Xiaohong Yang

Oxidized Phospholipids, Lipoprotein(a), and Progression of Calcific Aortic Valve Stenosis

BACKGROUNDnElevated lipoprotein(a) (Lp[a]) is associated with aortic stenosis (AS). Oxidized phospholipids (OxPL) are key mediators of calcification in valvular cells and are carried by Lp(a).nnnOBJECTIVESnThis study sought to determine whether Lp(a) and OxPL are associated with hemodynamic progression of AS and AS-related events.nnnMETHODSnOxPL on apolipoprotein B-100 (OxPL-apoB), which reflects the biological activity of Lp(a), and Lp(a) levels were measured in 220 patients with mild-to-moderate AS. The primary endpoint was the progression rate of AS, measured by the annualized increase in peak aortic jet velocity in m/s/year by Doppler echocardiography; the secondary endpoint was need for aortic valve replacement and cardiac death during 3.5 ± 1.2 years of follow-up.nnnRESULTSnAS progression was faster in patients in the top tertiles of Lp(a) (peak aortic jet velocity: +0.26 ± 0.26 vs. +0.17 ± 0.21 m/s/year; p = 0.005) and OxPL-apoB (+0.26 ± 0.26 m/s/year vs. +0.17 ± 0.21 m/s/year; p = 0.01). After multivariable adjustment, elevated Lp(a) or OxPL-apoB levels remained independent predictors of faster AS progression. After adjustment for age, sex, and baseline AS severity, patients in the top tertile of Lp(a) or OxPL-apoB had increased risk of aortic valve replacement and cardiac death.nnnCONCLUSIONSnElevated Lp(a) and OxPL-apoB levels are associated with faster AS progression and need for aortic valve replacement. These findings support the hypothesis that Lp(a) mediates AS progression through its associated OxPL and provide a rationale for randomized trials of Lp(a)-lowering and OxPL-apoB-lowering therapies in AS. (Aortic Stenosis Progression Observation: Measuring Effects of Rosuvastatin [ASTRONOMER]; NCT00800800).

Antisense oligonucleotide lowers plasma levels of apolipoprotein (a) and lipoprotein (a) in transgenic mice.

OBJECTIVESnThis study sought to assess whether an antisense oligonucleotide (ASO) directed to apolipoprotein (a) [apo(a)] reduces apo(a) and lipoprotein (a) [Lp(a)] levels in transgenic mouse models.nnnBACKGROUNDnElevated Lp(a) is a causal, independent, genetic risk factor for cardiovascular disease and myocardial infarction. Effective therapies to specifically lower plasma Lp(a) levels are lacking.nnnMETHODSnThree transgenic mouse models were utilized: 8K-apo(a) mice expressing 8 kringle IV (KIV) repeats with a single copy of KIV-2; 8K-Lp(a) mice expressing both the 8K apo(a) plus human apolipoprotein B-100; and 12K-apo(a) mice expressing a 12K apo(a) with 3 KIV-2 repeats. The mice were treated intraperitoneally with saline, a control ASO, or ASO 144367 directed to KIV-2 for 4 to 6 weeks. Apo(a), Lp(a), and oxidized phospholipids present on human apoB (OxPL/h-apoB) or apo(a) [OxPL/apo(a)] were measured at baseline and on and off therapy.nnnRESULTSnASO 144367 significantly reduced Lp(a) by 24.8% in 8K-Lp(a) mice, and reduced apo(a) levels by 19.2% in 8K-Lp(a) mice, 30.0% in 8K-apo(a) mice, and 86% in 12K-apo(a) mice; ASO 144367 also significantly reduced OxPL/apoB 22.4% in 8K-Lp(a) mice, and OxPL/apo(a) levels by 19.9% in 8K-Lp(a) mice, 22.1% in 8K-apo(a) mice, and 92.5% in 12K-apo(a) mice (p < 0.004, or less, for all). No significant changes occurred in Lp(a), apo(a), OxPL/apoB, or OxPL/apo(a) levels with control ASO or saline.nnnCONCLUSIONSnThis study documents the first specific therapy, to our knowledge, for lowering apo(a)/Lp(a) levels and their associated OxPL. A more potent effect was documented in mice expressing apo(a) with multiple KIV-2 repeats. Targeting liver expression of apo(a) with ASOs directed to KIV-2 repeats may provide an effective approach to lower elevated Lp(a) levels in humans.

