Identification of allo- or orthosteric VHH/single-domain antibodies that enhance or block pathogen binding to Siglec-1 on dendritic cells

Abstract

O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=87 SRC="FIGDIR/small/695420v1_ufig1.gif" ALT="Figure 1"> View larger version (20K): org.highwire.dtl.DTLVardef@e85819org.highwire.dtl.DTLVardef@1eff6b4org.highwire.dtl.DTLVardef@12daa92org.highwire.dtl.DTLVardef@11959e5_HPS_FORMAT_FIGEXP M_FIG C_FIG BackgroundSiglec-1 (Sialoadhesin/CD169) is expressed on myeloid cells and plays a key role in host defences by capturing incoming sialylated-pathogens such as Campylobacter jejuni. However, binding to Siglec-1 has also been exploited by pathogens such as SARS-CoV-2 for further dissemination. ResultsHere we identified high-affinity VHHs also known as single-domain antibodies or Nanobodies that bind to Siglec-1 and allo- or orthosterically modulate ligand binding. VHH 2C2 was shown to bind directly to the ligand binding site of Siglec-1 and blocked binding of ganglioside liposomes and Campylobacter jejuni to monocyte-derived dendritic cells (moDCs) and ex vivo Siglec-1+ DCs. VHH 2C2 also blocked SARS-CoV-2 binding of moDCs. In contrast, the VHHs 1B5 and 1C1 interacted with Siglec-1 outside the ligand binding site and acted as positive allosteric modulators of Siglec-1 ligand interactions, as was illustrated by increased ganglioside liposome and Campylobacter jejuni binding by moDCs. Our data suggests that mechanistically, the VHH 1B5 and 1C1 interfere with the cis-binding sialic acids present on the Siglec-1-expressing cell and thereby enhance trans-interactions with ligands. ConclusionIn conclusion, we have isolated VHH that enhance or block Siglec-1 ligand binding to a variety of sialylated-pathogens enabling further interrogation of Siglec-1 function. Moreover, unlike conventional blocking antibodies targeting specific pathogens, Siglec-1 binding VHH could potentially serve as broad-spectrum pathogen blocking agents.