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Infection and Immunity, July 2005, p. 4054-4061, Vol. 73, No. 7
0019-9567/05/$08.00+0     doi:10.1128/IAI.73.7.4054-4061.2005
Copyright © 2005, American Society for Microbiology. All Rights Reserved.

Characterization of a Human Monoclonal Antibody against Shiga Toxin 2 Expressed in Chinese Hamster Ovary Cells

D. E. Akiyoshi, C. M. Rich, S. O'Sullivan-Murphy, L. Richard, J. Dilo, A. Donohue-Rolfe, A. S. Sheoran, S. Chapman-Bonofiglio, and S. Tzipori*

Division of Infectious Diseases, Tufts University School of Veterinary Medicine, North Grafton, Massachusetts 01536

Received 17 September 2004/ Returned for modification 26 October 2004/ Accepted 2 March 2005

Shiga toxin-producing Escherichia coli infections can often lead to the development of hemolytic-uremic syndrome (HUS) in a small percentage of infected humans. Patients with HUS receive only supportive treatment as the benefit of antibiotic therapy remains uncertain. We have previously reported the generation and preclinical evaluation of neutralizing human monoclonal antibodies (HuMAbs) against the Shiga toxins (Stx). In this paper, we describe the expression in Chinese hamster ovary (CHO) cells of 5C12 HuMAb, which is directed against the A subunit of Stx2. The cDNAs of the light and heavy chain immunoglobulin (Ig) variable regions of 5C12 HuMAb were isolated and cloned into an expression vector containing human IgG1 constant regions. The vector was transfected into CHO cells, and transfectants secreting Stx2-specific antibody were screened by an Stx2-specific enzyme-linked immunosorbent assay. The CHO-produced recombinant 5C12 (r5C12) showed similar specificity and binding affinity to Stx2 as the parent hybridoma-produced 5C12. More significantly, the r5C12 displayed the same neutralizing activity as the parent 5C12 in vitro and in vivo. In the mouse toxicity model, both antibodies significantly and equally prolonged survival at a dose of 0.312 µg/mouse. The data showed that since r5C12, produced in CHO cells, was equally effective as the parent 5C12, it is our choice candidate as a potential prophylactic or therapeutic agent against hemolytic-uremic syndrome.

* Corresponding author. Mailing address: Tufts University School of Veterinary Medicine, 200 Westboro Road, Building 20, North Grafton, MA 01536. Phone: (508) 839-7955. Fax: (508) 839-7977. E-mail: saul.tzipori{at}tufts.edu.

Editor: J. D. Clements

Infection and Immunity, July 2005, p. 4054-4061, Vol. 73, No. 7
0019-9567/05/$08.00+0     doi:10.1128/IAI.73.7.4054-4061.2005
Copyright © 2005, American Society for Microbiology. All Rights Reserved.





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