Target Selectivity of FimH Antagonists

Mannose-based FimH antagonists are considered new therapeutics for the treatment of urinary tract infections (UTIs). They prevent the adhesion of uropathogenic <i>Escherichia coli</i> (UPEC) to urothelial cell surfaces triggered by the lectin FimH, which is located at the tip of bacterial type 1 pili. Because all reported FimH antagonists are α-d-mannosides, they are also potential ligands of mannose receptors of the human host system. We therefore investigated the selectivity range of five FimH antagonists belonging to different compound families by comparing their affinities for FimH and eight human mannose receptors. On the basis of the detected selectivity range of approximately 5 orders of magnitude, no adverse side effects resulting from nonselective binding to the human receptors have to be expected. FimH antagonists can therefore be further considered as potential therapeutics for the treatment of UTI

Nanomolar E‑Selectin Antagonists with Prolonged Half-Lives by a Fragment-Based Approach

Selectins, a family of C-type lectins, play a key role in inflammatory diseases (e.g., asthma and arthritis). However, the only millimolar affinity of sialyl Lewis^x (sLe^x), which is the common tetrasaccharide epitope of all physiological selectin ligands, has been a major obstacle to the development of selectin antagonists for therapeutic applications. In a fragment-based approach guided by NMR, ligands binding to a second site in close proximity to a sLe^x mimic were identified. A library of antagonists obtained by connecting the sLe^x mimic to the best second-site ligand via triazole linkers of different lengths was evaluated by surface plasmon resonance. Detailed analysis of the five most promising candidates revealed antagonists with <i>K</i>_D values ranging from 30 to 89 nM. In contrast to carbohydrate–lectin complexes with typical half-lives (<i>t</i>_1/2) in the range of one second or even less, these fragment-based selectin antagonists show <i>t</i>_1/2 of several minutes. They exhibit a promising starting point for the development of novel anti-inflammatory drugs

Label-Free FimH Protein Interaction Analysis Using Silicon Nanoribbon BioFETs

The detection of biomarkers at very low concentration and low cost is increasingly important for clinical diagnosis. Moreover, monitoring affinities for receptor-antagonist interactions by time-resolved measurements is crucial for drug discovery and development. Biosensors based on ion-sensitive field-effect transistors (BioFETs) are promising candidates for being integrated into CMOS structures and cost-effective production. The detection of DNA and proteins with silicon nanowires has been successfully demonstrated using high affinity systems such as the biotin–streptavidin interaction. Here, we show the time-resolved label-free detection of the interaction of the bacterial FimH lectin with an immobilized mannose ligand on gold-coated silicon nanoribbon BioFETs. By comparing our results with a commercial state of the art surface plasmon resonance system, additional surface effects become visible when using this charge based detection method. Furthermore, we demonstrate the effect of sensor area on signal-to-noise ratio and estimate the theoretical limit of detection

Stabilization of Branched Oligosaccharides: Lewis^x Benefits from a Nonconventional C–H···O Hydrogen Bond

Although animal lectins usually show a high degree of specificity for glycan structures, their single-site binding affinities are typically weak, a drawback which is often compensated in biological systems by an oligovalent presentation of carbohydrate epitopes. For the design of monovalent glycomimetics, structural information regarding solution and bound conformation of the carbohydrate lead represents a valuable starting point. In this paper, we focus on the conformation of the trisaccharide Le^x (Gal­[Fucα(1–3)]­β(1–4)­Glc<i>N</i>Ac). Mainly because of the unfavorable tumbling regime, the elucidation of the solution conformation of Le^x by NMR has only been partially successful so far. Le^x was therefore attached to a ¹³C,¹⁵N-labeled protein. ¹³C,¹⁵N-filtered NOESY NMR techniques at ultrahigh field allowed increasing the maximal NOE enhancement, resulting in a high number of distance restraints per glycosidic bond and, consequently, a well-defined structure. In addition to the known contributors to the conformational restriction of the Le^x structure (exoanomeric effect, steric compression induced by the <i>N</i>HAc group adjacent to the linking position of l-fucose, and the hydrophobic interaction of l-fucose with the β-face of d-galactose), a nonconventional C–H···O hydrogen bond between H–C(5) of l-fucose and O(5) of d-galactose was identified. According to quantum mechanical calculations, this C–H···O hydrogen bond is the most prominent factor in stabilization, contributing 40% of the total stabilization energy. We therefore propose that the nonconventional hydrogen bond contributing to a reduction of the conformational flexibility of the Le^x core represents a novel element of the glycocode. Its relevance to the stabilization of related branched oligosaccharides is currently being studied

Antiadhesion Therapy for Urinary Tract InfectionsA Balanced PK/PD Profile Proved To Be Key for Success

