Selectins in Biology and Human Disease: Opportunity in E-selectin Antagonism

Abstract

Selectins are cell adhesion proteins discovered in the 1980s. As C-type lectins, selectins contain an essential calcium ion in the ligand-binding pocket and recognize the isomeric tetrasaccharides sialyl Lewis^x (sLe^x) and sialyl Lewis^a (sLe^a). Three selectins, E-selectin, P-selectin, and L-selectin, play distinct, complementary roles in inflammation, hematopoiesis, and tumor biology. They have been implicated in the pathology of diverse inflammatory disorders, and several selectin antagonists have been tested clinically. E-selectin plays a unique role in leukocyte activation, making it an attractive target for intervention, for example, in sickle cell disease (SCD). This review summarizes selectin biology and pathology, structure and ligand binding, and selectin antagonists that have reached clinical testing with an emphasis on E-selectin.

Keywords: acute lung injury; adhesion; antagonism; inflammation; leukocyte activation; selectin; sickle cell disease; thrombosis; vaso-occlusion.

Figure 1. Selectin genes and protein domain structure

(a) Selectin gene cluster on chromosome 1q. (b) Selectin protein structure. The N-terminal CRD is labeled "lectin" at the right side of the figure, followed by an EGF domain (oval) and varying numbers of SCR domains (rectangles). Intracellular domains are represented by green rectangles, and glycosylation sites are represented by tridents. CRD: carbohydrate-recognition domain, EGF: epidermal growth factor, SCR: short consensus repeat Image credits: Dr. Drew Provan using Adobe Illustrator

Figure 2. Selectin binding ligand structures

(a) Structures of sialyl Lewis^x and sialyl Lewis^a​​​​​​​: the GlcNAc residue is highlighted in red to indicate the isomeric nature of the structure. (b) Structure of synthetic P-selectin ligand SGP-3, comprising the terminal 19 amino acids of PSGL-1, including three sulfated tyrosine residues (sY5, sY7, and sY10) (highlighted in red), an O-linked saccharide at T16 containing a sialyl Lewis^x​​​​​​​ (highlighted in red) moiety, and an enterokinase cleavage sequence from R20 to K28 [16]. PSGL-1: P-selectin glycoprotein ligand-1 Image credits: Dr. Drew Provan using Adobe Illustrator

Figure 3. Inflammatory cascade leading to cell migration

(1) Inflammation activates local endothelium to express P-selectin that initiates tethering and rolling of circulating leukocytes along the endothelial wall. (2) Inflammatory cytokines induce de novo synthesis and surface expression of E-selectin that also captures circulating neutrophils. (3) E-selectin binds to sLe^x residues expressed by L-selectin on captured neutrophils, inducing conformational changes of LFA-1 from an LA to an intermediate affinity, then an HA state. (4) LFA-1 in its HA conformation binds to ICAM-1 expressed by endothelial cells, leading to firm arrest. (5) Following firm arrest, leukocytes extravasate into inflamed tissue by the process of diapedesis. sLe^x: sialyl Lewis^x, LFA-1: leukocyte function-associated antigen-1, LA: low affinity, HA: high affinity, ICAM-1: intercellular adhesion molecule-1, PSGL-1: P-selectin glycoprotein ligand-1 Image credits: Dr. Drew Provan using Adobe Illustrator

Figure 4. Neutrophil extravasation and acute lung injury or pneumonia

In severe lung inflammation, activated endothelial surface selectin expression leads to circulating neutrophil capture, extravasation into alveoli, and activation leading to alveolar fluid accumulation and impaired gas exchange. To eliminate redundancy, the binding cascade here omits PSGL-1 interactions with P-selectin and E-selectin, as well as E-selectin interactions with L-selectin. PSGL-1: P-selectin glycoprotein ligand-1, NETS: neutrophil extracellular traps, HA: high affinity, LFA-1: leukocyte function-associated antigen-1, ICAM-1: intercellular adhesion molecule-1 Image credits: Dr. Drew Provan using Adobe Illustrator

