Preclinical, randomized phase 1, and compassionate use evaluation of REGN4461, a leptin receptor agonist antibody for leptin deficiency.

Deficiency in the adipose-derived hormone leptin or leptin receptor signaling causes class 3 obesity in individuals with genetic loss-of-function mutations in leptin or its receptor LEPR and metabolic and liver disease in individuals with hypoleptinemia secondary to lipoatrophy such as in individuals with generalized lipodystrophy. Therapies that restore leptin-LEPR signaling may resolve these metabolic sequelae. We developed a fully human monoclonal antibody (mAb), REGN4461 (mibavademab), that activates the human LEPR in the absence or presence of leptin. In obese leptin knockout mice, REGN4461 normalized body weight, food intake, blood glucose, and insulin sensitivity. In a mouse model of generalized lipodystrophy, REGN4461 alleviated hyperphagia, hyperglycemia, insulin resistance, dyslipidemia, and hepatic steatosis. In a phase 1, randomized, double-blind, placebo-controlled two-part study, REGN4461 was well tolerated with an acceptable safety profile. Treatment of individuals with overweight or obesity with REGN4461 decreased body weight over 12 weeks in those with low circulating leptin concentrations (<8 ng/ml) but had no effect on body weight in individuals with higher baseline leptin. Furthermore, compassionate-use treatment of a single patient with atypical partial lipodystrophy and a history of undetectable leptin concentrations associated with neutralizing antibodies to metreleptin was associated with noteable improvements in circulating triglycerides and hepatic steatosis. Collectively, these translational data unveil an agonist LEPR mAb that may provide clinical benefit in disorders associated with relatively low leptin concentrations.

Symptoms and Dietary Impact in Hypertriglyceridemia-Associated Pancreatitis: Development and Content Validity of Two New Measures.

BACKGROUND: Severe hypertriglyceridemia (sHTG) is a rare condition, complicated by episodes of acute pancreatitis (AP), which can cause pain and/or life-threatening multi-organ dysfunction. Currently, there are no disease-specific patient-reported outcome (PRO) measures evaluating symptoms or dietary impact for this condition. OBJECTIVE: The objective of this study was to explore patient-reported symptoms and impacts of sHTG and AP and develop new measures to capture the symptoms and dietary impacts of this condition using patient language. METHODS: In-depth, semi-structured concept elicitation interviews were conducted with 12 US-based participants to explore their experience and identify key symptoms and impact on dietary behavior, both during and between episodes of AP. Participants had a range of AP severity with a previous triglyceride reading > 1000 mg/dL, and at least one attack of AP within the last 12 months. Transcripts were coded using thematic analysis. RESULTS: Qualitative data analysis revealed the substantial burden of AP associated with sHTG. Participants reported experiencing symptoms, especially abdominal pain, both during and between attacks of AP, and discussed considerable diet changes to prevent or minimize future attacks. A conceptual model was refined, based on patient input, and reviewed by clinical experts to determine key concepts for inclusion within two PRO measures, one evaluating symptoms and another evaluating impact on dietary behavior. Items were drafted using patient-derived language. A 19-item symptoms measure [Hypertriglyceridemia and Acute Pancreatitis Symptom Scale (HAP-SS)] and a 6-item dietary impact measure (Hypertriglyceridemia and Acute Pancreatitis Dietary Behavior (HAP-DB) measure) were developed, both with a 24-h recall period. CONCLUSIONS: The qualitative analysis confirmed the substantial burden of AP associated with sHTG. This research resulted in development of two disease-specific PRO measures for use during and between attacks of AP. These measures are being utilized in a clinical trial, which will confirm content, structure, and psychometric properties.

Inhibition of Angiopoietin-Like Protein 3 With a Monoclonal Antibody Reduces Triglycerides in Hypertriglyceridemia.

