This is Mark Penn, co-founder and chief medical officer of Cleveland Heart Lab and medical director for the cardiovascular metabolic endocrine franchise of Quest Diagnostics. And I’m going to talk today about the inflammatory markers for defining cardiovascular risk and in particular, how they’re useful in identifying those patients at increased risk with respiratory viral syndromes or a coronavirus pandemic. We’ll talk about the cardiovascular risk and COVID 19 outcomes, discuss the role of vascular inflammation and cardiovascular risk assessment and review how viral infections impact vascular inflammation and plaque rupture. The presence of underlying risk for cardiovascular disease portends a poor outcome in patients who develop or get exposed to COVID 19. We look at the risk factors that predict poor outcomes. It’s hypertension, obesity, diabetes, prior cardiovascular disease and chronic lung disease, as well as aging above 65 years of age. The mortality rates of hypertensives is 6%, diabetics 7%, prior CAD 10%. With no comorbidities associated with cardiovascular disease, the mortality has been found to be relatively low. In Wuhan, China, a study of about 400 patients identified about 20% of the patients had cardiac injury and of that 19%, 51% died. The mechanisms of this injury are not yet well defined. However, there’s also a coronary microvascular disease microvascular dysfunction, as well as macrovascular disease that leads to a poor outcome in patients with COVID. And again, these macrovascular microvascular disease states are associated with risks of cardio metabolic syndrome and cardio metabolic disease, namely obesity, diabetes, hypertension and prior coronary artery disease. Vascular Inflammation is really the etiology of myocardial infarction and microvascular dysfunction. Cholesterol doesn’t cause heart attacks. It drives vascular inflammation leading to heart attacks. So does hypertension, diabetes, depression, poor dental, hygiene, stress, anxiety as well as we’ve discussed respiratory viral infections. When we started Cleveland HeartLab, our goal was through increasing our understanding of the inflammatory pathways involved in disease. We wanted to define novel biomarkers that could enable us physicians to better stratify patients for cardiovascular risk, identify the source and cause of that risk, and provide tools to monitor the therapeutic response to make sure we’re mitigating that risk with the goal is obviously to help practitioners reduce cardiovascular events and improve our patients outcomes. You can take measures of cardiovascular risk and in general divide them into one of three buckets. The ATTHA guidelines tell us to start testing our patients in their 20s for like a protein or lipid risk. Clearly, if a 23 year old has an LDL of 400, they have a lifelong problem trying to get to more precise measures, people looked at advanced lipid testing and fundamentally are looking for the discordance between A high or B, yet a controlled LDL cholesterol. Given there’s a single B per LDL molecule, if you have a high will be in a low cholesterol. These particles must be numerous and they should be small and dense or atherogenic and advanced lipid testing really gives us a measure of risk on orders of a decade or so. And then there’s the near term risk, which is really defined by inflammatory markers which can give us evidence of risk of an event within a handful of years. Atherosclerosis and inflammation is an old story and not a new one. Our markers are relatively new, but the story is old. This an update by Russell Ross in 1999 about his response to injury hypothesis of atherosclerosis originally proposed in the mid 1970s. The concept here is the response syndrome hypothesis drives atherosclerosis and injury is what starts the process, and that injury can be cholesterol mediated, can be diabetes, hyper glycemic mediated and other causes. But it’s the response, the inflammatory response that really drives the disease leading to clinical events. We as a field have focused for 40 years on the injury leg, if you will, by controlling lipids and bringing cholesterol levels down on a population basis to the point where 50% or more of the patients who are now presenting to hospital with myocardial infarction actually have controlled or low LDL cholesterol. So for us to have the next profound impact on improving the health of heart disease in America and other developing countries, we need to look at the second leg of Dr. Ross’s hypothesis, which is really response. The Jupiter trial published about a decade ago, was really the first to demonstrate this in a therapeutic strategy. Patients enrolled in Jupiter at the time did not belong on statin therapy based on guidelines, but they all had a high density CRP greater than 2. They were randomized to either rosuvastatin 40 milligrams or placebo. It was a five year trial that showed benefit in 19 months, and that benefit was survival. What we see here is if patients were on ROSUVASTATIN but their LDL remained above 70 or their CRP and their CRP remained above 2 they looked like placebo. But if their LDL was less than 70 or their CRP was less than 2 they did better. And if their LDL was less than 70 and their CRP was less than 2, they did get better. These data and other data like them really demonstrate and bring home the point. But if we choose to define our patients lipid risk or inflammatory risk, we’re really choosing to define half the risk profile. At Cleveland HeartLab and Quest Diagnostics, we’ve developed an approach to defining cardiovascular risk based on oxidation and inflammation going from F to isoprostane which is the gold standard for oxidation in the body and really defining lifestyle risk, oxidized LDL that’s defining to