So I have been tasked with a straightforward kind of topic of what is cardiac amyloidosis, setting the table for all the great speakers you're gonna hear later today, and you're gonna hear a number of themes that I'm gonna show in my presentation that echo, no pun or pun intended, what Dr. Hayes just showed you. So here's my research support. I didn't mention I'm from Boston University in Boston, Massachusetts. So you just saw this echocardiogram about two minutes ago. This is a patient who is an 85-year-old African-American gentleman who has heart failure. And as you can see here on the peristronal, long and short axis view, that the heart is thickened and the ejection fraction is normal, the left atrium is big. And for those who aren't accustomed to looking at cardiographic images, this is what a normal heart looks like. So we've got wall thickening here, and the key is basically that the ejection fraction's normal, there's marked increased wall thickness. So you go ahead and get electrocardiogram, and your electrocardiogram looks like this. The patient is in atrial fibrillation and really has prominently reduced voltages in the limb leads, and also looks like they've had an anterior infarct before. So this is the first clue that amyloid may be present, and that is discordance of the echocardiographic wall thickness and the predicted voltage. Most patients with hypertensive heart disease or LVH will have increased voltage. And so anybody with disproportionately reduced voltage and increased wall thickness should be a red flag. And in this particular case, this patient has an infarct pattern, but has not actually had an infarction. So I've already named two of the differential diagnoses here, the primary differential diagnoses. We're going to talk a lot about number three, and then finally a more uncommon cause of wall thickening, which would be a storage disease like fibrillates. But by and large, the differential diagnosis is going to involve hypertensive heart disease, hypertrophic cardiomyopathy, or infiltrative disease like amyloid. So what is amyloid? Amyloid basically is derived from the Latin word amylom, which means starch. And starch or starch like substance was what Virchow in 1854 first described as amyloidosis. Amyloid is a protein folding disorder. Protein folding, so all amyloid protein, all proteins need to fold normally, excuse me. But in the case of amyloid, proteins misfold. And the diagnosis of amyloid is made primarily by histology. And that's where aggregates of beta sheets that stain with a particular stain called Congo red are identified. We're going to talk a little about this in the next slide. And this is an example of a cardiac amyloid biopsy specimen showing Congo red staining that has green birefringence under light microscopy. Now the systemic amyloidosis are classified by their precursor protein. And amyloid, the precursor protein is the protein that misfolds. And amyloid can deposit in soft tissue and in visceral organs like the heart and in the peripheral nervous system. And the nomenclature is A for amyloid followed by an abbreviation of the protein that misfolds. So Congo red actually is a textile dye. It was developed in the 1880s, has nothing to do with histology, at least at its discovery. But it was discovered to actually dye clothes red. And because it was discovered in Germany at a time when there was a lot of interest in Africa and it was a continent that had a lot of mystique. So they chose the name Congo to make it seem exotic. But it really has nothing to do with the continent of Africa and they selected the dye to basically increase its commercial value. We learned ultimately that based upon the charge properties of amyloid and the affinity of Congo red dye, it sticks really well to amyloid. And amyloid is not a starch, it's a protein, even though the name suggests that it would be a starch. This is an electron micrograph showing amyloid fibrils, these needle-like structures that you can see here in this specimen, biopsy specimen. It was actually discovered at my institution many years before I was born by Cohen and Kalkins and showing that amyloid are these beta sheets of protein. So there are over 30 different proteins that misfold that can cause amyloid. For cardiac amyloidosis, you really only need to know two of them and that's transthyretin, which we're gonna talk about in detail today, and light chain, which you really have to know about and think about as you're thinking about transthyretin disease. Over 95% of cases, or maybe even more that you would see in clinical scenarios are related to these two proteins. We then subdivide a transthyretin protein into two genetic types. And you're gonna hear a lot about that in the next presentation. There's hereditary amyloidosis or variant amyloid where the genotype is abnormal and there's non-hereditary or sporadic cases. We call that wild type. This used to be called senile cardiac amyloidosis or age-related cardiac amyloidosis and it's abbreviated ATTR wild type. So epidemiology of AL amyloidosis is an uncommon disease. This is a rare disease that really affects maybe one to 50 and 100,000 people. This is a picture of the light chain, sorry, of the immunoglobulin and this is the light chain region that misfolds. Even in the best case scenario, we're talking about maybe 7,500 at most maybe 10,000 new cases a year. A rare disease, you will see it if you look for it and you may have already seen it in your clinical practice but it's quite uncommon. But it's so severe, it needs to be ruled out as you will hear in later presentations. Now, this is the most common disease, wild type transthyretin. And as I will show you, maybe 10% of men over the age of 60 with heart failure and increased wall thickness could have undiagnosed ATTR wild type. Maybe 10 to 15% of patients that you send for aortic valve replacement have undiagnosed ATTR wild type. And this relates to maybe 100,000, 200,000 or more undiagnosed cases in the United States at the present time. Now, variant amyloidosis is a bit of a wild card because the variants occur from birth but amyloid develops over time and we call that age-dependent penetrance. So the older you are, the more likely you are to have amyloid manifestations even though you have a variant. There's a particular risk variant that's much more common in people who identify as black and we'll talk about that. 3.4% of people, in fact, which makes it a common variant and there may be 50 or 100,000 and that estimate is really variable as I will show you in future slides. But this is why transthyretin is the much more relevant disease as a cardiologist that you need to know about. But you have to respect AL because it is the most rapidly progressive and likely to be more imminently lethal to your patient. AL amyloidosis results from a bone marrow disorder. So in the bone marrow, a plasma cell or a B cell clone which typically is a plasma cell becomes clonally proliferates like a multiple myeloma scenario but not necessarily myeloma per se and it produces light chains and those light chains then become unstable and then deposit as amyloid fibrils in the heart or in other organs. And these amyloid fibrils themselves cause problems but the light chains themselves also cause direct toxicity to the heart. The heart, in fact, is the most common organ affected by light chain amyloidosis and the second most common organ is the kidney. So it's actually typical that a patient with AL amyloidosis will have cardiac involvement although it is also possible to not have cardiac involvement and that depends obviously on how you make that definition. Now, it's different for transthyretin amyloidosis. In this case, the liver makes circulating transthyretin and that's a four subunit protein that's made by the liver and the name TTR comes from thyroid hormone and from retinol because this protein basically is a transporter of thyroid hormone and retinol or vitamin A. But under certain scenarios, the protein breaks apart and misfolds and then aggregates as these needle-like structures called fibrils and causes a restrictive cardiomyopathy as was illustrated in the first case by Dr. Hayes but also very importantly, a peripheral or autonomic polyneuropathy and you'll have a whole presentation about that later today to understand the significance of that. So as I mentioned, circulating TTR is produced by the liver. It's also produced by the choroid plexus in the brain and the retina and so in certain rare variants, you can have problems associated with hereditary amyloidosis but the most important production is in the liver and circulating TTR is the principal cause of systemic amyloid disease and as I illustrated in the previous slide, the TTR breaks apart and then misfolds and then causes amyloid fibrils. There's also other pathways that lead to amyloid pathogenesis but that's not gonna be the topic of today's presentation. The clinical manifestations are arrhythmias such as atrial fibrillation or heart block, restrictive cardiomyopathy, congestive heart failure and as I mentioned, conduction disease. But amyloid is a multisystem disease. It affects not only the heart and the nervous system as we discussed, it can also cause gastrointestinal problems resulting from say dysmotility and this can manifest as diarrhea or constipation or symptoms of early satiety. There's also musculoskeletal implications of amyloid which are very important. It can cause spinal stenosis or carpal tunnel syndrome because of deposition of amyloid and ligamentous structures or spontaneous ligament rupture and that can be an early sign that systemic amyloid is present. It also can cause kidney disease but typically not through direct kidney involvement in TTR unlike AL, but more through hemodynamic or kidney manifestations like cardiorenal syndrome or low cardiac output. So in that respect, the symptoms of cardiac amyloidosis are really nonspecific and it requires an astute clinician to think about the disease and then put together a bunch of different clinical clues to make the diagnosis. And this slide basically illustrates the cardiovascular manifestations that we've just discussed and the neuropathic manifestations and as well highlights the carpal tunnel syndrome and spinal stenosis that one can see with this disease. Now you probably all remember from medical school that amyloid patients get macroglossia and this is called raccoon eyes or periorbital purpura. Those are actually signs of AL amyloidosis and they're not seen, I should say almost never seen in ATTR amyloidosis. So if you see these manifestations, you really have to be suspicious of AL amyloidosis whereas ATTR is more likely to present with congestive heart failure and if they have variant ATTR more likely with a neuropathy syndrome as well. So the tricky thing about ATTR amyloidosis is that there's something about aging that leads to ATTR misfolding and accumulation because if you just look at the hearts of people who have died, irrespective of whether they've got heart failure or not, you're going to find amyloid deposits. And I think about this like calcifications in the vasculature as people age were indicative of ongoing atherosclerosis but not necessarily atherosclerotic occlusion of vessels. And so in this particular case, you've got amyloid deposition in the heart in patients as they age, but in some patients that process accelerates and that process can increase as they age. So as people get older, maybe only a small percentage of them have moderate to severe amyloid, but many of them do have amyloid in their hearts. And this is actually well illustrated in this slide over here. This is a study from Mayo Clinic where they looked at patients with heart failure and control. These are patients who have all died. So these are all autopsy specimens. They looked at the age of death and you can see nobody in their 60s had any amyloid, 50s or 40s had any amyloid in their hearts. But when you start getting over the age of 70, these bars indicate the presence of amyloid deposits and the black bar indicates HFPAF. And you can see that even though people without HFPAF have amyloid deposits, those who have HFPAF have much more likely have amyloid deposits. And the blue are men and the red are women. And this is a male dominant disease. Maybe two to one or three to one men have cardiac amyloidosis from ATTR compared to women. A quick word about variant disease. You're going to hear a lot about this in the future presentation. So I just wanted to touch on it. I mentioned to you that the variant or hereditary ATTR is one cause of transthyretin amyloidosis. And these are single nucleotide switches. So one amino acid is different and that messes up the entire protein. And those are abbreviated by this nomenclature here where you've got the amino acid substitution preceded by the original amino acid. And this is the position of the protein where that substitution occurs. So the most common variant is V142I, which is also called V122I. It's the same thing or Val122Ile because it's a valine substitution for isoleucine at position 122 of the molecule. This variant occurs in 3.4% of black individuals in the United States. That's 1.5 million people, rendering it a common variant, not an uncommon variant that one has to respect, as I'll show you in future slides. It predominantly causes cardiac amyloidosis and it is on your differential diagnosis for amyloid in an older African-American patient. This slide illustrates this table from a recent review that I wrote with Matt Maurer, basically showing the different genotypes and their estimated frequency. And I told you just to highlight the importance of wild-type ATTR, maybe 12% of patients with HEF-PEF, 10% in HEF-REF because as the disease advances, ejection fraction drops. Patients with carpal tunnel syndrome or hypertrophic cardiomyopathy, especially as they're older, a new diagnosis, 7% of those patients could have undiagnosed amyloid. Aortic stenosis, I already mentioned to you. And as I said, this prevalence of amyloid as people age. Now the V122i variant, we really don't know what the penetrance is because it depends on how long people live. And as people live longer, they're much more likely to develop penetrant disease and amyloid manifestations. And these other variants you'll hear about in the next presentation, they have geographic predilections as to where they originated. I wanna call your attention again to this variant because it's super important. This is an important publication from last year from Selvaraj and Scott Solomon and a recent publication in JAMA accentuated this point, that as people age with this variant, their risk for heart failure or death increases as they get older. So compared to variants, patients who don't have the variant, the age at risk, the age at which, I