Study of effect modifiers of genetically predicted CETP reduction.

Genetic variants in drug targets can be used to predict the long-term, on-target effect of drugs. Here, we extend this principle to assess how sex and body mass index may modify the effect of genetically predicted lower CETP levels on biomarkers and cardiovascular outcomes. We found sex and body mass index (BMI) to be modifiers of the association between genetically predicted lower CETP and lipid biomarkers in UK Biobank participants. Female sex and lower BMI were associated with higher high-density lipoprotein cholesterol and lower low-density lipoprotein cholesterol for the same genetically predicted reduction in CETP concentration. We found that sex also modulated the effect of genetically lower CETP on cholesterol efflux capacity in samples from the Montreal Heart Institute Biobank. However, these modifying effects did not extend to sex differences in cardiovascular outcomes in our data. Our results provide insight into the clinical effects of CETP inhibitors in the presence of effect modification based on genetic data. The approach can support precision medicine applications and help assess the external validity of clinical trials.

Adenylate cyclase type 9 antagonizes cAMP accumulation and regulates endothelial signalling involved in atheroprotection.

AIMS: The adenylate cyclase type 9 (ADCY9) gene appears to determine atherosclerotic outcomes in patients treated with dalcetrapib. In mice, we recently demonstrated that Adcy9 inactivation potentiates endothelial function and inhibits atherogenesis. The objective of this study was to characterize the contribution of ADCY9 to the regulation of endothelial signalling pathways involved in atherosclerosis. METHODS AND RESULTS: We show that ADCY9 is expressed in the endothelium of mouse aorta and femoral arteries. We demonstrate that ADCY9 inactivation in cultured endothelial cells paradoxically increases cAMP accumulation in response to the adenylate cyclase activators forskolin and vasoactive intestinal peptide (VIP). Reciprocally, ADCY9 overexpression decreases cAMP production. Using mouse femoral artery arteriography, we show that Adcy9 inactivation potentiates VIP-induced endothelial-dependent vasodilation. Moreover, Adcy9 inactivation reduces mouse atheroma endothelial permeability in different vascular beds. ADCY9 overexpression reduces forskolin-induced phosphorylation of Ser157-vasodilator-stimulated phosphoprotein (VASP) and worsens thrombin-induced fall of RAP1 activity, both leading to increased endothelial permeability. ADCY9 inactivation in thrombin-stimulated human coronary artery endothelial cells results in cAMP accumulation, increases p-Ser157-VASP, and inhibits endothelial permeability. MLC2 phosphorylation and actin stress fibre increases in response to thrombin were reduced by ADCY9 inactivation, suggesting actin cytoskeleton regulation. Finally, using the Miles assay, we demonstrate that Adcy9 regulates thrombin-induced endothelial permeability in vivo in normal and atherosclerotic animals. CONCLUSION: Adcy9 is expressed in endothelial cells and regulates local cAMP and endothelial functions including permeability relevant to atherogenesis.

Dalcetrapib and anacetrapib increase apolipoprotein E-containing HDL in rabbits and humans.

The large HDL particles generated by administration of cholesteryl ester transfer protein inhibitors (CETPi) remain poorly characterized, despite their potential importance in the routing of cholesterol to the liver for excretion, which is the last step of the reverse cholesterol transport. Thus, the effects of the CETPi dalcetrapib and anacetrapib on HDL particle composition were studied in rabbits and humans. The association of rabbit HDL to the LDL receptor (LDLr) in vitro was also evaluated. New Zealand White rabbits receiving atorvastatin were treated with dalcetrapib or anacetrapib. A subset of patients from the dal-PLAQUE-2 study treated with dalcetrapib or placebo were also studied. In rabbits, dalcetrapib and anacetrapib increased HDL-C by more than 58% (P < 0.01) and in turn raised large apo E-containing HDL by 66% (P < 0.001) and 59% (P < 0.01), respectively. Additionally, HDL from CETPi-treated rabbits competed with human LDL for binding to the LDLr on HepG2 cells more than control HDL (P < 0.01). In humans, dalcetrapib increased concentrations of large HDL particles (+69%, P < 0.001) and apo B-depleted plasma apo E (+24%, P < 0.001), leading to the formation of apo E-containing HDL (+47%, P < 0.001) devoid of apo A-I. Overall, in rabbits and humans, CETPi increased large apo E-containing HDL particle concentration, which can interact with hepatic LDLr. The catabolism of these particles may depend on an adequate level of LDLr to contribute to reverse cholesterol transport.

