Association of global coagulation profiles with cardiovascular risk factors and atherosclerosis: A sex disaggregated analysis from the BioHEART-CT study

Background: Although the association between dysregulated coagulation and atherosclerosis is well recognized, individual assays have been of minimal value in understanding disease susceptibility. Here we investigated the association of global coagulation profiles with coronary artery disease with consideration of sex differences. Methods and Results: The study included patients from the BioHEART-CT (The BioHEART Study: Assessing Patients With Suspected Cardiovascular Disease for New Disease Markers and Risk Factors) biobank who had computed tomography coronary angiograms scored for coronary artery calcium score (CACS) and Gensini score. The cohort included 206 adult patients who were referred for clinically indicated computed tomography coronary angiography and had a median of 2 major cardiac risk factors; 50% were women and the average age was 62.6 years (±9.9 years). The overall hemostatic potential (OHP) and calibrated automated thrombography generation assays were performed on platelet-poor plasma. CACS and Gensini score in men were significantly correlated in bivariate analysis with measures from the OHP assay, and regression models predicting disease severity by CACS or Gensini score were improved by adding the OHP assay variables in men but not in women. The calibrated automated thrombography generation assay demonstrated a more hypercoagulable profile in women than in men. The OHP assay showed hypercoagulable profiles in women with hyperlipidemia and men with obesity. Conclusions: The OHP assay identified hypercoagulable profiles associated with different risk factors for each sex and was associated with CACS and Gensini score severity in men, emphasizing the associations between increased fibrin generation and reduced fibrinolysis with cardiac risk factors and early atherosclerosis. Registration Information: www.anzctr.org.au. Identifier: ACTRN12618001322224.Katharine A. Kott, Marie-Christine Morel-Kopp, Stephen T. Vernon, Yuki Takagi, Belinda A. Di Bartolo, Karlheinz Peter, Jean Y. Yang, Stuart M. Grieve, Christopher Ward, Gemma A. Figtre

Incorporating a polygenic risk score-triaged coronary calcium score into cardiovascular disease examinations to identify subclinical coronary artery disease (ESCALATE): Protocol for a prospective, nonrandomized implementation trial

Background: Identifying and targeting established modifiable risk factors has been a successful strategy for reducing the burden of coronary artery disease (CAD) at the population-level. However, up to 1-in-4 patients who present with ST elevation myocardial infarction do so in the absence of such risk factors. Polygenic risk scores (PRS) have demonstrated an ability to improve risk prediction models independent of traditional risk factors and self-reported family history, but a pathway for implementation has yet to be clearly identified. The aim of this study is to examine the utility of a CAD PRS to identify individuals with subclinical CAD via a novel clinical pathway, triaging low or intermediate absolute risk individuals for noninvasive coronary imaging, and examining the impact on shared treatment decisions and participant experience. Trial Design: The ESCALATE study is a 12-month, prospective, multicenter implementation study incorporating PRS into otherwise standard primary care CVD risk assessments, to identify patients at increased lifetime CAD risk for noninvasive coronary imaging. One-thousand eligible participants aged 45 to 65 years old will enter the study, which applies PRS to those considered low or moderate 5-year absolute CVD risk and triages those with CAD PRS ≥80% for a coronary calcium scan. The primary outcome will be the identification of subclinical CAD, defined as a coronary artery calcium score (CACS) >0 Agatston units (AU). Multiple secondary outcomes will be assessed, including baseline CACS ≥100 AU or ≥75th age-/sexmatched percentile, the use and intensity of lipid- and blood pressure-lowering therapeutics, cholesterol and blood pressure levels, and health-related quality of life (HRQOL). Conclusion: This novel trial will generate evidence on the ability of a PRS-triaged CACS to identify subclinical CAD, as well as subsequent differences in traditional risk factor medical management, pharmacotherapy utilization, and participant experience.Michael P. Gray, Yemima Berman, Giordano Bottà, Stuart M. Grieve, Amy Ho, Jessica Hu, Karice Hyun, Jodie Ingles, Garry Jennings, Gary Kilov, Jean-Frederic Levesque, Peter Meikle, Julie Redfern, Tim Usherwood, Stephen T. Vernon, Stephen J. Nicholls, and Gemma A. Figtree, On behalf of the PPP-CAD Collaborator

Patient endothelial colony-forming cells to model coronary artery disease susceptibility and unravel the role of dysregulated mitochondrial redox signalling

