HDLForum

Here we present the first in a series of commentaries focusing on emerging therapies for raising HDL cholesterol.

MicroRNAs: novel therapeutic strategy for raising HDL cholesterol?

IMicroRNAs (miRNAs) are small RNA molecules, typically 20 to 25 nucleotides in length. These do not encode proteins but instead regulate gene expression. miRNAs repress or destabilise messenger RNAs by binding to target sites in their untranslated regions, thereby influencing a range of physiological processes. Each miRNA may simultaneously target a number of genes involved in a pathway. Consequently, targeting specific miRNAs with 'antisense' oligonucleotide inhibitors may have potential as a novel class of therapeutics.

Recent studies have focused attention on two related miRNAs, miR-33a and miR-33b. These are located in the sterol-regulatory element-binding factor-2 (Srebf-2) and Srebf-1 genes, respectively. These genes encode for sterol-regulatory element-binding protein-2 (SREBP-2) and SREBP-1, which play key roles in transcriptional regulation of cholesterol uptake and synthesis, and fatty acid oxidation.

Experimental studies have shown that miR-33 post transcriptionally represses key genes involved in cellular cholesterol export and HDL metabolism, fatty acid oxidation and glucose metabolism. HDL Forum Editor Professor Kerry-Anne Rye, The Heart Research Institute, Sydney, Australia discusses some of the latest findings of relevance to HDL.

Rayner KJ, Suarez Y, Davalos A et al. miR-33 contributes to the regulation of cholesterol homeostasis. Science 2010;328:1570-3.

This study used both in vitro and in vivo models to investigate the regulation of miR-33 expression. In mouse peritoneal macrophages, miR-33 and Srebf-2 expression was down-regulated by cholesterol loading, and up-regulated when the macrophages were depleted of cholesterol by treatment with simvastatin. In vivo, in a mouse model, miR-33 levels were inversely correlated with cholesterol levels and positively correlated with Srebf-2 expression. In vitro binding studies to investigate the function of miR-33 showed that it specifically repressed the expression of ABCA1 (ATP-binding cassette A1).

ABCA1 is a cholesterol transporter that plays a key role in the efflux of excess cholesterol from cells, including macrophages in the artery wall, to lipid-free/lipid-poor apolipoprotein (apo)A-I in the extracellular space. It is also important in initiating HDL formation. By increasing miR-33 levels in mice, there was a corresponding decrease in hepatic ABCA1 expression and plasma levels of HDL. Alternatively, inhibition of miR-33 expression led to increases in both hepatic ABCA1 expression and plasma HDL levels.

Together these data suggest that miR-33 plays a role in the regulation of cholesterol homeostasis, by targeting two key pathways that control plasma levels of HDL control:

• HDL biosynthesis that is mediated by expression of ABCA1 in the liver and
• Cellular cholesterol efflux from macrophages, the first step in reverse cholesterol transport.

From the perspective of the HDL hypothesis, these findings have generated much interest in the therapeutic potential of antagonists of miR-33 to raise plasma levels of HDL cholesterol and reduce or slow atherosclerosis.

Rayner KJ, Sheedy FJ, Esau CC et al. Antagonism of miR-33 in mice promotes reverse cholesterol transport and regression of atherosclerosis. J Clin Invest 2011;121:2921–31.

Here, mice deficient in the LDL receptor were fed a Western diet to establish atherosclerosis, and then treated with an oligonucleotide inhibitor of miR-33 for 4 weeks. In this model, miR-33 inhibition increased plasma levels of HDL cholesterol by 35%, and this was associated with a 35% reduction in both plaque size and lipid content. Additionally, the lesions in these treated mice showed increased markers of plaque stability, including reduced macrophage accumulation and inflammatory gene expression.

Studies have also focused on the effect of miR-33a/b on key enzymes involved in the oxidation of fatty acids.¹ Of note, miR-33b has been shown to inhibit expression of 5′ adenosine monophosphate-activated protein kinase (AMPK), which promotes hepatic fatty acid β-oxidation and inhibits cholesterol and triglyceride synthesis. However, because mice lack miRNA-33b, but it is present in humans, there is a need to investigate this in a more appropriate model, such as the non-human primate.

Rayner KJ, Esau CC, Hussain FN et al. Inhibition of miR-33a/b in non-human primates raises plasma HDL and lowers VLDL triglycerides. Nature 2011;478:404-7. doi: 10.1038/nature10486.

This study investigated the effect of an anti-miR-33 oligonucleotide that targets both miR-33a and miR-33b in the African green monkey. Anti-miR-33 was administered at a clinically relevant dose of 5 mg/kg subcutaneously twice weekly for the first 2 weeks and once weekly for the remaining 10 weeks of the study.

Treatment with anti-miR-33 increased plasma levels of HDL cholesterol by 50% after 8 weeks, with levels sustained for the remainder of the study. Triglyceride-rich lipoprotein levels were also reduced by 50%.

The authors concluded that pharmacological inhibition of miR-33a/b may represent a promising therapeutic strategy to concomitantly raise plasma levels of HDL and lower triglycerides. This approach may benefit individuals with the dyslipidemia that is characteristic of type 2 diabetes and the metabolic syndrome, and is associated with high cardiovascular risk.²

Professor Rye summarises the implications of these findings.

Anti-miR-33 inhibits key genes involved in HDL synthesis, cholesterol homeostasis and fatty acid metabolism. Emerging data suggest that targeting miR-33 with an ‘antisense’ inhibitor may have therapeutic potential for raising plasma levels of HDL cholesterol. The latest study in a primate model, of relevance to humans, also showed that inhibition of miR 33 reduced triglycerides in addition to raising HDL cholesterol.

These data suggest that this agent may have future potential in targeting atherogenic dyslipidemia, against which statins are only modestly effective. Further findings are awaited with interest.

References

1. Dávalos A, Goedeke L, Smibert P et al. miR-33a/b contribute to the regulation of fatty acid metabolism and insulin signaling. Proc Natl Acad Sci 2011; 108:9232-7.

2. Chapman MJ, Ginsberg HN, Amarenco P et al. Triglyceride-rich lipoproteins and high-density lipoprotein cholesterol in patients at high risk of cardiovascular disease: evidence and guidance for management. Eur Heart J 2011;32:1345-61.