BACKGROUND: Hypertriglyceridemia is associated with increased cardiovascular risk and may be caused by impaired lipoprotein clearance. Angiopoietin-like protein 3 (ANGPTL3) inhibits lipoprotein lipase activity, increasing triglycerides and other lipids. Evinacumab, an ANGPTL3 inhibitor, reduced triglycerides in healthy human volunteers and in homozygous familial hypercholesterolemic individuals. Results from 2 Phase 1 studies in hypertriglyceridemic subjects are reported here. METHODS: Subjects with triglycerides >150 but ≤450 mg/dL and low-density lipoprotein cholesterol ≥100 mg/dL (n=83 for single ascending dose study [SAD]; n=56 for multiple ascending dose study [MAD]) were randomized 3:1 to evinacumab:placebo. SAD subjects received evinacumab subcutaneously at 75/150/250 mg, or intravenously at 5/10/20 mg/kg, monitored up to day 126. MAD subjects received evinacumab subcutaneously at 150/300/450 mg once weekly, 300/450 mg every 2 weeks, or intravenously at 20 mg/kg once every 4 weeks up to day 56 with 6 months of follow-up. The primary outcomes were incidence and severity of treatment-emergent adverse events. Efficacy analyses included changes in triglycerides and other lipids over time. RESULTS: In the SAD, 32 (51.6%) versus 9 (42.9%) subjects on evinacumab versus placebo reported treatment-emergent adverse events. In the MAD, 21 (67.7%) versus 9 (75.0%) subjects on subcutaneously evinacumab versus placebo and 6 (85.7%) versus 1 (50.0%) on intravenously evinacumab versus placebo reported treatment-emergent adverse events. No serious treatment-emergent adverse events or events leading to death or treatment discontinuation were reported. Elevations in alanine aminotransferase (7 [11.3%] SAD), aspartate aminotransferase (4 [6.5%] SAD), and creatinine phosphokinase (2 [3.2%) SAD, 1 [14.3%] MAD) were observed with evinacumab (none in the placebo groups), which were single elevations and were not dose-related. Dose-dependent reductions in triglycerides were observed in both studies, with maximum reduction of 76.9% at day 3 with 10 mg/kg intravenously (P<0.0001) in the SAD and of 83.1% at day 2 with 20 mg/kg intravenously once every 4 weeks (P=0.0003) in the MAD. Significant reductions in other lipids were observed with most evinacumab doses versus placebo. CONCLUSION: Evinacumab was well-tolerated in 2 Phase 1 studies. Lipid changes in hypertriglyceridemic subjects were similar to those observed with ANGPTL3 loss-of-function mutations. Because the latter is associated with reduced cardiovascular risk, ANGPTL3 inhibition may improve clinical outcomes. CLINICAL TRIAL REGISTRATION: https://www.clinicaltrials.gov. Unique identifiers: NCT01749878 and NCT02107872.

Genetic and Pharmacologic Inactivation of ANGPTL3 and Cardiovascular Disease.

BACKGROUND: Loss-of-function variants in the angiopoietin-like 3 gene (ANGPTL3) have been associated with decreased plasma levels of triglycerides, low-density lipoprotein (LDL) cholesterol, and high-density lipoprotein (HDL) cholesterol. It is not known whether such variants or therapeutic antagonism of ANGPTL3 are associated with a reduced risk of atherosclerotic cardiovascular disease. METHODS: We sequenced the exons of ANGPTL3 in 58,335 participants in the DiscovEHR human genetics study. We performed tests of association for loss-of-function variants in ANGPTL3 with lipid levels and with coronary artery disease in 13,102 case patients and 40,430 controls from the DiscovEHR study, with follow-up studies involving 23,317 case patients and 107,166 controls from four population studies. We also tested the effects of a human monoclonal antibody, evinacumab, against Angptl3 in dyslipidemic mice and against ANGPTL3 in healthy human volunteers with elevated levels of triglycerides or LDL cholesterol. RESULTS: In the DiscovEHR study, participants with heterozygous loss-of-function variants in ANGPTL3 had significantly lower serum levels of triglycerides, HDL cholesterol, and LDL cholesterol than participants without these variants. Loss-of-function variants were found in 0.33% of case patients with coronary artery disease and in 0.45% of controls (adjusted odds ratio, 0.59; 95% confidence interval, 0.41 to 0.85; P=0.004). These results were confirmed in the follow-up studies. In dyslipidemic mice, inhibition of Angptl3 with evinacumab resulted in a greater decrease in atherosclerotic lesion area and necrotic content than a control antibody. In humans, evinacumab caused a dose-dependent placebo-adjusted reduction in fasting triglyceride levels of up to 76% and LDL cholesterol levels of up to 23%. CONCLUSIONS: Genetic and therapeutic antagonism of ANGPTL3 in humans and of Angptl3 in mice was associated with decreased levels of all three major lipid fractions and decreased odds of atherosclerotic cardiovascular disease. (Funded by Regeneron Pharmaceuticals and others; ClinicalTrials.gov number, NCT01749878 .).

Evaluation of CETP activity in vivo under non-steady-state conditions: influence of anacetrapib on HDL-TG flux.

