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Trastuzumab-monomethyl auristatin E conjugate exhibits potent cytotoxic activity in vitro against HER2-positive human breast cancer

Abdollahpour-Alitappeh, M. and Lotfinia, M. and Bagheri, N. and Sineh Sepehr, K. and Habibi-Anbouhi, M. and Kobarfard, F. and Balalaie, S. and Foroumadi, A. and Abbaszadeh-Goudarzi, G. and Abbaszadeh-Goudarzi, K. and Abolhassani, M. (2018) Trastuzumab-monomethyl auristatin E conjugate exhibits potent cytotoxic activity in vitro against HER2-positive human breast cancer. Journal of Cellular Physiology.

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Abstract

Targeted therapy using specific monoclonal antibodies (mAbs) conjugated to chemotherapeutic agents or toxins has become one of the top priorities in cancer therapy. Antibody–drug conjugates (ADCs) are emerging as a promising strategy for cancer-targeted therapy. In this study, trastuzumab, a humanized monoclonal anti-HER2 antibody, was reduced by dithiothreitol and conjugated to the microtubule-disrupting agent monomethyl auristatin E (MMAE) through a valine-citrulline peptide linker (trastuzumab-MC-Val-Cit-PABC-MMAE trastuzumab-vcMMAE). After conjugation, ADCs were characterized by using UV–vis, sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE), and flow cytometry. The antitumor activity of the ADC was evaluated in breast cancer cells in vitro. In addition, ADCs were further characterized using purification by the protein A chromatography, followed by assessment using apoptosis and MTT (3-(4,5-dimethylthiazol-2-yl)-2, 5-diphenyltetrazolium bromide) assays. Hydrophobic interaction chromatography was used to determine drug-to-antibody ratio species of ADCs produced. Our finding showed that approximately 5.12 drug molecules were conjugated to each mAb. H2L2, H2L, HL, H2, H, and L forms of ADCs were detected in nonreducing SDS-PAGE. The binding of trastuzumab-vcMMAE to HER2-positive cells was comparable with that of the parental mAb. The MTT assay showed that our ADCs induced significant cell death in HER2-positive cells, but not in HER2-negative cells. The ADCs produced was a mixture of species, unconjugated trastuzumab (14.147%), as well as trastuzumab conjugated with two (44.868%), four (16.886%), six (13.238%), and eight (10.861%) molecules of MMAE. These results indicated that MMAE-conjugated trastuzumab significantly increases the cytotoxic activity of trastuzumab, demonstrating high affinity, specificity, and antitumor activity in vitro. Trastuzumab-vcMMAE is an effective and selective agent for the treatment of HER2-positive breast tumors. © 2018 Wiley Periodicals, Inc.

Item Type: Article
Additional Information: cited By 0; Article in Press
Subjects: بیماریهای زنان WP
QU بیوشیمی
فارماکولوژی QV
> دانشکده داروسازی > فارماکولوژی QV

آسیب شناسی QZ
مقالات نمایه شده محققین دانشگاه در سایت ,Web of Science ,Scopus
Divisions: UNSPECIFIED
Depositing User: GOUMS
Date Deposited: 24 Dec 2018 08:28
Last Modified: 24 Dec 2018 08:28
URI: http://eprints.goums.ac.ir/id/eprint/9787

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