Low-dose colchicine and high-sensitivity C-reactive protein after myocardial infarction: A combined analysis using individual patient data from the COLCOT and LoDoCo-MI studies.

BACKGROUND: Low-dose colchicine is effective in reducing the risks of recurrent cardiovascular events following an acute myocardial infarction (MI). However, the influence of colchicine on inflammation remains inconclusive. In the current study, we conducted a combined analysis using individual patient data from the COLCOT and LoDoCo-MI trials to assess the effect of low-dose colchicine on high-sensitivity C reactive protein (hs-CRP) in patients with acute MI. METHODS: We performed a combined analysis of individual patient data from two clinical trials (COLCOT, LoDoCo-MI). Paired pre-treatment and post-treatment hs-CRP (mg/L) were available in 222 patients for LoDoCo-MI and 207 patients for COLCOT (npooled = 429). We evaluated the effect of colchicine vs. placebo on post-treatment hs-CRP coded continuously and ≤ 1.0 mg/L in adjusted mixed-model multi-level regression analyses. RESULTS: Colchicine was not significantly associated with post-treatment hs-CRP when it was considered as a continuous variable (beta: -0.41, P = 0.429). However, the intervention was significantly associated with increased odds of achieving post-treatment hs-CRP values ≤1.0 mg/L compared to placebo (odds ratio: 1.64, 95% confidence interval: 1.07 to 2.51, P = 0.024). CONCLUSIONS: Reduction of inflammation may be a key component in the clinical efficacy of low-dose colchicine with respect to decreased risk of recurrent cardiovascular events following MI. Systematic sampling of hs-CRP before and after treatment with colchicine may be relevant.

Lipoprotein (a): When to Measure and How to Treat?

PURPOSE OF REVIEW: The purpose of this article is to review current evidence for lipoprotein (a) (Lp(a)) as a risk factor for multiple cardiovascular (CV) disease phenotypes, provide a rationale for Lp(a) lowering to reduce CV risk, identify therapies that lower Lp(a) levels that are available clinically and under investigation, and discuss future directions. RECENT FINDINGS: Mendelian randomization and epidemiological studies have shown that elevated Lp(a) is an independent and causal risk factor for atherosclerosis and major CV events. Lp(a) is also associated with non-atherosclerotic endpoints such as venous thromboembolism and calcific aortic valve disease. It contributes to residual CV risk in patients receiving standard-of-care LDL-lowering therapy. Plasma Lp(a) levels present a skewed distribution towards higher values and vary widely between individuals and according to ethnic background due to genetic variants in the LPA gene, but remain relatively constant throughout a person's life. Thus, elevated Lp(a) (≥50 mg/dL) is a prevalent condition affecting >20% of the population but is still underdiagnosed. Treatment guidelines have begun to advocate measurement of Lp(a) to identify patients with very high levels that have a family history of premature CVD or elevated Lp(a). Lipoprotein apheresis (LA) efficiently lowers Lp(a) and was recently associated with a reduction of incident CV events. Statins have neutral or detrimental effects on Lp(a), while PCSK9 inhibitors significantly reduce its level by up to 30%. Specific lowering of Lp(a) with antisense oligonucleotides (ASO) shows good safety and strong efficacy with up to 90% reductions. The ongoing CV outcomes study Lp(a)HORIZON will provide a first answer as to whether selective Lp(a) lowering with ASO reduces the risk of major CV events. Given the recently established association between Lp(a) level and CV risk, guidelines now recommend Lp(a) measurement in specific clinical conditions. Accordingly, Lp(a) is a current target for drug development to reduce CV risk in patients with elevated levels, and lowering Lp(a) with ASO represents a promising avenue.

