title: Surface plasmon resonance and NMR analyses of anti Tn-antigen MLS128 monoclonal antibody binding to two or three consecutive Tn-antigen clusters.
abstract: Tn-antigens are tumour-associated carbohydrate antigens that are involved in metastatic processes and are associated with a poor prognosis. MLS128 monoclonal antibody recognizes the structures of two or three consecutive Tn-antigens (Tn2 or Tn3). Since MLS128 treatment inhibits colon and breast cancer cell growth [Morita, N., Yajima, Y., Asanuma, H., Nakada, H., and Fujita-Yamaguchi, Y. (2009) Inhibition of cancer cell growth by anti-Tn monoclonal antibody MLS128. Biosci. Trends 3, 32-37.], understanding the interaction between MLS128 and Tn-clusters may allow us to the development of novel cancer therapeutics. Although MLS128 was previously reported to have specificity for Tn3 rather than Tn2, similar levels of Tn2/Tn3 binding were unexpectedly observed at 37°C. Thus, thermodynamic analyses were performed via surface plasmon resonance (SPR) using synthetic Tn2- and Tn3-peptides at 10, 15, 20, 25 and 30°C. SPR results revealed that MLS128's association constants for both antigens were highly temperature dependent. Below 25°C MLS128's association constant for Tn3-peptide was clearly higher than that for Tn2-peptide. At 30°C, however, the association constant for Tn2-peptide was higher than that for Tn3-peptide. This reversal of affinity is due to the sharp increase in K(d) for Tn3. These results were confirmed by NMR, which directly measured MLS128-Tn binding in solution. This study suggested that thermodynamic control plays a critical role in the interaction between MLS128/Tn2 and MLS128/Tn3.
authors.affiliation: Department of Applied Biochemistry, Tokai University School of Engineering, 4-1-1 Kitakaname, Hiratsuka, Kanagawa 259-1292, Japan.
authors.forename: Ayano
authors.initials: A
authors.lastname: Matsumoto-Takasaki
authors.forename: Shinya
authors.initials: S
authors.lastname: Hanashima
authors.forename: Ami
authors.initials: A
authors.lastname: Aoki
authors.forename: Noriyuki
authors.initials: N
authors.lastname: Yuasa
authors.forename: Haruhiko
authors.initials: H
authors.lastname: Ogawa
authors.forename: Reiko
authors.initials: R
authors.lastname: Sato
authors.forename: Hiroko
authors.initials: H
authors.lastname: Kawakami
authors.forename: Mamoru
authors.initials: M
authors.lastname: Mizuno
authors.forename: Hiroshi
authors.initials: H
authors.lastname: Nakada
authors.forename: Yoshiki
authors.initials: Y
authors.lastname: Yamaguchi
authors.forename: Yoko
authors.initials: Y
authors.lastname: Fujita-Yamaguchi
mesh_terms: D000911:Antibodies, Monoclonal / Q000276:immunology / Q000378:metabolism*; D000937:Antigen-Antibody Reactions; D015295:Antigens, Tumor-Associated, Carbohydrate / Q000276:immunology / Q000378:metabolism*; D002151:Calorimetry; D004797:Enzyme-Linked Immunosorbent Assay; D006020:Glycopeptides / Q000378:metabolism; D006801:Humans; D009682:Magnetic Resonance Spectroscopy / Q000379:methods*; D011485:Protein Binding; D020349:Surface Plasmon Resonance / Q000379:methods*; D013696:Temperature; D013816:Thermodynamics
chemical_list: D000911:Antibodies, Monoclonal; D015295:Antigens, Tumor-Associated, Carbohydrate; D006020:Glycopeptides; C040286:Tn antigen