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111In-Bn-DTPA-nimotuzumab with/without modification with nuclear translocation sequence (NLS) peptides: an Auger electron-emitting radioimmunotherapeutic agent for EGFR-positive and trastuzumab (Herceptin)-resistant breast cancer

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Abstract

Increased expression of epidermal growth factor receptors (EGFR) in breast cancer (BC) is often associated with trastuzumab (Herceptin)-resistant forms of the disease and represents an attractive target for novel therapies. Nimotuzumab is a humanized IgG1 monoclonal antibody that is in clinical trials for treatment of EGFR-overexpressing malignancies. We show here that nimotuzumab derivatized with benzylisothiocyanate diethylenetriaminepentaacetic acid for labelling with the subcellular range Auger electron-emitter, 111In and modified with nuclear translocation sequence (NLS) peptides (111In-NLS-Bn-DTPA-nimotuzumab) was bound, internalized and transported to the nucleus of EGFR-positive BC cells. Emission of Auger electrons in close proximity to the nucleus caused multiple DNA double-strand breaks which diminished the clonogenic survival (CS) of MDA-MB-468 cells that have high EGFR density (2.4 × 106 receptors/cell) to less than 3 %. 111In-Bn-DTPA-nimotuzumab without NLS peptide modification was sevenfold less effective for killing MDA-MB-468 cells. 111In-Bn-DTPA-nimotuzumab with/without NLS peptide modification were equivalently cytotoxic to MDA-MB-231 and TrR1 BC cells that have moderate EGFR density (5.4 × 105 or 4.2 × 105 receptors/cell, respectively) reducing their CS by twofold. MDA-MB-231 cells have intrinsic trastuzumab resistance due to low HER2 density, whereas TrR1 cells have acquired resistance despite HER2 overexpression. Biodistribution and microSPECT/CT imaging revealed that 111In-NLS-Bn-DTPA-nimotuzumab exhibited more rapid elimination from the blood and lower tumour uptake than 111In-Bn-DTPA-nimotuzumab. Tumour uptake of the radioimmunoconjugates in mice with MDA-MB-468 xenografts was high (8–16 % injected dose/g) and was blocked by administration of an excess of unlabelled nimotuzumab, demonstrating EGFR specificity. We conclude that 111In-Bn-DTPA-nimotuzumab with/without NLS peptide modification are promising Auger electron-emitting radioimmunotherapeutic agents for EGFR-positive BC, but 111In-Bn-DTPA-nimotuzumab may be preferred due to its higher tumour uptake in vivo.

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Acknowledgments

This study was supported by a grant from the Canadian Breast Cancer Research Alliance (No. 019513) with funds from the Canadian Cancer Society and by a grant from the Ontario Institute for Cancer Research (1 mm Cancer Challenge) with funds from the Province of Ontario. Humphrey Fonge was the recipient of a post-doctoral fellowship from the Ontario Ministry of Research and Innovation. Nimotuzumab was provided by YM Biosciences, Inc. through a Materials Transfer Agreement. Parts of this study were presented at the Breast Cancer Imaging: State-of-the-Art 2011 conference, National Institutes of Health, Bethesda, MD, April 21–22, 2011 and at the American Association for Cancer Research 101st annual meeting, April 17–21, 2010.

Conflicts of interest

RMR has a Materials Transfer Agreement with YM Biosciences, Inc. IT was an employee of YM Biosciences, Inc.

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Correspondence to Raymond M. Reilly.

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Aisha Fasih and Humphrey Fonge contributed equally to this study.

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Fasih, A., Fonge, H., Cai, Z. et al. 111In-Bn-DTPA-nimotuzumab with/without modification with nuclear translocation sequence (NLS) peptides: an Auger electron-emitting radioimmunotherapeutic agent for EGFR-positive and trastuzumab (Herceptin)-resistant breast cancer. Breast Cancer Res Treat 135, 189–200 (2012). https://doi.org/10.1007/s10549-012-2137-y

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  • DOI: https://doi.org/10.1007/s10549-012-2137-y

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