Oxidized phospholipid dynamics in the early post-infarction period: Effects of PCSK9 inhibition with evolocumab.

We measured oxidized phospholipids (OxPL), lipoprotein (a) [Lp(a)], and lipoprotein-associated phospholipase A(2) (Lp-PLA(2)) pre- and postapheresis in 18 patients with familial hypercholesterolemia (FH) and with low(∼10 mg/dl; range 10-11 mg/dl), intermediate (∼50 mg/dl; range 30-61 mg/dl), or high (>100 mg/dl; range 78-128 mg/dl) Lp(a) levels. By using enzymatic and immunoassays, the content of OxPL and Lp-PLA(2) mass and activity were quantitated in lipoprotein density fractions plated in microtiter wells, as well as directly on apoB-100, Lp(a), and apoA-I immunocaptured within each fraction (i.e., OxPL/apoB and Lp-PLA(2)/apoB). In whole fractions, OxPL was primarily detected in the Lp(a)-containing fractions, whereas Lp-PLA(2) was primarily detected in the small, dense LDL and light Lp(a) range. In lipoprotein capture assays, OxPL/apoB and OxPL/apo(a) increased proportionally with increasing Lp(a) levels. Lp-PLA(2)/apoB and Lp-PLA(2)/apoA-I levels were highest in the low Lp(a) group but decreased proportionally with increasing Lp(a) levels. Lp-PLA(2)/apo(a) was lowest in patients with low Lp(a) levels and increased proportionally with increasing Lp(a) levels. Apheresis significantly reduced levels of OxPL and Lp-PLA(2) on apoB and Lp(a) (50-75%), particularly in patients with intermediate and high Lp(a) levels. In contrast, apheresis increased Lp-PLA(2)-specific activity (activity/mass ratio) in buoyant LDL fractions. The impact of apheresis on Lp(a), OxPL, and Lp-PLA(2) provides insights into its therapeutic benefits beyond lowering apoB-containing lipoproteins.

Introduction: Increased platelet activity during the peri- and early post-infarction period may contribute to adverse outcomes during that time. Platelet factor 4 (PF4) promotes coagulation and inflammation. The trajectory of PF4 and the impact of proprotein convertase subtilisin/kexin type 9 (PCSK9) inhibition on that trajectory are unknown. Hypothesis: Early administration of the PCSK9 inhibitor evulocumab, added to dual anti-platelet therapy, inhibits a rise in PF4 during the early post-infarction period. Methods: Participants from the Evolocumab in Acute Coronary Syndrome trials (EVACS; NCT03515304, NCT04082442), with a NSTEMI and a troponin-I of > 5 ng/ml or a STEMI were randomized to placebo or to 420 mg SQ of evolocumab within 24 hours of hospitalization. We performed serial ex vivo serum PF4 levels obtained at baseline (prior to study drug administration), and at days 1 and 30 and PF4 values were normalized to total platelet count. Normally distributed data were summarized using means and standard deviations and non-normally distributed data using medians and interquartile ranges. Results: Twenty-three participants were randomized to placebo and 23 to evolocumab. Mean age was 60±13 years, 48% were women, 22% were African American and 96% were on dual anti platelet therapy. Baseline PF4 concentrations (expressed as ng/1k platelets) did not differ between the two groups (placebo: 9.3 [4-12] vs evolocumab 8.0 [4-12], p=0.8). There was no significant change in PF4 in the placebo group from baseline to day 1 but a significant increase to 13.0 [11-14], p<0.01 (baseline vs 30 days) at 30 days (+32%). In contrast, there were no significant changes from baseline in the evolocumab group at either of the timepoints. At 30 days PF4 was higher in the placebo group 13.06 [11-14] than the evolocumab group 10.7 [6-13], p=0.04. Further analysis of results in the evolocumab group did not show any significant changes in PF4 based on the type of P2Y12 inhibitor used and no association with the change in LDL-cholesterol. Conclusion: A rise in PF4 levels was observed during the early post-infarction period in acute myocardial infarction patients. This rise was reversed by a single 420 mg SQ dose of evolocumab administered within 24 hours of hospital admission.

