Heart failure biomarkers: The importance of cardiac specificity Piek, Arnold
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10.33612/diss.146698618
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Piek, A. (2020). Heart failure biomarkers: The importance of cardiac specificity. University of Groningen. https://doi.org/10.33612/diss.146698618
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Chapter 1
Introduction
8
INTRODUCTION
Heart failure (HF) is a clinical syndrome characterized by a heart that is unable to adequately circulate blood to meet the metabolic demands of peripheral organs and tissues. Together, this results in typical HF signs and symptoms, including amongst others fatigue, dyspnea, peripheral edema and pulmonary congestion1. Besides being a disease with high morbidity, HF is also a disease with high mortality, and 5- and 10-year mortality rates are 50% and 90%, respectively2. Currently, 8% of the population aged > 75 years is diagnosed with HF, and, due to the ageing population, it is expected that the prevalence of HF will increase in the upcoming decades3. Though HF is always characterized by an insufficient cardiac output, underlying etiology and pathophysiology is different amongst HF patients, resulting in a heterogeneous population with a wide range of symptoms and comorbidities. Currently, stratification is based on physical examination and cardiac imaging, but additional tools are needed that allow better stratification and understanding of underlying molecular mechanisms, ultimately resulting in patient-tailored therapies.
Cardiac remodeling is defined as any structural, morphological or functional alteration of the heart, and myocardial fibrosis, cardiac hypertrophy and inflammation are important processes that contribute to cardiac remodeling1,3,4. At first, these are compensatory processes necessary for the heart to cope with stress conditions5. However, with sustained stress, for example due to myocardial infarction, hypertension, arrhythmias or valve insufficiencies, cardiac remodeling may become pathological and ultimately lead to HF development5-8. Proteins and other substances involved in or associated with these pathophysiological processes may end up in the circulation. Potentially, their blood plasma levels could serve as biomarkers for the severity of these processes in the heart, and for example markers specific for different types of fibrosis or hypertrophy could possibly be identified. Different etiologies of HF are associated with the development of different subtypes of HF. For example, myocardial infarction is a known driver of inflammation, replacement fibrosis and eccentric hypertrophy, and may result in heart failure with reduced ejection fraction (HFrEF left entric lar ejection fraction LVEF 40%). The other main subtype of HF is heart failure with preserved ejection fraction (HFpEF, LVEF 50%)1,9. Important risk factors for HFpEF are obesity, hypertension and aging, and these comorbidities are thought to induce a systemic proinflammatory state, eventually culminating into concentric hypertrophy and interstitial fibrosis, and hence diastolic dysfunction4,9. Finally, a relatively new subtype is heart failure with mid-range ejection fraction (HFmrEF, 40 LVEF 49%), which is thought to be an intermediate between HFrEF and HFpEF, resembling characteristics of both HFrEF and HFpEF1. Besides categorizing patients based on LVEF, which is still rather rough, biomarkers could possibly play a role in further stratifying subtypes of HF by distinguishing different types of remodeling.
Introduction
9
Not surprisingly, circulating substances have gained increased attention, and the diagnostic and prognostic quality of numerous of such proteins and substances as HF biomarkers were investigated. Criteria for the ideal biomarker have been postulated previously10-12. In particular, the ideal biomarker is accessible through noninvasive methods, is sensitive and specific for the respective disease, has a long half-life after obtaining the sample, allows early disease detection, is sensitive to changes in disease severity or other relevant changes within this disease, and is capable of rapid and accurate detection10-12. Blood plasma biomarkers have the potential to meet most of these criteria. Moreover, costs for measuring are relatively low and the lab infrastructure is available in most modern hospitals. Blood biomarker measurement can be quick and many tests can be performed on a daily basis. Therewith, the implementation of biomarkers in daily clinical practice, alongside established diagnostic means like electrocardiography and echocardiography, is highly feasible.