alter physiology and risk of metabolic syndrome, asymmetric dimethylarginine (ADMA) microbial binary or high ccrp really defining the presence of disease and Myeloperoxidase or MPO and Lp-PLA2 activity as the measures of vulnerable plaque formation. A recent study by Williams and colleagues demonstrates the utility of CT angiography and what type of plaque truly correlates with the risk of myocardial infarction or clinical cardiovascular event. On 1800 patients who were enrolled in this trial, there was a significant number of diabetics hypertensives, a few who had prior myocardial infarction. And what they did was they looked at the CT angio and defined divided into 4 measures of plaque, total plaque burden, non calcified plaque burden, fatty streaks, fatty lesions, if you will, low attenuation, plaque burden and then calcifications calcified plaque burden, which is really the coronary calcium score that we’re used to. We see that based on 10 year cardiovascular risk, coronary calcium score or calcified plaque burden correlates nicely with the risk going from 0 to 1.6 or less than 10% risk or greater than 20 low attenuation plaque burden is increased in patients or subjects who had cardiovascular risk based on the risk calculator but didn’t correlate as strongly with the degree of risk ending with non calcified plaque burden and similar to total plaque burden. Further, the same correlations were seen in total calcium where the calcium score or calcified plaque burden correlated nicely with the amount of calcium. But the other measures were increased in the presence of calcium but did not correlate. And similarly in obstructive disease, when a univariate analysis was looked at or prediction of cardiovascular events, either fatal or not fatal at MI what we see here is all 4 types or 4 measures of plaque correlated with the risk of myocardial infarction. However, in a multivariate analysis, when all 4 were included in the model, only low attenuation plaque burden predicted the risk or correlated with the risk of state or nonfatal myocardial infarction. And in the graph on the right, you’ll appreciate that greater or less than 4% low attenuation plaque burden define 2 different populations. One had a significant risk of having a myocardial infarction over the ensuing years versus those who had a much lower risk. But what is low attenuation plaque? Low attenuation plaque is inflamed plaque with an activated thin fibrous cap and activated endothelium or a large inflamed, necrotic core for biomarkers that were on the artery wall I showed you before, endothelial activation of the thin fibrous cap is really defined by Myeloperoxidase and the large inflamed necrotic core is defined by Lp-PLA2 activity. So we can use markers to define cardiovascular risk. And we’re beginning to understand specifically the types of lesions that these markers are defining. Based on the data I just shared with you, where’s the value in this? And I think this is shown here by Wong and colleagues. Again, a study done about a decade ago. But important. Based on more recent data, Wong and colleagues looked at patients with calcium score and stratified the calcium score based on Myeloperoxidase. so what you see is the higher the calcium score, the higher the Myeloperoxidase level, which makes sense. The more disease you have, the more likely you are to have vulnerable plaques. And they showed, as we have shown and many others have shown, the higher the Myeloperoxidase, the higher the risk of having a cardiovascular event. But what’s interesting is when you divide the patient population to those who have a low or controlled MPO versus those who have a high MPO, you’ll appreciate that a high calcium score predicts a high risk of cardiovascular events here 7.1%. But that’s in the setting of a normal MPO or low amount of low attenuation plaque. However, if you have a high MPO, you double that 7.1% risk of 14 because not only do have a lot of calcium and a lot of disease, but you also have a lot of low attenuation plaque burden because the MPO is high and reflecting the presence of a vulnerable plaque. So biomarkers blood based biomarkers in this case and anatomical measures based on calcium scoring or CT angiography are synergistic, they are not redundant. And importantly, the blood based biomarkers would modulate to effective preventive strategies and go down, whereas the calcium score will actually go up if you start somebody on a statin as those arteries leach out of cholesterol and calcify, thus rendering calcium scores as ineffective to a determine if we’ve actually improved the patient’s outcome and whether preventive strategies are identifying the risk that that specific patient may have. So why is defining cardiovascular risk and vascular inflammation critical, especially in these days of a viral pandemic? An important study by Kwong and colleagues 2 years ago in the New England Journal looked at the risk of myocardial infarction in the 7 days following a positive viral titer compared to the 52 weeks prior to that positive viral titer and the 51 weeks following the week of a positive viral titer. It’s important to remember that the CDC used to define a flu epidemic or increase the amount of flu in a region not based on viral titers in the 80s and 90s, where we didn’t measure them, but rather an excess of myocardial infarction and mortality in the elderly in that region. These data are really, as you’ll see, highly supportive of that concept. Looking at the patient characteristics in this study, about 300 patients, median age of 77 higher risk if you’re above 65, as we’re seeing with COVID, no difference really between men and women. Interestingly, if patients had a prior hospitalization for a myocardial infarction, there was less risk, no risk, but less risk likely because they were on aspirin and other preventive strategies. 