**7. Lp-PLA2 as a therapeutic target**

in population-based Framingham Heart Study identified one locus associated with Lp-PLA2 mass, and four loci associated with Lp-PLA2 activity [80]. Twelve SNPs in the region of chromosome 6p12.3 near the gene for PLA2G7 were associated with Lp-PLA2 mass at a genome-wide level of significance. The top hit SNP rs1805017, is a nonsynonymous change (H92R) within the PLA2G7 gene. It was found that T allele that corresponds to the amino acid histidine was associated with higher Lp-PLA2 mass. On the other side, four loci achieving genome-wide significance for association with Lp-PLA2 activity was identified: a) first within the APOE/APOC1 gene cluster on chromosome 19q13.32 (rs41377151); b) second locus on chromosome 1p13.3, which includes the genes PSRC1 (rs599839), CELSR2 (rs4970834); c) third locus within an intron of SCARB1 on chromosome 12q24.31 (rs10846744); d) fourth locus in ZNF259 gene and BUD13 gene on chromosome 11q23.3 near the apolipoprotein gene cluster APOA5/APOA4/APOC3/APOA1 (rs12286037, rs11820589). Investigation from Suchindran's study was extended by Grallert et al. who made meta-analysis with additional four cohorts [81]. They performed genome-wide association study as part of the Cohorts for Heart and Aging Research in Genomic Epidemiology (CHARGE) Consortium in five population-based studies with the aim to identify genetic loci related to Lp-PLA2 mass/activity [82]. This study, in the popoulation of 13,664 subjects, revealed the association of PLA2G7 loci variants with Lp-PLA2 mass/activity, and genetic variants (APOC1, CELSR2, LDL, ZNF259, SCARB1)

related to low-density lipoprotein cholesterol levels with Lp-PLA2 activity.

significance [79].

120 Lipoproteins - From Bench to Bedside

exon 4 (Arg92His; rs1805017) [84-86].

Also, Lp-PLA2 mass was associated with SNP rs247616 on chromosome 16 within the choles‐ teryl ester transfer protein gene. The T allele of rs247616 was associated with higher HDL-C concentration and higher mean Lp-PLA2 mass [83]. The additional value of this study was the revealed significant association of four polymorphisms (in APOC1, CELSR2, SCARB1, ZNF259), but not PLA2G7 with coronary heart disease. Large-scale analysis focusing on variants in PLA2G7 did not identify any SNP that achieved experiment-wide statistical

The most frequently studied SNPs within PLA2G7 are: variants in exon 9 (Val279Phe; rs45619133), exon 11 (Val379Ala; rs1051931), exon 7 (Ile198Thr, Iso195Thr; rs1805018), and

The V279F SNP was found in subjects of Asian ancestry. The Val279Phe substitution is located within the catalytical domain of Lp-PLA2 and leads to reduction of enzyme activity in V/F heterozygous individuals or complete loss of enzymatic activity in homozygous F/F individ‐ uals. The 279F null allele is relatively frequent in Japan, with approximately 25% and 2% of the population carrying one or two copies. Its prevalence declines toward the West, has intermediate frequencies in China and Korea, is rare in the Middle East, and almost completely absent in European populations. The results from the association studies on this V279F variant and coronary artery disease have thus far been inconclusive. Li et al. [87] in Chinese Han population, found significant association of V279F and coronary artery disease, indicating that carrier of F allele increases the risk of coronary artery disease. This result was consistent with the Yamada et al. [88] study (850 cases/1,684 controls), which found that subjects carrying the mutant allele are at higher risk of arterial events (MI or stroke), and the Shimokata study (3,085 subjects with coronary artery disease/2,163 controls) [89]. The Jang et al. [90] study in two

It is well known that lipid-altering medications, including statins, fenofibrate, prescription of omega-3 fatty acids, for weight loss, have been shown to reduce Lp-PLA2 levels. The degree of its reduction correlates with the extent of lipid lowering.

Lp-PLA2 was identified as a potential novel target of therapy. The most used therapy targeting Lp-PLA2 in plasma in advanced stages of clinical investigation is darapladib. Darapladib is a selective, potent, reversibile, oral inhibitor of lipoprotein-associated phospholipase A2. The basic idea of applying darapladib is to improve patient outcomes in addition to evidence-based treatments and potentially reduce cardiovascular and cerebrovascular events by decreasing cytokines concentrations, stabilizing atherosclerotic plaque, inhibiting macrophage infiltra‐ tion, and thickening of the connective tissue cap.

