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Research Articles: Therapeutics, Targets, and Development

Target-specific cytotoxic activity of recombinant immunotoxin scFv(MUC1)-ETA on breast carcinoma cells and primary breast tumors

¹ Protein Engineering and Biotherapy, Tata Memorial Centre, Advanced Centre for Treatment Research and Education in Cancer, Navi Mumbai, India; ² Laboratory Nuclear Medicine Centre, Tata Memorial Hospital, Parel, Mumbai, India; and ³ Chemotherapeutisches Forschungsinstitut Georg-Speyer-Haus, Frankfurt am Main, Germany

Requests for reprints: Dilip Bandyopadhyay, Protein Engineering and Biotherapy, Advanced Centre for Treatment Research and Education in Cancer, Tata Memorial Centre, Navi Mumbai-410 208, India. E-mail: dilip.bandyopadhyay{at}gmail.com

Abstract

MUC1 is a mucin family protein, overexpressed in more than 90% of breast cancers in an underglycosylated form, exposing the core peptides of the extracellular domain that act as a potential target for antibody-mediated therapy. We have developed an anti-MUC1 scFv antibody from a phage library of mice immunized with synthetic peptide MUC1-variable number of tandem repeats. MUC1 binding phages were affinity selected through biopanning using a biotin-streptavidin pull-down method. The selected phage clones showed target-specific binding to MUC1-expressing cells. Fusion of truncated Pseudomonas aeruginosa exotoxin A (ETA) to a high binder, phage-derived scFv clone and bacterial expression and purification of recombinant scFv(MUC1)-ETA immunotoxin were done with good yield and purity. In vitro target-specific cytotoxic activity and target-specific binding of immunotoxin were shown on MUC1-expressing cells and primary breast tumor samples. A truncated ETA fusion protein expressed from the same vector but lacking scFv did not show cytotoxic effects, confirming target specificity. Our results suggest that the scFv(MUC1)-ETA immunotoxin has therapeutic potential and deserves further development and characterization for MUC1-specific breast cancers treatment. [Mol Cancer Ther 2007;6(2):562–9]

Grant support: Indian Council of Medical Research, Government of India; International Bureau of the Federal Ministry of Education and Research (BMBF), Government of Germany; Tata Memorial Centre (Mumbai, India); and Georg-Speyer-Haus (Frankfurt am Main, Germany).

The costs of publication of this article were defrayed in part by the payment of page charges. This article must therefore be hereby marked advertisement in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.

⁴ R. Singh and D. Bandyopadhyay, unpublished data.

Received 10/ 2/06; revised 11/ 4/06; accepted 12/15/06.