Determinants of binding of oxidized phospholipids on apolipoprotein (a) and lipoprotein (a)

Oxidized phospholipids (OxPLs) are present on apolipoprotein (a) [apo(a)] and lipoprotein (a) [Lp(a)] but the determinants influencing their binding are not known. The presence of OxPLs on apo(a)/Lp(a) was evaluated in plasma from healthy humans, apes, monkeys, apo(a)/Lp(a) transgenic mice, lysine binding site (LBS) mutant apo(a)/Lp(a) mice with Asp55/57→Ala55/57 substitution of kringle (K)IV10)], and a variety of recombinant apo(a) [r-apo(a)] constructs. Using antibody E06, which binds the phosphocholine (PC) headgroup of OxPLs, Western and ELISA formats revealed that OxPLs were only present in apo(a) with an intact KIV10 LBS. Lipid extracts of purified human Lp(a) contained both E06- and nonE06-detectable OxPLs by tandem liquid chromatography-mass spectrometry (LC-MS/MS). Trypsin digestion of 17K r-apo(a) showed PC-containing OxPLs covalently bound to apo(a) fragments by LC-MS/MS that could be saponified by ammonium hydroxide. Interestingly, PC-containing OxPLs were also present in 17K r-apo(a) with Asp57→Ala57 substitution in KIV10 that lacked E06 immunoreactivity. In conclusion, E06- and nonE06-detectable OxPLs are present in the lipid phase of Lp(a) and covalently bound to apo(a). E06 immunoreactivity, reflecting pro-inflammatory OxPLs accessible to the immune system, is strongly influenced by KIV10 LBS and is unique to human apo(a), which may explain Lp(a)’s pro-atherogenic potential.

Oxidation-specific biomarkers and risk of peripheral artery disease

OBJECTIVESnThe goal of this study was to examine the prospective association between oxidation-specific biomarkers, primarily oxidized phospholipids (OxPL) on apolipoprotein B-100-containing lipoproteins (OxPL/apoB) and lipoprotein (a) [Lp(a)], and risk of peripheral artery disease (PAD). We examined, as secondary analyses, indirect measures of oxidized lipoproteins, including autoantibodies to malondialdehyde-modified low-density lipoprotein (MDA-LDL) and apolipoprotein B-100 immune complexes (ApoB-IC).nnnBACKGROUNDnBiomarkers to predict the development of PAD are lacking. OxPL circulate in plasma, are transported by Lp(a), and deposit in the vascular wall and induce local inflammation.nnnMETHODSnThe study population included 2 parallel nested case-control studies of 143 men within the Health Professionals Follow-up Study (1994 to 2008) and 144 women within the Nurses Health Study (1990 to 2010) with incident confirmed cases of clinically significant PAD, matched 1:3 to control subjects.nnnRESULTSnLevels of OxPL/apoB were positively associated with risk of PAD in men and women: pooled relative risk: 1.37, 95% confidence interval: 1.19 to 1.58 for each 1-SD increase after adjusting age, smoking, fasting status, month of blood draw, lipids, body mass index, and other cardiovascular disease risk factors. Lp(a) was similarly associated with risk of PAD (pooled adjusted relative risk: 1.36; 95% confidence interval: 1.18 to 1.57 for each 1-SD increase). Autoantibodies to MDA-LDL and ApoB-IC were not consistently associated with risk of PAD.nnnCONCLUSIONSnOxPL/apoB were positively associated with risk of PAD in men and women. The major lipoprotein carrier of OxPL, Lp(a), was also associated with risk of PAD, reinforcing the key role of OxPL in the pathophysiology of atherosclerosis mediated by Lp(a).

MCP-1 binds to oxidized LDL and is carried by lipoprotein(a) in human plasma

Lipoprotein oxidation plays an important role in pathogenesis of atherosclerosis. Oxidized low density lipoprotein (OxLDL) induces profound inflammatory responses in vascular cells, such as production of monocyte chemoattractant protein-1 (MCP-1) [chemokine (C-C motif) ligand 2], a key chemokine in the initiation and progression of vascular inflammation. Here we demonstrate that OxLDL also binds MCP-1 and that the OxLDL-bound MCP-1 retains its ability to recruit monocytes. A human MCP-1 mutant in which basic amino acids Arg-18 and Lys-19 were replaced with Ala did not bind to OxLDL. The MCP-1 binding to OxLDL was inhibited by the monoclonal antibody E06, which binds oxidized phospholipids (OxPLs) in OxLDL. Because OxPLs are carried by lipoprotein(a) [Lp(a)] in human plasma, we tested to determine whether Lp(a) binds MCP-1. Recombinant wild-type but not mutant MCP-1 added to human plasma bound to Lp(a), and its binding was inhibited by E06. Lp(a) captured from human plasma contained MCP-1 and the Lp(a)-associated endogenous MCP-1 induced monocyte migration. These results demonstrate that OxLDL and Lp(a) bind MCP-1 in vitro and in vivo and that OxPLs are major determinants of the MCP-1 binding. The association of MCP-1 with OxLDL and Lp(a) may play a role in modulating monocyte trafficking during atherogenesis.