Figure 5. Vaso-occlusion in sickle cell disease

(1) Inflammatory triggers, such as hypoxia and hemolysis products, activate endothelium to express P-selectin, which captures circulating leukocytes. (2) E-selectin is expressed following de novo synthesis, capturing more leukocytes. (3) E-selectin binding to sLe^x on L-selectin (not shown in the figure), followed by the activation of LFA-1, induces endothelial capture of inflammatory cells, as well as secondary capture of platelets and red blood cells, leading to vascular occlusion. To eliminate redundancy, this binding cascade omits PSGL-1 interactions with P-selectin and E-selectin, as well as E-selectin interactions with L-selectin. sLe^x: sialyl Lewis^x, LFA-1: leukocyte function-associated antigen-1, PSGL-1: P-selectin glycoprotein ligand-1, LA: low affinity, HA: high affinity Image credits: Dr. Drew Provan using Adobe Illustrator

Figure 6. Thrombosis

(1) Endothelium is activated via hypoxia or inflammatory triggers leading to the expression of P-selectin, E-selectin, and vWF. These proteins capture circulating platelets, leukocytes, and TF+MV. (2) Activation of bound leukocytes leads to the expression of tissue factors on their surface. Captured platelets also release tissue factor. (3) Tissue factor then activates the clotting cascade, leading to thrombus formation, including rbc. vWF: von Willebrand factor, TF+MV: tissue factor expressing microvesicles, rbc: red blood cells, PSGL-1: P-selectin glycoprotein ligand-1 Image credits: Dr. Drew Provan using Adobe Illustrator

Figure 7. Selectin family X-ray crystal structures

Tertiary structures of E-selectin, P-selectin, and L-selectin CRD and EGF-like domains from X-ray crystallography. RCSB Protein Data Bank reference numbers are shown. CRD: carbohydrate-recognition domain, EGF: epidermal growth factor, RCSB: Research Collaboratory for Structural Bioinformatics Image credits: Dr. Drew Provan using Pymol

Figure 8. Bound and unbound selectin structures

(a) Crystal structures of CRD and EGF-like domains of E-selectin unbound and bound to sLe^x. (b) Crystal structures of P-selectin CRD and EGF-like domains unbound and bound to the PSGL-1 derived sulfoglycopeptide, SGP-3. RCSB Protein Data Bank reference numbers are shown. CRD: carbohydrate-recognition domain, EGF: epidermal growth factor, sLe^x: sialyl Lewis^x, PSGL-1: P-selectin glycoprotein ligand-1, RCSB: Research Collaboratory for Structural Bioinformatics Image credits: Dr. Drew Provan using Pymol

Figure 9. E-selectin bound to sLex (4CSY)

(a) Interactions between E-selectin and the fucose residue of sLe^x. (b) Interactions between E-selectin and the galactose and NeuAc residues of sLe^x. sLe^x: sialyl Lewis^x Image credits: Dr. Drew Provan using Pymol

Figure 10. P-selectin bound to PSGL-1 derived glycosulfopeptide SGP-3

(a) Interactions between P-selectin and the fucose residue of SGP-3. (b) Interactions between P-selectin and SGP-3 galactose and NeuAc residues. (c) Interactions between P-selectin and SGP-3 sulfotyrosine residues 7 and 10. Image credits: Dr. Drew Provan using Pymol

Figure 11. Structures of selectin antagonists that have been reported

Image credits: Dr. Drew Provan using Adobe Illustrator

Figure 12. Docking of uproleselan into E-selectin crystal structure 4CSY

(a) Predicted interactions in the fucose region of uproleselan. (b) Predicted interactions in the GalNAc-cyclohexylmethyl lactic acid region of uproleselan [109]. Image credits: Dr. Drew Provan using Pymol