Despite heparin being the most widely used macromolecular drug, the design of small-molecule ligands to modulate its effects has been hampered by the structural properties of this polyanionic polysaccharide. Now a dynamic covalent selection approach is used to identify a new ligand for heparin, assembled from extremely simple building blocks. The amplified molecule strongly binds to heparin (K in the low μm range, ITC) by a combination of electrostatic, hydrogen bonding, and CH-π interactions as shown by NMR and molecular modeling. Moreover, this ligand reverts the inhibitory effect of heparin within an enzymatic cascade reaction related to blood coagulation. This study demonstrates the power of dynamic covalent chemistry for the discovery of new modulators of biologically relevant glycosaminoglycans.

Semaphorin3A (Sema3A) is a vertebrate-secreted protein that was initially characterized as a repulsive-guidance cue. Semaphorins have crucial roles in several diseases; therefore, the development of Sema3A inhibitors is of therapeutic interest. Sema3A interacts with glycosaminoglycans (GAGs), presumably through its C-terminal basic region. We used different biophysical techniques (i.e., NMR, surface plasmon resonance, isothermal titration calorimetry, fluorescence, and UV-visible spectroscopy) to characterize the binding of two Sema3A C-terminus-derived basic peptides (FS2 and NFS3) to heparin and chondroitin sulfate A. We found that these peptides bind to both GAGs with affinities in the low-micromolar range. On the other hand, a peptoid named SICHI (semaphorin-induced chemorepulsion inhibitor), which is positively charged at physiological pH, was first identified by our group as being able to block Sema3A chemorepulsion and growth-cone collapse in axons at the extracellular level. To elucidate the direct target for the reported SICHI inhibitory effect in the Sema3A signaling pathway, we looked first to the protein-protein interaction between secreted Sema3A and the Nrp1 receptor. However, our results show that SICHI does not bind directly to the Sema3A sema domain or to Nrp1 extracellular domains. We evaluated a new, to our knowledge, hypothesis, according to which SICHI binds to GAGs, thereby perturbing the Sema3A-GAG interaction. By using the above-mentioned techniques, we observed that SICHI binds to GAGs and competes with Sema3A C-terminus-derived basic peptides for binding to GAGs. These data support the ability of SICHI to block the biologically relevant interaction between Sema3A and GAGs, thus revealing SICHI as a new, to our knowledge, class of inhibitors that target the GAG-protein interaction.

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The 1,2,3-triazole ring has recently attracted a renewed interest as a structural scaffold with many applications in different fields. However, very often, the unambiguous assignment of the correct structure is not an easy task, and the development of robust characterization methodologies is needed. Herein, the three possible isomeric forms of disubstituted 1,2,3-triazole (1,4- or 1,5- or 2,4-disubstituted derivatives) have been characterized and distinguished by routine (1)H/(15)N gHMBC experiments at (15)N natural abundance. The calculated (GIAO-B3LYP/6-311++G**) (15)N NMR chemical shifts showed good agreement with the experimental values, further supporting their unambiguous structural characterization.

Apoptosis is a biological process relevant to different human diseases that is regulated through protein-protein interactions and complex formation. Peptidomimetic compounds based on linear peptoids and cyclic analogues with different ring sizes have been previously reported as potent apoptotic inhibitors. Among them, the presence of cis/trans conformers of an exocyclic tertiary amide bond in slow exchange has been characterized. This information encouraged us to perform an isosteric replacement of the amide bond by a 1,2,3-triazole moiety, in which different substitution patterns would mimic different amide rotamers. The syntheses of these restricted analogues have been carried out through an Ugi multicomponent reaction followed by an intramolecular cyclization. The unexpected formation of a β-lactam scaffold prompted us to study the course of the intramolecular cyclization of the Ugi adducts. In order to modulate this cyclization, a small library of compounds bearing both heterocyclic scaffolds has been synthesized and their activities as apoptosis inhibitors have been evaluated.