can't speak, the age at which the risk for atrial fibrillation increases compared to non-variant carriers is 65. And it's 70 for heart failure and it's 75 for death. And the curves you can see diverge at those ages such that as people really get older into their 80s, there's actually a 24% increased risk of death or heart failure hospitalization. A recent publication in JAMA suggested that maybe a million years of life are lost for patients who carry this variant. And by identifying them and getting them on therapy, as you'll hear by Dr. Solomon's presentation later today, we can potentially bend that curve. Now, as I mentioned to you, we really don't know the burden of disease of this particular variant in the population. And this is something that I'm studying in a study called SCAN-MP, which we hope to release results later this year. But it basically is looking at patients at three different Northeastern cities who are older, who identify as black race or Caribbean Hispanic ethnicity and have increased wall thickness and heart failure. And we hope to report the prevalence of disease as well as important features of sex versus, male versus female, as well as other, or hereditary and wild type in this population. So, as I mentioned to you, I showed you the EKG showing low voltage. That's a pretty late manifestation of disease. You saw Dr. Hayes' presentation, diastolic dysfunction, also a fairly late manifestation of disease. As you go back in time and in the progression of disease, and as you'll hear using imaging from Dr. Dorbala, that we can use things like cardiac MR and centigraphy and PET to identify disease earlier and earlier using sensitive tracers and special techniques. And we're moving to a point where we may even identify people at risk who have this ligamentous manifestation of disease. And just to kind of tantalize you with that point, this is a study that was published in Jack last year, where they looked in Denmark of patients who had bilateral carpal tunnel syndrome, and they brought them back five to 15 years later after that bilateral carpal tunnel surgery and screened them for amyloid. And lo and behold, they found amyloid in a sizable population of people, up to 25% of men over the age of 70, who actually in this case, they were not overweight compared to those who didn't. So it's really important to think about carpal tunnel syndrome and spinal stenosis as an indicator. And you see that in a patient with heart failure, you have to think that this potentially could be amyloid. And that's because this disease kind of percolates for a long time before it actually reaches clinical manifestation. We cardiologists see the people, see patients when they start having heart failure symptoms. But there's a whole period of time, in wild-type disease, maybe 10 years or so, and in variant disease since birth, where they potentially could be identified and monitored and then got on good therapy that could potentially interrupt the progression of this restrictive cardiomyopathy and ultimately to heart failure. And then as was illustrated in this review by Jan Griffin, kind of a point of no return. At this point, you can't really do anything for people, but when you want to get them when they're here or here, as you'll hear in later presentations today, where these disease-modifying therapies and our heart failure therapies can actually make a difference. So just to wrap things up, you should think about amyloidosis in patients who have increased wall thickness that's really above the normal range, and that is a lot of people. And the most common diagnosis is going to be hypertensive heart disease, but it should be in your differential, especially if they have other features. For example, those who have aortic stenosis, maybe a third of them with low flow, low gradient AS actually have amyloidosis. And maybe just, but by all comers, maybe 10% of them have undiagnosed ATTR. People who have hypotension when they used to be hypertensive, corrected hypotension, intolerance to beta blockers, peripheral neuropathy and carpal tunnel syndrome, the spinal stenosis, biceps tendon rupture, spontaneous tendon rupture, or this family history of unexplained cardiomyopathy or peripheral neuropathy that's really important. And then finally, you want to put that together with some other diagnostic clues, the low voltage pattern that I mentioned. And then as you'll hear through the subsequent presentations today, some of these imaging clues, diastolic dysfunction, abnormal global longitudinal strain, or abnormal cardiac MR findings, for example, that might prompt you to proceed to diagnostic nuclear imaging that can be used to make the diagnosis. So with that, I hope that I have set the table appropriately for my subsequent speakers, and I look forward to answering questions in the Q&A. Thank you very much for your attention. And it's really a pleasure and an honor to speak with you today. At this point, I'd like to introduce our next speaker, Dr. Pomanowski, who will be speaking with you about variant transdermal amyloidosis. Dr. Pomanowski. Hello, thank you for having me. I'm from University of Mississippi Medical Center. We have adult genetics program whereby we have neurogenetics, cardiovascular genetics, as well as cancer genetics and other clinics. And so we see amyloid patients, and I think we're in the area where we should be seeing a lot more of them. And so I thank you for coming and listening to this, because I think just as Dr. Rueberg alluded to, I think we're not picking up a lot of the patients that we can be. I have no disclosures. Now, when I think of amyloidosis, I think of really primarily three subtypes. There's amyloid associated, which is really rare in here, but it's due to a chronic disease. It's under the transcriptional regulation of cytokines, interleukins one and six and TNF. And that we see more with something like tuberculosis and outside of this country, and in here, mainly rheumatoid arthritis. And that's SSA, that's the serum amyloid A that's really due to inflammatory state. And the other major group is what already has been mentioned, but it's the amyloid light. It's inappropriate formation of the light chains, and that's produced in the bone marrow. And for that, we give chemotherapy. That is a very different amyloid. And again, it's aggressive and it is toxic in addition to that producing abnormal light chain. So this is an urgent intervention if it's there and really hematologists are the ones to deal with this condition in addition to cardiology, but on the end of progression of the disease hematology as well. And then we have abnormal folding of the proteins again, and this is what's been mentioned before, but this is where we have this in the senile or the wild type TTR, and where we have inherited mutant forms of transdiurin. So there are many different proteins, but transdiurin is by far the most common one, more than 95% of the cases, but they all can be amyloidogenic. They can all cause amyloidosis, if you will, with the wild type without a mutation, but the protein changes shape and therefore deposits. And with the inherited subtype where you have a mutation and that mutation subsequently predisposes an individual to having a change in the protein that then subsequently is deposited in different tissues. So at this point, this is on the left, we have an old table that was, that's how we used to classify it and just descriptively have that. Now the Europeans actually, it's much easier to use, but they have it segregated so that we can look at each variant, each change in that amino acid and see if it has been seen, what has been seen with it, what is the clinical phenotype? How does the patient look when we have that genotype, that change in a particular protein, whichever protein that may be, and transdiurin is one of them. So we now follow these patients and we follow the mutations, and then we try to see whether or not there's a phenotype genotype association. In other words, if you have this particular change in the DNA of transdiurin or other protein that's amyloidogenic, what does the patient look like? What organs may be involved? And so really they're all autosomal dominant in nature. So really one allele is all that is needed. We always have two alleles and one allele is all that is needed to cause disease. And therefore transmission of that allele is 50-50 because we give either a normal or an abnormal copy of the gene to our kids, and then our partner gives the other one. And that's how we have our kids being a combination of us. So the transmission of it is 50-50. And the penetrance, it's been a moving target right now. It's at around 37%, but I think penetrance really is defined by two things. One is, do we really test these patients thoroughly and say, we've looked at the neurovascular deficits and neurological deficits, and we looked at the cardiac deficits and others, and have we really identified, have we really interrogated everything before we say someone is exhibiting some forms of disease or not? And so I think that's a primary thing to sort of determine penetrance. And at this point in time, we think that transdiurin penetrance is around 37% or so. And again, this slide just shows you different proteins. And again, when they change form to this beta-plated sheet, you almost have these fibrils that form, and then subsequently these fibrils is what deposits in different tissues and the disease develops. So again, this is a slide that we've seen before, but the same concept, right? In here, we just have the heart and we just have this polyneuropathy. And that is true, but amyloidosis can also involve, the kidneys can also involve, GI, so a number of organ systems that may be involved. However, the heart is the most commonly involved along with the nervous system. The other important caveat to this is that the involvement of the heart determines usually the survival of the patient. So that's a critical component of it. And as you look at this table, one of the things that I want you to think about is that when we have therapies, and that will be discussed later, they act at different points of what we're seeing in here. Some of them are stabilizers, so they stabilize this TTR tetramer. Some of them are mRNA knockdown, so they knock down the mRNA in production of transthyretin, et cetera. So it's useful to sort of visualize this as to what's going on and why people develop amyloidosis. So TTR is a minor carrier of tyroxin and retinal binding protein, but despite the fact that it's a carrier protein, last, I discussed this with a number of drug developers, even if you knock it down to about 95%, even totally, they didn't see any side effects from that not being around, perhaps because it's a minor carrier. The transmission is dominant. And as I mentioned, that means that one allele, one affected copy is enough to cause disease. And the age range varies. Again, some of the mutations are a little bit earlier than others, but even that is not 100%. So even though most of the patients are later, and we see them when they're past 60, 70, 80s, really when we see them, my youngest one, prior to my coming here to UMMC, I was in Delaware, the youngest patient that I had was 18. And she actually had the most common African-American mutation. She had neuropathy, not cardiomyopathy, but nevertheless was symptomatic. 35%, it's important to note that 35% have positive family history. So even though this is a dominant disorder, which means that half of the kids will be affected, half of the siblings will be affected, because again, because of that allele being affected, and it's a 50% that you give the affected allele, we only see it at around 35% of the time. We used to think the disease is really rare. We used to think one in 100,000, one in a million, but again, it's now known to be at 3.4% of the African population on average. And that means that in Mississippi, in this region, we probably have just statistically about five and a half to 6,000 patients that are TTR positive, mutation positive. And that's a huge number. Just to give you a perspective on this, one of the largest studies that analyzed amyloidosis came out of Britain, and they looked at four and a half thousand patients, and that was over 25 years time. They found about 17% of TTR, that would have translated to about 1,600 patients. So in here, we probably have triple of what they had looking at over 25 years, and they were referral center. So again, TTR presents in two forms. There's wild type, as we've mentioned before, and that's a protein that over time degenerates and then forms these fibrils, and hereditary TTR, which is inherited form of amyloid. The average age diagnosis, again, is roughly around 74, and the disease increases with age, with prevalence as high as, again, mentioned, 20 to 25% in those that are 80 or higher. Median survival, that's important, because the median survival for wild type, if in untreated patients, is approximately three and a half years if it starts to involve the myocardium. And the same thing, median survival for hereditary TTR is even worse, especially for the V122I. It's about two and a half years to three and a half years if another variant is involved. With neuropathy or polyneuropathy, it is better eight to 10 years, but again, depending on the diagnosis is made. If someone who's 18 has polyneuropathy as a consequence of amyloid, eight to 10 years is not very long. So when we look at the protein and we look at the distribution of different mutations, again, there are some things that are purely genetic, but they're, and they've been described. So the gene dosage makes a difference. So people that have two alleles, two copies of the same gene, are affected more so than the ones that just have one copy affected. There's certain areas of the protein, a certain loop, like 51 to 56, which are highly amyloidogenic. So when we see mutations in those regions, we can make some predictions. Anticipation to disease, having earlier onset in families, not very much seen in here, but certainly described in Portuguese, Japanese, Swedish families in the past. And there are some heterogenic modification that was mentioned in some of the papers, and that is a mutation within the gene that may have some role whereby the disease severity is modulated. So when we look at the mutation spectrum, we have the neurological presentation, and then we have cardiovascular presentation. And so some mutations are more likely to have cardiac presentation. So like valine 122 isoleucine, and cardiomyopathy predominates. Again, not all the time. And again, there's exceptions. Like I mentioned that 18 year old girl that we had, but by and large, that is the case. And that is what is seen. And again, that particular variant is the variant that's most highly