Targeting MicroRNA-33 To Reduce Intracranial Atherosclerosis and Other Neurovascular Hallmarks of Vascular Cognitive Impairment and Dementia

Atherosclerosis is a chronic, maladaptive inflammatory disease that can affect extra- and intracranial arteries. The combination of endothelial cell dysfunction, lipoprotein retention, monocyte/macrophage infiltration, intracellular and extracellular cholesterol accumulation, impaired apoptotic cell clearance, and extracellular matrix degradation leads to formation of advanced, unstable atherosclerotic plaques that can limit or occlude blood flow to tissues causing acute or chronic tissue damage. Intracranial atherosclerosis (ICAS) is a causative factor for ischemic stroke and has been associated with increased risk for Alzheimer’s disease (AD). Intracranial compared to extracranial atherosclerosis has a delayed onset of ~20 years but increases in prevalence and severity in individuals 60 years or older. With the steady rise in the percentage of US citizens above the age of 60, ICAS will play an ever-growing role in the morbidity and mortality caused by neurodegenerative diseases. Decreasing the concentration of low-density lipoprotein cholesterol with statins is the primary approach for blocking atherosclerosis progression and lowering the risk of an ischemic event. However, statins only reduce stroke risk by ~30% thus making it imperative to find additional therapies that regress or stabilize intracranial atheromas. The approach of our lab and others to reduce atherosclerosis is the inhibition of the microRNA-33 (miR-33) family. MiR-33a and miR-33b coordinately regulate lipid metabolism with their host genes sterol regulatory element binding transcription factor 2 (SREBF2) and SREBF1. By derepressing target genes involved in cholesterol trafficking, autophagy, and fatty acid oxidation, inhibiting miR-33 in macrophages increases cholesterol efflux, apoptotic cell clearance, and alternative M2 polarization. Through these combined mechanisms, pharmacological antagonism of miR-33 in mice reduces formation and stimulates regression of aortic atherosclerosis. However, murine studies are of limited translational value since mice express only miR-33a and do not to get ICAS. In contrast, nonhuman primates (NHP) like humans express both miR-33 family members and develop ICAS. Using a unique set of NHP samples, we will test the hypothesis that miR-33a/b antagonism stabilizes ICAS. We also hypothesize that the efficacy of miR-33 antagonists in the treatment of human ICAS is dependent upon the macrophage content of the atheromas. To evaluate whether human ICAS would be responsive to miR-33 antagonism, markers of macrophage content and function will be measured in human intracranial arteries available from the University of Kentucky AD Center. In the course of an NIH funded project (R01 HL111932) testing the hypothesis that miR-33a/b antagonism stabilizes coronary artery atherosclerosis, we found that our cynomolgus monkey model developed ICAS. While other studies have reported that NHPs develop ICAS, our study is unique in that we collected intracranial arteries from NHPs under conditions that should not only promote but also stabilize ICAS. Using our unique set of NHP samples, we will test the hypothesis that macrophage miR-33a/b antagonism in combination with aggressive plasma lipid lowering stabilizes ICAS by promoting cholesterol efflux, efferocytosis of apoptotic cells, and polarization towards a pro-resolving program that includes collagen deposition. We hypothesize that in combination with statins, miR-33a/b antagonism will stabilize intracranial atherosclerotic lesions by reducing macrophage content and directing macrophages towards a resolving phenotype. However, there is no research to date on the effects of statins on macrophage number and function within human intracranial atheromas. Therefore, we propose to determine the macrophage content of basilar arteries (BA) from elderly individuals who were treated without or with statins. For macrophage positive samples, we will further analyze the atheromas for markers of macrophage polarization, autophagy, cholesterol efflux, and efferocytosis, all of which are regulated by miR-33a/b. While statin treatment should have positive effects on macrophage content and function, we anticipate that statins will not eliminate pro-atherogenic macrophages from the intracranial atheromas thus setting the stage for the development and use of anti-inflammatory co-therapies such as miR-33 antagonists. We believe the innovation and significance of the project are high because it will: 1) Define the ability of miR-33a/b antagonists to regress or stabilize ICAS in a unique NHP model. 2) Assess the clinical potential of miR-33a/b antagonism for treating ICAS by characterizing macrophage content and function in human basilar arteries from humans treated statins. 3) Generate first of its kind information on the composition of ICAS from a translationally relevant human cohort and a highly controlled set of NHPs.