Mechanisms involved in the individual susceptibility to atherosclerotic coronary artery disease (CAD) beyond traditional risk factors are poorly understood. Here, we describe the utility of cultured patient-derived endothelial colony-forming cells (ECFCs) in examining novel mechanisms of CAD susceptibility, particularly the role of dysregulated redox signalling. ECFCs were selectively cultured from peripheral blood mononuclear cells from 828 patients from the BioHEART-CT cohort, each with corresponding demographic, clinical and CT coronary angiographic imaging data. Spontaneous growth occurred in 178 (21.5%) patients and was more common in patients with hypertension (OR 1.45 (95% CI 1.03–2.02), p = 0.031), and less likely in patients with obesity (OR 0.62 [95% CI 0.40–0.95], p = 0.027) or obstructive CAD (stenosis > 50%) (OR 0.60 [95% CI 0.38–0.95], p = 0.027). ECFCs from patients with CAD had higher mitochondrial production of superoxide (O2 −–MitoSOX assay). The latter was strongly correlated with the severity of CAD as measured by either coronary artery calcium score (R2 = 0.46; p = 0.0051) or Gensini Score (R2 = 0.67; p = 0.0002). Patient-derived ECFCs were successfully cultured in 3D culture pulsatile mini-vessels. Patient-derived ECFCs can provide a novel resource for discovering mechanisms of CAD disease susceptibility, particularly in relation to mitochondrial redox signalling.Marie Besnier, Meghan Finemore, Christine Yu, Katharine A. Kott, Stephen T. Vernon, Nicole A. Seebacher, Elijah Genetzakis, Anamarija Furman, Owen Tang, Ryan L. Davis, Thomas Hansen, Peter J. Psaltis, Kristen J. Bubb, Steven G. Wise, Stuart M. Grieve, Belinda A. Di Bartolo, and Gemma A. Figtre

Taking the next steps to implement polygenic risk scoring for improved risk stratification and primary prevention of coronary artery disease

Coronary artery disease (CAD) remains the leading cause of death worldwide. The role of hypertension, cholesterol, diabetes mellitus, and smoking in driving disease has been well recognized at a population level and has been the target of primary prevention strategies for over 50 years with substantial impact. However, in many cases, these factors alone do not provide enough precision at the individual level to allow physicians and patients to take appropriate preventive measures and many patients continue to suffer acute coronary syndromes in the absence of these risk factors. Recent advances in user-friendly chip designs, high speed throughput, and economic efficiency of genome-wide association studies complemented by advances in statistical analytical approaches have facilitated the rapid development of polygenic risk scores (PRSs). The latest PRSs combine data regarding hundreds of thousands of single-nucleotide polymorphisms to predict chronic diseases including CAD. Novel CAD PRSs are strong predictors of risk and may have application, in a complementary manner with existing risk prediction algorithms. However, there remain substantial controversies, and ultimately, we need to move forward from observational studies to prospectively and rigorously assess the potential impact if widespread implementation is to be aspired to. Consideration needs to be made of ethnicity, sex, as well as age, and risk estimate based on existing non-genomic algorithms. We provide an overview and commentary on the important advances in deriving and validating PRSs, as well as pragmatic considerations that will be required for implementation of the new knowledge into clinical practice.Gemma Alexandra Figtree, Stephen Thomas Vernon, and Stephen James Nicholl

Progression of coronary atherosclerosis in patients without standard modifiable risk factors

Background and aims: The outcome of patients with clinical coronary artery disease despite traditional risk factors is poorly understood. Methods: Clinical characteristics and plaque burden on serial intravascular ultrasonography were compared in patients without (n ¼ 165) and with (n ¼ 492) standard modifiable risk factors after matching on age, sex and use of statins from a database of 5823 patients participating in clinical trials of anti-atherosclerotic therapies. Results: Patients without standard modifiable risk factors had lower baseline systolic blood pressure (118 ± 12 vs. 129 ± 17 mmHg, p < 0.001), low-density lipoprotein cholesterol (87 ± 21 vs. 104 ± 34 mg/dl, p < 0.001), triglycerides [106 vs. 136 mg/dl, p < 0.001)] and C-reactive protein [1.5 vs. 2.1 mg/l, p ¼ 0.001]. At baseline, patients without modifiable risk factors had a lower percent atheroma volume (35.7 ± 8.6 vs. 38 ± 8.8%, p ¼ 0.004) and total atheroma volume (174.7 ± 80 vs. 190.9 ± 84 mm³, p ¼ 0.03) and less images with calcification (22.2 vs. 26.5%, p ¼ 0.025). The use of aspirin and statin prior to and during the trials was similar. The use of ACE inhibitors and beta blockers was lower in the no risk factor group prior to and during the trials. The change in percent atheroma volume (-0.2 ± 2.8 vs. -0.1 ± 3.6%, p ¼ 0.71), total atheroma volume (-5.5 ± 23.4 vs. -3.8 ± 22.7 mm³, p ¼ 0.42), and the percentage of patients demonstrating any degree of progression (50.9% vs 45.1%, p ¼ 0.20) were similar in those without and with standard modifiable risk factors, respectively. Conclusion: Patients who develop clinical coronary atherosclerosis without standard modifiable risk factors have similar rates of plaque progression to those with traditional risk factors.Jawad Mazhar, Gemma Figtree, Stephen T. Vernon, Keyvan Karimi Galougahi, Julie Carlo, Steven E. Nissen, Stephen J. Nicholl

