ALBERT

Description: Uproleselan (GMI-1271), an E-selectin antagonist, has been shown in preclinical models to disrupt activation of cell survival pathways in acute myeloid leukemia (AML), enhance chemotherapy efficacy, and improve survival. Uproleselan received FDA breakthrough therapy designation for adult relapsed/refractory AML in 2017 and Phase III studies are ongoing. In the present studies we report on the in vitro and in vivo comparative activities of an innovative high potency E-selectin antagonist, GMI-1687, a potential subcutaneously administered follow-on drug candidate to Uproleselan. The binding constant, association and dissociation rates of GMI-1687 to immobilized recombinant human (rh) E-selectin were determined by surface plasmon resonance (SPR) at 25oC. The KD of GMI-1687 was 2.4 nM, with Kon = 3 x 106 M-1s-1 and Koff = 1 x10-2 s-1. Under similar experimental conditions the KD of Uproleselan was 520 nM with Kon = 0.02 x 106 M-1s-1 and Koff = 1 x10-2 s-1. GMI-1687 was evaluated for its ability to inhibit binding of sialyl Lea to immobilized rh E-selectin. The median IC50 (n=6 independent assays) of GMI-1687 and Uproleselan in this assay was 15 and 550 nM, respectively. The in vitro activity of GMI-1687 to release adherent KG1a AML cells from E-selectin coated wells was also determined. GMI-1687 at 100 nM detached approximately 55% of adherent AML cells and was significantly different from Uproleselan at an identical concentration (38% detachment, P=0.0216). The percent bioavailability (%F) of GMI-1687 was evaluated in male Sprague-Dawley rats following intravenous (IV) and subcutaneous (SC) routes of administration at 5 mg/kg. The mean (+/- SD) SC %F for GMI-1687 was 126 +/- 3.8%. GMI-1687 also showed high bioavailability in CD-1 mice after SC administration of 0.58 mg/kg with %F = 132 +/-38. The in vivo therapeutic activity of GMI-1687 following SC administration was assessed in an acute model of inferior vena cava (IVC) thrombosis and a tumor model of AML.Immediately following the induction of a non-occlusive thrombosis via electrical stimulation (250 mAmp) of the IVC, cohorts of male C57BL/6J mice (n=5/group) were given a single SC injection of saline (0.1 mL); Uproleselan (40 mg/kg); or GMI-1687 (4 mg/kg, 0.4 mg/kg or 0.04 mg/kg), and twenty-four hrs post thrombus induction the IVC was harvested from all mice and thrombus weights were determined. Treatment with GMI-1687 decreased thrombus formation with significant inhibition at 0.04 mg/kg (92%, P

Description: E-selectin functions in venous thrombosis by binding and activating host cells to initiate the coagulation cascade. The E-selectin antagonist, Uproleselan (GMI-1271), has been shown to attenuate thrombus formation following electrical stimulation in a preclinical inferior vena cava (IVC) model without significantly affecting hemostasis (Culmer DL et al. Thromb Haemost. 117:1171-1181, 2017). A recent study has reported that galectin-3 (gal-3) and gal-3 binding protein are associated with murine thrombogenesis where they interact at the thrombus-vein wall interface, and that gal-3 may be contributing to thrombosis through proinflammatory mechanisms (Elise P et al. Blood 125:1813-1821, 2015). Collectively these studies suggest that pharmacologic targeting of both E-selectin and gal-3 function may afford a new class of therapeutics for treatment of venous thrombosis. In the present studies, we report on the activity of a novel glycomimetic compound with dual functional antagonism of E-selectin and gal-3, and demonstrate its anti-thrombotic activity in an IVC model. Binding affinities of GMI-1757 in solution or to immobilized E-selectin or gal-3 were determined by surface plasmon resonance (SPR) or microscale thermophoresis (MST), respectively. By SPR the mean (+/- SD) KD of GMI-1757 was 61 +/- 4 nM and 276 +/- 98 nM for E-selectin and gal-3, respectively (N=2 determinations). By MST the mean KD of GMI-1757 was 563 +/- 158 nM and 83 +/- 9 nM for E-selectin and gal-3, respectively (N=2-3 determinations). GMI-1757 was subsequently evaluated for its ability to inhibit binding of (a) sialyl Lea to immobilized human E-selectin and (b) gal-3 to an immobilized Galb1-3GlcNAc carbohydrate structure. The median IC50 of GMI-1757 for ligand binding of E-selectin or gal-3 was 800 nM (n=9 independent assays) and 110 nM (n=7 independent assays), respectively. The pharmacokinetics of GMI-1757 was evaluated following intravenous (IV) administration in male CD-1 mice at 5 mg/kg. Blood samples were collected up to 24 hours post dose, and GMI-1757 concentrations were determined with a qualified LC-MS/MS method. Following IV dosing of GMI 1757 at 5 mg/kg, the average half-life was 3.5 hours. Its average clearance rate was 1.39 +/- 0.180 L/hr/kg and the average volume of distribution was 1.35 +/- 0.651 L/kg. The in vivo activity of GMI-1757 following intraperitoneal (IP) administration was assessed in an acute model of IVC thrombosis.Immediately following the induction of a non-occlusive thrombosis via electrical stimulation (250 mAmp) of the IVC, cohorts of male C57BL/6J mice were given a single IP injection of saline (0.1 mL, n=9); Uproleselan (40 mg/kg, n=7); or GMI-1757 (40 mg/kg, n=6), and twenty-four hrs post thrombus induction the IVC was harvested from all mice and thrombus weights were determined. All treatments were well tolerated. The median thrombus weight from mice treated with Uproleselan was 0.0138 g and was significantly reduced from mice receiving saline (0.0208 g, P=0.004). Treatment with GMI-1757 resulted in a further reduction in median thrombus weight (0.0098 g) which was statistically differentiated from both saline and Uproleselan treatment (P= 0.002 and 0.049, respectively). Mechanism-of-action studies continue to be pursued to fully understand the impact of E-selectin and gal-3 inhibition in this model and other models where disease progression is dependent on both inflammation and fibrosis. In summary, an innovative dual antagonist of E-selectin and gal-3, GMI-1757, has been produced that demonstrates marked inhibition of thrombus formation in a murine IVC model following electrical stimulation. Figure. Figure. Disclosures Peterson: GlycoMimetics: Employment, Equity Ownership. Vohra:GlycoMimetics: Employment, Equity Ownership. Locatelli-Hoops:GlycoMimetics: Employment, Equity Ownership. Lee:GlycoMimetics: Employment, Equity Ownership. Deng:GlycoMimetics: Employment, Equity Ownership. Smith:GlycoMimetics: Employment, Equity Ownership. Fogler:GlycoMimetics: Employment, Equity Ownership. Magnani:GlycoMimetics: Employment, Equity Ownership, Membership on an entity's Board of Directors or advisory committees.