affecting the African-American population, 3.4%. The second most common mutation in the U.S. is the 360 alanine. It's basically mostly of European origin. Patients from Ireland or of Irish origin frequently are found to have this. And carpal tunnel is particularly frequent in that subgroup. And most common mutation worldwide is the valine 30 methionine, which mainly has polyneuropathic form. And it is endemic in Portugal, Japan, and Sweden. And again, when you look at some populations, for instance, Swedish, not Swedish population, but rather Portuguese population. In Portuguese population, some of the mutations are as common as one in 500. So similar to what we see in hypertrophic cardiomyopathy. So that's very common. And we have some of the ancestry in the U.S. so we have to pay attention to that. So penetrance. The term penetrance is different from just a dominant allele or a recessive allele. Penetrance is essentially proportion of individuals carrying a particular pathogenic variant of a gene. So a gene that's mutated, but that also express an associated trait or a phenotype. So does the patient who have the mutation, do they express disease? That is a penetrance. How penetrant is that disease? What does it mean that you carry a mutation? Will you get that disease? That is the core concepts of penetrance. And when we look at different subtypes, different mutations within a transdiuretan, we see that there's a variability in that. So the most common, the valine 30 methionine, that's very penetrant. More than 90% of the people will develop neuropathy over time. We think that the valine 122 isoleucine is much less. So it's about 40% or so, but I think that that's not well-defined. And I agree with Dr. Ruppert that that depends because if you look at patients and you say, well, they're symptomatic because they developed heart failure, but really 10 years ago, no one checked them if they had neuropathy. Maybe some of them do have neuropathy and we don't know. They never developed cardiomyopathy. Maybe they never see a physician. So that's a moving target because it depends as to did you really evaluate that patient thoroughly? Did you do a full neurological evaluation, cardiovascular evaluation and others? Did you address it? And then do they have that mutation? So I think that's the goal of our clinic. When we see patients in our genetics clinic, we make sure that they have a full neurological evaluation and a full cardiovascular evaluation. And then subsequent to that, if they carry the variant, then we can determine what systems are involved or maybe not, maybe they are truly non-penetrant. So the way we look at, or at least I think of TTR and when do I need to worry about it, is I look at a few main features. Number one, why is there a concern for amyloidosis? So is there an abnormal echo? Is there abnormal physical exam? Is the MRI, cardiac MRI abnormal? Then afterwards, the next thing that I look at is I look at Kappa Lambda chains. Because if I have abnormal echo and an abnormal Kappa to Lambda chain ratio, then I know what I'm thinking of. And again, at the same time of doing free light chains, we do immunofixation. The ratio of free light chains is much more specific than doing a UPEP and SPEP. And that's highly discouraged, it's not a substitute. So really, if there is a concern for amyloidosis, we really ought to do the free light chain ratio. And then we do a PYP scan and together sort of in conglomerate, it helps to determine what kind of amyloidosis is there and if there is amyloidosis. So if the scintigraphy is really, if the PYP is really positive, two or three, and we have a normal Kappa to Lambda ratio, then we do think of genetic testing for ATTR because we think we'll most likely either have the hereditary or the wild type. So at this point, please refer them to a genetic center and have that testing done. That testing is by the way, most places free of charge. So it's not going to cost the patient anything. And the sooner we diagnose them, the earlier the stage of heart failure, the earlier the stage of neuropathy, all the data shows that the outcomes are better. The later we diagnose them, the less we can do. So it's important to recognize that they have a positive PYP, normal Kappa Lambda ratio, and they have another abnormality that's supportive of amyloid. That's a patient that needs to get genetic testing. That's a patient that may need treatment. If on the other hand, scintigraphy, so PYP is grade two or three, and we have an increased Kappa Lambda chain ratio, we have to consider, do we need a bone biopsy or a cardiac biopsy? Because it's not clear, is it the AL? Is it the TTR? And we have to figure that out because the treatment is different. The treatment for light chains is chemo. The treatment for cardiac is defamitis and others, so stabilizers or knockdowns. That's very different treatment. And then if the PYP is grade zero or one, and Kappa Lambda ratio is increased, we always think of AL amyloid, and we need to get hematology involved and determine that this is not the light chain amyloidosis that needs chemotherapy. And finally, when our scintigraphy is zero, one, and Kappa Lambda is normal, we do think, but there's other reasons to suspect amyloidosis. So CMR is highly suggestive, or an echo shows strain, or a physical exam has significant neuropathy, bilateral neuropathy that we can't explain. We do think of other amyloidogenic proteins, and I think it's reasonable then to consider those. And this slide basically emphasizes that this is a multi-system disease. So when we have someone with heart failure at an appropriate age, so male more than 65, female more than 70, and there are other red flags. What are those red flags? Well, carpal tunnel, we mentioned several times. So polyneuropathy, and consistent increase in troponin on repeated occasions, or BNP, an echo abnormality, or a CMR that's abnormal, or a discrepancy in voltage, as mentioned earlier. Those things together, in conglomerate, should raise our index of suspicion. And again, if there's any question whatsoever within the area of Mississippi, or where our viewers may be, please do refer them to us at UMMC. We'll do a full evaluation and see whether or not we can be of help. But neuropathy of unknown cause, abnormal strain on an echo, neuropathy with bilateral carpal tunnel, positive troponins, elevated BNP, those are the things that that I encourage people to pay attention to, providers to pay attention to, because we may be able to catch amyloid a lot earlier, and the outcomes, when we look at the data, when we look at the Tefamata study, the outcomes are much better when there's an earlier involvement of the myocardium than later. When they're already, you know, stage four or three, that's not where we can do much in a way of treatment. And lastly, we need to follow patients who are genotype positive, but phenotype negative. So, in other words, let's say we have a patient who comes in, and we do a full evaluation, they're genotype positive for TTR, but they don't have any symptoms. We do an echo, the echo is normal. We do a neurological evaluation, neurological evaluation is normal. There's a particular one, NISP plus seven, that's more on the research side, but it's very much utilized in the world of amyloid. If they're all normal, then there are some guidelines that we have that would allow us to follow those patients. So, follow them over time, because they may be non-penetrant today, and that doesn't mean that they're non-penetrant five, six years from now. So, that's important to appreciate. It's important to appreciate that if we have someone who is genotype positive, it's not enough to say, hey, you're genotype positive, but you don't have any symptoms. So, therefore, you're non-penetrant, and all the best, you're all set, because that disease may still develop. We're still searching for that, where does most of it develop? Well, maybe it's after 60, but what about neuropathy? Does that presentation start earlier? And I don't think it's well-defined. I, therefore, caution everyone to say, well, the penetrance is 37, it's 40, you know, we can guess on that. I don't think we have enough data. I think it's important to make sure that we do our fair share, and make sure we follow these patients longitudinally, because things might change. So, this is our team, if someone is interested at UMMC, and we'll be happy to see patients, and help with evaluation of amyloidosis, and getting the treatment that they need. Those are the people I work with. So, thank you very much for your attention. I would now like to introduce Dr. Tawfiq Al-Kham. Thank you. Thank you. Hi, my name is Tawfiq Al-Kham. I'm a neurologist with a neuromuscular specialty in the University of Pittsburgh. I'm partnering with Dr. Soman in our amyloid center over here, and I'm happy to talk to you today about the polyneuropathy in amyloidosis. Now, to start, as a neurologist, we like to keep things vague a little bit. So, I'm going to try to clear some of this terminology that we use, and I'm going to just start by kind of how we approach and evaluate patients with polyneuropathy. Now, you hear these patterns and words before. So, when we talk about the stocking glove distribution, we are talking about the length-dependent peripheral neuropathy that will start distally in the feet and gradually over months and months spread proximally, and by the time it gets to the mid-shins, it starts also in the fingers, and usually it's the symmetric type. So, that's the most common type of peripheral neuropathy, the length-dependent peripheral neuropathy. Now, we have a different type that we call mononeuritis multiplex, which basically it will be an asymmetric presentation where patients will have symptoms on median nerve on the right side, ulnar nerve on the left, peroneal on one side or another. So, it's kind of the very striking, complete asymmetric, what we can even call patchy distribution, but usually it follows a mononeurose distribution. Now, we jump to much simpler things, the mononeuropathy, which you will see whenever patients present with carpal tunnel, that they have symptoms only on one median nerve on one side or another. Now, radiculopathy is the common kind of knowledge of the root involvement, which, as we recall from medical school, that dermatome distribution picture of an L4 or an L5 or CSAT. So, stocking glove, the length-dependent symmetric neuropathy, the asymmetric, the mononeuropathy, and radiculopathy. Now, the interesting thing is amyloid going to present in any of those or all together. So, this is a picture here of the length-dependent symmetric stocking glove distribution where it starts and be more dense distally and spread proximally, get to the hands and spreads for farther. The mononeuritis multiplex, as we said, it's going to be an asymmetric different nerve, so an ulnar nerve in one side, median nerve on the other side. So, the first way to approach polyneuropathy is the distribution. The other way to approach it will be, or the second step will be, so what type of nerves are we facing problems here? Is it a motor or sensory or sensory-motor together? Now, the most common one will be the both together. So, we will call it a length-dependent symmetric sensory and motor polyneuropathy. That's the most common type. Rarely, we might see only a motor neuropathy by itself and just kind of for like a knowledge here, that's where we'll see it with some of the hereditary Charcot-Marie-Tooth or something like lead poisoning or even porphyria. So, that's something that's much, much rarer than the common neuropathy that we all can see in our clinics. Now, when we get to the point of just talking about pure sensory neuropathy, which can present also with amyloidosis here, we have to break it down to, are we talking about a large-fiber sensory neuropathy or a small-fiber neuropathy? And the small fiber is the one that's going to give me a pain and temperature sensation, so the symptoms would be more involving these two modalities, while large-fiber peripheral neuropathy is going to give me the paresthesia, the ataxia, the unsteady gait, the large-fiber position sensation. So, the most common type, as we said, it's both the sensory-motor, less common would be the pure sensory, and much, much more less common, uncommon, would be the motor neuropathy. After we talk about distribution and then what type of nerves do we have, now, when we do our evaluation, we get to narrow it down to the pathology. Are we talking about an axonal loss, which is the most common one, or are we talking about a demyelinating neuropathy? Canonically, and when we are doing the physical evaluation, knowing that the axonal is the most common one, we can call it that way. Now, it's unlikely that we can be definite from our clinical approach to say this is definitely demyelinating, unless we are talking about a clinical presentation of more an acute, like Guillain-Barré syndrome or AIDP. So, but this kind of classification we'll get when we get to the work of doing something like nerve conduction studies and EMGs. To make things a little bit more interesting, not too much complicated, we have a different nervous system that we have to definitely approach and look into it, and that's the autonomic nervous system. And interestingly, we're going to find that it controls mostly everything and all of these things that we have no control on. So, sweating, heart rate, all these GI and GU symptoms that we can expect to see to even like how much you can have different dilated or constricted pupils. So, even visual symptoms might be a presentation here. So, interestingly, these symptoms might present in a mixture, because as we said, we have a sympathetic and parasympathetic function. So, some patients might present with dry skin, some patients might present with increased sweating. So, but the most common symptoms that we usually screen for when we are approaching patients with neuropathy, when we are looking for dysautonomia, is gastroparesis. So, things like constipation, interesting patient might present with diarrhea, as we said, they will present with early anxiety, and that's a neuropathy symptom. Urinary retention, which is more than what we usually expect to see with the normal aging group that we deal with patients with neuropathy. Resting tachycardia, orthostatic intolerance, impotence, and as we said, dry skin or paradoxical hyperhidrosis. So, too many symptoms that we have to put on our list of screening when we are evaluating patients with neuropathy, but when we narrow it down to distribution, to what kind of symptoms, is there any dysautonomia with that, we can really pinpoint exactly what type of neuropathy are we dealing with. As we said, the most common type of neuropathy will be the length-dependent sensory