Studies in lipoprotein kinetics almost exclusively rely on steady-state approaches to modeling. Herein, we have used a non-steady-state experimental design to examine the role of cholesteryl ester transfer protein (CETP) in mediating HDL-TG flux in vivo in rhesus macaques, and therefore, we developed an alternative strategy to model the data. Two isotopomers ([(2)H11] and [(13)C18]) of oleic acid were administered (orally and intravenously, respectively) to serve as precursors for labeling TGs in apoB-containing lipoproteins. The flux of a specific TG (52:2) from these donor lipoproteins to HDL was used as the measure of CETP activity; calculations are also presented to estimate total HDL-TG flux. Based on our data, we estimate that the peak total postprandial TG flux to HDL via CETP is ∼ 13 mg · h(-1) · kg(-1) and show that this transfer was inhibited by 97% following anacetrapib treatment. Collectively, these data demonstrate that HDL TG flux can be used as a measure of CETP activity in vivo. The fact that the donor lipoproteins can be labeled in situ using well-established stable isotope tracer techniques suggests ways to measure this activity for native lipoproteins in free-living subjects under any physiological conditions.

Effects of anacetrapib on plasma lipids in specific patient subgroups in the DEFINE (Determining the Efficacy and Tolerability of CETP INhibition with AnacEtrapib) trial.

BACKGROUND: In the Determining the Efficacy and Tolerability of cholesteryl ester transfer protein (CETP) INhibition with AnacEtrapib (DEFINE) trial, anacetrapib added to statin produced robust low-density lipoprotein cholesterol (LDL-C)-lowering and high-density lipoprotein cholesterol (HDL-C)-raising vs placebo in patients with coronary heart disease (CHD). Predictors of the degree of LDL-C and HDL-C responses to anacetrapib, however, are poorly understood. OBJECTIVE: Lipid effects of anacetrapib in patient subgroups within the DEFINE trial (clinicaltrials.gov: NCT00685776) are reported. METHODS: The percent of placebo-corrected changes from baseline for LDL-C (estimated by Friedewald calculation [Fc-LDL-C]) and HDL-C after 24 weeks of anacetrapib 100 mg/day were compared among patients by age, gender, race, diabetes status, type of concomitant statin with or without other lipid therapies, and baseline HDL-C, Fc-LDL-C, and triglyceride (TG) levels. RESULTS: Percent decreases in Fc-LDL-C and increases in HDL-C with anacetrapib were similar (magnitude of difference generally <1/5 of the overall treatment effect) across subgroups by age, gender, diabetes status, lipid-modifying regimen, and baseline Fc-LDL-C, HDL-C, or TG. On the other hand, anacetrapib effects on Fc-LDL-C (-24% vs -41%) and HDL-C (+75% vs +139%) appeared to be less in black vs white patients, respectively. CONCLUSION: Effects of anacetrapib on Fc-LDL-C and HDL-C were generally comparable across subgroups, including being relatively independent of baseline Fc-LDL-C, HDL-C, or TG levels. The clinical impact of the lipid-modifying effects of anacetrapib is being evaluated in the cardiovascular disease outcomes trial, Randomized EValuation of the Effects of Anacetrapib though Lipid-modification (REVEAL).

Changes in LDL particle concentrations after treatment with the cholesteryl ester transfer protein inhibitor anacetrapib alone or in combination with atorvastatin.

OBJECTIVES: Our aim was to assess the effects of the cholesteryl ester transfer protein (CETP) inhibitor anacetrapib and atorvastatin, both as monotherapy and in combination, on particle concentrations of low-density lipoproteins (LDL), very low-density lipoproteins (VLDL), and intermediate-density lipoproteins in dyslipidemic patients. BACKGROUND: Although increases in high-density lipoproteins with CETP inhibition are well-documented, effects on atherogenic lipoprotein particle subclasses in dyslipidemic patients have not been extensively characterized. METHODS: Ion mobility was performed on stored plasma samples collected from patients before and after treatment with anacetrapib alone (150 and 300 mg/d) or in combination with atorvastatin (20 mg/d) in a previously conducted 8-week phase IIb study. RESULTS: Anacetrapib produced significant placebo-adjusted reductions of total LDL particles and all subfractions except for increases in very small LDL 4a and 4b. Atorvastatin reduced all LDL subfractions except LDL 4b. Results were generally additive for anacetrapib + atorvastatin. For patients treated with anacetrapib, the placebo-adjusted reduction in LDL 3a was attenuated and there was an increase in LDL 3b and 4a for those with low vs high triglyceride (TG) levels. For the atorvastatin alone vs placebo treatment comparison, there were small reductions in LDL 3a, 3b, and 4a for those with low vs high TG levels. CONCLUSIONS: Anacetrapib and atorvastatin produced similar reductions from baseline in total LDL particles, but did not have comparable effects on all LDL particle subfractions, and neither drug reduced the smallest LDL 4b particles. The clinical significance of these changes and the differential effects on very small LDL 4a in patients with higher vs lower TG remain to be determined (clinicaltrials.gov, NCT00325455).