Role of Adenylate Cyclase 9 in the Pharmacogenomic Response to Dalcetrapib: Clinical Paradigm and Molecular Mechanisms in Precision Cardiovascular Medicine.

Following the neutral results of the dal-OUTCOMES trial, a genome-wide study identified the rs1967309 variant in the adenylate cyclase type 9 (ADCY9) gene on chromosome 16 as being associated with the risk of future cardiovascular events only in subjects taking dalcetrapib, a CETP (cholesterol ester transfer protein) modulator. Homozygotes for the minor A allele (AA) were protected from recurrent cardiovascular events when treated with dalcetrapib, while homozygotes for the major G allele (GG) had increased risk. Here, we present the current state of knowledge regarding the impact of rs1967309 in ADCY9 on clinical observations and biomarkers in dalcetrapib trials and the effects of mouse ADCY9 gene inactivation on cardiovascular physiology. Finally, we present our current model of the interaction between dalcetrapib and ADCY9 gene variants in the arterial wall macrophage, based on the intracellular role of CETP in the transfer of complex lipids from endoplasmic reticulum membranes to lipid droplets. Briefly, the concept is that dalcetrapib would inhibit CETP-mediated transfer of cholesteryl esters, resulting in a progressive inhibition of cholesteryl ester synthesis and free cholesterol accumulation in the endoplasmic reticulum. Reduced ADCY9 activity, by paradoxically leading to higher cyclic AMP levels and in turn increased cellular cholesterol efflux, could impart cardiovascular protection in rs1967309 AA patients. The ongoing dal-GenE trial recruited 6145 patients with the protective AA genotype and will provide a definitive answer to whether dalcetrapib will be protective in this population.

Pharmacogenomics of the Efficacy and Safety of Colchicine in COLCOT.

BACKGROUND: The randomized, placebo-controlled COLCOT (Colchicine Cardiovascular Outcomes Trial) has shown the benefits of colchicine 0.5 mg daily to lower the rate of ischemic cardiovascular events in patients with a recent myocardial infarction. Here, we conducted a post hoc pharmacogenomic study of COLCOT with the aim to identify genetic predictors of the efficacy and safety of treatment with colchicine. METHODS: There were 1522 participants of European ancestry from the COLCOT trial available for the pharmacogenomic study of COLCOT trial. The pharmacogenomic study's primary cardiovascular end point was defined as for the main trial, as time to first occurrence of cardiovascular death, resuscitated cardiac arrest, myocardial infarction, stroke, or urgent hospitalization for angina requiring coronary revascularization. The safety end point was time to the first report of gastrointestinal events. Patients' DNA was genotyped using the Illumina Global Screening array followed by imputation. We performed a genome-wide association study in colchicine-treated patients. RESULTS: None of the genetic variants passed the genome-wide association study significance threshold for the primary cardiovascular end point conducted in 702 patients in the colchicine arm who were compliant to medication. The genome-wide association study for gastrointestinal events was conducted in all 767 patients in the colchicine arm and found 2 significant association signals, one with lead variant rs6916345 (hazard ratio, 1.89 [95% CI, 1.52-2.35], P=7.41×10-9) in a locus which colocalizes with Crohn disease, and one with lead variant rs74795203 (hazard ratio, 2.51 [95% CI, 1.82-3.47]; P=2.70×10-8), an intronic variant in gene SEPHS1. The interaction terms between the genetic variants and treatment with colchicine versus placebo were significant. CONCLUSIONS: We found 2 genomic regions associated with gastrointestinal events in patients treated with colchicine. Those findings will benefit from replication to confirm that some patients may have genetic predispositions to lower tolerability of treatment with colchicine.

