E-selectin and transferrin receptor functionalized synthetic miRNA nanoshuttles with therapeutic potential in stroke

Current acute ischemic stroke treatment is limited to reperfusion therapies within the first hours after the onset, underscoring the urgent need for novel therapeutic strategies to mitigate the progression of the brain lesion. Here, we describe the development of bio-inspired synthetic nanoshuttles-lipid nanoparticles (LNPs)-designed to deliver miRNAs and target post-stroke inflammation, a key contributor to secondary brain damage. miRNAs isolated from human induced pluripotent stem cells (hiPSCs) were encapsulated into LNPs prepared by a microfluidic technique. We produced small sized, uniform nanoparticles with high miRNA encapsulation efficiency. LNPs were functionalized with anti-transferrin receptor (TfR) and anti-E-selectin antibodies to target the inflamed cerebral vasculature. We assayed LNPs' effect in a model of vascular inflammation in vitro and in vivo following brain ischemia/reperfusion in mice. In cultured human brain endothelial cells, empty LNPs, dual-targeted empty LNPs, and dual-targeted LNP encapsulating miRNAs (DT-LNP) reduced lipopolysaccharide-induced interleukin-6 production, indicating a modulatory effect of the lipid carrier under our experimental conditions. Mice received, via i.v., either empty LNPs (control) or two different doses of DT-LNPs at reperfusion following brain ischemia. The low dose of DT-LNPs reduced infarct volume and tended to improve the neurological deficit, and to downregulate the expression of IL-1β and E-selectin mRNAs, whereas the high dose did not. Altogether, these findings provide proof-of-concept that TfR-E-selectin-targeted LNPs encapsulating hiPSC-derived miRNAs represent a promising therapeutic approach for stroke.