Glycosylation inhibitors as novel vascular therapeutics for sickle cell disease

A salient pathophysiological change of sickle cell disease (SCD) is vascular inflammation, which is associated with overexpression of multiple adhesion molecules. Recently cell-free heme, which is resulting from intravascular hemolysis, is shown to be responsible for vascular inflammation. Although cell-free heme is toxic to cells and is a potent inducer of inflammation, paroxysmal nocturnal hemoglobinuria is characterized by higher intravascular cell-free heme levels than SCD but demonstrates normal levels of adhesion molecules in blood. This suggests to us that in addition to cell-free heme, other mechanisms also contribute to vascular inflammation. Our recent intravital microscopic studies demonstrated that hypoxia and low nitric oxide bioavailability enhance the adhesion of sickle red blood cells (RBCs) in a synergistic manner, and that P-selectin is responsible for the synergistic sickle RBC adhesion; importantly, we showed for the first time that p38 mitogen-activated protein kinase pathways contribute to the elevated expression of P-selectin on endothelial cells. While PSI-697, a P-selectin binding antagonist, was developed for treating vascular thrombosis and other disorders, this agent was unable to inhibit platelet'monocyte aggregate formation in a clinical trial, which suggested the necessity to develop novel P-selectin inhibitors. Because P-selectin antagonists unlikely have regulatory effects on adhesion molecule expression, we hypothesized that chemicals which inhibit the expression of adhesion molecules in endothelial cells might have potent clinical activities for vascular inflammation in SCD. We recently found that glycosylation inhibitors efficiently inhibit both P-selectin expression in endothelial cells as well as the adhesion of sRBCs and leukocytes to endothelial cells, and that these inhibitors downregulate P-selectin expression on endothelial cells by inhibiting phosphatidylinositol 3 kinase (PI3K)/AKT signaling. The goal of this application is to determine whether glycosylation inhibitors and PI3K/AKT inhibitors are able to improve vascular inflammation in SCD model mice. Specific Aim1 is to investigate the roles for glycosylation inhibitors in cell adhesion and vascular inflammation in SCD. In Specific Aim 2, we will investigate the molecular and signaling mechanisms by which glycosylation inhibitors downregulate the expression of adhesion molecules in endothelial cells; we will focus on the roles of PI3K/AKT signaling. (AHA Program: Grant-in-Aid)