Synthesis and Binding Mode Predictions of Novel Siglec-7 Ligands.

Siglec-7 regulates immune cell activity and is a promising target for immunomodulation. Here, we report the discovery of novel sialic acid derivatives binding to Siglec-7. Synthesis and affinity measurements are complemented by high-quality models of sialoside-Siglec-7 complexes based on molecular dynamics (MD) simulations on the microsecond time scale. We provide details for the predicted binding modes for the new ligands, e.g., that an extension of the carbon backbone leads to a different molecular interaction pattern with the receptor and the nearby water structure than found for known Siglec-7 ligands. Further on, we uncover some shortcomings of the GLYCAM06 and GAFF2 force fields when used for the simulation of sialoside-based glycomimetics. Our results open new opportunities for the rational design of Siglec-7 inhibitors. In addition, we provide strategies on how to use and visualize MD simulations to describe and investigate sialoside-Siglec complexes in general.

Targeting Human CD22/Siglec-2 with Dimeric Sialosides as Novel Oligosaccharide Mimetics.

Significant interest in the development of high-affinity ligands for Siglecs exists due to the various therapeutically relevant functions of these proteins. Here, we report a new strategy to develop and design Siglec ligands as disialyl-oligosaccharide mimetics exemplified on Siglec-2 (CD22). We report insights into development of dimeric ligands with high affinity and avidity to cell surface-expressed CD22, assay development, tool compounds, structure activity relationships, and biological data on calcium flux regulation in B-cells. The binding modes of selected ligands have been modeled based on state-of-the-art molecular dynamics simulations on the microsecond timescale, providing detailed views on ligand binding and opening a new perspective on drug design efforts for Siglecs. High-avidity dimeric ligands containing a linker opening the way towards bispecifics are presented as well.

New Human CD22/Siglec-2 Ligands with a Triazole Glycoside.

CD22 is a member of the Siglec family. Considerable attention has been drawn to the design and synthesis of new Siglec ligands to explore target biology and innovative therapies. In particular, CD22-ligand-targeted nanoparticles with therapeutic functions have proved successful in preclinical settings for blood cancers, autoimmune diseases, and tolerance induction. Here we report the design, synthesis and affinity evaluation of a new class of Siglec ligands: namely sialic acid derivatives with a triazole moiety replacing the natural glycoside oxygen atom. In addition, we describe important and surprising differences in binding to CD22 expressed at the cell surface for compounds with distinct valences. The new class of compounds might serve as a template for the design of ligands for other members of the Siglec family and next-generation CD22-ligand-based targeted therapies.

Design, Synthesis, and Biological Evaluation of Small, High-Affinity Siglec-7 Ligands: Toward Novel Inhibitors of Cancer Immune Evasion.

Natural killer cells are able to directly lyse tumor cells, thereby participating in the immune surveillance against cancer. Unfortunately, many cancer cells use immune evasion strategies to avoid their eradication by the immune system. A prominent escape strategy of malignant cells is to camouflage themselves with Siglec-7 ligands, thereby recruiting the inhibitory receptor Siglec-7 expressed on the NK cell surface which subsequently inhibits NK-cell-mediated lysis. Here we describe the synthesis and evaluation of the first, high-affinity low molecular weight Siglec-7 ligands to interfere with cancer cell immune evasion. The compounds are Sialic acid derivatives and bind with low micromolar Kd values to Siglec-7. They display up to a 5000-fold enhanced affinity over the unmodified sialic acid scaffold αMe Neu5Ac, the smallest known natural Siglec-7 ligand. Our results provide a novel immuno-oncology strategy employing natural immunity in the fight against cancers, in particular blocking Siglec-7 with low molecular weight compounds.

Synthesis and biological evaluation of 9-N-oxamyl sialosides as Siglec-7 ligands.

Siglecs (sialic acid recognizing immunoglobulin like lectins) are a family of lectins with specificity for sialic acid containing carbohydrates. Synthetic sialic acid derivatives with high affinity proved useful to unravel the biological role of the ligand binding domain, although many of their functions in immunity remain unknown. Here we present design, synthesis, affinity evaluation and molecular modeling of novel 9-N-oxamoyl modified sialosides as Siglec-7 ligands.

Discovery of multifold modified sialosides as human CD22/Siglec-2 ligands with nanomolar activity on B-cells.

Sialic acids are abundant in higher domains of life and lectins recognizing sialosaccharides are heavily involved in the regulation of the human immune system. Modified sialosides are useful tools to explore the functions of those lectins, especially members of the Siglec (sialic acid binding immunoglobulin like lectin) family. Here we report design, synthesis, and affinity evaluation of novel sialoside classes with combined modification at positions 2, 4, and 9 or 2, 3, 4, and 9 of the sialic acid scaffold as human CD22 (human Siglec-2) ligands. They display up to 7.5 × 10(5)-fold increased affinity over αMe Neu5Ac (the minimal Siglec ligand). CD22 is a negative regulating coreceptor of the B-cell receptor (BCR). In vitro experiments with a human B-lymphocyte cell line showed functional blocking of CD22 upon B-cell receptor (BCR) stimulation in the presence of nanomolar concentrations of the novel ligands. The observed increased Ca(2+) response corresponds to enhanced cell activation, providing an opportunity to therapeutically modulate B-lymphocyte responses, e.g., in immune deficiencies and infections.