Several cardiac markers are included in the guidelines for HF treatment of the European Society of Cardiology (ESC) and the American Heart Association (AHA), and these include the natriuretic peptides and cardiac troponins1,13. Atrial natriuretic peptide (ANP) and brain natriuretic peptide (BNP), the most important variants of natriuretic peptides, are mainly produced in the atria and ventricles, respectively14. Production of both ANP and BNP increases upon mechanical strain in cardiomyocytes, and thus natriuretic peptides are myocardial stretch markers15-17. Blood plasma levels of natriuretic peptides have become standard in the evaluation of HF patients, and normal levels can rule out the presence of HF1,18-21. Cardiac troponins are released into the circulation upon cardiomyocyte damage, and elevated blood plasma levels of cardiac troponin hint towards ongoing cardiac injury1,22. Troponins are also present in skeletal muscle, but high sensitivity cardiac troponin (hsTn) assays specifically measure cardiac troponins and, moreover, these tests can detect low grade cardiac injury in the absence of acute myocardial damage1,22-24. Besides natriuretic peptides and hsTn, two other promising markers have been mentioned in the American HF guidelines, namely galectin-3 (Gal-3) and soluble suppression of tumorigenicity 2 (sST2), which are both associated with fibrosis13,25-28. However, their exact clinical value is still debated. Examples of other suggested HF biomarkers are interleukin-6 (IL-6), myeloperoxidase (MPO) and growth differentiation factor 15 (GDF-15) for inflammation, metabolite profiles and insulin-like growth factor binding protein 7 (IGFBP-7) for metabolic dysfunction, and heart-type fatty acid binding protein (H-FABP) for cardiomyocyte injury29-37. How these markers may improve patient stratification is schematically depicted in Figure 1. Current available pharmacological HF treatments predominantly focus on reducing cardiac workload, thereby preventing HF progression. Angiotensin-converting enzyme inhibitors (ACE-inhibitors) or angiotensin receptor blockers (ARBs), combined with beta blockers and diuretics, is the standard HF drug regime according to the European and American HF guidelines1,13. These drugs have improved outcomes of HFrEF patients, but not of HFpEF patients1,13,38, possibly explained by differences in etiology and cardiac remodeling between
10
Figure 1. Patient stratification based on HF biomarkers. Currently, HF cases (indicated in red) are selected
from the patient pool with HF symptoms using natriuretic peptides, imaging and ECG, and thus separated from patients with no HF (indicated in blue). Novel HF biomarkers could improve further stratification of the heterogeneous total HF population (indicated in different shades of red), here shown for myocardial fibrosis, inflammation, metabolic dysfunction, and cardiomyocyte injury. More markers and processes could be added to extend this figure. ECG=Electrocardiography. Gal-3=Galectin-3. GDF-15=Growth differentiation factor 15. H-FABP=Heart-type fatty acid binding protein. HsTn=High sensitive troponin. IGFBP-7=Insulin-like growth factor binding protein 7. IL-6=Interleukin-6. MPO=Myeloperoxidase. sST2=Soluble suppression of tumorogenicity 2.
these HF subtypes. Since HF biomarkers can be involved in the modulation of pathophysiological processes of HF, drugs that target biomarkers could improve current HF treatment strategies. An example of a drug that modulates biomarker levels is sacubitril, a drug which targets the degradation of natriuretic peptides. Natriuretic peptides reduce cardiac workload by reducing blood pressure and peripheral vascular resistance, since they promote a fluid shift from the intravascular to the extravascular compartment, promote
Gal-3 sST2 IL-6 GDF-15 IGFBP-7 Metabolites HsTn H-FABP Imaging Imaging ECG ECG MPO Natriuretic peptides Natriuretic peptides
Introduction
11
diuresis and reduce vascular sympathetic tonus14,39. Neprilysin is responsible for the degradation of natriuretic peptides, and by inhibition of neprilysin activity by sacubitril, part of the novel drug Entresto, the positive effects of natriuretic peptides can be enhanced40-42. In mice, inhibition of galectin-3 (Gal-3), a biomarker involved in myocardial fibrosis, resulted in reduced cardiac remodeling26. For HFpEF, which is thought to be inflammatory driven through HF risk factors like obesity, hypertension and diabetes mellitus, interference with inflammatory pathways could be beneficial4. Genetic disruption of myeloperoxidase (MPO), an inflammatory enzyme with increased plasma levels in HF, was associated with reduced cardiac remodeling and attenuated the severity of HF comorbidities, including obesity and diabetes, and might possibly be a HF treatment target35,37,43-46.
Taken together, plasma biomarkers have the potential to improve current diagnostic, prognostic and therapeutic strategies in HF patients, either by using their plasma concentrations to distinguishing different types of remodeling, or by altering the modulating effect of biomarkers on cardiac remodeling. Ideally, a large panel including multiple markers, herewith covering a wide variety of processes involved in HF development, should be measured in every HF patient, resulting in a detailed disease profile of each patient. These kinds of multi-marker panels could potentially improve treatment and prognosis of HF patients, and could result in a more personalized kind of medicine.
AIMS OF THIS THESIS
In this thesis, we aimed to investigate several aspects of HF biomarkers, including HF biomarker biology, the clinical performance of novel HF biomarkers and HF biomarkers as pharmacological treatment targets. As discussed, HF biomarkers have great potential to improve the stratification of HF patients, and could serve as therapeutic targets.
Figure 2. Approaches used in this thesis to study HF biomarkers. In all studies included in this thesis, either
one or a combination of the above approaches is used to study HF biomarkers. Each approach has its own advantages, as listed in the figure above each panel. HF=Heart failure.