motor axonal neuropathy, and that's kind of the typical neuropathy picture that we see with the hereditary amyloidosis. Now, when we are talking about ATTR polyneuropathy, it's part of this multi-system involvement, as we heard so far in all the discussion that we had so far. Peripheral nervous system will be involved with this accumulation of misfolded protein, and patients will present with a sensory motor polyneuropathy, and they will have dysautonomia. So, the symptoms that we are talking about would be the sensory loss, the tingling, burning sensation. So, kind of two things. Patients will be talking about the pins and needles, burning sensation, but on the same time, they have lost sensation. They don't feel the cold floor or the hot water, and when you do your exam, you'll find that they have loss of sensation while patients complaining about hypersensitivity. Unsteady gait, as we talk about the large fiber neuropathy, will present with this ataxia and weakness, because while the sensory symptoms are the prominent one, but motor neuropathy also would be a component here. Now, with dysautonomia, the common presentation would be positional dizziness and, as we say, GI symptoms and even neurology symptoms. Now, moving to other organs that will have amyloid deposition and present with neurological symptoms, we hear about the radiculopathies, and that's from leptomeningeal involvement. We hear about the connective tissue involvement, and patients will present with the carpal tunnel syndrome and with biceps tendon rupture, which usually will be kind of the incidental thing that even patients will forget talking about that. They will be presenting to our clinics, cardiology or neurology, in their 50s or 60s, and by getting deeper in the story, we found that, yeah, they had bilateral carpal tunnel surgeries 10 years ago, and when we dig deep, we find that these are people that are not working too much with their hands, that we're expecting like a carpenter to have a carpal tunnel, and also we hear about this spontaneous rupture. They were doing some mild physical activity and not very much straining, and they have that spontaneous rupture on the biceps, and these will be the red flags that will tell us about it's more than just a typical neuropathy. So, we say that peripheral neuropathy with ATTR will be the typical neuropathy, the length dependent, sensory, more than motor, pins and needles, loss of sensation, some weakness, unsteady gait, but when we hear the red flags of bilateral carpal tunnel, polyradiculopathies, the dysautonomia, and the biceps tendons, all these are red flags to say we are not dealing with just the typical neuropathy. Now, a few words to finish with peripheral neuropathy. The most common cause of neuropathy is diabetes. About a third of patients will be diabetes, that kind of type of neuropathy that we're talking about, and over the years, we are evolving from, we used to say a third of patients are idiopathic neuropathy, so to times that we are picking up things that they are under the surface to say, no, this is not just idiopathic neuropathy because we didn't find diabetes or we didn't find any paraproteinemia, this might be something like amyloidosis. Now, the clinical phenotypes for the hereditary ATTR patients who are presenting with peripheral neuropathy, we talked about the sensory motor, small and large fiber axonal neuropathy, so the painful pins and needles, loss of sensation, ataxia with the large fiber, and the dysautonomia that usually are slowly progressive over two to three years. So, they will never be this rapid within two, three months. If we find the patients who had paraproteinemia and their symptoms present very rapidly over a few months, that's likely multiple myeloma or MGUS or something else, while amyloidosis would be usually the slower one, away from the different, which light chain are we talking about. Now, they are symmetric, they are length dependent, and by length dependent, kind of that picture that we had in the first few slides, starting in the toes and spread proximally, then it get to the hands and spread even more proximally. And mainly, it would be sensory symptoms with the sensory loss and pain, but as we said, we'll have also some motor findings. When we are doing our physical exam, we narrow it down, is it small fiber, large fiber, do we have loss of reflexes, and also we do the questionnaire about all this autonomic dysfunction. Back again, let's kind of list it. Orthostatic hypotension, gastrointestinal dysmotility, the early sciatica, constipation, or even diarrhea, erectile dysfunction, hypohidrosis, or sometimes the other way around, and skin vascular changes. So, patients will be talking about the cold hands, cold feet, purple or bluish discoloration of the skin, and all of these things are dysautonomia features. Now, that's the most common clinical phenotype, but we have others. Patients might present with ATTR with neuropathy, they might present with painless neuropathy, they might present with no dysautonomia symptoms, or actually they might present with just or prominent dysautonomia symptoms. Now, as we heard about the different genes, in neuropathy, we might actually have different type of neuropathy, the rare one with different genes. So, we talked about the exona neuropathy, but some of these genes might present with demyelinating polyneuropathy. Some of these genes might present with more of the mononeuritis multiplex, that we said it would be the patchy asymmetric distribution. Some of these genes will present with the polyradiculopathy. Now, some of these genes, or actually even the wild type, they might not present with much of dysautonomia, they might not present with at all any polyradiculopathies, but beside the very prominent cardiac symptoms, they might have just the kind of silent, painless neuropathy. So, different genes might make things more interesting for us. When we are doing our clinical evaluation with neuropathy, yeah, initially we're going to look for what type distribution, what type of fibers, and any other nervous system involvement with the dysautonomia, but when we start following our confirmed diagnosis patients, we try to kind of have different stages and scores to kind of evaluate the progression. So, the familiar amyloid polyneuropathy staging, or the peripheral neuropathy disability scores, will be a good way to assess the progression or the response to the treatment, and we are talking about from stage zero, where they are asymptomatic carrier of like a feminine members, to very severe bedridden or wheelchair-bound patients, and somewhere in between. So, the checklist to identify ATTR peripheral neuropathy. Sensory loss, pins and needles, tingling sensation, burning sensation, that, as you say, that's the length-dependent stalking glove pattern, unsteady gait, feeling off-balance, and need to hold on the rails, something like that, or having trouble taking a shower, standing up, so anything about being off-balance. Positional dizziness with orthostatic hypotension, the dysautonomia symptoms, that also will include GI symptoms, and that might be, as we said, it can take either one of the sides, either constipation or diarrhea, or a mix sometime. And the red flags, history of bilateral carpal tunnel surgery and surgeries, history of spinal stenosis and polyauriculopathies, and that history of spontaneous biceps tendon rupture because of the connective tissue involvement. So, the clinical presentation might sound like the typical peripheral neuropathy that you might hear and say, all right, patients have diabetes, like whatever, but when we have these red flags, especially that there is not really a big story, they are not football wearer, they are not carpenter to have bilateral carpal tunnel, so why should they have these things? We have to really be careful and kind of try to think about ATTR. And I think that will be my last slide, and I will move to Dr. Dorvala, and I'm sure I will take some questions at the end. Thank you. Thank you so much, Dr. Laham. Let me share my screen. All right, hi, good afternoon and good evening, everyone. I'm really honored to be here and talk about nuclear scintigraphy with bone avid tracers. These are my disclosures. As an outline, I will begin with a brief background, talk about when to suspect cardiac amyloidosis, how to diagnose it, then spend more time about radio tracers, protocols and scan interpretation, describe briefly the pitfalls of nuclear scintigraphy, share one of the many algorithms that are out there to evaluate cardiac amyloidosis, and then provide some summary or key points. So why are we all talking about nuclear scintigraphy? This is what was discovered in the late 70s, early 80s, that when patients underwent bone scans and cardiac uptake was detected on the bone scans, using biopsy and other techniques, they figured out that this represented amyloid in the heart. So this is a very old technique that has now taken a rebirth, especially with improved detection using SPECT-CT and with availability of new tracers. The biggest advantage or the biggest reason we are seeing a big uptake is this multi-center study from eight major amyloidosis centers in five different countries, which showed that bone-avoid tracer scintigraphy using either pyrophosphate, BPD or HMDP is nearly 100% specific in select patients, particularly if they have what is called grade two or grade three myocardial uptake, and if a clonal process is excluded using serum-free light chain assay, serum and urine immunofixation electrophoresis. So based on the results of this study, it was felt we could now avoid biopsy and make a non-biopsy diagnosis of this particular disease using imaging techniques. Soon after these results were published, multiple drugs became available for directed therapy of transparent in cardiac amyloidosis, and this therapy has taken off in clinical practice. When do you suspect cardiac amyloidosis? As you heard from Dr. Ruberg, a number of different clinical scenarios can lead one to suspect cardiac amyloidosis. In general, we talk about an older adult with heart failure with preserved or even reduced ejection fraction, as Dr. Ruberg mentioned, with clinical echocardiographic or magnetic resonance imaging features that suggest amyloidosis. Either a TTR variant, low flow, low gradient aortic stenosis, neuropathy with heart failure, musculoskeletal manifestations, carpal tunnel syndrome, biceps tendon rupture, lumbar spinal stenosis, intolerance to heart failure medications, and of course, classical features on echocardiography or magnetic resonance imaging, which typically includes biventricular thickening, diastolic dysfunction, reduced global longitudinal strain, atrial enlargement, and on MRI, diffused late cadelinium enhancement and expanded extracellular volume. So let me start off with a case, a 74-year-old man presenting with new onset heart failure. His main complaint was progressive osteoarthritis of the knee, and he was evaluated preoperatively prior to total knee replacement. He had complained of being fatigued after walking a flight or two of stairs, and his past history included surgery for bilateral carpal tunnel symptoms. Here's his echocardiogram, and you can see left ventricular thickening, and here you can see right ventricular thickening as well. Biatrial enlargement, and then a very classical pattern of global longitudinal strain where the apical myocardial segments contract better compared to the mid and basal segments. This appearance has been variably called cherry-on-top appearance on echocardiography. So overall, his ejection fraction was preserved, but he had restrictive filling with marked thickening of both ventricles and a classical global longitudinal strain. He also was referred for a cardiac MRI, and the MRI confirmed the echocardiographic findings, showed diffuse late cardilineum enhancement, all this white, and expansion of the extracellular map with an ECV of more than 0.41 overall, normal being less than 0.33. Laboratory evaluation was abnormal with abnormal cardiac biomarkers, antiprobian P as well as troponin T. Evaluation for a plasma cell dyscrasia was negative, and a fat pad biopsy was negative, not helpful. At that point, he was referred for a technetium 99m-labeled SPECT-CT study, and these are the results. So we perform what is called SPECT with CT, and the grayscale images in the bottom image are the CT, and here you can see the color image, which is myocardial uptake of pyrophosphate in the myocardium with a very clear absence of tracer in the cavity, all right? So this is a classical, what we would call grade three uptake, where your myocardial uptake is much higher than rib uptake, and you can see rib uptake here. Basically, what next? As you just heard, patient requires a genetic test once a decision is made, once a diagnosis is made of transthyretin amyloidosis. So a genetic test was sent off, and he had no variants in the TTR gene. Final diagnosis of transthyretin wild-type cardiac amyloidosis was made, and he was treated with tefamidase. So how do you diagnose cardiac amyloidosis? So one would think for a restrictive cardiomyopathy with infiltration of the myocardium, endomyocardial biopsy could be the way to go, and yes, it can be, but the yield of endomyocardial, there is a risk of endomyocardial biopsy, and by the time someone considers a biopsy, the patient typically has advanced disease. You can have sulfated algin blue staining, all this blue-green showing amyloid deposition between the cardiomyocytes, and you definitely need mass spectrometry, immunohistochemistry, or one of the other methods to confirm the type of amyloid. So congurate staining and sulfated algin blue show you amyloid, presence of amyloid, but you need to type the amyloid as either transthyretin or AL. The other approach is to have a systemic biopsy, either a fat pad or involved organ biopsy, and combine that with typical imaging features for a diagnosis of transthyretin or AL amyloidosis, because here you have a diagnosis of systemic amyloid and typing, and the imaging will confirm that the patient has cardiac involvement, or in the case of AL amyloidosis, if you have abnormal antiprobian p-entroponin T with no other reason for their elevation, then that could also help you diagnose cardiac AL amyloid. Of course, transthyretin cardiac amyloidosis can now be diagnosed without biopsy. You can have a patient with heart failure and typical imaging features, exclude a plasma cell dyscrasia, and then you have a grade two or grade three uptake on the pyrophosphate imaging, and that can tell you that this patient has transthyretin cardiac amyloidosis. So let me go into some details about more specifics about these technique. So amyloid imaging with SPECT radiotracers, I-123-SAP has been used in UK and