Reduction in lipoprotein-associated apoC-III levels following volanesorsen therapy: phase 2 randomized trial results

Elevated apoC-III levels predict increased cardiovascular risk when present on LDL and HDL particles. We developed novel high-throughput chemiluminescent ELISAs that capture apoB, lipoprotein (a) [Lp(a)], and apoA-I in plasma and then detect apoC-III on these individual lipoproteins as apoCIII-apoB, apoCIII-Lp(a), and apoCIII-apoAI complexes, respectively. We assessed the effects on these complexes of placebo or 100–300 mg volanesorsen, a generation 2.0+ antisense drug that targets apoC3 mRNA in patients with hypertriglyceridemia, including familial chylomicronemia syndrome (n = 3), volanesorsen monotherapy (n = 51), and as add-on to fibrate (n = 26), treated for 85 days and followed for 176 days. Compared with placebo, volanesorsen was associated with an 82.3 ± 11.7%, 81.3 ± 15.7%, and 80.8 ± 13.6% reduction in apoCIII-apoB, apoCIII-Lp(a), and apoCIII-apoA-I, respectively (300 mg dose; P < 0.001 for all), at day 92. Strong correlations in all assay measures were noted with total plasma apoC-III, chylomicron-apoC-III, and VLDL-apoC-III. In conclusion, novel high-throughput ELISAs were developed to detect lipoprotein-associated apoC-III, including for the first time on Lp(a). Volanesorsen uniformly lowers apoC-III on apoB-100, Lp(a), and apoA-I lipoproteins, and may be a potent agent to reduce triglycerides and cardiovascular risk mediated by apoC-III.

Relationship of Oxidized Phospholipids on Apolipoprotein B-100 to Cardiovascular Outcomes in Patients Treated With Intensive Versus Moderate Atorvastatin Therapy: The TNT Trial

BACKGROUNDnOxidized phospholipids on apolipoprotein B-100 (OxPL-apoB) is a biomarker of increased risk for major adverse cardiovascular events (MACE) in community cohorts, but its role in patients with stable coronary heart disease (CHD) is unknown.nnnOBJECTIVESnThis study sought to examine the relationship between these oxidative biomarkers and cardiovascular outcomes in patients with established CHD.nnnMETHODSnIn a random sample from the TNT (Treating to New Targets) trial, OxPL-apoB levels were measured in 1,503 patients at randomization (after an 8-week run-in period taking atorvastatin 10 mg) and 1 year after being randomized to atorvastatin 10 or 80 mg. We examined the association between baseline levels of OxPL-apoB and MACE, defined as death from CHD, nonfatal myocardial infarction, resuscitation after cardiac arrest, and fatal/nonfatal stroke, as well as the effect of statin therapy on OxPL-apoB levels and MACE.nnnRESULTSnPatients with events (n = 156) had higher randomization levels of OxPL-apoB than those without events (p = 0.025). For the overall cohort, randomization levels of OxPL-apoB predicted subsequent MACE (hazard ratio [HR]: 1.21; 95% confidence interval: 1.04 to 1.41; p = 0.018) per doubling and tertile 3 versus tertile 1 (hazard ratio: 1.69; 95% confidence interval [CI]: 1.14 to 2.49; p = 0.01) after multivariate adjustment for age, sex, body mass index, among others, and treatment assignment. In the atorvastatin 10-mg group, tertile 3 was associated with a higher risk of MACE compared to the first tertile (HR: 2.08; 95% CI: 1.20 to 3.61; p = 0.01) but this was not significant in the atorvastatin 80-mg group (HR: 1.40; 95% CI: 0.80 to 2.46; p = 0.24).nnnCONCLUSIONSnElevated OxPL-apoB levels predict secondary MACE in patients with stable CHD, a risk that is mitigated by atorvastatin 80 mg. (A Study to Determine the Degree of Additional Reduction in CV Risk in Lowering LDL Below Minimum Target Levels [TNT]; NCT00327691).