Lipids, safety parameters, and drug concentrations after an additional 2 years of treatment with anacetrapib in the DEFINE study.

Anacetrapib is a cholesteryl ester transfer protein (CETP) inhibitor that has previously been shown to reduce low-density lipoprotein cholesterol (LDL-C) and raise high-density lipoprotein cholesterol (HDL-C) in patients with or at high risk of coronary heart disease in the 76-week, placebo-controlled, Determining the Efficacy and Tolerability of CETP Inhibition with Anacetrapib (DEFINE) trial. Here, we report the results of the 2-year extension to the DEFINE study where patients (n = 803) continued on the same assigned treatment as in the original 76-week study. Treatment with anacetrapib during the 2-year extension was well tolerated with a safety profile similar to patients on placebo. No clinically important abnormalities in liver enzymes, blood pressure, electrolytes, or adverse experiences were observed during the extension. At the end of the extension study, relative to the original baseline value, anacetrapib reduced Friedewald-calculated LDL-C by 39.9% and increased HDL-C by 153.3%, compared to placebo. The apparent steady state mean plasma trough concentration of anacetrapib was ∼640 nmol/L. Geometric mean plasma concentrations of anacetrapib did not appear to increase beyond week 40 of the 2-year extension of the 76-week DEFINE base study. In conclusion, an additional 2 years of treatment with anacetrapib were well tolerated with durable lipid-modifying effects on LDL-C and HDL-C.

Effects of anacetrapib on plasma lipids, apolipoproteins and PCSK9 in healthy, lean rhesus macaques.

Inhibition of cholesteryl ester transfer protein (CETP) has been vigorously pursued as a potential therapy to treat patients who are at an elevated risk for coronary artery disease. Anacetrapib, a novel CETP inhibitor, has been shown clinically to raise HDL cholesterol and reduce LDL cholesterol when provided as monotherapy or when co-administered with a statin. Preclinically, the effects of anacetrapib on the functionality and composition of HDL have been extensively studied. In contrast, the effects of anacetrapib on other parameters related to lipoprotein metabolism and cardiovascular risk have been difficult to explore. The aim of the present investigation was to evaluate the effects of anacetrapib in rhesus macaques and to compare these to effects reported in dyslipidemic humans. Our results from two separate studies show that administration of anacetrapib (150 mg/kg q.d. for 10 days) to rhesus macaques results in alterations in CETP activity (reduced by more than 70%) and HDL cholesterol (increased by more than 110%) which are similar to those reported in dyslipidemic humans. Levels of LDL cholesterol were reduced by more than 60%, an effect slightly greater than what has been observed clinically. Treatment with anacetrapib in this model was also found to lead to statistically significant reductions in plasma PCSK9 and to reduce cholesterol excursion in the combined chylomicron and remnant lipoprotein fraction isolated from plasma by fast protein liquid chromatography. Collectively, these data suggest that rhesus macaques may be a useful translational model to study the mechanistic effects of CETP inhibition.

Evaluation of lipids, drug concentration, and safety parameters following cessation of treatment with the cholesteryl ester transfer protein inhibitor anacetrapib in patients with or at high risk for coronary heart disease.

The aim of this study was to assess the effects on lipids and safety during a 12-week reversal period after 18 months of treatment with anacetrapib. The cholesteryl ester transfer protein inhibitor anacetrapib was previously shown to reduce low-density lipoprotein cholesterol by 39.8% (estimated using the Friedewald equation) and increase high-density lipoprotein (HDL) cholesterol by 138.1%, with an acceptable side-effect profile, in patients with or at high risk for coronary heart disease in the Determining the Efficacy and Tolerability of CETP Inhibition With Anacetrapib (DEFINE) trial. A total of 1,398 patients entered the 12-week reversal-phase study, either after completion of the active-treatment phase or after early discontinuation of the study medication. In patients allocated to anacetrapib, placebo-adjusted mean percentage decreases from baseline were observed at 12 weeks off the study drug for Friedewald-calculated low-density lipoprotein cholesterol (18.6%), non-HDL cholesterol (17.6%), and apolipoprotein B (10.2%); placebo-adjusted mean percentage increases were observed for HDL cholesterol (73.0%) and apolipoprotein A-I (24.5%). Residual plasma anacetrapib levels (about 40% of on-treatment apparent steady-state trough levels) were also detected 12 weeks after cessation of anacetrapib. No clinically important elevations in liver enzymes, blood pressure, electrolytes, or adverse experiences were observed during the reversal phase. Preliminary data from a small cohort (n = 30) revealed the presence of low concentrations of anacetrapib in plasma 2.5 to 4 years after the last anacetrapib dose. In conclusion, after the cessation of active treatment, anacetrapib plasma lipid changes and drug levels decreased to approximately 40% of on-treatment trough levels at 12 weeks after dosing, but modest HDL cholesterol elevations and low drug concentrations were still detectable 2 to 4 years after the last dosing.