primarily for AL amyloid. It does not help in imaging cardiac amyloid. Bone-avid tracer compounds, PYP, DPD, and HMDP have been used, and these are the structures. How do we do this test? We inject the radiotracer. We wait for about two to three hours. We wait for two and a half to three hours at the Brigham. Then we perform a SPECT CT scan. So CT is used for attenuation correction, and more importantly, for localization of the tracer to the myocardium. Extremely important. If you have a SPECT CT, you should be using SPECT CT for this technique because it is what is called hotspot imaging. You need the anatomic definition of the CT to localize the hotspot or the uptake in the myocardium. We perform a 360-degree study, so all around, and we look at the images visually. So the recommended radiotracers are PYP, DPD, and HMDP with a long delay. This is how the scans are interpreted currently. Visually, grade zero is no myocardial uptake with normal bone uptake. Grade one, myocardial uptake less than rib. Two, uptake equal to rib. And three is greater than rib. In addition to visual grading, which you see here, grade zero, no uptake, uptake less than rib, equal to rib, and greater than rib, we can, with advanced techniques, especially using SPECT CT, we can quantify the absolute radiotracer uptake in the myocardium in terms of standardized uptake value. And we can measure volumetrically amyloid in the entire myocardial volume, come up with cardiac amyloid activity, or what is called percent injected dose as volumetric measures. Planar imaging-based ratios of heart-to-contralateral ratios, as well as heart-to-whole body ratios with DPD, have been shown to be prognostically relevant in that patients with a ratio of less than a certain number, 1.6, or here in this case, less than 7.5, had better outcomes compared to patients with a ratio greater than that. Using DPD with visual scoring, the NAC group showed that patients with a negative scan had better uptake compared to patients with grade one, two, or three. But in this study, they could not distinguish prognostically any difference between grade one, two, or three. So basically, no amyloid versus amyloid was helpful in prognostication. So a more recent study comes from a multi-center NAC plus Italian centers in a large cohort, 1,400 patients. And what they did was they looked at all patients who were positive for transthyretin based on imaging, and they looked at right ventricular uptake, focal uptake versus diffuse uptake. And what they showed was in the overall cohort, patients with focal RV uptake had better outcome compared to diffuse RV uptake, and this held true for both grade two and grade three. So basically, this is one study which suggests that on top of visual grading of left ventricular uptake, right ventricular uptake may offer some prognostic benefit. So what are the pitfalls of this particular technique? There's some pitfalls which we all need to be careful about to make an accurate diagnosis, especially an accurate non-biopsy diagnosis. First thing, MDP is not recommended. It's important to recognize blood pool, and for that, you need delayed imaging and spec. Certain forms of hereditary ATTR amyloidosis have a low sensitivity, so important to recognize that. And then important to recognize that with AL amyloidosis, you can have any grade uptake in the myocardium, grade zero, one, two, or three. So here's an example of an older adult with prostate cancer and a bone scan with MDP that was negative. But he had heart failure, and the question was, does he have transthyretin amyloid? Because MRI was suggestive of cardiac amyloidosis. So we repeated the test with PYP, and here you can see clearly positive. So remember, MDP has low sensitivity and is not recommended. PYP, DPD, or HMDP are recommended. Here's an example of an older patient with heart failure with preserved ejection fraction referred to us for management of newly diagnosed transthyretin amyloidosis based on what is called a planar image and a contralateral ratio. So for this particular problem, we do need to look at delayed imaging and SPECT imaging. And I know this is early imaging because I see no rib uptake on the planar. So these images were obtained very early on suggesting this could be blood pool activity. So basically, if you did SPECT CT, this is how blood pool would look like. This was the repeat imaging from our institution. Contrast that on the right side with a positive case where you see myocardial uptake, but no blood pool uptake. Recognize how by using CT, you're able to see the negative myocardial shadow around the uptake in the heart. You didn't have this, then this could be read as positive on SPECT alone, but CT definitely tells you that it's not. But of course, when you have overt florid cases, even SPECT alone may be sufficient. So important to recognize blood pool. A similar example here where SPECT was performed, it was not just planar, but it was called positive. And clearly again, no rib uptake. This is blood pool activity. And this is the patient example that you saw earlier. So always remember the quality of the PYP images. If there's no rib uptake, it's very early image. And even though SPECT was done, it's important to recognize that myocardial uptake should be identified and delayed imaging will help with preventing these errors in a lot of cases. Hereditary ATTR cases. Here's a study, a small number of patients, 10 patients and five controls. Alternate rows, C11 Pittsburgh B compound PET with DPD and healthy volunteers. What you see here is DPD was positive in patients with group A or fragmented fibrils, but negative in those with full length fibrils. But C11 Pittsburgh B compound, which is a PET amyloid tracer was positive in both cases. Again, suggesting that certain hereditary forms, particularly those patients of full length fibrils, this test may be falsely negative. So something to keep in mind. Here's an example of a patient who was referred to us as positive. And this is truly positive when you look at SPECT, but this patient ended up having AL amyloidosis. So his plasma cell evaluation was positive. So we sent the patient for endomyocardial biopsy and he had AL lambda light chain with immunofluorescence analysis. How do you exclude plasma cell dyscrasia? Look at serum and urine immunofixation electrophoresis and serum free light chain assay. Extremely important to do this in all patients all the way from grade zero to grade three before you make a non-biopsy diagnosis of this condition. So finally, let me share with you an algorithm and conclude with some key points. Patient with suspected amyloidosis, one proposal could be to start off by looking at a monoclonal process. If that is completely negative, go ahead with bone abitracer SPECT-CT if you have it or SPECT. Grade zero, you're done. Grade two or three, send off a genetic test to make a diagnosis of wild type or variant. Grade one cases and in some of the grade zero cases where you have high clinical suspicion, further evaluation with MRI plus or minus biopsy could be considered for making early ATTRCM or rare forms of amyloid diagnosis. If the plasma cell dyscrasia evaluation is positive and if it's an isolated free light chain abnormality, use EGFR adjusted free light chain levels and then proceed this way if they're all within normal limits here. If not, or if it's not an isolated free light chain abnormality, immediately refer to a hematologist who may perform further evaluation including involved organ biopsy to make a final diagnosis. So let me end by key points. So remember specific considerations about AL, hereditary ATTR and equivocal cases. Use appropriate tracers. Do not use MDP. Exclude a plasma cell dyscrasia. Perform SPECT and if available SPECT-CT. Image late, two to three hours post injection. Grade the myocardial uptake visually on SPECT images. Do not look at planar images alone. And quantitative SPECT-CT and amyloid PET are new novel emerging techniques. Thank you very much for your attention. I now would like to introduce Dr. Charles Moore. He'll speak about heart failure management and supportive therapies. So this evening, I wanted to talk about the heart failure management of cardiac amyloidosis and supportive therapies. I don't have any disclosures pertinent to this discussion. So in the next 20 minutes or so, I'd like to give you a brief overview of the general treatment of symptomatic cardiac amyloidosis. First of all, with the management of heart failure syndrome. Secondly, the treatment of arrhythmias and thromboembolic complications. And lastly, we'll touch on treatment of the combination of aortic stenosis and cardiac amyloidosis. So in terms of the heart failure management, the first thing to know is that it's quite challenging, particularly as the disease progresses into more advanced stages. These patients have what you could term very brittle volume status because of the degree of diastolic dysfunction, the very narrow range of optimal volume status. A little bit of excess volume creates significant congestive symptoms, both systemic and pulmonary congestion. And on the other hand, excessive diuresis or volume depletion can lead to worsening orthostatic hypotension and other issues. In terms of the guideline medical therapy, it's a little bit of an uncertain quantity, and we'll touch on why that's the case. And lastly, as patients proceed on to more advanced stages of disease, there are some potential advanced heart failure treatment options to consider. So the volume status traditionally will be controlled with loop diuretics. These patients typically have both right and left ventricular involvement and thus have both right-sided and left-sided heart failure symptoms and signs. Because of the gut congestion and low cardiac output, they often have poor bioavailability of loop diuretics, and so agents such as torcimide or bumetanide are sometimes used instead of furosemide. In addition, we often have to resort to adding other agents to loop diuretics in order to get controlled volume status. Adding a thiazide such as metolazone is an option, but that has to be done with caution because these patients are prone to excessive diuresis leading to worsening renal function and worsening hypotension. The same is true for adding a mineralocorticoid receptor antagonist like spironolactone because they often have concomitant kidney disease and low GFRs. There's concern for hyperkalemia and worsening renal function with those agents as well. In terms of trying to keep them at an optimal volume status, monitoring is critical, and obviously we try to do that outpatient, and so that has to rely on various forms of telemonitoring and home health monitoring to try to closely follow daily weights. But there are other more advanced techniques for trying to optimize volume status and maintain it as well. You can look at serial natriuretic peptide levels. These patients tend to have quite excessively elevated BNP levels, and that complicates things, but looking at them in a serial way can still be useful in trying to maintain the desired volume status. In the minority of patients who happen to have an implanted electrical device like a pacemaker or ICD, some of those devices have thoracic impedance monitoring capability, and you can rely on that as additional data to try to help you with monitoring their volume status. There is also some anecdotal experience with using ambulatory pulmonary artery pressure monitoring, and obviously that allows you to get a daily reading of both diastolic and systolic and mean pulmonary artery pressures and allows you to try to maintain the patient at their optimal weight and volume status. So what about guideline medical, directed medical therapy? Well, these patients typically start off with diastolic dysfunction, and then later in their course they can develop systolic dysfunction with worsening LV and RV ejection fractions, but in the beginning they have a predominantly diastolic dysfunction, and until recently we didn't really have much in the way of guideline directed medical therapy for HFPF, but that's no longer the case with the recent data on SGLT2 inhibitors as well as also data on the mineralocorticoid receptor antagonists and the ARNI drugs. So if we look at the beta blockers and the ACE, ARB, ARNI groups, these would be predominantly for patients who've developed reduced ejection fraction. The problem is that, number one, we don't have randomized data to inform us about the efficacy of these agents in this patient population, and secondly, and perhaps even more importantly, we know through experience that they tend to be poorly tolerated, and that's due to a number of anatomical and physiologic issues. First of all, these patients tend to have small left ventricular cavity size. They have the severe diastolic dysfunction, as I've already mentioned. They also have impaired atrial contraction and also, in many cases, autonomic neuropathy, which leads to orthostatic hypotension. So the combination of all these factors tends to cause marked tendency towards hypotension, particularly orthostatic hypotension, and also heart rate dependence, and so these patients typically feel worse on beta blockers, and when patients are diagnosed who are already on beta blockers, they're often discontinuing or at least reducing the dose of those agents. What about the SGLT2 inhibitors? Particularly important because they're indicated throughout the ejection fraction spectrum, and so until recently, we didn't have a lot of data, but just this year, there was a paper published about SGLT2 inhibitor therapy, at least in a retrospective study, and so what they did was that they looked at a database of about 2,400 patients with ATTR cardiomyopathy, and they identified out of that database about 220 patients who were being treated with SGLT2 inhibitors. They then, through propensity score matching, looking at 16 different variables, they tried to do matching with 220 untreated controls, and then they compared outcomes in those two groups, and they were able to show that, in fact, the all-cause mortality, cardiovascular mortality, heart failure hospitalization, and the combination of cardiovascular mortality and heart failure hospitalization were all improved by treatment with SGLT2 inhibitors, and you had a number of secondary benefits as well in terms of slowing the progression of renal dysfunction, less rise in natriotic peptide levels, lower diuretic requirement, and they did not have issues with blood pressure, and so the discontinuation rate was less than 5% over two years of follow-up. So, again, this is retrospective data, but at least encouraging regarding potential efficacy and tolerability for the SGLT2 inhibitors for this patient population. So, what about the MRA drugs, spironolactone and epilarinone? We