Establishment of new potential biomarkers for cardiometabolic diseases

Abstract

Cardiometabolic diseases is a growing health issue worldwide. New knowledge about the pathophysiological processes have been gained and several research discoveries have contributed to better management of the diseases. Even so, the prognosis, diagnosis, and treatment of cardiometabolic diseases needs to be further improved. For this, the discovery and implementation of additional biomarkers is extremely relevant. New advancing technologies have increased the opportunities to perform discovery studies and to identify new potential biomarkers. However, for several reasons, only few biomarkers survive the long journey from discovery into clinical implementation and there is a need to make this process more efficient. The objective of the thesis was to evaluate new potential biomarkers to improve the diagnosis and management of cardiometabolic diseases. An additional aim was to establish an efficient process for rapid transfer of new potential biomarkers, identified in discovery studies into evaluation in the routine care setting. Four studies in different patient cohorts, characterized by separate designs, and reflecting diverse phases in the implementation of biomarkers for cardiometabolic diseases are presented. Lipoprotein (a) [Lp(a)] is a modified low-density lipoprotein (LDL) particle and its concentration in plasma is mainly genetically determined. High levels of Lp(a) is associated with an increased risk for cardiovascular diseases (CVD). Despite the fact that it is not yet clear whether reduction of plasma Lp(a) levels translate into a reduced CVD risk, more knowledge about its role as a risk factor in different cohorts and diagnoses is needed to better understand how patients with high Lp(a) levels should be managed.

In Paper I we investigated the distribution of plasma Lp(a) levels and its association with CVD in a large cohort of patients who had their plasma levels of Lp(a) determined in routine care. Laboratory data from 23 398 patients was linked to data retrieved from National Board of Health and Welfare registers and National Quality registers. Lp(a) levels had a skewed distribution, increased with age, and was higher in females. Patients with Lp(a) levels in the 4th quartile had a 1.36-fold (95% Confidence Interval (CI) 1.14-1.61, p=0.001) increased risk for ischemic heart disease compared to patients belonging to the 1st quartile. The risk was independent of age, previous CVD, diabetes, and LDL-cholesterol levels. Hence, Lp(a) is an important risk factor for ischemic heart disease also in patients referred from hospitals, out-patient clinics, and general practitioners in the Region Stockholm.

Very little is known about the role of Lp(a) as a risk factor for CVD in patients with type 1 diabetes. Therefore, in Paper II we investigated the association of Lp(a) with cardiovascular complications and metabolic control in 1860 subjects with type 1 diabetes. Lp(a) levels had a skewed distribution, increased with age, and was not influenced by sex. Patients with poor metabolic control (HbA1c >52 mmol/mol) had higher Lp(a) levels compared to patients with good metabolic control. Patients with high Lp(a) levels (>120 nmol/L) had a 1.51-fold (95 % CI 1.01-2.28, p=0.048) increased risk for any macrovascular diseases, a 1.68-fold (95% CI 1.12-2.50, p=0.01) increased risk for albuminuria, and a 2.03-fold (95 % CI 1.02-4.01, p=0.043) increased risk for calcified aortic valve disease compared to patients with very low levels (<10 nmol/L). In summary, Lp(a) is a relevant risk factor also in patients with type 1 diabetes.

In Paper III we aimed to establish an efficient process for transfer of newly discovered potential biomarkers into evaluation in the routine care setting. The prototype was based on the evaluation of chemokine ligand 16 (CCL-16), previously identified as interesting biomarker for acute coronary syndrome (ACS) in a discovery project called Vinncardio, initiated by the Science for Life laboratory, Royal Institute of Technology (KTH), Stockholm, Sweden. Patients eligible for inclusion were identified when their plasma was analyzed for high sensitive Troponin T at the Karolinska University Laboratory, Stockholm, Sweden. The plasma samples were temporarily stored and meanwhile the patients received a letter of invitation to participate in the study. A positive response was retrieved from ~40 % and 1631 patients were included. No significant differences in CCL-16 were observed between patients with ACS and other diagnosis and CCL-16 do not appear to be a valid biomarker for ACS. Despite this negative result, we manage to establish a process for early evaluation of new potential biomarkers in routine care settings and to rapidly create a biobank and include patients referred to the hospital with an acute medical condition.

Clinical randomized trials have shown that addition of ezetimibe to simvastatin treatment further improve the reduction of CVD events, especially in patients with type 2 diabetes where elevation of remnant-cholesterol is characteristic. Remnant-cholesterol is a new and interesting biomarker and mendelian randomization studies have identified it as an independent risk factor for CVD, also promoting and sustaining low grade inflammation. In Paper IV we aimed to in detail study how the lipoprotein metabolism is affected by simvastatin and ezetimibe treatment, alone or in combination, to gain further understanding of the molecular effects of these two widely used lipid lowering drugs. Forty patients eligible for cholecystectomy were randomized to four-week treatment before surgery to placebo, simvastatin (80 mg daily), ezetimibe (10 mg daily), or to combination of both. The combination of simvastatin and ezetimibe resulted in further reduction of cholesterol and cholesteryl esters in remnant- and LDL-particles, as well as reduction of apolipoprotein B (apoB) containing particles, and reduced apoB-containing lipoprotein affinity for arterial proteoglycans compared to simvastatin. These additional positive effects on atherogenic lipoproteins and especially remnant-particles can possibly explain the further reduction of CVD events previously observed, and the combination of ezetimibe and simvastatin seems to be the optimal treatment in conditions with elevated remnant-cholesterol.

In conclusion, these four studies have provided further knowledge about the different biomarkers investigated. Also, they can contribute to an improved management of patients with cardiometabolic diseases and indicate the way to a rapid recruitment of patients in clinical studies. Hence, this thesis adds to a deeper understanding of the complexity in the process to validate and implement new biomarkers.