Intensification of statin therapy results in a rapid reduction in atherosclerotic inflammation: results of a multicenter fluorodeoxyglucose-positron emission tomography/computed tomography feasibility study.

OBJECTIVES: The study sought to test whether high-dose statin treatment would result in greater reductions in plaque inflammation than low-dose statins, using fluorodeoxyglucose-positron emission tomography/computed tomographic imaging (FDG-PET/CT). BACKGROUND: Intensification of statin therapy reduces major cardiovascular events. METHODS: Adults with risk factors or with established atherosclerosis, who were not taking high-dose statins (n = 83), were randomized to atorvastatin 10 versus 80 mg in a double-blind, multicenter trial. FDG-PET/CT imaging of the ascending thoracic aorta and carotid arteries was performed at baseline, 4, and 12 weeks after randomization and target-to-background ratio (TBR) of FDG uptake within the artery wall was assessed while blinded to time points and treatment. RESULTS: Sixty-seven subjects completed the study, providing imaging data for analysis. At 12 weeks, inflammation (TBR) in the index vessel was significantly reduced from baseline with atorvastatin 80 mg (% reduction [95% confidence interval]: 14.42% [8.7% to 19.8%]; p < 0.001), but not atorvastatin 10 mg (% reduction: 4.2% [-2.3% to 10.4%]; p > 0.1). Atorvastatin 80 mg resulted in significant additional relative reductions in TBR versus atorvastatin 10 mg (10.6% [2.2% to 18.3%]; p = 0.01) at week 12. Reductions from baseline in TBR were seen as early as 4 weeks after randomization with atorvastatin 10 mg (6.4% reduction, p < 0.05) and 80 mg (12.5% reduction, p < 0.001). Changes in TBR did not correlate with lipid profile changes. CONCLUSIONS: Statin therapy produced significant rapid dose-dependent reductions in FDG uptake that may represent changes in atherosclerotic plaque inflammation. FDG-PET imaging may be useful in detecting early treatment effects in patients at risk or with established atherosclerosis.

Measurement of LDL-C after treatment with the CETP inhibitor anacetrapib.

Estimation of low-density lipoprotein cholesterol (LDL-C) using the Friedewald (FR) formula is often inaccurate when triglycerides are elevated or VLDL particle composition is altered. We hypothesized that LDL-C estimation by the FR formula and other measurement methods might also be inaccurate in individuals treated with a cholesteryl ester transfer protein (CETP) inhibitor. An assay comparison study was conducted using pre and posttreatment serum samples from 280 of the 811 patients treated with the CETP inhibitor anacetrapib in the DEFINE study (determining the efficacy and tolerability of CETP inhibition with anacetrapib). After 24 weeks of treatment with anacetrapib, mean LDL-C values by FR formula, Roche direct method (RDM) and Genzyme direct method (GDM) deviated from that measured by the ß-quantification (BQ) reference method by -12.2 ± 7.5, -10.2 ± 6.6, -10.8 ± 8.8 mg/dl, respectively. After treatment with anacetrapib, the FR formula and detergent-based direct methods provided lower LDL-C values than those obtained by the BQ reference method. The bias by the FR formula appeared to be due to an overestimation of VLDL-C by the TG/5 component of the formula. Evaluation of the clinical significance of these findings awaits comprehensive lipid and cardiovascular outcome data from ongoing Phase III clinical studies of anacetrapib.

Correlation between arterial FDG uptake and biomarkers in peripheral artery disease.