Background/Introduction Although inflammation is commonly present and often beneficial in the period following tissue injury, prolonged inflammation may impair healing and repair following acute ischemic injury. In addition to its impact on cholesterol metabolism PCSK9 is elevated in patients with inflammatory diseases and is thought to be a pro-inflammatory mediator. The presence and early trajectory of myocardial inflammation, and the impact of PCSK9 inhibition on that trajectory, in patients with an acute myocardial infarction have not been characterized. Purpose Our goals were to assess myocardial inflammation in patients with an acute myocardial infarction using 18F-FDG-PET scans obtained at the time of hospitalization for the infarction and at 30-days following the infarction and the impact of PCSK9 inhibition with evolocumab on inflammation in a placebo-controlled, randomized trial. Methods Fifty-five participants from the Evolocumab in Acute Coronary Syndrome trials, with a NSTEMI and a troponin-I of > 5 ng/ml or with a STEMI were randomized to placebo (n=30) or to 420 mg SQ of evolocumab (n=25) within 24 hours of admission. 18F-FDG-PET scans were performed within five days (2 [1-4]) of randomization and at 30 days. The myocardial mean Standardized Uptake Value (SUV) in the two randomized groups at baseline and the changes from baseline to 30 days were compared. All participants received guideline-directed therapies for acute infarction. Results Mean±SD age of the cohort was 57±13 years, 22% were African American, and 36% were women. There were no significant differences in demographic or clinical characteristics between the two groups. Mean SUV at baseline did not differ between the placebo (2.4±0.5) and the evolocumab (2.7±0.7) groups, p=0.25 for the between groups comparison. Mean SUV decreased in both groups, from 2.4±0.5 to 2.1±0.7 in the placebo group (p=0.05) and from 2.7±0.7 to 1.9±0.5 in the evolocumab group (p <0.0001), and the decrease was less in the placebo group, p=0.039 for the between groups comparison. The percent-change in the placebo group (-10.4%±36%) was also less than the percent change in the evolocumab group (-26.7%±20%), p = 0.04 for the between groups comparison. In addition, the percent of participants in whom the mean SUV units decreased was lower in the placebo group (60%) than in the evolocumab group (88%,), p=0.03 for the between groups comparison. Conclusion Myocardial inflammation assessed using 18F-FDG-PET decreases but is still present 30 days following an acute myocardial infarction. PCSK9 inhibition with evolocumab given within the first 24 hours of admission improves resolution of the inflammation. In addition to its marked impact on lipid levels, PCSK9 inhibition has significant anti-inflammatory effects in the early post-infarction setting.

Low-fat diets have been shown to increase plasma concentrations of lipoprotein(a) [Lp(a)], a preferential lipoprotein carrier of oxidized phospholipids (OxPLs) in plasma, as well as small dense LDL particles. We sought to determine whether increases in plasma Lp(a) induced by a low-fat high-carbohydrate (LFHC) diet are related to changes in OxPL and LDL subclasses. We studied 63 healthy subjects after 4 weeks of consuming, in random order, a high-fat low-carbohydrate (HFLC) diet and a LFHC diet. Plasma concentrations of Lp(a) (P < 0.01), OxPL/apolipoprotein (apo)B (P < 0.005), and OxPL-apo(a) (P < 0.05) were significantly higher on the LFHC diet compared with the HFLC diet whereas LDL peak particle size was significantly smaller (P < 0.0001). Diet-induced changes in Lp(a) were strongly correlated with changes in OxPL/apoB (P < 0.0001). The increases in plasma Lp(a) levels after the LFHC diet were also correlated with decreases in medium LDL particles (P < 0.01) and increases in very small LDL particles (P < 0.05). These results demonstrate that induction of increased levels of Lp(a) by an LFHC diet is associated with increases in OxPLs and with changes in LDL subclass distribution that may reflect altered metabolism of Lp(a) particles.

Oxidized Phospholipids Are Present on Plasminogen, Affect Fibrinolysis, and Increase Following Acute Myocardial Infarction