50% were diabetics, a third were dyslipidemia and 85% were hypertensive. So similar to the risk factors that we’re experiencing in the COVID pandemic, the primary finding of the study is really on the first line here. There was a 6.05 fold increased risk of a myocardial infarction in the 7 days following a positive viral titer compared to the 52 weeks before, in the 51 weeks after, essentially 7 days of a positive viral titer is the time in which patients are at greatest risk of having a myocardial infarction over a 2 year period. And interestingly, this risk goes down rather rapidly by day 8 where you see a 0.6 incident ratio that is not statistically significant. The rest of this table focuses on the sensitivity analysis to try and demonstrate that the 6.05 fold increase in zero ratio is real. And towards the bottom of this slide, you’ll see that other respiratory viruses also cause this increased risk, whether it’s a respiratory syncytial virus or respiratory virus other than influenza or RSV, that is a negative control, if you will. They looked at hospitalizations for diabetes and associated complications and found not a significant increase in MI. So it’s not just being admitted to the hospital, it causes a risk. It’s really the respiratory viral syndrome. When they look at a subgroup analysis, again, similar to what we’re seeing with COVID, greater than 65 years of age has a significantly increased risk, although less than 65 still has some risk. No difference between men and women. They were both Influenza A and B, although B tended to have a higher risk here, both did what you were vaccinated or not. If you got sick, you got sick and had an increased risk and a history of prior MI. Again, so had a significant increased risk but less of a risk likely due to the fact that there was presence of some preventive strategies on board. Looking at this figure from the European Society of Cardiology and one of their papers, there is an activation of multiple inflammatory pathways by COVID and frankly by most respiratory viral syndromes that ultimately impacts on two things leading to a poor outcome based on cardiovascular risk. One is if they have plaque instability, they’re at risk of rupturing, leading to a macrovascular event and a myocardial infarction. Consistent with what Kwong and colleagues showed us and an MPO or a positive Lp-PLA2 two can identify patients prior to developing a respiratory viral syndrome or an increased risk of a poor outcome and give us the opportunity to try and passivate that vascular inflammation in an attempt to minimize their risk of a poor outcome. If they were to develop COVID 19 or other respiratory viral syndromes. The second risk is really a microvascular dysfunction, which seems to have a higher predilection with coronavirus, and Myeloperoxidase consumes nitric oxide and asymmetric dimethylarginine inhibits nitric oxide generation. So again, identifying folks who have a high MPO or a high ADMA and increasing these levels should increase nitric oxide bioavailability to the microvasculature and lead to less of the microvascular dysfunction when they develop a respiratory viral syndrome or coronavirus. This is the approach we’ve developed. Specifically ADMA, MPO and Lp-PLA2 activity offer us an insight into our patients current vascular health in an attempt to optimize that health to minimize the risk of a poor outcome if they were to develop COVID 19 or other respiratory viral syndromes. Why 2 measures of Vulnerable Plaque? We’ve previously published these data of about 2800 patients from an executive health and preventive cardiology clinic looking at the positivity of Lp-PLA2 and Myeloperoxidase again, Lp-PLA2 is reflecting that activated, necrotic or developing vulnerable plaque and in this population, about 5% were positive. MPO is the finding that thinning fibrous cap and activated ilium and this population had a 6% positivity despite having about 2800 patients in front of you. You’ll appreciate that only about 6 patients had both markers up, so MPO and Lp-PLA2 are defining different risk and are synergistic in defining the total population of risk. And if you had a high healthy Lp-PLA2 and MPO, you can appreciate it’s more a reflection of plaque rupture in a patient who is particularly high risk. So the conclusions of May 2020 really are vascular Inflammation is the cause of plaque rupture, lab cardio infarction, not hyperlipidemia or diabetes, but hyperlipidemia and diabetes clearly dry vascular inflammation. It should have been anticipated that patients with cardiovascular disease or CV risk, age, obesity, diabetes and hyperlipidemia are at the greatest risk of death from COVID 19. It’s consistent with every other viral substrate viral syndrome we have and tens of thousands of deaths we see in this country every year due to influenza and other respiratory viral syndromes. We can reassure patients and optimize outcomes by assessing and addressing their level of vascular inflammation in an attempt to define those who may be at the greatest risk and try to passivate that risk prior to them getting sick. And our data suggest identifying patients with vascular inflammation increases the likelihood of greater investigation into and treatment of the underlying risk drivers and those drivers of risk once address resulted in a lowering of an MPO and Lp-PLA2 and ADMA and all of our partners, quite frankly, defining patients who have a decreased risk. Once again, I thank you very much for your time and your interest in what we’re trying to achieve at Cleveland Heartlab and Quest Diagnostics. We’re happy to answer any questions you may have in the future. Thank you.