OBJECTIVES: A prospective, multicenter (18)fluorine-fluorodeoxyglucose ((18)F-FDG) positron emission tomography (PET)/computed tomography (CT) imaging study was performed to estimate the correlations among arterial FDG uptake and atherosclerotic plaque biomarkers in patients with peripheral artery disease. BACKGROUND: Inflammation within atherosclerotic plaques is associated with instability of the plaque and future cardiovascular events. Previous studies have shown that (18)F-FDG-PET/CT is able to quantify inflammation within carotid artery atherosclerotic plaques, but no studies to date have investigated this correlation in peripheral arteries with immunohistochemical confirmation. METHODS: Thirty patients across 5 study sites underwent (18)F-FDG-PET/CT imaging before SilverHawk atherectomy (FoxHollow Technologies, Redwood City, California) for symptomatic common or superficial femoral arterial disease. Vascular FDG uptake (expressed as target-to-background ratio) was measured in the carotid arteries and aorta and femoral arteries, including the region of atherectomy. Immunohistochemistry was performed on the excised atherosclerotic plaque extracts, and cluster of differentiation 68 (CD68) level as a measure of macrophage content was determined. Correlations between target-to-background ratio of excised lesions, as well as entire arterial regions, and CD68 levels were determined. Imaging was performed during the 2 weeks before surgery in all cases. RESULTS: Twenty-one patients had adequate-quality (18)F-FDG-PET/CT peripheral artery images, and 34 plaque specimens were obtained. No significant correlation between lesion target-to-background ratio and CD68 level was observed. CONCLUSIONS: There were no significant correlations between CD68 level (as a measure of macrophage content) and FDG uptake in the peripheral arteries in this multicenter study. Differences in lesion extraction technique, lesion size, the degree of inflammation, and imaging coregistration techniques may have been responsible for the failure to observe the strong correlations with vascular FDG uptake observed in previous studies of the carotid artery and in several animal models of atherosclerosis.

Changes in lipoprotein subfraction concentration and composition in healthy individuals treated with the CETP inhibitor anacetrapib.

We investigated the effects of the cholesteryl ester (CE) transfer protein inhibitor anacetrapib (ANA) on plasma lipids, lipoprotein subfraction concentrations, and lipoprotein composition in 30 healthy individuals. Participants (n = 30) were randomized to ANA 20 mg/day, 150 mg/day, or placebo for 2 weeks. Changes in concentration of lipoprotein subfractions were assessed using ion mobility, and compositional analyses were performed on fractions separated by density gradient ultracentrifugation. ANA 150 mg/day versus placebo resulted in significant decreases in LDL-cholesterol (26%) and apo B (29%) and increases in HDL-cholesterol (82%). Concentrations of medium and small VLDL, large intermediate density lipoprotein (IDL), and medium and small LDL (LDL2a, 2b, and 3a) decreased whereas levels of very small and dense LDL4b were increased. There was enrichment of triglycerides and reduction of CE in VLDL, IDL, and the densest LDL fraction. Levels of large buoyant HDL particles were substantially increased, and there was enrichment of CE, apo AI, and apoCIII, but not apoAII or apoE, in the mid-HDL density range. Changes in lipoprotein subfraction concentrations and composition with ANA 20 mg/day were similar to those for ANA 150 mg/day but were generally smaller in magnitude. The impact of these changes on cardiovascular risk remains to be determined.

Efficacy and safety after cessation of treatment with the cholesteryl ester transfer protein inhibitor anacetrapib (MK-0859) in patients with primary hypercholesterolemia or mixed hyperlipidemia.

This report describes the lipid and safety data collected during an off-drug period that followed 8 weeks of treatment with the cholesteryl ester transfer protein inhibitor, anacetrapib (ANA). A total of 589 patients with primary hypercholesterolemia or mixed hyperlipidemia were randomized to placebo, atorvastatin (ATV) 20 mg, and varying doses of ANA, provided as monotherapy or coadministered with ATV 20 mg daily. Patients were treated for 8 weeks, followed by an 8-week follow-up period, during which ANA was switched to placebo. At week 16 (8 weeks after ANA was stopped), persistent reductions in low-density lipoprotein cholesterol (LDL-C) were evident for the monotherapy groups receiving ANA 150 and 300 mg (-9.3% and -15.3%, respectively), and residual increases in high-density lipoprotein cholesterol (HDL-C) were observed for the monotherapy groups receiving ANA 40 mg (18.6%), 150 mg (40.5%), and 300 mg (43.4%). The effects on apolipoprotein B and apolipoprotein A-I were consistent with the changes observed for LDL-C and HDL-C, respectively. Corresponding residual changes in LDL-C and HDL-C were also noted in the ATV coadministration groups at the similar doses of ANA compared with ATV 20 mg alone. Residual plasma drug levels accompanied by reductions in cholesteryl ester transfer protein activity were observed at week 16 and may account for the alterations in plasma lipids 8 weeks after cessation of ANA.

Safety of anacetrapib in patients with or at high risk for coronary heart disease.