BackgroundLipoprotein(a) [Lp(a)], a major carrier of oxidized phospholipids (OxPL), is associated with an increased incidence of aortic stenosis (AS). However, it remains unclear whether elevated Lp(a) and OxPL drive disease progression and are therefore targets for therapeutic intervention. ObjectivesThis study investigated whether Lp(a) and OxPL on apolipoprotein B-100 (OxPL-apoB) levels are associated with disease activity, disease progression, and clinical events in AS patients, along with the mechanisms underlying any associations. MethodsThis study combined 2 prospective cohorts and measured Lp(a) and OxPL-apoB levels in patients with AS (Vmax >2.0 m/s), who underwent baseline 18F-sodium fluoride (18F-NaF) positron emission tomography (PET), repeat computed tomography calcium scoring, and repeat echocardiography. In vitro studies investigated the effects of Lp(a) and OxPL on valvular interstitial cells. ResultsOverall, 145 patients were studied (68% men; age 70.3 ± 9.9 years). On baseline positron emission tomography, patients in the top Lp(a) tertile had increased valve calcification activity compared with those in lower tertiles (n = 79; 18F-NaF tissue-to-background ratio of the most diseased segment: 2.16 vs. 1.97; p = 0.043). During follow-up, patients in the top Lp(a) tertile had increased progression of valvular computed tomography calcium score (n = 51; 309 AU/year [interquartile range: 142 to 483 AU/year] vs. 93 AU/year [interquartile range: 56 to 296 AU/year; p = 0.015), faster hemodynamic progression on echocardiography (n = 129; 0.23 ± 0.20 m/s/year vs. 0.14 ± 0.20 m/s/year] p = 0.019), and increased risk for aortic valve replacement and death (n = 145; hazard ratio: 1.87; 95% CI: 1.13 to 3.08; p = 0.014), compared with lower tertiles. Similar results were noted with OxPL-apoB. In vitro, Lp(a) induced osteogenic differentiation of valvular interstitial cells, mediated by OxPL and inhibited with the E06 monoclonal antibody against OxPL. ConclusionsIn patients with AS, Lp(a) and OxPL drive valve calcification and disease progression. These findings suggest lowering Lp(a) or inactivating OxPL may slow AS progression and provide a rationale for clinical trials to test this hypothesis.

Effect of Evolocumab on Atherogenic Lipoproteins During the Peri- and Early Postinfarction Period: A Placebo-Controlled, Randomized Trial.

Little is known about the heterogeneity in low-density lipoprotein cholesterol levels (LDL-C) lowering with proprotein convertase subtilisin kexin 9 (PCSK9) inhibitor medications. To evaluate the interindividual variability in LDL-C reduction with the PCSK9 inhibitor drug evolocumab. We examined the percentage change in LDL-C levels from baseline in the Further Cardiovascular Outcomes Research With PCSK9 Inhibition in Subjects With Elevated Risk (FOURIER) trial, a placebo-controlled randomized clinical trial of the PCSK9 inhibitor evolocumab in patients with stable atherosclerotic cardiovascular disease who were taking statin medications. Patients in either treatment arm who had high baseline LDL-C variability during screening and either did not receive the study drug, altered their background lipid-lowering therapy regimen, or had no LDL-C level sample in week 4 were excluded from the primary analysis. Analyses in the patients were stratified by treatment arm. Data was collected from 2013 to 2016, and data were analyzed from January 2018 to November 2018. Interindividual variation in percent reduction in LDL-C with evolocumab. There were 27 564 individuals in the cohort; after exclusions for baseline variability (n = 3524) or alterations in background lipid therapy and other causes (n = 2272), 21 768 patients remained. At week 4, the median percent reduction in LDL-C levels from baseline was 66% (interquartile range, 54%-76%; median [interquartile range] baseline value, 90 [79-105] mg/dL; postchange value, 31 [21-44] mg/dL) with evolocumab. During the first year, a total of 10 325 of 10902 patients in the evolocumab group (94.7%) had a reduction 50% or greater in LDL-C levels, 10 669 of 10 902 (97.9%) had a reduction 30% or more, and 10 849 of 10 902 (99.5%) had any reduction in LDL-C levels. Fifty-three patients (0.5%) had no apparent reduction in LDL-C levels. In the placebo arm, the median LDL-C reduction was 4% (interquartile range, 6% increase to 13% reduction; baseline median [IQR] value, 90 [79-106] mg/dL; postchange value, 87 [74-103] mg/dL) at 4 weeks. Waterfall plots showed notable variability in the top and bottom 5% of patients for both evolocumab and placebo groups, with large changes in LDL-C levels in the placebo group (increases of ≥25%, 531 patients [4.9%]; decreases of ≥25%, 985 patients [9.1%]). At 4 weeks, the placebo-adjusted reductions in LDL-C levels with evolocumab were 50% or greater in 9839 of 10 866 patients (90.5%) and 30% or greater in 10 846 of 10 866 patients (99.8%). Results were consistent across clinically relevant subgroups. There appears to be a highly consistent robust reduction in LDL-C levels with evolocumab use. ClinicalTrials.gov identifier: NCT01764633.

