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Background: Coronary allograft vasculopathy (CAV) is an important cause of mortality after cardiac transplantation. High density lipoprotein (HDL) cholesterol efflux capacity has been inversely associated with coronary artery disease and is impaired in cardiac transplant recipients. We performed a single center case-cohort study to test the hypothesis that reduced efflux capacity is a risk factor for mortality and a second, multi-center retrospective study to test if efflux capacity is associated with CAV progression. Methods: We designed a single center case-cohort study in which we identified cardiac transplant patients who died between 2009-2012 (cases, n=34) and controls as cardiac transplant patients who were alive as of the fourth quarter of 2013 (n=57). Efflux capacity was measured by incubating apolipoprotein B-depleted serum with macrophages in a validated ex vivo system. In a second study, we utilized pre-transplant samples from the Clinical Trials in Organ Transplantation 5 (CTOT5) study to determine the association between ATP-binding-cassette (ABC) A1-dependent cholesterol efflux and CAV progression at 1 year. Results: In our single center study, the average time from transplant to study entry was well-matched between cases and controls (7.6±1.0 vs 7.7±0.8 years, respectively, p=0.48). Multivariable Cox proportional hazard ratios demonstrated that higher levels of HDL cholesterol efflux capacity were associated with survival (HR 0.61, 95% CI 0.43-0.85), even after adjustment for HDL cholesterol mass. To determine whether excess mortality observed in subjects with reduced efflux could be attributable to CAV progression, we tested the relationship between intravascular ultrasound (IVUS) progression of CAV and cholesterol efflux capacity using linear regression. ABCA1-dependent efflux and IVUS progression were significantly associated (β = -0.90, 95% CI [-1.73 - -0.07], p = 0.037, R2 = 0.37). Conclusion: Reduced efflux capacity is an important mediator of CAV progression and mortality in cardiac transplant recipients. This finding suggests that interventions to increase HDL cholesterol efflux capacity may provide clinical benefit in cardiac transplant recipients.

Background: Although plasma HDL-C levels are strongly associated with risk of coronary heart disease (CHD), interventions that raise HDL-C have not been shown to reduce CHD risk. A prototypical measure of HDL function, namely HDL cholesterol efflux capacity, has been shown to be associated with prevalent CHD even after adjusting for HDL-C, but its association with incident CHD events remains uncertain. Methods: We measured HDL cholesterol efflux capacity and assessed its relationship to incident CHD events in a nested case control sample within a prospective study (EPIC-Norfolk) of 25,639 individuals aged 40-79 years examined in 1993-1997 and followed up to 2009. We quantified efflux capacity in 1895 incident CHD cases and 2474 control participants free of any cardiovascular disorders, by using a validated ex-vivo radiotracer assay that involved incubation of J774 macrophages with apoB-depleted serum from the study participants. Results: HDL cholesterol efflux capacity was positively correlated with HDL-C levels (r2=0.27; P-value Conclusions: HDL cholesterol efflux capacity is significantly and inversely associated with incident CHD events, independent of established vascular risk factors and after adjusting for HDL-C and apoA-I levels.

Genome-wide association studies (GWAS) have identified 95 loci in the human genome that harbor common variants associated with plasma lipid traits. Of the 95 loci, 17 harbor genes known to cause monogenic lipid disorders and collectively a third of them contain genes with characterized roles in lipid metabolism. Therefore in the majority of loci the causal genes are unknown. We selected 32 genes, not previously implicated in lipid metabolism and representing a total of 26 loci, to test for their ability to modify plasma lipid concentrations upon somatic overexpression in vivo. We utilized adeno-associated virus serotype 8 (AAV8) to overexpress the selected genes specifically in the livers of both C57BL/6 mice and in an appropriate humanized mouse model (either mice expressing human apolipoprotein A-I for HDL loci or Apobec1-knockout, Ldlr haploinsufficient mice expressing human apolipoprotein B-100 for triglyceride and LDL loci). Approximately half of the genes tested reproducibly affected plasma lipids. For 13 of the interrogated loci the lipid-associated variants also correlated with expression variations of the respective genes in liver (liver expression quantitative trait loci-eQTLs). We demonstrate a causal role for 7 of these 13 genes. The overexpression of these 7 genes not only affected the predicted lipid class, but additionally exerted its effect in the predicted direction in 6 of 7 cases (Tmem57, Slc39a8, Ppp1r3b, Vkorc1, Tbkbp1 and Ube2l3). Additionally for a subset of the examined genes we proceeded to develop small interfering RNA (siRNA) nanoparticles that were particularly targeted to the liver. We were able to obtain robust knockdown for a significant number of genes and, in several cases, observe reciprocal effects on plasma lipids from our overexpression and knockdown studies. This work has identified several novel lipid regulators, whose further investigation can uncover novel mechanisms and pathways controlling plasma lipids.