Pharmacological strategies to inhibit intra-plaque angiogenesis in atherosclerosis.

Atherosclerosis is an inflammatory disease of the arterial wall leading to myocardial infarction, stroke and peripheral arterial disease. We published that elastin fragmentation, due to a mutation (C1039G+/-) in the fibrillin-1 (Fbn1) gene, promotes atherogenesis and a highly unstable plaque phenotype in apolipoprotein E deficient (ApoE-/-) mice on a Western diet. Interestingly, ApoE-/-Fbn1C1039G+/- mice reveal substantial intraplaque (IP) neovascularization, which is a typical feature of advanced human atherosclerotic plaques, but rarely observed in animal models. Because clinical evidence links IP angiogenesis with progressive and unstable vascular disease, we investigate whether inhibition of IP neovascularization has a beneficial effect on atherogenesis and plaque stability. Hitherto, blocking VEGF was the primary strategy to reduce neovascularization. Unfortunately, limited efficacy and adverse effects have downsized its success, even when multiple blockers were used simultaneously. In view of the above mentioned findings, we will investigate the following pharmacological approaches to inhibit IP angiogenesis and to stabilise atherosclerotic plaques: a nitric oxide (NO) donor (WP 1), glycolysis inhibitors (WP 2&3) and the olive polyphenol hydroxytyrosol (WP 4). WP 1 and 2 have already been performed in the framework of the Horizon 2020 MSCA-ITN project Moglynet, which started in May 2016. WP 3 and 4 will be finalised through the present DOCPRO 1. WP1: Because it has been shown that NO is an endogenous antiangiogenic mediator, we investigated the effect of the NO donor molsidomine on IP angiogenesis, atherogenesis and plaque stability in the ApoE-/-Fbn1C1039G+/- mouse model. We found that molsidomine favoured some features of atherosclerotic plaque stability and reduced myocardial infarction. However, the occurrence of IP neovascularization, the number of microvessels and the occurrence of haemorrhages in plaques were not affected (manuscript submitted). WP 2: Given that endothelial cells (ECs) rely on glycolysis for up to 85% of their energy demand, targeting the glycolytic pathway represents an attractive novel strategy to inhibit IP angiogenesis. Studies in the oncology field already showed that transient and partial inhibition of glycolysis in proliferating ECs inhibits pathological angiogenesis without interfering in the metabolism of healthy cells. In WP 2, we investigated the effects and the mechanism of action of glycolysis inhibitor 3PO [3-(3-pyridinyl)-1-(4-pyridinyl)-2-propen-1-one]. We found that in atherosclerotic ApoE−/−Fbn1C1039G+/− mice 3PO (50 μg/g, i.p.) reduced IP neovascularization and haemorrhages by 50% in a preventive regimen and by 38% in a curative regimen. However, the reduction in IP microvessels did not exert a significant effect on plaque composition. Nevertheless, 3PO reduced plaque formation, indicating that 3PO already has an effect in the onset of atherosclerosis. We could demonstrate that 3PO does not bind to PFKFB3, a key regulating enzyme in glycolysis, but inhibits glycolysis indirectly by lowering the intracellular pH due to blockade of monocarboxylate transporter 4 (manuscript in preparation). WP 3: Subsequently, we initiated WP 3 in order to evaluate in vitro and in ApoE-/-Fbn1C1039G+/- mice the effect of compound AZ67, which is a very potent inhibitor of PFKFB3. These experiments are ongoing and will be finalised during the duration of the DOCPRO1 grant. WP 4: Because it has recently been shown that the olive polyphenol hydroxytyrosol can inhibit angiogenesis both ex vivo and in vivo, we will investigate in WP 4 the effects of hydroxytyrosol on IP angiogenesis, atherogenesis and plaque stability in ApoE-/- Fbn1C1039G+/- mice. We already obtained interesting preliminary in vitro data. Further in vitro and in vivo experiments will be carried out during the duration of the DOCPRO1 grant.

Keywords:ATHEROSCLEROSIS, ENDOTHELIAL CELLS, NEOVASCULARISATION

Disciplines:Vascular diseases, Pharmacology not elsewhere classified