Introduction: Lipoproteins carrying pro-inflammatory oxidized phospholipids (OxPL), exemplified by lipoprotein(a) [Lp(a)], contribute to residual risk for major adverse cardiovascular events (MACE) in patients with chronic coronary syndrome (CCS) despite optimal medical treatment. Hypothesis: Anti-inflammatory colchicine therapy modifies MACE in patients with elevated Lp(a), OxPL on apo(a) [OxPL-apo(a)], and OxPL on apoB containing lipoproteins (OxPL-apoB). Aims To study the relationship of Lp(a), OxPL-apo(a) and OxPL-apoB with MACE in 1777 LoDoCo2 trial participants with CCS randomized to colchicine or placebo. Methods: End of study but not baseline samples were available. Because Lp(a), OxPL-apo(a) and OxPL-apoB levels are strongly genetically determined, we assumed that end of study values would reflect baseline values. The primary endpoint was a composite of MI, ischemic CVA or ischemia driven coronary revascularization. Cox regression was used to compare the incidence of the primary endpoint by Lp(a) dichotomized at 125 nmol/L and OxPL-apo(a) and OxPL-apoB in tertiles. Results: At study end, median (IQR) on-colchicine Lp(a), OxPL-apo(a), and OxPL-apoB levels in nmol/L were 22.4 (7.2-102.7), 7.8 (2.0-34.5) and 3.1 (1.3-6.3), similar to on-placebo levels of 23.9 (9.9-95.7), 8.8 (2.7-34.0), and 3.1 (1.4-5.9), respectively (p = 0.20, 0.98 and 0.18). MACE reduction with colchicine was independent of Lp(a) and OxPL-apo(a) (P interaction = 0.92 and 0.66 respectively). However, the largest MACE reduction was present in the highest vs lowest OxPL-apoB tertile ( Figure , P interaction &lt; 0.05). Conclusions: The benefit of colchicine in reducing MACE was highest in subjects with elevated OxPL-apoB, suggesting colchicine may be most effective in subjects with heightened oxidation-driven inflammation. These findings may be hypothesis-generating and require validation in larger trials with baseline biomarker assessment including CRP and IL-6.

Objectives: Lipoprotein(a)[Lp(a)] is a causal, genetic risk factor for cardiovascular disease(CVD) and is enriched in pro-inflammatory oxidized phospholipids(OxPL) and lipoprotein-associated phospholipase A2(Lp-PLA2). Epidemiologic outcome studies have shown that the OxPL, Lp(a) and Lp-PLA2 collectively mediate additive risk for CVD. Methods: To derive insights into the relationships between Lp(a), OxPL and Lp-PLA2, 18 patients with familial hypercholesterolemia(FH) and Low(&lt;10mg/dl), Intermediate (∼50mg/dl) or High(&gt;100mg/dl) Lp(a) levels on chronic LDL apheresis were evaluated. Twenty-five isopycnic density fractions(density &lt;1.015 to &gt;1.189 g/ml) were isolated from pre and post apheresis plasma(n=50 fractions per patient). OxPL and Lp-PLA2 mass and activity were quantitated with a variety of techniques under 3 conditions: in plasma, in each density fraction with direct plating techniques and on individual lipoproteins within each density fraction by using antibody capture assays to apoB-100(OxPL/apoB and Lp-PLA2/apoB), Lp(a)[OxPL/apo(a) and Lp-PLA2/apo(a)] and apoA-I(OxPL/apoA-I and Lp-PLA2/apoA-I)(total of 900 fractions per measure) and detecting OxPL and Lp-PLA2 with monoclonal antibodies E06 and 4B4, respectively. Results: In whole density gradients by direct plating techniques, OxPL were almost exclusively detected in the Lp(a) containing fractions(density 1.059-1.094 g/ml) and Lp-PLA2 mass and activity were present between the apoB and Lp(a) density fractions. In lipoprotein capture assays, OxPL/apoB and OxPL/apo(a) increased proportionally with increasing Lp(a) levels; Lp-PLA2/apoB and Lp-PLA2/apoA-I levels were highest in patients with Low Lp(a) but decreased proportionally with increasing Lp(a) levels; Lp-PLA2/apo(a) was lowest in patients with Low Lp(a) levels and increased proportionally with increasing Lp(a) levels. LDL apheresis significantly reduces levels of OxPL and Lp-PLA2 on apoB and Lp(a), particularly in patients with high Lp(a) levels Conclusions: This study documents that OxPL are primarily present on Lp(a) and Lp-PLA2 primarily on apoB, but the proportion of each increases on Lp(a) and decreases on apoB as plasma Lp(a) levels increase. These data suggest a pathophysiological relationship between Lp(a), OxPL and Lp-PLA2 and provide a rationale for understanding how they collectively mediate CVD in humans.