ApoA-I mimetic does not improve left ventricular diastolic dysfunction in rabbits without aortic valve stenosis.

BACKGROUND: We previously demonstrated that high-density lipoprotein (HDL) infusions may improve left ventricular diastolic dysfunction (LVDD) in an aortic valve stenosis (AVS) model. Whether the benefit was direct or mediated by the observed reduction in AVS severity is not clear. Here, we aimed to test the direct effect of an ApoA-I mimetic on LVDD in the absence of AVS. METHODS: Rabbits were exposed to three different protocols to develop LVDD. First, rabbits were exposed to 0.5% cholesterol-rich diet for an average of 17 weeks. Second, rabbits were subjected to surgical ascending aortic constriction (AAC), to mimic the effect of fixed reduced aortic valve area, and studied after 10 weeks. The third model combined both cholesterol-enriched diet (for 12 weeks) and surgical AAC. The control group consisted of age-matched rabbits under normal diet. After development of LVDD, rabbits were randomized to receive infusions of saline or apoA-I mimetic (25 mg/kg) 3 times per week for 4 weeks. Detailed cardiac structure and function measurements were assessed at baseline and weekly during treatment period. Histological and molecular analyses were performed on LV samples. RESULTS: In the three models, echocardiographic results showed development of LVDD over time, with preserved LV systolic and aortic valve functions versus controls. ApoA-I mimetic infusions did not significantly improve echocardiographic parameters nor molecular markers of cardiac inflammation, oxidative stress and fibrosis. CONCLUSION: ApoA-I mimetic therapy did not directly improve LVDD. These results indicate that previously observed changes of LVDD were caused by AVS improvement induced by this treatment.

Colchicine reduces lung injury in experimental acute respiratory distress syndrome.

The acute respiratory distress syndrome (ARDS) is characterized by intense dysregulated inflammation leading to acute lung injury (ALI) and respiratory failure. There are no effective pharmacologic therapies for ARDS. Colchicine is a low-cost, widely available drug, effective in the treatment of inflammatory conditions. We studied the effects of colchicine pre-treatment on oleic acid-induced ARDS in rats. Rats were treated with colchicine (1 mg/kg) or placebo for three days prior to intravenous oleic acid-induced ALI (150 mg/kg). Four hours later they were studied and compared to a sham group. Colchicine reduced the area of histological lung injury by 61%, reduced lung edema, and markedly improved oxygenation by increasing PaO2/FiO2 from 66 ± 13 mmHg (mean ± SEM) to 246 ± 45 mmHg compared to 380 ± 18 mmHg in sham animals. Colchicine also reduced PaCO2 and respiratory acidosis. Lung neutrophil recruitment, assessed by myeloperoxidase immunostaining, was greatly increased after injury from 1.16 ± 0.19% to 8.86 ± 0.66% and significantly reduced by colchicine to 5.95 ± 1.13%. Increased lung NETosis was also reduced by therapy. Circulating leukocytosis after ALI was not reduced by colchicine therapy, but neutrophils reactivity and CD4 and CD8 cell surface expression on lymphocyte populations were restored. Colchicine reduces ALI and respiratory failure in experimental ARDS in relation with reduced lung neutrophil recruitment and reduced circulating leukocyte activation. This study supports the clinical development of colchicine for the prevention of ARDS in conditions causing ALI.

Efficacy and Safety of Low-Dose Colchicine after Myocardial Infarction.