Bioinforma ic
Total body gene expression Cardiac specificity Biomarker identifiction
HF animal model
In-depth molecular investigations Multi-organ biomarker biology Pharmacological biomarker targeting
H man HF cohor
Clinical relevance Prognostic quality Correlations with clinical variables
12
Nevertheless, for many suggested novel HF biomarkers their clinical value and biological function remains unclear. Preclinical studies are lacking, whilst these could provide further insights. Therefore, in this thesis we combined the strengths and advantages of three different approaches to study HF biomarkers: Bioinformatics, HF animal models and human clinical cohorts (Figure 2). Bioinformatics are useful to screen for potential novel biomarkers. HF animal models allow in depth molecular investigations and pharmacological studies. By investigating HF biomarkers in human HF cohorts, a translation towards the human situation can be made and these cohorts can also be used to determine the clinical relevance and applicability of a potential novel marker.
THESIS OUTLINE
An overview of the current literature on cardiac fibrosis, an important pathological process in cardiac remodeling, is provided in chapter 2. Whilst myocardial fibrosis is a key pathological process, till date no adequate blood plasma biomarker has been identified to mark for this process. In this review, we describe the molecular processes involved in fibrosis and its effect on cardiac tissue and function. We postulate the existence of a vicious cycle of fibrosis and cell death, which ultimately leads to HF. Finally, we discuss ways to target myocardial fibrosis and to eliminate the sustained fibrotic response in the heart.
Chapter 3 describes a study in which a potential novel fibrotic HF biomarker, human
epididymis protein 4 (HE4) is investigated in a cohort of patients with chronic heart failure. HE4 blood plasma levels in patients were compared to healthy controls. Next, we show the associations of blood plasma HE4 with several clinical and laboratory variables. Finally, we describe the predictive value of this novel marker for HF outcome.
In chapter 4, the current literature on novel HF biomarkers is reviewed. The usefulness of established HF biomarkers, including natriuretic peptides and troponins, is described. Furthermore, the impasse and problems within the field of HF biomarkers are explained, and a lack of cardiac specificity is suggested as the main reason why, despite elaborate research, most suggested novel biomarkers do not make it to the clinic.
To further investigate the cardiac specificity of novel HF biomarkers, in chapter 5 we describe an animal study in which the origin of several blood plasma HF biomarkers is investigated. In this mouse study several animal models are used, including a myocardial infarction model, a pressure overload model (transverse aortic constriction (TAC)) and an obesity/hypertension mouse model with characteristics of HFpEF. Investigated biomarkers are atrial natriuretic peptide (ANP), galectin-3 (Gal-3), growth differentiation factor-15 (GDF-15) and tissue inhibitor of metalloproteinase-1 (TIMP-1). The levels of these markers are investigated in all three models at three different levels: Cardiac gene expression, cardiac
Introduction
13
protein levels and blood plasma levels. Moreover, gene expression and protein levels are investigated in several non-cardiac organs and tissues. The associations between cardiac expression levels of these four markers and their circulating levels are described, also in relation to the cardiac phenotype observed in these models. Moreover, we investigated the effect of several co-morbidities on circulating levels of these markers.
In chapter 6, we further investigate the cardiac specificity of novel HF biomarkers. In a bioinformatics analysis, we determined the cardiac specific expression of the human genome with a focus on genes encoding secreted proteins. Next, using this dataset, we describe the lack of cardiac specificity of several suggested HF biomarkers. Finally, based on the bioinformatics analysis, one promising cardiac specific gene, namely Dickkopf-3 (DKK3) was selected for further analysis. DKK3 biomarker biology was investigated in three different HF mouse models, and its performance as a HF biomarker was investigated in a human HF patient cohort.
Whilst in chapter 6 DKK3 was investigated in a HF population, in chapter 7 DKK3 is investigated as a biomarker in the general population. We studied the associations of DKK3 with cardiovascular risk factors, and history of cardiovascular disease and kidney disease. Moreover, we investigated the predictive value of plasma DKK3 concentrations for new-onset cardiovascular disease and chronic kidney disease.
Whereas in the previous chapters HF biomarkers were studied for their diagnostic and prognostic qualities, in chapter 8 a study is described in which a plasma biomarker is pharmacologically targeted. We pharmacologically inhibited the inflammatory enzyme myeloperoxidase (MPO), which is a suggested HF biomarker, in a mouse model that incorporates inflammatory risk factors of HF, including obesity and hypertension. High fat diet (HFD) and angiotensin II (AngII) were administered to mice, resulting in a mouse model characterized by obesity, hypertension, non-alcoholic steatohepatitis (NASH) and HFpEF characteristics. In this model, we investigated the effects of the novel MPO inhibitor AZM198 on adipose tissue, liver tissue and cardiac tissue.
Finally, in chapter 9, the main findings of this thesis are discussed. Moreover, we reflect on future perspectives for HF biomarker research.
14
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