Mortality and Cardiovascular Outcomes in Patients Presenting With Non-ST Elevation Myocardial Infarction Despite No Standard Modifiable Risk Factors: Results From the SWEDEHEART Registry.

BACKGROUND: A significant proportion of patients with ST-segment– elevation myocardial infarction (MI) have no standard modifiable cardiovascular risk factors (SMuRFs) and have unexpected worse 30-day outcomes compared with those with SMuRFs. The aim of this article is to examine outcomes of patients with non–ST- segment– elevation MI in the absence of SMuRFs. METHODS AND RESULTS: Presenting features, management, and outcomes of patients with non–ST- segment– elevation MI without SmuRFs (hypertension, diabetes, hypercholesterolemia, smoking) were compared with those with SmuRFs in the Swedish MI registry SWEDEHEART (Swedish Web-System for Enhancement and Development of Evidence-Based Care in Heart Disease Evaluated According to Recommended Therapies; 2005–2018). Cox proportional hazard models were used. Out of 99 718 patients with non–ST- segment– elevation MI, 11 131 (11.2%) had no SMuRFs. Patients without SMuRFs had higher all-cause and cardiovascular mortality at 30 days (hazard ratio [HR], 1.20 [95% CI, 1.10–1.30], P<0.0001; and HR, 1.25 [95% CI, 1.13–1.38]), a difference that remained after adjustment for age and sex. SMuRF-less patients were less likely to receive secondary prevention statins (76% versus 82%); angiotensin-converting enzyme inhibitors/angiotensin receptor blockade (54% versus 72%); or β-blockers (81% versus 87%, P for all <0.0001), with lowest rates observed in women without SMuRFs. In patients who survived to 30 days, rates of all-cause and cardiovascular death were lower in patients without SMuRFs compared with those with risk factors, over 12 years. CONCLUSIONS: One in 10 patients presenting with non–ST- segment– elevation MI present without traditional risk factors. The excess 30-day mortality rate in this group emphasizes the need for both improved population-based strategies for prevention of MI, as well as the need for equitable evidence-based treatment at the time of an MI.Gemma A. Figtree, MBBS, DPhil, Oxon, FRACP, FAHA, Stephen T. Vernon, B.Med Sci, MBBS, PhD, FRACP, Nermin Hadziosmanovic, MSc, Johan Sundström, MD, PhD, Joakim Alfredsson, MD, Stephen J. Nicholls, MBBS, PhD, FRACP, Clara K. Chow, MBBS, PhD, FRACP, Peter Psaltis, MBBS, PhD, FRACP, Helge Røsjø, MD, PhD, Margrét Leósdóttir, MD, Emil Hagström, MD, PhD, FES

Single-cell immune profiling in coronary artery disease: the role of state-of-the-art immunophenotyping with mass cytometry in the diagnosis of atherosclerosis

Coronary artery disease remains the leading cause of death globally and is a major burden to every health system in the world. There have been significant improvements in risk modification, treatments, and mortality; however, our ability to detect asymptomatic disease for early intervention remains limited. Recent discoveries regarding the inflammatory nature of atherosclerosis have prompted investigation into new methods of diagnosis and treatment of coronary artery disease. This article reviews some of the highlights of the important developments in cardioimmunology and summarizes the clinical evidence linking the immune system and atherosclerosis. It provides an overview of the major serological biomarkers that have been associated with atherosclerosis, noting the limitations of these markers attributable to low specificity, and then contrasts these serological markers with the circulating immune cell subtypes that have been found to be altered in coronary artery disease. This review then outlines the technique of mass cytometry and its ability to provide high‐dimensional single‐cell data and explores how this high‐resolution quantification of specific immune cell subpopulations may assist in the diagnosis of early atherosclerosis in combination with other complimentary techniques such as single‐cell RNA sequencing. We propose that this improved specificity has the potential to transform the detection of coronary artery disease in its early phases, facilitating targeted preventative approaches in the precision medicine era.Katharine A. Kott, Stephen T. Vernon, Thomas Hansen, Macha de Dreu, Souvik K. Das, Joseph Powell, Barbara Fazekas de St Groth, Belinda A. Di Bartolo, Helen M. McGuire, Gemma A. Figtre