The initial step for the successful establishment of urinary tract infections (UTIs), predominantly caused by uropathogenic Escherichia coli, is the adhesion of bacteria to urothelial cells. This attachment is mediated by FimH, a mannose-binding adhesin, which is expressed on the bacterial surface. To date, UTIs are mainly treated with antibiotics, leading to the ubiquitous problem of increasing resistance against most of the currently available antimicrobials. Therefore, new treatment strategies are urgently needed, avoiding selection pressure and thereby implying a reduced risk of resistance. Here, we present a new class of highly active antimicrobials, targeting the virulence factor FimH. When the most potent representative, an indolinylphenyl mannoside, was administered in a mouse model at the low dosage of 1 mg/kg (corresponding to approximately 25 μg/mouse), the minimal therapeutic concentration to prevent UTI was maintained for more than 8 h. In a treatment study, the colony-forming units in the bladder could be reduced by almost 4 orders of magnitude, comparable to the standard antibiotic treatment with ciprofloxacin (8 mg/kg, sc)

Low Molecular Weight Antagonists of the Myelin-Associated Glycoprotein: Synthesis, Docking, and Biological Evaluation

The injured adult mammalian central nervous system is an inhibitory environment for axon regeneration due to specific inhibitors, among them the myelin-associated glycoprotein (MAG), a member of the Siglec family (sialic-acid binding immunoglobulin-like lectin). In earlier studies, we identified the lead structure 5, which shows a 250-fold improved in vitro affinity for MAG compared to the tetrasaccharide binding epitope of GQ1bα (1), the best physiological MAG ligand described so far. By modifying the 2- and 5-position, the affinity of 5 could be further improved to the nanomolar range (→19a). Docking studies to a homology model of MAG allowed the rationalization of the experimental binding properties. Finally, pharmacokinetic parameters (stability in the cerebrospinal fluid, logD and permeation through the BBB) indicate the drug-like properties of the high-affinity antagonist 19a

Sweet Drugs for Bad Bugs: A Glycomimetic Strategy against the DC-SIGN-Mediated Dissemination of SARS-CoV‑2

The C-type lectin receptor DC-SIGN is a pattern recognition receptor expressed on macrophages and dendritic cells. It has been identified as a promiscuous entry receptor for many pathogens, including epidemic and pandemic viruses such as SARS-CoV-2, Ebola virus, and HIV-1. In the context of the recent SARS-CoV-2 pandemic, DC-SIGN-mediated virus dissemination and stimulation of innate immune responses has been implicated as a potential factor in the development of severe COVID-19. Inhibition of virus binding to DC-SIGN, thus, represents an attractive host-directed strategy to attenuate overshooting innate immune responses and prevent the progression of the disease. In this study, we report on the discovery of a new class of potent glycomimetic DC-SIGN antagonists from a focused library of triazole-based mannose analogues. Structure-based optimization of an initial screening hit yielded a glycomimetic ligand with a more than 100-fold improved binding affinity compared to methyl α-d-mannopyranoside. Analysis of binding thermodynamics revealed an enthalpy-driven improvement of binding affinity that was enabled by hydrophobic interactions with a loop region adjacent to the binding site and displacement of a conserved water molecule. The identified ligand was employed for the synthesis of multivalent glycopolymers that were able to inhibit SARS-CoV-2 spike glycoprotein binding to DC-SIGN-expressing cells, as well as DC-SIGN-mediated trans-infection of ACE2+ cells by SARS-CoV-2 spike protein-expressing viruses, in nanomolar concentrations. The identified glycomimetic ligands reported here open promising perspectives for the development of highly potent and fully selective DC-SIGN-targeted therapeutics for a broad spectrum of viral infections

FimH Antagonists for the Oral Treatment of Urinary Tract Infections: From Design and Synthesis to in Vitro and in Vivo Evaluation

Urinary tract infection (UTI) by uropathogenic Escherichia coli (UPEC) is one of the most common infections, particularly affecting women. The interaction of FimH, a lectin located at the tip of bacterial pili, with high mannose structures is critical for the ability of UPEC to colonize and invade the bladder epithelium. We describe the synthesis and the in vitro/in vivo evaluation of α-d-mannosides with the ability to block the bacteria/host cell interaction. According to the pharmacokinetic properties, a prodrug approach for their evaluation in the UTI mouse model was explored. As a result, an orally available, low molecular weight FimH antagonist was identified with the potential to reduce the colony forming units (CFU) in the urine by 2 orders of magnitude and in the bladder by 4 orders of magnitude. With FimH antagonist 16b, the great potential for the effective treatment of urinary tract infections with a new class of orally available antiinfectives could be demonstrated