The first study that showed as a secondary endpoint a change in coronary artery plaque necrotic core with darapladib after 12 months of treatment was Integrated Biomarkers and Imaging Study (IBIS)-2 [98]. It was a multicenter, randomized, double-blind placebo-control‐ led study that included 330 patients with angiographically confirmed coronary artery disease. Inhibition of Lp-PLA2 with darapladib also prevented necrotic core expansion of coronary plaque as measured on intravascular ultrasound.

One multicenter, randomized, double-blind placebo-controlled study examined the effects of darapladib on biomarkers of cardiovascular risk in 959 CAD and CAD-risk equivalent patients who were previously randomized to atorvastatin 20 mg or 80 mg and then randomized to oral darapladib 40, 80, 160 mg, or placebo for 12 weeks. Overall dose-dependent inhibition of Lp-PLA2 activity was sustained over the study period and was present in both atorvastatin dose groups, at different baseline LDL cholesterol < or ≥70 mg/dl, and high-density lipoprotein cholesterol HDL-C < or ≥40 mg/dl [99].

The first study that examined the effects of darapladib on Lp-PLA2 activity in Japanese dyslipidemic patients with/without the Val279Phe single-nucleotide polymorphism (SNP) of the *PLA2G7* gene showed that darapladib produced sustained inhibition of Lp-PLA2 activity [100].

Two large-scale studies with hard clinical endpoints were completed: the Stabilization of Atherosclerotic Plaque by Initiation of Darapladib Therapy (STABILITY) trial, with 15,828 randomized patients, and the Stabilization of Plaques using Darapladib-Thrombolysis In Myocardial Infarction 52 Trial (SOLID-TIMI 52), with an estimated recruitment of 13,026 patients [101-103].

The SOLID-TIMI 52 was a randomized, double-blind, placebo-controlled, multicenter, and event-driven trial that determined the clinical benefit of direct inhibition of Lp-PLA2 activity with darapladib in patients after an acute coronary syndrome (non-ST-elevation or STelevation myocardial infarction). Subjects were randomized to darapladib (160 mg entericcoated tablet daily) or matching placebo within 30 days after acute coronary syndrome. The primary endpoint was the composite of coronary heart disease death, nonfatal myocardial infarction, or nonfatal stroke, and secondary endpoints were major and total coronary events, individual components of the primary endpoint, and all-cause mortality. Patients were followed up for a median of 2.5 years. In patients who experienced an ACS event, direct inhibition of Lp-PLA2 with darapladib added to optimal medical therapy and initiated within 30 days of hospitalization did not reduce the risk of major coronary events.

The STabilization of Atherosclerotic plaque By Initiation of darapLadIb TherapY (STABILITY) trial was a double-blind trial in patients randomized more than one month after a myocardial infarction to double-blind darapladib (160 mg daily) or placebo (daily). The primary endpoint was a composite of cardiovascular death, myocardial infarction, or stroke. Secondary end‐ points included the components of the primary endpoint as well as major coronary events (death from coronary heart disease, myocardial infarction, or urgent coronary revasculariza‐ tion for myocardial ischemia) and total coronary events (death from coronary heart disease, myocardial infarction, hospitalization for unstable angina, or any coronary revascularization). Patients were followed up for a median of 3.7 years. In patients with stable coronary heart disease, darapladib did not significantly reduce the risk of the primary composite endpoint of cardiovascular death, myocardial infarction, or stroke.

Darapladib is a selective Lp-PLA2 inhibitor that is under investigation for its potential to stabilize high-risk atherosclerotic plaques and potentially reduce cardiovascular events [104,105]. A phase 2 clinical trial in 959 patients with CHD or CHD-risk equivalents demon‐ strated that darapladib was effective at producing sustained inhibition of plasma Lp-PLA2 activation in patients on atorvastatin therapy. This study was a post hoc analysis of this phase 2 trial studying high-risk patients with a diagnosis of peripheral arterial disease (PAD). Despite a more aggressive baseline risk factor profile, darapladib was equally effective at reducing Lp-PLA2 in patients with and without PAD [106]. Johnson et al. [107] assessed the effects of darapladib on both plasma and plaque Lp-PLA2 activity in patients undergoing elective carotid endarterectomy randomized to darapladib 40 mg (n = 34), 80 mg (n = 34), or placebo (n = 34) for 14 days. Patients were followed by carotid endarterectomy 24 hours after the last dose of study medication. Darapladib reduced plasma and plaque Lp-PLA2 activity compared with placebo.