Inflammatory lipoprotein(a) [Lp(a)] and oxidized phospholipids (OxPLs) on lipoproteins convey residual cardiovascular disease risk. The low-dose colchicine 2 (LoDoCo2) trial showed that colchicine reduced the risk of cardiovascular events occurring on standard therapies in patients with chronic coronary syndrome (CCS). We explored the effects of colchicine on Lp(a)- and oxidized lipoprotein-associated risk in a LoDoCo2 biomarker subpopulation. Lipoprotein(a) and OxPLs on apolipoprotein(a) [OxPL-apo(a)] and apolipoprotein B-100 (OxPL-apoB) levels were determined in the biomarker population of the LoDoCo2 trial (n = 1777). The Cox regression analysis was used to compare the risk of the primary endpoint, consisting of myocardial infarction, ischaemic stroke, or ischaemia-driven revascularization by biomarker levels. Interactions between treatment, Lp(a), and OxPL levels were evaluated. Lipoprotein(a), OxPL-apo(a), and OxPL-apoB levels were similar between the colchicine and placebo groups. Consistent risk reduction by colchicine was observed in those with Lp(a) < 125 nmol/L and ≥125 nmol/L and the highest OxPL-apo(a) tertile compared with the lowest (Pinteraction = 0.92 and 0.66). The absolute risk reduction for those with Lp(a) ≥ 125 nmol/L appeared higher compared with those with Lp(a) < 125 nmol/L (4.4% vs. 2.4%). A treatment interaction for colchicine was found in those with the highest OxPL-apoB tertile vs. the lowest (Pinteraction = 0.04). In patients with CCS, colchicine reduces cardiovascular disease risk in those with and without elevated Lp(a) but absolute benefits appeared higher in those with Lp(a) ≥ 125 nmol/L. Patients with higher levels of OxPL-apoB experienced greater benefit of colchicine, suggesting that colchicine may be more effective in subjects with heightened oxidation-driven inflammation.

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

The Trajectory of Lipoprotein(a) During the Peri- and Early Postinfarction Period and the Impact of Proprotein Convertase Subtilisin/Kexin Type 9 Inhibition

BackgroundLp(a) (lipoprotein(a)) is the major lipoprotein carrier of oxidized phospholipids (OxPL) and this function mediates Lp(a) atherogenicity. However, the relationship between OxPL, Lp(a), and genetic and biological characteristics remains poorly understood. We assessed the relationship between Lp(a)‐bound OxPL, apolipoprotein(a) (apo(a)) size, age, and family structure in 2 racial groups.Methods and ResultsHealthy Black and White families were recruited from the general population (age: 6–74 years, n=267). OxPL and Lp(a) levels were assayed enzymatically; apo(a) isoform, LPA allele sizes, and allele‐specific Lp(a) levels were determined. Lp(a)‐OxPL levels did not differ significantly by racial and age groups. Lp(a)‐OxPL levels were associated with total plasma Lp(a) in all participants and in race‐specific analyses. Further, OxPL levels were significantly associated with allele‐specific Lp(a) levels carried by the smaller apo(a) size in all participants (β=0.33, P=0.0003) as well as separately for Black (β=0.50, P=0.0032) and White (β=0.26, P=0.0181) participants. A significant association of OxPL with allele‐specific Lp(a) levels for larger apo(a) sizes was seen only in Black participants (β=0.53, P=0.0076). In this group, Lp(a)‐OxPL levels were also heritable (h2=0.29, P=0.0235), resulting in a significant interracial difference in heritability between Black and White people (P=0.0352).ConclusionsLp(a)‐OxPL levels were associated with allele‐specific Lp(a) level carried on smaller apo(a) sizes and among Black participants also for larger apo(a) sizes. The heritability estimates for Lp(a)‐bound OxPL differed by race.

Lp(a) and SARS-CoV-2: A conspiracy of two mysteries.