have even less data here, but there was a paper published about three years ago when they did sort of a retrospective look at the TopCat study, and as you recall, that was a HIF-PEF trial looking at spironolactone for patients that had an injection fraction greater than or equal to 45%. So, what they did here was basically, they looked at a subgroup of about 600 of those patients who had both echo and tissue doppler data at study entry, and they basically developed four quartiles based on two echo parameters, the first being left ventricular wall thickness of greater than or equal to 12 millimeters, and the second being tissue doppler parameter of systolic mitral annular velocity of less than or equal to 6 centimeters per second. And so, if you look at the quartile who had both increased wall thickness and tissue doppler abnormalities, that is what you see in the red line, and this is looking at survival over four years follow-up, and you can see that that subgroup had the worst survival in the study. They were able to then look at outcomes in that group, and they were able to demonstrate that that group still demonstrated equal efficacy and benefit as the other groups with treatment with spironolactone, so there was no signal that this group did less well with spironolactone than the others. Now, the problem with this is, of course, that even though you potentially enriched the subgroup who may have had cardiac amyloidosis, undiagnosed obviously, we really don't know. We don't know the specificity of those echo findings in this study population, so while, you know, unselected HEF-PEF populations, maybe 16, 20 percent in some studies have been demonstrated to have cardiac amyloidosis, we really don't know in this particular top cat subgroup how many patients really had cardiac amyloidosis. The other problem with MRAs is, again, as I mentioned before, concomitant kidney disease, low GFRs, potential hyperkalemia, and worsening renal function, and so tolerability is definitely an issue. So, the problem with diagnosing cardiac amyloidosis is that it's often done late, as you've already heard, and so even though you may start therapy, these patients may already be in an advanced stage or still progress to more advanced stages of symptomatology and poor prognosis, and so you often do need to look at advanced treatment options beyond just medical therapy or basic device therapy. In terms of heart transplantation, there are select patients with cardiac amyloidosis, in particular the ATTR variety, who can be candidates for cardiac transplantation. Age and comorbidities tend to be the limiting factors, and then with AL amyloidosis, it's more complicated because it's difficult to get control of the underlying primary AL abnormality, and then patients tend to have to have the transplant of their heart before they can have their bone marrow transplant to try to control the AL amyloidosis. In terms of mechanical circulatory support in the ambulatory setting, LVADs tend to be a poor option for the vast majority of patients for a couple reasons. Number one, I've already mentioned the small left ventricular cavity, and so occlusion of the left ventricular assist device inflow cannula is a problem, and even more importantly, because of the biventricular involvement, they tend to have right ventricular dysfunction, and so supporting the left ventricle alone is not a good option because the right ventricle is not able to keep up its side of the bargain. Having said that, palliative care is a good option to get involved early in the decision-making, regardless of what you decide about these other advanced options, to do shared decision-making with the patient about what their goals are, and more informed decisions about the treatment options you're going to select. So, beyond heart failure, these patients are also prone to arrhythmias, and that's basically of all types, because of the diffuse nature of cardiac involvement with the amyloid, they can get this conduction system disease, sinus node disease, and so sinus node dysfunction, baby node dysfunction are not uncommon. These patients may require standard pacemaker for bradyarrhythmias. If they do have AV block and have a high degree of right ventricular pacing, and they have developed systolic dysfunction, then definitely can consider cardiac resynchronization therapy with biventricular pacing as an option. Ventricular tachyarrhythmias are also fairly common, particularly non-sustained ventricular tachycardia, and certainly implantable cardiac defibrillators, ICDs, can be used for secondary prevention in patients who've had aborted sudden cardiac death or symptomatic sustained ventricular tachycardia. At least in those patients who are expected to have otherwise survival of over a year. The role for primary prevention in patients who've developed low ejection fraction is less clear. There's no data to really guide us there. There's some interesting ambulatory electrocardiographic monitoring data that really shows that a lot of the sudden cardiac death in patients with cardiac amyloidosis is not necessarily ventricular tachyarrhythmias, but rather bradyarrhythmias and really electromechanical dissociation. And so calls into question whether ICDs would be as effective as you normally would expect for primary prevention in these cardiac amyloid patients with systolic dysfunction. So that's definitely a situation for shared decision-making in those patients. Atrial fibrillation, quite common. These patients tend to be older to start with. They tend to have a lot of comorbidities that have a high incidence of atrial fibrillation, and then you add the amyloidosis, and it even makes the incidence that much higher. In terms of rhythm control, really amiodarone is the first-line agent for most of these patients. It's a little unclear how effective ablation is as an option. There's definitely data to suggest that if it is an option, it's going to be more effective earlier in the course of the disease than later when the disease is more advanced. Rain control, fortunately, because of the concomitant conduction system disease, rapid ventricular response is often less of an issue. So that's good because beta-blockers, as we've already mentioned, are not very well tolerated. And for that matter, the non-dihydropyridine calcium channel blockers are not tolerated well either because of their negative inotropic effects. We used to really have a prescription against using digoxin. There was in vitro evidence that amyloid protein, amyloid fibrils in myocardial tissue can bind digoxin, and so it was concerned about unusually high local tissue levels of DIG having a high degree of toxicity. Subsequent experience and data out of Cleveland Clinic and other centers has demonstrated that if you use caution and low doses and appropriate drug level monitoring and rhythm monitoring, DIG can be used in select cases of atrial fibrillation for rate control with caution. A lot of times, though, we end up using amiodarone not only for rhythm but also for rate control, for these patients. With the high incidence of atrial fibrillation, of course, thromboembolism must enter the equation, and these patients are indeed at increased risk for thromboembolic complications. That's even more so because of the already mentioned reduced atrial systolic function. So, not only do you have the atrial fibrillation, but the atrial systolic function was poor to start with. For that reason, anticoagulation in atrial fibrillation with cardiac amyloidosis is indicated regardless of what the chance to VASC score is. In addition, in patients who are going to attempt DC cardioversion for their atrial fibrillation, you should do a TEE before even, it doesn't matter how long they've been on therapeutic anticoagulation, a TEE is indicated prior to cardioversion. We really don't have any particular data on comparative efficacy between Coumadin or Warfarin and the newer oral anticoagulant agents, so all of the above have been used in various patients. So, if you've actually looked at some of these patients who are still in sinus rhythm, you can discover that this reduced atrial systolic function can be quite profound, and so what you see here is a mitral-doppler flow of a patient with cardiac amyloidosis, and you can see they have the early diastolic filling wave, the E wave, but during systole, you can see they have no A wave whatsoever, so they functionally have so-called atrial standstill or atrial electromechanical dissociation. So, for this reason, patients with this degree of poor systolic atrial function, in some cases, it's been advocated that they be treated with oral anticoagulant agents, even in sinus mechanism to try to reduce the risk of thromboembolic complications. Lastly, I will mention the treatment of aortic stenosis, which there's an increased incidence of this combination. We know from observational data that the combination leads to a higher mortality than would be otherwise expected for either disorder alone. In the series that have looked, specifically done evaluation looking for cardiac amyloidosis in TAVR patients, the series has demonstrated that 16% of those patients did, in fact, have cardiac amyloidosis, so it's an enriched population. Fortunately, there is evidence that TAVR transcutaneous aortic valve replacement does improve mortality and morbidity compared with just medical therapy in this patient population. However, there does appear to be a higher risk of procedural stroke and probably AV block as well. You can see here in this observational study that looked at a nationwide database over three years of TAVR patients, they were able to identify 273 patients who had both aortic stenosis with TAVR and who had amyloidosis ATTR compared to the full database of over 244,000. So, obviously, based on the other 16%, you can see this is far less than that, so you could presume that there are a lot of undiagnosed cardiac amyloid patients in the comparison group, but be that as it may, they were able to show that there was no difference in early mortality and morbidity in the amyloid patients compared to the other standard patients, but they did have a threefold increase in the risk of acute ischemic stroke during the hospitalization. We also have sort of anecdotal experience that these patients do less well chronically after TAVR, so that's one of the sort of red flags to consider cardiac amyloidosis is if you have a TAVR patient who doesn't seem to respond as well to their procedure over the subsequent months of follow-up than you otherwise would expect, then you might suspect maybe they have the cardiac amyloidosis as a secondary disorder that's leading to their poor response. So, this is a nice summary from the European Society of Cardiology that basically summarizes what I've already explained to you over the course of this evening. So, you can look at that at your leisure, and now we'll move on to the next group. So, the next talk is Dr. Prem Soman. He'll be talking about evolving therapeutic landscape in ATTR. Thank you very much, and very nice talk. Good evening, everybody. These are my disclosures. Now, the therapeutic developments in ATTR amyloidosis have paralleled our understanding of structure-function relationships in this disease, and you may have seen this slide or a similar scheme in an earlier talk today, but we know that the critical step in the development of cardiac amyloidosis, TTR amyloidosis, is the dissociation of the very stable tetrameric form of the TTR molecule into its constituent monomers. Now, when these monomers are misfolded, process of tetramer dissociation, monomer aggregation, and the formation of the amyloid fibril is associated with the clinical manifestations of amyloidosis, but it does not tell us the association between these individual stages and the specific manifestations of this disease. So, we know that, at least in cardiac amyloidosis, there is myocardial infiltration, but we also know that certainly in AL amyloidosis and likely in TTR amyloidosis, there is also an element of myotoxicity, direct myotoxicity, possibly by some of the prefibrillar oligomer stages. So, cardiac amyloidosis, TTR amyloidosis, is not just an infiltrative cardiomyopathy but most definitely a toxic infiltrative cardiomyopathy. The current therapies, the specific therapies for TTR amyloidosis, can be divided into precursor therapies, therapies that reduce the formation of the amyloid fibril, and antifibrillar therapies, which are directed towards removing the fibrils that have been already formed and deposited. Note that the precursor therapies reduce not just the final amyloid fibril, but all the intermediate prefibrillar stages that are formed before the amyloid fibril. So, the success of these precursor therapies don't help us identify the specific stages that produce the pathology. Now, pharmacotherapeutic efforts have paralleled, if you will, the lifespan of the amyloid fibril, and they consisted of approaches or they consist of approaches to reduce the formation of TTR, and the earliest approach in this regard in wild-type disease was liver transplantation. We can also turn off the production of TTR using the RNA silencers. We can remove or repair the TTR gene using gene editing techniques, and all of these approaches will dramatically reduce the level of TTR that is produced. The next approach is to stabilize the TTR and to prevent it from dissociating into its constituent monomers, and these are the stabilizers. Now, for a number of years, we have recognized the minor effect of agents like doxycycline, which act to disrupt the TTR fibrils that are already in existence, and more recently, a number of drugs have been developed to attach to the misfolded amyloid fibrils and increase the phagocytosis. Now, these are very interesting approaches and target epitopes on the amyloid fibril that are usually not exposed when the TTR fibril is normally folded. Now, when the TTR fibril misfolds, these epitopes are exposed, and these antibodies can therefore act. Now, note that because they're monoclonal antibodies, these antibodies have to be specific against ATTR and AL fibrils. Alternatively, they can be a pan-amyloid agent targeting the SAP or the serum amyloid protein, which is ubiquitous in all types of amyloid, like the trellis compound AT03. So, and as you know, last year, one of the TTR monoclonal antibodies reported very encouraging phase one data. So, I will not talk about liver transplantation today, partly because a lot more liver transplantation was done in Europe, and much more for neuropathy than for cardiomyopathy, and for a number of reasons that are beyond the scope of this talk, liver transplantation has largely fallen out of favor as a therapeutic option for this disease. I will focus mostly on the RNA silencers and TTR stabilizers, but