β-D-Glucosyl Conjugates of Highly Potent Inhibitors of Blood Coagulation Factor Xa Bearing 2-Chorothiophene as a P1 Motif

We synthesized a novel O-glucoside of the recently reported potent factor Xa (fXa) inhibitor 1, which bears a 5-chlorothien-2-yl moiety and 1-isopropylpiperidine as fragments that bind the S1 and S4 enzyme pockets, respectively. A β-D-glucosyl unit was conjugated through an ether-linked C3-alkyl spacer to the central phenyl ring of 1. The synthesized β-D-glucose-based compound 16 achieved picomolar inhibitory potency against human fXa (K_i=60pM) and high selectivity over thrombin and other serine proteases. In addition to the chlorothienyl S1 binder, a large gain in ΔG resulted from the addition of protonated 1-isopropylpiperidine (ΔΔG=29.7-30.5kJmol^-1), which should bind to the aromatic S4 pocket through efficient cation-π and CH{dot operator}{dot operator}{dot operator}π interactions. Instead, the C3-alkyl-linked glucose fragment, which is likely directed toward the solvent outside the enzyme binding site, improves ΔG by an average of 2.9-3.8kJmol^-1. Compound 16 showed sub-micromolar invitro anticoagulant activity, as assessed by prothrombin time (PT) and activated thromboplastin time (aPTT) clotting assays in pooled human plasma (PT₂ and aPTT₂ equal to 0.135 and 0.389μM, respectively). Although compound 16 was 1.4-fold less active than parent compound 1 in the exvivo anticoagulant assay in mice, it showed a significant (1.6-fold) prolongation of PT relative to controls (P<0.05) 60min after oral dosing (75mgkg^-1).