A novel approach to determining the affinity of protein-carbohydrate interactions employing adherent cancer cells grown on a biosensor surface.

Abstract

The development of biological agents for the treatment of solid tumours is an area of considerable activity. We are pursuing carbohydrate-binding proteins (lectins) in a strategy aimed at targeting cancer-associated changes in glycosylation. To evaluate lectin-cancer cell interactions we developed a novel cell biosensor in which binding events take place at the cell surface, more closely mimicking an in vivo system. Metastatic, SW620, and non-metastatic, SW480, colorectal cancer cells were grown on the surface of a tissue-culture compatible polystyrene coated biosensor chip and housed in a quartz crystal microbalance (QCM) apparatus, the kinetics of binding of a diverse range of lectins was evaluated. The lectin Helix pomatia agglutinin (HPA) has been shown to bind aggressive metastatic cancer and was produced in recombinant form (His- and RFP-tagged). The affinity of HPA was in the nanomolar range to the metastatic SW620 cells but was only in the micromolar range to the non-metastatic SW480. Overall, the dissociation constant (K(D)) of the lectins tested in the new cell biosensor system was an order of magnitude lower (nanomolar range) than has generally been reported with systems such as QCM/SPR. This new cell-biosensor enables molecular interactions to be studied in a more relevant environment. An intrinsic problem with developing new biological therapies is the difficulty in determining the affinity with which proteins will interact with intact cell surfaces. This methodology will be of interest to researchers developing new biological approaches for targeting cell surfaces in a wide range of diseases, including cancer.

• Authors: Diluka Peiris, Anatoliy Markiv, G Paul Curley, Miriam V Dwek
• Journal: Biosensors & bioelectronics (Biosens Bioelectron) Vol. 35 Issue 1 Pg. 160-166 (May 15 2012) ISSN: 1873-4235 [Electronic] England
• PMID: 22424753 (Publication Type: Journal Article, Research Support, Non-U.S. Gov't)
• Copyright: Crown Copyright © 2012. Published by Elsevier B.V. All rights reserved.

Chemical References

• Helix lectin
• Lectins
• Polystyrenes
• Recombinant Proteins
• Acetylgalactosamine

Topics

• Acetylgalactosamine (metabolism)
• Biosensing Techniques (methods)
• Carbohydrate Metabolism
• Cell Adhesion
• Cell Line, Tumor
• Cells, Immobilized
• Colorectal Neoplasms (metabolism, pathology)
• Humans
• Kinetics
• Lectins (metabolism)
• Microscopy, Confocal
• Polystyrenes
• Protein Binding
• Quartz Crystal Microbalance Techniques (methods)
• Recombinant Proteins (metabolism)
• Surface Properties