BACKGROUND: Anacetrapib is a cholesteryl ester transfer protein inhibitor that raises high-density lipoprotein (HDL) cholesterol and reduces low-density lipoprotein (LDL) cholesterol. METHODS: We conducted a randomized, double-blind, placebo-controlled trial to assess the efficacy and safety profile of anacetrapib in patients with coronary heart disease or at high risk for coronary heart disease. Eligible patients who were taking a statin and who had an LDL cholesterol level that was consistent with that recommended in guidelines were assigned to receive 100 mg of anacetrapib or placebo daily for 18 months. The primary end points were the percent change from baseline in LDL cholesterol at 24 weeks (HDL cholesterol level was a secondary end point) and the safety and side-effect profile of anacetrapib through 76 weeks. Cardiovascular events and deaths were prospectively adjudicated. RESULTS: A total of 1623 patients underwent randomization. By 24 weeks, the LDL cholesterol level had been reduced from 81 mg per deciliter (2.1 mmol per liter) to 45 mg per deciliter (1.2 mmol per liter) in the anacetrapib group, as compared with a reduction from 82 mg per deciliter (2.1 mmol per liter) to 77 mg per deciliter (2.0 mmol per liter) in the placebo group (P<0.001)--a 39.8% reduction with anacetrapib beyond that seen with placebo. In addition, the HDL cholesterol level increased from 41 mg per deciliter (1.0 mmol per liter) to 101 mg per deciliter (2.6 mmol per liter) in the anacetrapib group, as compared with an increase from 40 mg per deciliter (1.0 mmol per liter) to 46 mg per deciliter (1.2 mmol per liter) in the placebo group (P<0.001)--a 138.1% increase with anacetrapib beyond that seen with placebo. Through 76 weeks, no changes were noted in blood pressure or electrolyte or aldosterone levels with anacetrapib as compared with placebo. Prespecified adjudicated cardiovascular events occurred in 16 patients treated with anacetrapib (2.0%) and 21 patients receiving placebo (2.6%) (P = 0.40). The prespecified Bayesian analysis indicated that this event distribution provided a predictive probability (confidence) of 94% that anacetrapib would not be associated with a 25% increase in cardiovascular events, as seen with torcetrapib. CONCLUSIONS: Treatment with anacetrapib had robust effects on LDL and HDL cholesterol, had an acceptable side-effect profile, and, within the limits of the power of this study, did not result in the adverse cardiovascular effects observed with torcetrapib. (Funded by Merck Research Laboratories; ClinicalTrials.gov number, NCT00685776.).

Design of the DEFINE trial: determining the EFficacy and tolerability of CETP INhibition with AnacEtrapib.

BACKGROUND: Residual cardiovascular (CV) risk often remains high despite statin therapy to lower low-density lipoprotein cholesterol (LDL-C). New therapies to raise high-density lipoprotein cholesterol (HDL-C) are currently being investigated. Anacetrapib is a cholesteryl ester transfer protein (CETP) inhibitor that raises HDL-C and reduces LDL-C when administered alone or with a statin. Adverse effects on blood pressure, electrolytes, and aldosterone levels, seen with another drug in this class, have not been noted in studies of anacetrapib to date. METHODS: Determining the EFficacy and Tolerability of CETP INhibition with AnacEtrapib (DEFINE) is a randomized, double-blind, placebo-controlled trial to assess the efficacy and safety profile of anacetrapib in patients with coronary heart disease (CHD) or CHD risk equivalents (clinical trials.gov NCT00685776). Eligible patients at National Cholesterol Education Program-Adult Treatment Panel III LDL-C treatment goal on a statin, with or without other lipid-modifying medications, are treated with anacetrapib, 100 mg, or placebo for 18 months, followed by a 3-month, poststudy follow-up. The primary end points are percent change from baseline in LDL-C and the safety and tolerability of anacetrapib. Comprehensive preplanned interim safety analyses will be performed at the 6- and 12-month time points to examine treatment effects on key safety end points, including blood pressure and electrolytes. A preplanned Bayesian analysis will be performed to interpret the CV event distribution, given the limited number of events expected in this study. RESULTS: A total of 2,757 patients were screened at 153 centers in 20 countries, and 1,623 patients were randomized into the trial. Lipid results, clinical CV events, and safety outcomes from this trial are anticipated in 2010.

Novel strategy using F1-congenic mice for validation of QTLs: studies at the proximal chromosome 10 atherosclerosis susceptibility locus.