BACKGROUND: Experimental and clinical evidence supports the role of inflammation in atherosclerosis and its complications. Colchicine is an orally administered, potent antiinflammatory medication that is indicated for the treatment of gout and pericarditis. METHODS: We performed a randomized, double-blind trial involving patients recruited within 30 days after a myocardial infarction. The patients were randomly assigned to receive either low-dose colchicine (0.5 mg once daily) or placebo. The primary efficacy end point was a composite of death from cardiovascular causes, resuscitated cardiac arrest, myocardial infarction, stroke, or urgent hospitalization for angina leading to coronary revascularization. The components of the primary end point and safety were also assessed. RESULTS: A total of 4745 patients were enrolled; 2366 patients were assigned to the colchicine group, and 2379 to the placebo group. Patients were followed for a median of 22.6 months. The primary end point occurred in 5.5% of the patients in the colchicine group, as compared with 7.1% of those in the placebo group (hazard ratio, 0.77; 95% confidence interval [CI], 0.61 to 0.96; P = 0.02). The hazard ratios were 0.84 (95% CI, 0.46 to 1.52) for death from cardiovascular causes, 0.83 (95% CI, 0.25 to 2.73) for resuscitated cardiac arrest, 0.91 (95% CI, 0.68 to 1.21) for myocardial infarction, 0.26 (95% CI, 0.10 to 0.70) for stroke, and 0.50 (95% CI, 0.31 to 0.81) for urgent hospitalization for angina leading to coronary revascularization. Diarrhea was reported in 9.7% of the patients in the colchicine group and in 8.9% of those in the placebo group (P = 0.35). Pneumonia was reported as a serious adverse event in 0.9% of the patients in the colchicine group and in 0.4% of those in the placebo group (P = 0.03). CONCLUSIONS: Among patients with a recent myocardial infarction, colchicine at a dose of 0.5 mg daily led to a significantly lower risk of ischemic cardiovascular events than placebo. (Funded by the Government of Quebec and others; COLCOT ClinicalTrials.gov number, NCT02551094.).

Investigational drugs in development for hypertriglyceridemia: a coming-of-age story.

Introduction: Elevated triglyceride (TG) level is a prevalent condition in the general population and in patients with cardiovascular (CV) risk even under statin therapy. Severe hypertriglyceridemia (HTG) puts patients at risk for acute pancreatitis. Several TG-lowering drugs failed in clinical trials, but subgroup analyses suggest that high-risk patients, such as those with atherogenic dyslipidemia or diabetes, benefit from TG lowering.Areas covered: We review advances for TG-lowering drugs in clinical development. These include selective PPARα modulators, omega-3 fatty acid formulations that have been approved for severe HTG, and inhibitors of apolipoprotein C-III, angiopoietin-like-3 or microsomal transfer protein. Lessons learned from the success of the phase 3 trial REDUCE-IT with high-dose icosapent ethyl are also reviewed.Expert opinion: We believe that TG-lowering therapies are coming of age as they will allow to treat patients with high CV risk and moderate HTG, including T2D subjects, as well as patients with severe HTG or even homozygous familial hypercholesterolemia, all of which being 'optimally' treated with a statin. More studies on the impact of therapy on quality of life in patients with severe HTG should be conducted with the help of patient registries.

From HDL-cholesterol to HDL-function: cholesterol efflux capacity determinants.

PURPOSE OF REVIEW: The validity of HDL-cholesterol (HDL-C) elevation as a therapeutic target has been questioned, in comparison to enhancing HDL functionality. Cholesterol efflux capacity (CEC) is an in-vitro assay that measures the ability of an individual's HDL to promote cholesterol efflux from cholesterol donor cells such as macrophages. CEC of HDL is a predictor of cardiovascular risk independent of HDL-C levels. However, molecular determinants of CEC and the effects of diseases and therapeutic interventions on CEC have not been completely defined. RECENT FINDINGS: We review here recent findings on elevated HDL-C and disease risk, as well as determinants of CEC, from genetics and proteomics to pathophysiology and therapeutic interventions that contribute to our understanding of CEC as a biomarker of HDL functionality. SUMMARY: Elevated HDL-C levels are not always protective against cardiovascular disease and mortality. CEC is a heritable trait, and genetic polymorphisms in genes involved in HDL and triglycerides metabolism are associated with CEC. Multiple HDL proteins correlate positively with CEC levels and inversely with noncalcified plaque burden. Differences in CEC assays that make comparisons between studies difficult are also emphasized. CEC should be measured in clinical trials of lipid-modifying and anti-inflammatory therapies to determine whether increases are cardioprotective.