Immunoglobulin E sensitization to mammalian oligosaccharide galactose-alpha-1,3 (alpha-Gal) is associated with noncalcified plaque, obstructive coronary artery disease, and ST-segment-elevated myocardial infarction

Background: Treating known risk factors for coronary artery disease (CAD) has substantially reduced CAD morbidity and mortality. However, a significant burden of CAD remains unexplained. Immunoglobulin E sensitization to mammalian oligosaccharide galactose-α-1,3-galactose (α-Gal) was recently associated with CAD in a small observational study. We sought to confirm that α-Gal sensitization is associated with CAD burden, in particular noncalcified plaque. Additionally, we sort to assess whether that α-Gal sensitization is associated with ST-segment–elevated myocardial infarction (STEMI) Methods: We performed a cross-sectional analysis of participants enrolled in the BioHEART cohort study. We measured α-Gal specific-immunoglobulin E antibodies in serum of 1056 patients referred for CT coronary angiography for suspected CAD and 100 selected patients presenting with STEMI, enriched for patients without standard modifiable risk factors. CT coronary angiograms were assessed using coronary artery calcium scores and segmental plaque scores. Results: α-Gal sensitization was associated with presence of noncalcified plaque (odds ratio, 1.62 [95% CI, 1.04–2.53], P=0.03) and obstructive CAD (odds ratio, 2.05 [95% CI, 1.29-3.25], P=0.002), independent of age, sex, and traditional risk factors. The α-Gal sensitization rate was 12.8-fold higher in patients with STEMI compared with matched healthy controls and 2.2-fold higher in the patients with STEMI compared with matched stable CAD patients (17% versus 1.3%, P=0.01 and 20% versus 9%, P=0.03, respectively). Conclusions: α-Gal sensitization is independently associated with noncalcified plaque burden and obstructive CAD and occurs at higher frequency in patients with STEMI than those with stable or no CAD. These findings may have implications for individuals exposed to ticks, as well as public health policy.Stephen T. Vernon, Katharine A. Kott, Thomas Hansen, Meghan Finemore, Karl W. Baumgart, Ravinay Bhindi, Jean Yang, Peter S. Hansen, Stephen J. Nicholls, David S. Celermajer, Michael R. Ward, Sheryl A. van Nunen, Stuart M. Grieve, Gemma A. Figtre

Mass cytometry analysis reveals altered immune profiles in patients with coronary artery disease

Objective. The importance of inflammation in atherosclerosis is well accepted, but the role of the adaptive immune system is not yet fully understood. To further explore this, we assessed the circulating immune cell profile of patients with coronary artery disease (CAD) to identify discriminatory features by mass cytometry. Methods. Mass cytometry was performed on patient samples from the BioHEART-CT study, gated to detect 82 distinct cell subsets. CT coronary angiograms were analysed to categorise patients as having CAD (CAD+) or having normal coronary arteries (CAD ). Results. The discovery cohort included 117 patients (mean age 61 12 years, 49% female); 79 patients (68%) were CAD+. Mass cytometry identified changes in 15 T-cell subsets, with higher numbers of proliferating, highly differentiated and cytotoxic cells and decreases in na€ıve T cells. Five T-regulatory subsets were related to an age and gender-independent increase in the odds of CAD incidence when expressing CCR2 (OR 1.12), CCR4 (OR 1.08), CD38 and CD45RO (OR 1.13), HLA-DR (OR 1.06) and Ki67 (OR 1.22). Markers of proliferation and differentiation were also increased within B cells, while plasmacytoid dendritic cells were decreased. This combination of changes was assessed using SVM models in discovery and validation cohorts (area under the curve = 0.74 for both), confirming the robust nature of the immune signature detected. Conclusion. We identified differences within immune subpopulations of CAD+ patients which are indicative of a systemic immune response to coronary atherosclerosis. This immune signature needs further study via incorporation into risk scoring tools for the precision diagnosis of CAD.Katharine A Kott, Adam S Chan, Stephen T Vernon, Thomas Hansen, Taiyun Kim, Macha de Dreu, Bavani Gunasegaran, Andrew J Murphy, Ellis Patrick, Peter J Psaltis, Stuart M Grieve, Jean Y Yang, Barbara Fazekas de St Groth, Helen M McGuire, and Gemma A Figtre