OBJECTIVE: We have previously identified a quantitative trait locus (QTL) for atherosclerosis susceptibility on proximal chromosome 10 (Chr10) (Ath11) in independent crosses of FVB and C57BL/6 (B6) mice on the apolipoprotein E (ApoE-/-) and LDL receptor (LDLR-/-) deficient backgrounds. The aims of the current study were to (1) test a novel strategy for validating QTLs using interval-specific congenic strains that were heterozygous (F1) across the genome, (2) validate the Chr10 QTL, and (3) to assess whether the phenotype is transferable by bone marrow transplantation. METHODS AND RESULTS: We generated Chr10 (0 to 21 cM) interval-specific mice on the F1.ApoE-/- background by crossing congenic FVB.ApoE-/-Chr10(B6/FVB) with B6.ApoE-/-, and B6.ApoE-/-Chr10(B6/FVB) with FVB.ApoE-/- mice. Lesion size was significantly larger in the resultant F1.ApoE-/-Chr10(FVB/FVB) mice compared to F1.ApoE-/-Chr10(B6/FVB) and F1.ApoE-/-Chr10(B6/B6) mice, validating the Chr10 QTL. The effect of the congenic interval was more robust on the F1.ApoE-/- than on the FVB.ApoE-/- and B6.ApoE-/- backgrounds. Bone marrow transplantation in congenic mice showed that the effect of the proximal Chr10 interval was not transferable by bone marrow-derived cells. CONCLUSIONS: A novel strategy of congenic strains on an F1 background proved useful to validate an atherosclerosis susceptibility QTL on mouse proximal Chr10.

Leptin-enhanced neointimal hyperplasia is reduced by mTOR and PI3K inhibitors.

Despite the use of the sirolimus (rapamycin) drug-eluting coronary stent, diabetics are at increased risk of developing in-stent restenosis for unclear reasons. Hyperleptinemia, which often coexists with diabetes and metabolic syndrome, is an independent risk factor for progression of coronary artery disease. It has not been determined whether elevated circulating leptin decreases the efficacy of the sirolimus drug-eluting stent in inhibiting neointimal hyperplasia, the process underlying restenosis after stenting. Here we show that leptin activates the mammalian target of rapamycin (mTOR) signaling pathway in primary murine vascular smooth muscle cells (VSMC) and stimulates VSMC proliferation in a PI3K-dependent fashion. Exogenous leptin, administered at levels comparable to those found in obese humans, promotes neointimal VSMC hyperplasia in a murine femoral artery wire injury model. Leptin significantly increases the dose of the mTOR inhibitor sirolimus that is required for effective inhibition of neointimal formation. Combination therapy with LY294002, a PI3K inhibitor, and sirolimus effectively inhibits leptin-enhanced neointimal hyperplasia. These data show that, in the setting of hyperleptinemia, higher doses of an mTOR inhibitor, or combination therapy with mTOR and PI3K inhibitors, inhibits neointimal hyperplasia after arterial injury. These studies may explain the higher rates of restenosis observed in diabetics treated with a sirolimus-eluting coronary stent and suggest a potential novel therapeutic approach for inhibiting in-stent restenosis in such patients.

Neuronatin: a new inflammation gene expressed on the aortic endothelium of diabetic mice.

OBJECTIVE: Identification of arterial genes and pathways altered in obesity and diabetes. RESEARCH DESIGN AND METHODS: Aortic gene expression profiles of obese and diabetic db/db, high-fat diet-fed C57BL/6J, and control mice were obtained using mouse Affymetrix arrays. Neuronatin (Nnat) was selected for further analysis. To determine the function of Nnat, a recombinant adenovirus (Ad-Nnat) was used to overexpress the Nnat gene in primary endothelial cells and in the mouse aorta in vivo. RESULTS: Nnat, a gene of unknown vascular function, was upregulated in the aortas of db/db and high-fat diet-fed mice. Nnat gene expression was increased in db/db mouse aorta endothelial cells. Nnat protein was localized to aortic endothelium and was selectively increased in the endothelium of db/db mice. Infection of primary human aortic endothelial cells (HAECs) with Ad-Nnat increased expression of a panel of nuclear factor-kappaB (NF-kappaB)-regulated genes, including inflammatory cytokines, chemokines, and cell adhesion molecules. Infection of mouse carotid arteries in vivo with the Ad-Nnat increased expression of vascular cell adhesion molecule 1 protein. Nnat activation of NF-kappaB and inflammatory gene expression in HAECs was mediated through pathways distinct from tumor necrosis factor-alpha. Nnat expression stimulated p38, Jun NH(2)-terminal kinase, extracellular signal-related kinase, and AKT kinase phosphorylation. Phosphatidylinositol 3-kinase and p38 inhibitors prevented Nnat-mediated activation of NF-kappaB-induced gene expression. CONCLUSIONS: Nnat expression is increased in endothelial cells of obese and diabetic mouse blood vessels. The effects of Nnat on inflammatory pathways in vitro and in vivo suggest a pathophysiological role of this new gene in diabetic vascular diseases.