along the way will allude to the mechanism of action of these other agents. Now, this is a very simplistic scheme of the mechanisms by which the information in the DNA is used to produce protein. So, the DNA, as you know, within the nucleus is transcribed into messenger RNA within the nucleus that is then taken out into the cytoplasm and translated into protein via the endoplasmic reticulum. Now, the small molecules that are RNA silencers can be antisense oligonucleotides or small interfering RNAs. Petiseran was the first small interfering RNA that was approved for clinical use. Now, these agents behave, have a slightly different mechanism of action. The antisense oligonucleotides are DNA-based and have to be delivered into the nucleus, where the small interfering RNAs are double-stranded RNAs that form risk complexes and act in the cytoplasm. Nevertheless, the end result of both of these approaches is to dramatically reduce the amount of RNA that is available to translate the production of TTR gene, TTR protein, and therefore, they greatly reduce the amount of TTR, circulating TTR. Now, as you very well know, both of these agents, Petiseran, the antisense, the small interfering RNA, and inotersin, the antisense oligonucleotide, were both established in large multicenter studies to have efficacy for retarding the progression of hereditary amyloid polyneuropathy, and both of these agents are now approved for the use in hereditary polyneuropathy. Now, the initial offerings of this class of drugs were either based on a lipid nanoparticle or an unconjugated DNA strand and were either associated with significant side effects like thrombocytopenia and glomerulonephritis in case of inotersin or required pretreatment to avoid allergic reactions, as in the case of Petiseran, which was delivered via a lipid nanoparticle. The next iterations of these are galnacmoiety-based and it has a much greater affinity for the liver, and therefore, greater amounts can be delivered effectively, and the administration sequences or the frequencies are much more favorable for the patient, and these do not require premedication and are not associated with major side effects. So, vitruciran is the second generation small interfering RNA and aplanterosin, the antisense oligonucleotide. Now, it must be remembered that when the production of TTR is greatly reduced, patients require vitamin A supplementation. This is because the retinol binding protein, which is a small molecule, is bound to the TTR molecule, and if it is not bound, because of its small size, is rapidly cleared by the kidneys, and patients can theoretically develop vitamin A deficiency, although I don't think this has been reported clinically. An interesting observation of interesting clinical situation is that when you give patients supplementation, their serum levels will continue to be low because of the low TTR values, and so we don't have a good way of monitoring the efficacy of vitamin A supplementation in these patients. Now, although these agents were approved for wild-type disease, some of these agents are undergoing clinical trials currently for ATTR cardiomyopathy, both wild-type and variant. Now, you may recall that last year, there was an application for an expanded indication for patisserin, which was denied by the FDA, because they felt that even though the primary endpoints of the six-minute walk test and the cancer's KCCQ questionnaires were statistically significant in the active group, those differences were not clinically meaningful, and so the application for the use of patisserin in cardiomyopathy was denied in 2023. As you all know, the Helios B trial looking at vitrucerin for this indication recently reported, and I'll talk about that, and in 2025, we can expect results from the large cardio transform study looking at the efficacy of aplantericin, which is the antisense oligonucleotide in ATTR cardiomyopathy. So, these are the results as presented in the Alnylam press release just a couple of weeks ago, and it looks like they had very encouraging results from the Helios B phase 3 studies looking at vitrucerin in cardiomyopathy, achieving a 28% and 33% reduction in the composite of all-cause mortality and recurrent cardiovascular events in both the overall group, which included a percentage of patients who are also on tefamidus and in the monotherapy populations, and similarly, very powerful results looking at all-cause mortality alone in the overall and monotherapy populations. We have to wait for the publications to get a better handle on these results, but at first look, they look very, very encouraging. The TTR stabilizers, this class of drugs has had some very, very interesting developments, and again, they were based on the structure-function relationship of the TTR molecule. Here is again the tetrameric configuration, and shown here are some of the more common gene mutations that destabilize this molecule either kinetically or thermodynamically. Now, a very beautiful story here of a serendipitous observation by Teresa Coelho, who is a neurologist in Portugal and has contributed tremendously to the field of hereditary polyneuropathy, who noticed that in some of her patients with the VALTAT-EMED mutation, the clinical manifestations of polyneuropathy did not develop. Now, most of these point mutations in TTR are quite pathogenetic, and it is unusual for people with the V30 mutation to not develop disease, and when she sequenced the gene of these patients, she noticed that they were compound heterozygous for another mutation, the T119 mutation, which actually stabilized the TTR molecule and prevented the manifestations of V30M from occurring. So, these mutations, and there are a couple of other mutations, but the T119 mutation is the most well-studied, are called rescue mutations, and in a beautiful story of a translation of a serendipitous clinical observation into the development of a clinically useful drug, AG10 was developed to mimic the amino acid changes that are produced by the T119 mutation. So, the agents that are stabilizers, tefamidin and diflunasol, act by binding to the thyroxin binding site and therefore stabilizing the tetramer and AG10 by mimicking the stabilizing effects of the T119 mutation. Now, we know that there were two large trials that established the efficacy of stabilizers, ATTR Act, which led to the approval of tefamidin, which was published in 2018, and more recently, the ATTRIBUTE trial, which showed excellent results with AG10, which is now called ACORAMIDAS. Now, as you know, both of these trials used a hierarchical analysis of the composite endpoint and used the VIN ratio to represent the effect size of this analysis. So, the Finkelstein-Schoenfield method of pairwise comparison was used for the hierarchical analysis and the VIN ratio to represent the effect size, and they're shown here. What you can also see is that the endpoints included slightly different components for both of these trials, but let's for a moment look at the mortality data, and you can see here that both the absolute and the relative reduction in mortality was lower with the ACORAMIDAS trial than with ATTR Act, and in fact, the analysis looking at mortality alone did not achieve statistical significance with the ATTRIBUTE-CM. But note that the ATTRIBUTE-CM was published several years after the ATTR Act and recruited patients with a different degree of disease severity, because by the time ATTRIBUTE-CM came along, we were tuned into the diagnosis of TTR amyloidosis and were making this diagnosis at an earlier stage. So, if you look at the mortality in the active group in the ATTR Act, it was 29% versus 42% in the placebo group and 25% in the placebo group versus 14% in the active group in the ACORAMIDAS trial. So, the placebo rate in the ACORAMIDAS trial, the rate of mortality in the placebo group was lower than the mortality rate in the tefamidase group in ATTR Act, and the mortality rate in the ACORAMIDAS group actually was close to the mortality of age-match controls in the ATTRIBUTE trial. So, this raises a very important question of how difficult it is going to be for future trials looking at ATTR amyloidosis, and we're going to have to recruit a larger number of patients, follow them up for a longer period of time, and also contend with the fact that mortality-changing therapy is now going to be ubiquitous for this disease. Now, this is a very important slide from three different studies, all emphasizing the fact that early diagnosis and therapy leads to a mortality benefit. So, the first slide, the top panel on the left, is from ATTR Act, showing you that the reduction of death is dependent on the NYHA class of the patient, and you can see that there is a significant reduction in the mortality benefit as the NYHA class increases. The top right panel is from the ACORAMIDAS trial, looking at the composite of all-cause mortality and cardiovascular hospitalization, showing that the curves separate quite early, about three months after initiation of therapy, and the bottom panel is from an old neuropathy study, a European study, showing you that if you look at the percentage of patients who were stable or improved, it was 68 percent in the patients who were started on tefamidus, but only 48 percent, 46 percent in the group of patients who initially received placebo and then were put on tefamidus 18 months after being on placebo. That 18-month delay in the start of therapy resulted in a significant reduction in the percentage of patients who were stable or improved. So, very strong evidence from multiple trials of the benefit of early therapy. Now, this is, I think, very important data from Rick Rueberg's group. Diflunasol, which costs pennies and is an NSAID that is available over the counter, is also a fairly powerful stabilizer of TTR. Now, we have sort of generally ignored the fact that diflunasol exists, and because it is a very cheap drug, I doubt whether, I doubt that we're going to have large prospective trials. Now, being an NSAID, one of the concerns was that the use of diflunasol in patients with cardiomyopathy would result in the common side effects of sodium and water retention, reduction in GFR, and also gastric toxicity. And so, this observational data from Rick Rueberg's group shows us that in carefully selected patients, the majority of patients were able to tolerate the drug for the first couple of years. And I think this is really important, and most of us are now gaining confidence in using diflunasol in stable patients. And in this trial, even stable patients with NYHA class 3 disease were tolerated diflunasol, and we're gaining confidence using this in patients who are not able to afford to farm it, to farm it. So in this particular trial, there was a survival advantage of patients who were on diflunasol, but it has to be interpreted in the context of the fact that this was a non-randomized observational dataset. Now one of the most exciting developments has been the first in-man study of gene editing using the CRISPR-Cas9 system in patients with TTR amyloidosis. There was a large phase 1 slash phase 2 study conducted in England and Australia, and based on that, the company has already started and has begun recruiting for a phase 3 trial in the United States. The compound NTLA-2001 consists of a mRNA, which is a human-optimized streptopiogenous mRNA that produces the Cas9 system and a single guide RNA that is specific to the TTR sequence, and this is enclosed in a lipid nanoparticle and results in a greater than 95% TTR knockdown. This is data from the phase 1 study at day 28, and so far has not been associated with significant off-target effects or side effects. So this is very exciting data. So this again is a scheme from one of Rick Rueberg's excellent review articles from some years ago, which lays out the current treatment landscape for wild-type ATTR and hereditary ATTR. So as you can see here, if patients have hereditary ATTR with some component of neuropathy, whether they have cardiomyopathy or not, one of the RNA silencers can be used. For wild-type ATTR, it is only tefamidase that is approved for use, and diflunasol, of course, can be used as an off-label agent in all of these patients. But this landscape is going to change. We anticipate that Acquiramidase will be approved by the FDA, I think the PDUFA date is sometime in November in 2005. Vitruceran, with its very encouraging results, probably will be approved in the next year or so. And then ongoing trials with aplanteracin, the Cardiotransform trial, gene editing, and antifibrillotherapy, and all of these agents will be applicable to both wild-type and hereditary ATTR patients. Now I want to end with this slide, and that is to remind everybody that these agents have varying effects on the TTR levels. So agents that reduce the production of TTR will result in significant reductions in prealbumin levels, or TTR levels, in the blood, whereas agents that stabilize the TTR will increase the prealbumin levels in the blood. Now, as of now, there is no evidence to suggest that one of these approaches is better or worse than the other, but I think as we accrue more and more experience with these agents, these differences in the mechanism of action is something that we should keep in mind. Now I haven't spoken about several other issues related to specific therapy in TTR amyloidosis, including the cost of the drugs and how to equitably deliver the drug to white segments of the population, and very importantly, as these agents become, as more and more agents become available, we're all going to face choices of individual agents and combination therapy in these patients, and there's very little data as yet to guide us on those aspects. I'll stop there, and thank you very much for your attention. Excellent presentation. I want to thank all the faculty members for doing the presentations, and we want to go into the Q&A portion of the discussion. If you have a question, please enter into the Q&A button at the bottom of your screen, and we'll go ahead and start with a few. I'll read some of these. Some of these have already been answered, typed in, but I'll start with the last question. Can the PYP nucleus scan be picked up by the stomach or intestine, and will it be positive for amyloid? Maybe I'll get Dr. Durvala. You want to answer that question? Sure, yes. So that's a great question. PYP uptake is in the bone and in the heart muscle. If you see PYP uptake in the stomach, it actually indicates a problem with the quality control, which is 3-per-technetate, so we need to be careful. We cannot use PYP to image gastrointestinal amyloid, and in fact, we just looked at a case today from one of our clinical trials where there was a lot of uptake in the stomach, and then the question became, is the cardiac image now uninterpretable