Location matters: functional analysis of monoclonal antibodies to various regions in glypican-3

Tuesday, September 23, 2014 — Poster Session III
12:00 p.m. – 2:00 p.m.	FAES Academic Center	NCI	PHARM-5

Authors

• YF. Zhang
• M. Feng
• Z. Tang
• M. Ho

Abstract

Glypican-3 (GPC3) is a cell surface proteoglycan that is overexpressed in hepatocellular carcinoma and is a potential target for antibody therapy. Our lab has generated a group of monoclonal antibodies that recognize different epitopes in GPC3. One (HN3) targets a conformational epitope while several others (e.g., YP7) bind the C terminus. By comparing their Fv-pseudomonas exotoxin fusion proteins (immunotoxin), we found that the anti-C terminus immunotoxins had stronger tumor cell binding but weaker cytotoxicity than the HN3 immunotoxin. In addition, although anti-C terminus immunotoxin can bind GPC3 both inside and outside of the detergent resistant membrane (DRM), the HN3 antibody can only bind GPC3 that is outside of the DRM, suggesting that the endocytosis of HN3 may be mediated by clatharin rather than caveolin. In cell surface co-immunoprecipitation, the anti-C terminus immunotoxins can pull down unglycosylated, lightly glycosylated and heavily glycosylated GPC3, whereas the HN3 immunotoxin can only pull down lightly glycosylated surface GPC3, which are located outside of DRM. This evidence showed that the HN3 immunotoxin targets a unique portion of cell surface GPC3, which may have a higher internalization rate than other GPC3 molecules. This character makes the HN3 more suitable for immunotoxins or potentially antibody drug conjugates.

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