Variants at the APOE /C1/C2/C4 Locus Modulate Cholesterol Efflux Capacity Independently of High-Density Lipoprotein Cholesterol.

Background Macrophage cholesterol efflux to high-density lipoproteins ( HDLs ) is the first step of reverse cholesterol transport. The cholesterol efflux capacity ( CEC ) of HDL particles is a protective risk factor for coronary artery disease independent of HDL cholesterol levels. Using a genome-wide association study approach, we aimed to identify pathways that regulate CEC in humans. Methods and Results We measured CEC in 5293 French Canadians. We tested the genetic association between 4 CEC measures and genotypes at >9 million common autosomal DNA sequence variants. These analyses yielded 10 genome-wide significant signals ( P<6.25×10-9) representing 7 loci. Five of these loci harbor genes with important roles in lipid biology ( CETP , LIPC , LPL , APOA 1/C3/A4/A5, and APOE /C1/C2/C4). Except for the APOE /C1/C2/C4 variant ( rs141622900, P nonadjusted=1.0×10-11; P adjusted=8.8×10-9), the association signals disappear when correcting for HDL cholesterol and triglyceride levels. The additional 2 significant signals were near the PPP 1 CB / PLB 1 and RBFOX 3/ ENPP 7 genes. In secondary analyses, we considered candidate functional variants for 58 genes implicated in HDL biology, as well as 239 variants associated with blood lipid levels and/or coronary artery disease risk by genome-wide association study . These analyses identified 27 significant CEC associations, implicating 5 additional loci ( GCKR , LIPG , PLTP , PPARA , and TRIB 1). Conclusions Our genome-wide association study identified common genetic variation at the APOE /C1/C2/C4 locus as a major determinant of CEC that acts largely independently of HDL cholesterol. We predict that HDL -based therapies aiming at increasing CEC will be modulated by changes in the expression of apolipoproteins in this gene cluster.

Comprehensive and Reproducible Untargeted Lipidomic Workflow Using LC-QTOF Validated for Human Plasma Analysis.

The goal of this work was to develop a label-free, comprehensive, and reproducible high-resolution liquid chromatography-mass spectrometry (LC-MS)-based untargeted lipidomic workflow using a single instrument, which could be applied to biomarker discovery in both basic and clinical studies. For this, we have (i) optimized lipid extraction and elution to enhance coverage of polar and nonpolar lipids as well as resolution of their isomers, (ii) ensured MS signal reproducibility and linearity, and (iii) developed a bioinformatic pipeline to correct remaining biases. Workflow validation is reported for 48 replicates of a single human plasma sample: 1124 reproducible LC-MS signals were extracted (median signal intensity RSD = 10%), 50% of which are redundant due to adducts, dimers, in-source fragmentation, contaminations, or positive and negative ion duplicates. From the resulting 578 unique compounds, 428 lipids were identified by MS/MS, including acyl chain composition, of which 394 had RSD < 30% inside their linear intensity range, thereby enabling robust semiquantitation. MS signal intensity spanned 4 orders of magnitude, covering 16 lipid subclasses. Finally, the power of our workflow is illustrated by a proof-of-concept study in which 100 samples from healthy human subjects were analyzed and the data set was investigated using three different statistical testing strategies in order to compare their capacity in identifying the impact of sex and age on circulating lipids.

Elevated level of lecithin:cholesterol acyltransferase (LCAT) is associated with reduced coronary atheroma burden.

