On-Chip Synthesis and Screening of a Sialoside Library Yields a High Affinity Ligand for Siglec-7

Abstract

The Siglec family of sialic acid-binding proteins are differentially expressed on white blood cells of the immune system and represent an attractive class of targets for cell-directed therapy. Nanoparticles decorated with high-affinity Siglec ligands show promise for delivering cargo to Siglec-bearing cells, but this approach has been limited by a lack of ligands with suitable affinity and selectivity. Building on previous work employing solution-phase sialoside library synthesis and subsequent microarray screening, we herein report a more streamlined ‘on-chip’ synthetic approach. By printing a small library of alkyne sialosides and subjecting these to ‘on-chip’ click reactions, the largest sialoside analog library to date was generated. Siglec-screening identified a selective Siglec-7 ligand, which when displayed on liposomal nanoparticles, allows for targeting of Siglec-7+ cells in peripheral human blood. In silico docking to the crystal structure of Siglec-7 provides a rational for the affinity gains observed for this novel sialic acid analog.

Sialic Acid Glycoengineering Using an Unnatural Sialic Acid for the Detection of Sialoglycan Biosynthesis Defects and On-Cell Synthesis of Siglec Ligands

“…In an on-chip glycan array screen, Rillahan et al have found that clicking distinct azide groups to sialic acids can strongly enhance their binding affinity to Siglec-7. 49 From this on-chip screen, we have selected three azides derivatives (azide1−3) that either strongly enhance Siglec-7 binding (azide 1), led to an intermediate increase in Siglec-7 binding (azide 2), or did not enhance Siglec-7 binding (azide 3) (Figure 5a). 49 THP-1 or Jurkat cells were treated with Ac 4 ManNPoc or Ac 5 NeuNPoc, and the three different azides were clicked to the Poc-modified glycans using CuAAC.…”

Disubstituted sialic acid ligands targeting siglecs CD33 and CD22 associated with myeloid leukaemias and B cell lymphomas

“…cellular autofluorescence). For hCD33-ligand specificity analysis with overexpressing recombinant cell lines, 28, 31, 32 1% ligand-displaying liposomes were used. This panel of cell lines consists of CHO cells expressing mSn, hSig3, hSig5, hSig8, hSig9, and hSig10, Jurkat cells expressing hSig7, and Ramos cells expressing hCD22 (other B-cell lines were found to express additional siglecs, data not shown).…”

Glycoarrays: An Invaluable Tool for Glycomics

“…Modified sialic acids with alkyne groups in positions five and nine were immobilized in NHS modified glass slides, and subjected to CuAAC reactions with 47 different azido compounds. The modified sialic acids generated on chip were screened towards Siglec-7 and Siglec-10, permitting characterization of a high affinity ligand for Siglec-7 [49].…”

“…First examples in this direction are the enzymatic on-chip diversification of glycan scaffolds in nano-droplets [17], or the on-chip synthesis of potential Siglec antagonists employing click reactions for the introduction of a second library of structural substituents [49].…”