Perspectives on Anti-Glycan Antibodies Gleaned from Development of a Community Resource Database

Abstract

Antibodies are used extensively for a wide range of basic research and clinical applications. While an abundant and diverse collection of antibodies to protein antigens have been developed, good monoclonal antibodies to carbohydrates are much less common. Moreover, it can be difficult to determine if a particular antibody has the appropriate specificity, which antibody is best suited for a given application, and where to obtain that antibody. Herein, we provide an overview of the current state of the field, discuss challenges for selecting and using antiglycan antibodies, and summarize deficiencies in the existing repertoire of antiglycan antibodies. This perspective was enabled by collecting information from publications, databases, and commercial entities and assembling it into a single database, referred to as the Database of Anti-Glycan Reagents (DAGR). DAGR is a publicly available, comprehensive resource for anticarbohydrate antibodies, their applications, availability, and quality.

Figure 1

Applications of anticarbohydrate antibodies in research and clinical therapy. Antiglycan antibodies have been used in the detection and discovery of glycoantigens in various tumor samples. Antiglycan antibodies are also used as diagnostic tools (CA19–9 levels in pancreatic cancer patients) and as therapeutics, such as Unituxin (ch14.18) in the treatment of neuroblastoma.

Figure 2

Cartoon representations of the major mammalian biosynthetic carbohydrate families.

Figure 3

Structural variations of the sialyl Tn (STn) antigen. An antibody targeting the sialyl Tn (STn) antigen may target one of many structurally similar glycans. For example, the nitrogen on the sialic acid residue can be modified with an acetyl group (Ac) or a glycolyl group (Gc). Additionally, O-acetylated variations can occur at numerous hydroxyl positions. The GalNAc residue can be attached to either serine or threonine. Finally, there may be a single STn residue on a peptide chain or there maybe additional STn residues in close proximity on the peptide chain (clustered STn). All of these structures could be described generally as STn.

Figure 4

Graphical representations of the DAGR entries. (a) Antibodies that target branched glycan structures (blue) are well represented in the DAGR. Antibodies targeting sialic acid charged epitopes are well represented; however sulfated or uronic acid containing glycan epitopes are conspicuously underrepresented (red). Common monosaccharides are present in a significant percentage of the epitopes targeted (green). (b) Cell/cell lysate (blue) and glycoprotein (red) immunizations are the preferred means of generating antibodies to glycans. Glycolipid (green), tissue preparation (purple), and synthetic antigen (orange) are other immunizations strategies. Antibodies to glycans have also been prepared as human isolates (pink) and from infected animals (black).

Figure 5

Multivalency influence on binding affinity and specificity. For many antiglycan antibodies, formation of a multivalent complex (a) is required for a high avidity interaction. Antibody formats with a single binding site (b) often have poor affinity and/or specificity. If the target epitope is present with the wrong spacing (c) or orientation (d) to form a multivalent complex, an antiglycan antibody may not bind the sample of interest. These factors must be considered when selecting an antibody and interpreting results.

Figure 6

Snapshots of the Database for Anti-Glycan Reagents (DAGR) interface. (a) The advanced search option allows the user to search based upon a variety of criteria such as clone name, epitope name, sequence or family, and commercial availability. (b) Following the selection of a particular reagent, available information on immunogen, host species, and hybridoma availability are provided. References to significant literature in antibody production (not shown) are also provided.