BACKGROUND AND AIMS: Lecithin:cholesterol acyltransferase (LCAT), a key enzyme in high-density lipoprotein (HDL) metabolism and reverse cholesterol transport (RCT), has been associated with atheroprotection. However, its relation to plaque characteristics has not been confirmed to date. We aimed to determine the relationship between plasma LCAT mass concentration and plaque burden in a multi-center imaging study. METHODS: Two hundred sixty-seven patients with angiographically proven coronary artery disease (CAD) underwent intravascular ultrasonography (IVUS) imaging. Ninety-six patients without CAD served as controls for biochemistry assessments. RESULTS: Plasma LCAT mass concentration was higher in CAD patients as compared to controls (8.94 ±â€¯2.51 µg/mL vs. 7.89 ±â€¯2.99 µg/mL, p = 0.003), while cholesterol esterification rate (CER) was downregulated (253.6 ±â€¯83.9 µM/2 h vs. 315.3 ±â€¯115.0 µM/2 h, p<0.0001). Both parameters correlated inversely with total atheroma volume (r = -0.14, p = 0.027 and r = -0.14, p = 0.024, respectively), while only LCAT mass was found to be a significant predictor of atheroma volume (ß-coefficient -0.18, p = 0.0047) when tested in a stepwise linear regression model against known CAD risk factors as predictor variables. Accordingly, patients with LCAT mass in the highest quartile had significantly less atheroma burden than those in the lower quartiles (39.7 ±â€¯10.7% vs. 45.4 ±â€¯10.4%, p = 0.0014 for highest vs. lowest quartile of LCAT mass). CONCLUSIONS: Plasma LCAT mass concentration is upregulated in CAD patients and inversely related to plaque volume, suggesting atheroprotective effects. LCAT mass concentration outperformed LCAT activity in risk prediction models for atheroma burden, suggesting that LCAT mass is a key variable in atheroprotection. Further studies assessing LCAT as a therapeutic target in cardiovascular disease are warranted.

Lipids, Apolipoproteins, and Inflammatory Biomarkers of Cardiovascular Risk: What Have We Learned?

Cardiovascular diseases (CVD) are the first cause of death in the world. CVD risk is influenced by multiple factors, some nonmodifiable such as age, sex, and genetic background, and others modifiable. Great progress has been made over the last decades in the identification of biomarkers of incident or recurrent CV risk and surrogate endpoints of CV outcomes. We present the current state of knowledge for CV biomarkers in plasma including lipids, apolipoproteins, inflammation-related, and emerging omics-based biomarkers. Clinically validated surrogate endpoints for CV outcomes include plasma low-density lipoprotein-cholesterol reduction, and plasma triglyceride reduction is a likely relevant surrogate endpoint. High-density lipoprotein-cholesterol is not a validated surrogate endpoint, but is a useful biomarker of CV risk. CV risk biomarkers of interest include apolipoprotein B and non-HDL-cholesterol, lipoprotein (a), C-reactive protein, and recently, genetic and protein-based risk scores and gut microbiota-derived trimethylamine oxide levels.

Structure-Activity Relationship Study of Cyclic Pentapeptide Ligands for Atypical Chemokine Receptor 3 (ACKR3).

The atypical chemokine receptor 3 (ACKR3)/CXC chemokine receptor 7 (CXCR7) recognizes stromal cell-derived factor 1 (SDF-1)/CXCL12 and is involved in a number of physiological and pathological processes. Here, we investigated the SAR of the component amino acids in an ACKR3-selective ligand, FC313 [ cyclo(-d-Tyr-l-Arg-l-MeArg-l-Nal(2)-l-Pro-)], for the development of highly active ACKR3 ligands. Notably, modification at the l-Pro position with a bulky hydrophobic side chain led to improved bioactivity toward ACKR3.

ADCY9 (Adenylate Cyclase Type 9) Inactivation Protects From Atherosclerosis Only in the Absence of CETP (Cholesteryl Ester Transfer Protein).

