Rational design of heparin antagonist: Guanidine-based mimetics unveil key carbohydrate-carbohydrate interactions

Small-molecule inhibitors targeting heparin (HP)-protein interactions represent a promising strategy for developing therapeutic agents against serious bleeding complications. Herein, we report a rational design and synthesis of a library of eight trisaccharide HP mimetics incorporating positively charged guanidinium residues aimed at disrupting the ionic interactions of HP and modulating HP-mediated biological activities. The introduction of guanidine residue in HP backbone significantly influenced the conformational plasticity of l-idose and l-iduronic acid, shifting the major ⁴C₁-conformation to predominant ²S₀-geometries, akin to the role of high sulfation in native HS. Unlike aminoglycosides, the guanidine-based HP mimetics exhibited no antibacterial activity and demonstrated low cytotoxicity towards both cancerous and normal cell lines. When evaluated as potential antidotes for heparin and fondaparinux-mediated blood coagulation, the highly guanidine-substituted HP mimetics displayed sub-micromolar antagonist potency. NMR studies further confirmed the carbohydrate-carbohydrate interactions between fondaparinux and the HP mimetics, providing a mechanistic basis for the observed activity and introducing a new strategy to block HP-mediated biological functions.