Effect of therapeutic interventions on oxidized phospholipids on apolipoprotein B100 and lipoprotein(a)

BACKGROUNDnOxidized phospholipids (OxPL) on apolipoprotein B-100 (OxPL-apoB) reflect the biological activity of lipoprotein(a) (Lp[a]) and predict cardiovascular disease events. However, studies with statins and low-fat diets show increases in OxPL-apoB and Lp(a).nnnOBJECTIVEnThis study evaluated changes in OxPL-apoB and Lp(a) with extended-release niacin (N), ezetimibe/simvastatin (E/S) and combination E/S/N. A systematic literature review of previously published trials, measuring both OxPL-apoB and Lp(a) after therapeutic interventions, was also performed.nnnMETHODSnOxPL-apoB and Lp(a) were measured in 591 patients at baseline and 24xa0weeks after therapy with N, E/S, or E/S/N in a previously completed randomized trial of hypercholesterolemic patients. The literature review included 12 trials and 3896 patients evaluating statins, low-fat diets, antisense to apolipoprotein(a) and lipid apheresis.nnnRESULTSnNiacin decreased OxPL-apoB levels (median [interquartile range]; 3.5 [2.2-9.2] nM to 3.1 [1.8-7.2] nM, Pxa0<xa0.01) and Lp(a) (10.9 [4.6-38.4] to 9.3 [3.1-32.9] mg/dL, Pxa0<xa0.01). In contrast, E/S and E/S/N significantly increased OxPL-apoB (3.5 [2.1-7.8] to 4.9 [3.0-11.1] nM, Pxa0<xa0.01) and (3.3 [1.9-9.3] to 4.3 [2.6-11.2] nM, Pxa0<xa0.01), respectively and Lp(a) (11.5 [6.1-36.4] to 14.9 [6.6-54.6] mg/dL, Pxa0<xa0.01) and (11.3 [5.4-43.8] to 11.6 [5.9-52.8] mg/dL, Pxa0<xa0.01), respectively. The systematic review of statins and diet demonstrated 23.8% and 21.3% mean increases in OxPL-apoB and 10.6% and 19.4% increases in Lp(a), respectively. However 44.1% and 52.0% decreases in OxPL-apoB and Lp(a), respectively, were present with Lp(a)-lowering therapies.nnnCONCLUSIONSnThis study demonstrates differential changes in OxPL-apoB and Lp(a) with various lipid-lowering approaches. These changes in OxPL-apoB and Lp(a) may provide insights into the results and interpretation of recent cardiovascular disease outcomes trials.

Oxidized Phospholipids on Apolipoprotein B-100 and Recurrent Ischemic Events Following Stroke or Transient Ischemic Attack

BACKGROUNDnBiomarkers to predict recurrent stroke and targets of therapy to prevent stroke are lacking.nnnOBJECTIVESnThis study evaluated whether patients with prior cerebrovascular events and elevated levels of oxidized phospholipids on apolipoprotein B-100 (OxPL-apoB), but without prior coronary artery disease (CAD), are at risk for recurrent stroke and CAD events following high-dose statin therapy.nnnMETHODSnIn the SPARCL (Stroke Prevention by Aggressive Reduction in Cholesterol Levels) trial, OxPL-apoB levelsxa0were measured in 4,385 patients with stroke or transient ischemic attack at baseline and in 3,106 patients at 5xa0years following randomization to placebo or 80 mg atorvastatin. The primary endpoint was the time from randomization to a second nonfatal or fatal stroke. Secondary endpoints included first major coronary events and anyxa0cardiovascular event.nnnRESULTSnPatients with recurrent stroke had higher baseline median OxPL-apoB levels than patients without (15.5xa0nmol/lxa0vs. 11.6 nmol/l; pxa0< 0.0001). After multivariable adjustment, elevated baseline OxPL-apoB predictedxa0recurrent strokexa0(hazard ratio [HR]: 4.3; pxa0< 0.0001), first major coronary events (HR: 4.0; pxa0< 0.0001), and any cardiovascular event (HR: 4.4; pxa0< 0.0001). These comparisons for any endpoint did not differ by treatment, shown as a nonsignificant interaction test. The net reclassification improvement, integrated discrimination improvement, and area under the receiver-operating characteristic curve (AUC) were all significantly improved by adding OxPL-apoB to the models, with ΔAUCxa0+0.0505 (pxa0<xa00.0001) for recurrent stroke, ΔAUCxa0+0.0409 (pxa0< 0.0001) for first major coronary event, and ΔAUCxa0+0.0791 (pxa0<xa00.0001) for any cardiovascular event.nnnCONCLUSIONSnElevated OxPL-apoB levels predicted recurrent stroke and first major coronary events in patients with prior stroke or transient ischemic attack. The lack of statin-OxPL-apoB treatment interaction suggested thatxa0OxPLs might be statin-independent therapeutic targets to reduce risk of cardiovascular events. (Lipitor inxa0thexa0Prevention of Stroke, for Patients Who Have Had a Previous Stroke [SPARCL]; NCT00147602).

Lipoprotein(a)-Associated Molecules Are Prominent Components in Plasma and Valve Leaflets in Calcific Aortic Valve Stenosis

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