BACKGROUND: Pharmacogenomic studies have shown that ADCY9 genotype determines the effects of the CETP (cholesteryl ester transfer protein) inhibitor dalcetrapib on cardiovascular events and atherosclerosis imaging. The underlying mechanisms responsible for the interactions between ADCY9 and CETP activity have not yet been determined. METHODS: Adcy9-inactivated ( Adcy9Gt/Gt) and wild-type (WT) mice, that were or not transgenic for the CETP gene (CETPtg Adcy9Gt/Gt and CETPtg Adcy9WT), were submitted to an atherogenic protocol (injection of an AAV8 [adeno-associated virus serotype 8] expressing a PCSK9 [proprotein convertase subtilisin/kexin type 9] gain-of-function variant and 0.75% cholesterol diet for 16 weeks). Atherosclerosis, vasorelaxation, telemetry, and adipose tissue magnetic resonance imaging were evaluated. RESULTS: Adcy9Gt/Gt mice had a 65% reduction in aortic atherosclerosis compared to WT ( P<0.01). CD68 (cluster of differentiation 68)-positive macrophage accumulation and proliferation in plaques were reduced in Adcy9Gt/Gt mice compared to WT animals ( P<0.05 for both). Femoral artery endothelial-dependent vasorelaxation was improved in Adcy9Gt/Gt mice (versus WT, P<0.01). Selective pharmacological blockade showed that the nitric oxide, cyclooxygenase, and endothelial-dependent hyperpolarization pathways were all responsible for the improvement of vasodilatation in Adcy9Gt/Gt ( P<0.01 for all). Aortic endothelium from Adcy9Gt/Gt mice allowed significantly less adhesion of splenocytes compared to WT ( P<0.05). Adcy9Gt/Gt mice gained more weight than WT with the atherogenic diet; this was associated with an increase in whole body adipose tissue volume ( P<0.01 for both). Feed efficiency was increased in Adcy9Gt/Gt compared to WT mice ( P<0.01), which was accompanied by prolonged cardiac RR interval ( P<0.05) and improved nocturnal heart rate variability ( P=0.0572). Adcy9 inactivation-induced effects on atherosclerosis, endothelial function, weight gain, adipose tissue volume, and feed efficiency were lost in CETPtg Adcy9Gt/Gt mice ( P>0.05 versus CETPtg Adcy9WT). CONCLUSIONS: Adcy9 inactivation protects against atherosclerosis, but only in the absence of CETP activity. This atheroprotection may be explained by decreased macrophage accumulation and proliferation in the arterial wall, and improved endothelial function and autonomic tone.

Different contributions of chemokine N-terminal features attest to a different ligand binding mode and a bias towards activation of ACKR3/CXCR7 compared with CXCR4 and CXCR3.

BACKGROUND AND PURPOSE: Chemokines and their receptors form an intricate interaction and signalling network that plays critical roles in various physiological and pathological cellular processes. The high promiscuity and apparent redundancy of this network makes probing individual chemokine/receptor interactions and functional effects, as well as targeting individual receptor axes for therapeutic applications, challenging. Despite poor sequence identity, the N-terminal regions of chemokines, which play a key role in their activity and selectivity, contain several conserved features. Thus far little is known regarding the molecular basis of their interactions with typical and atypical chemokine receptors or the conservation of their contributions across chemokine-receptor pairs. EXPERIMENTAL APPROACH: We used a broad panel of chemokine variants and modified peptides derived from the N-terminal region of chemokines CXCL12, CXCL11 and vCCL2, to compare the contributions of various features to binding and activation of their shared receptors, the two typical, canonical G protein-signalling receptors, CXCR4 and CXCR3, as well as the atypical scavenger receptor CXCR7/ACKR3, which shows exclusively arrestin-dependent activity. KEY RESULTS: We provide molecular insights into the plasticity of the ligand-binding pockets of these receptors, their chemokine binding modes and their activation mechanisms. Although the chemokine N-terminal region is a critical determinant, neither the most proximal residues nor the N-loop are essential for binding and activation of ACKR3, as distinct from binding and activation of CXCR4 and CXCR3. CONCLUSION AND IMPLICATIONS: These results suggest a different interaction mechanism between this atypical receptor and its ligands and illustrate its strong propensity to activation.