because it's poor tagging of the radiotracer? So that's something to keep in mind. So we cannot image gastrointestinal amyloid with PYP. If you see PYP in the stomach, think of a quality control problem. Great, and maybe the next question, is there a non-biopsy pathway using MRI to make the ATTR diagnosis? Assume you can rule out AL in the usual manner. Maybe I'll throw that back to you, over to you again, Dr. Dorbala. Sure, I don't know if Dr. Rubak is still on, but yeah, so I think people have proposed, Dr. Rubak can chime in, people have proposed using a similar approach because MRI can be somewhat specific for amyloid in terms of gadolinium kinetics, and if you see this diffuse late gadolinium enhancement and diffuse ECV expansion. But the one thing in my mind that's different between MRI and with nuclear techniques is nuclear techniques are molecular imaging techniques. They tell us about a process, whereas MRI techniques currently are not nuclear. Maybe there will soon be some molecular tracers for amyloid using MRI as well. So just purely based on that, I would hesitate to say that you could make a definitive non-biopsy diagnosis without biopsy using MRI alone, but Dr. Rubak is an MRI expert, yeah. Yes, Dr. Rubak, would you care to comment? Sure, I would. Of course, as usual, I'll agree with my colleague, Dr. Dorbala. There is a publication that was published in EHJ CDI by Jeremy Slipnick from the University of Chicago that kind of tried to follow the same pathway that the consensus PYPDPD bone avid radiotracer paper followed by using a combination of serum testing for and urine testing for a monoclonal gammopathy clone as well as the imaging. But I think the proof is in what is the imaging that we call diagnostic. A lot of those scans, a lot of those categorizations are kind of subjective categorizations of typical versus non-typical because there are a lot of things that can cause an MRI to be abnormal, as I'm sure the attendees know, that can potentially look like amyloid and be kind of overlapping but not necessarily diagnostic of amyloid with the early myocardial nulling, the very high extracellular volume, and the diffuse nonspecific pattern in latent enhancement. So I think the problem is going to be that gray area, whereas I do think just like in nuclear imaging, a negative predictive value is probably pretty high. So an MRI that shows no latent enhancement in a normal extracellular volume fraction probably does not have, you know, clinically important and may not have any amyloid at all, similar to what you would say about a negative PYP or a negative PET scan. Thank you. Maybe Dr. Suman, I can answer this next question. Is anyone using combination therapy at this point? We don't routinely use combination therapy. Our approach is, at least for now, if the presentation is one of primary cardiomyopathy to use the stabilizers, if it's one of primary neuropathy to use one of the RNAi agents. Now, there are times when you put somebody on an RNAi agent and they clearly progress in their cardiomyopathy on the RNAi, and in a couple of patients like that, I've added on tefamidase or vice versa. They have cardiomyopathy and they're on tefamidase and the neuropathy is progressing, and then I've added on RNAis. But we do not routinely use combination therapy as of now. Dr. Al-Raham, this question for you. Is there any evidence for CNS symptoms with HTTR patients with seizures and mild early onset dementia along with cardiomyopathy? Not really, not so far. So, CNS is very unlikely to be affected with amyloidosis, at least in this kind of presentation with seizures or dementia. Okay. How about this question? How common is it when multiple indicators are present, such as carpal tunnel spinal stenosis, neuropathy, amyloidosis is not present? What else can cause all these symptoms concurrently other than amyloidosis? What else should we think of? That's a very good question. We had actually a few cases here with Dr. Solman, and initially patients had completely negative workup, including genetic testing, and down the road, in 5-10 years, they developed kind of the cardiac symptoms and their PYP scan was positive. So, I think I would have very low tendency, if I have that clinical presentation, even if their neurology workup is negative and they don't have the cardiac evaluation, it might be reasonable to establish care with cardiology. When we suspect a patient having amyloidosis, how should we interpret the FLC capital lambda ratio immunofixation results, especially when one or two are abnormal while some are normal? Maybe, Dr. Rueberg, you want to answer that one? Sure, that's a great question. I think, you know, I always, you know, tell my, you know, trainees, and when I give talks like this, nothing in our cardiology training has prepared us to interpret light chain testing. It is, it is something that is not, it's a nuanced interpretation. So, I think I would say, I mean, the evidence would suggest that if the serum immunofixation electrophoresis is negative, the urine electrophoresis is negative, and the red light chain ratio is in the normal range, there is a 98% or a 99% chance that this is not AL amyloidosis. It is still possible, but you're talking about a rare disease and a rare occurrence of a rare disease, so that virtually excludes light chain amyloidosis. However, there are some instances where the light chain ratio is elevated, and that's actually quite common as people get older because the kappa light chain, it's a kappa to lambda ratio, kappa to lambda ratio, and if the, as people get older and their EGFR drops because kappa is cleared primarily through the kidney, you can have an elevated kappa ratio that can get up to about three with an upper range of normal, typically 1.65 in the typically used free light assay. So, but someone with a ratio of three who is a 25-year-old, for example, with a normal EGFR would not be normal. So, you have to look at the adjustment based upon the kidney function. Also, the other pearl that I tell people is that if the lambda, if it's a lambda monoclonal hemopathy and the ratio is low, that is always abnormal and always should be followed up by a hematologist because about 70% of cases of AL amyloidosis are lambda. So, seeing a lambda abnormality on immunofixation or a lambda light chain number, which would be less than 0.26 on the free light assay, that should prompt further testing. But it is often the case where you'll have a normal ratio and an IFE abnormality that should go to hematology, or you'll have an abnormal ratio that's low that should go to hematology with no IFE, or if the kappa is elevated, say, above three or four, then you may have a kappa monoclonal hemopathy and should be seen by hematologist. I hope that that's all spelled out actually in the expert consensus decision pathway document that's been referenced by a couple of the speakers today. Great. Thank you for that. Maybe Dr. Pomenosky, you can answer this question. What is the male versus female prevalence of HTTR in African ancestry? Yeah, I think there's been some debate. I think the latest article from European Heart by Ritchie Patel really didn't find much of a difference, and that was in 2022. So, really, I don't think that there's a striking difference. It used to be that perhaps we're missing the females. I think the jury is still out, but nothing definitive and that I could come across. Okay. This is kind of open to the group. What diagnosis pathway would you recommend for elderly patients with multiple red flags but normal LV thickness? Dr. Moore, you want to answer that question? Maybe get your thoughts about that. Normal LV wall thickness? Yes. I guess it depends. It obviously reduces the likelihood, but doesn't exclude it. It could be early in the course of the disease, but then the question would be, is it really responsible for symptoms? So, I'd be curious what the others think about that. Yes. Dr. Rueberg. Yeah, that's a great question, and I think Dr. Moore's observation is absolutely right. I mean, as we try to look earlier and earlier to find people, especially when we start either screening people with bilateral corporal tunnel syndrome and spinal stenosis, you're going to find that wall thickness filter that we apply to identify people with more advanced disease is going to fall away. Dr. Dorbala is the imaging expert on this call, on this call, on this meeting, and I'm sure she has thoughts on this, but I would probably start with a cardiac MR in that particular case. It depends, though. I mean, sometimes you can't do a cardiac MR if the patient has a no implanted electrical device that's not MR compatible or a defibrillator already, or if their EGFR is too low, that can impair the interpretation of the gadolinium enhancement. But generally, the studies that have looked at PYP and imaging, and I know Prem and Tramila can comment on this, are generally looking at people who are of a higher likelihood of disease, and so that's where it's been validated. But when you start screening populations of patients who have normal wall thickness, you're more likely to have false positive. But I don't think there's really a good answer. This is where potentially the more sensitive specific pet agents that were alluded to by Dr. Dorbala might have utility, but they're not presently approved yet for those indications. So it also... Yeah, maybe I can add one comment to that. I fully agree with the comments so far. The one thing I want to point out is patients who have the light chain form of disease may not have significantly increased wall thickness. So if everything else is pointing to amyloid and your wall thickness is kind of close to normal or not very abnormal, maybe Dr. Rubag will be the expert, but I was thinking that maybe you should consider light chain amyloidosis in these patients. That's an excellent point. And that's where, again, that kind of would support the use of CMR as an initial diagnostic test, because the PYP scan would obviously likely not be diagnostic in that setting. Just a comment that everyone has said, that this is a multi-system disease. So it just depends on your index of suspicion, right? So they may have cardiac involvement, they may not have cardiac involvement. Sometimes we see neurological involvement, GI involvement, even though we do a thorough workup for cardiac. So the sort of index of suspicion is important. And furthermore, things like troponin and BNP are also at times useful if we do have that index at an earlier stage. Excellent. The next one, maybe Dr. Durbala, is anyone using HMDP and what about soft tissue uptake? Sorry, that's a good question. So in the United States, PYP is most widely used, but HMDP is available. And what we've seen over the past two years or so is that during periods of PYP shortage, people have been using HMDP. For instance, this week at the Brigham, we are using HMDP and because we have a PYP shortage. So the large clinical trial with the 1498 patients included DPD, PYP, and HMDP. And when you look at the results, they were very similar for all three tracers. So you could use any of the tracers technically, but for soft tissue uptake, you see more with DPD that has been described in Europe. We don't see much soft tissue uptake with PYP, and we see even less soft tissue uptake with HMDP because it just shows more bone and less blood pool and less soft tissue uptake. Next question for Dr. Laham. Similar natural history for AL dementia, amyloid accentuates in the brain long before symptoms appear. Looking at treating these patients before symptoms, how about the heart? Any studies looking at early therapy? Yeah, but the question here, is it connected or not? Especially that the symptoms that we are talking in the early stages, it might be just like any other neuropathies. So that would be more a specific question for people with family history and positive gene that even if they don't have the diagnosis, but they're going to develop it definitely. That's where I would be more aggressive. Right. Maybe, Dr. Rueberg, the next question, if CMR is abnormal in these patients with normal wall thickness, but they are asymptomatic, what would you do with this information? Would you treat them? It's an excellent question. So I think, so we're going down the pathway that it could be AL. It also could be early ATTR. The CMR, as we discussed earlier, is not sufficient to make the diagnosis alone. I definitely would proceed with light chain testing as we just discussed, as if you're kind of recapitulating the nuclear pathway. And if the light chain testing is abnormal, then that's helpful. I would not treat the patient. In the United States, this is different than elsewhere in the world, but in the United States, we still require, and I speak about the major treatment seminars that are represented on this call, we require a tissue diagnosis to make AL amyloidosis a diagnosis. So if your suspicion was high and you were not sure, then you would need to do a tissue biopsy. We do fat aspirates at our center. They're relatively benign procedures and their sensitivity for identifying AL amyloidosis is quite good. It's usually around 75 to 80% if you have a good specimen. But again, I think most people would tell you if your suspicion remains high, then you should move forward with a heart biopsy. And I hesitate to recommend heart biopsy in people with normal wall thickness. I don't do that without caution because obviously the risk of perforation is higher. But if your suspicion remains high and you think that the patient is highly likely to have AL amyloidosis if their light chain testing is abnormal, but you have no other way to make the diagnosis, that's the way you have to go. I'm curious. I'm sure, you know, I'm sure, I'm sure. I just wanted to add there. I think this is where the PET tracers may have an important role because they seem to work really well for the light chain form of disease. Although the atrialis compound, it was amyotide seems to work well for both ATTR and AL, but the F-18 brain amyloid PET tracers work really well for amyloid. So we hope that in future they'll be tested and these are the scenarios where we may pick up patients who have uptake in the heart and then they could go on to biopsy if needed. Well, we're at the time of the end of the program. I certainly want

Cardiac amyloidosis diagnosis

Biomarkers for cardiac amyloidosis

Troponin for diagnosis

NT-proBNP for diagnosis

Echocardiography for diagnosis

Cardiac MRI for diagnosis

Nuclear scintigraphy for diagnosis

99M T-centered pyrophosphates

Technetium bis-phosphate HMDP

MIPP for imaging contrast

AAVC1195 for cardiac amyloidosis

Pitfalls of nuclear scintigraphy

Therapeutic algorithm for cardiac amyloidosis

Imaging modalities for cardiac amyloidosis

Multi-organ involvement in cardiac amyloidosis