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Imaging of EGFR expression in murine xenografts using site-specifically labelled anti-EGFR 111In-DOTA-ZEGFR:2377 Affibody molecule: aspect of the injected tracer amount

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Abstract

Introduction

Overexpression of epidermal growth factor receptor (EGFR) is a prognostic and predictive biomarker in a number of malignant tumours. Radionuclide molecular imaging of EGFR expression in cancer could influence patient management. However, EGFR expression in normal tissues might complicate in vivo imaging. The aim of this study was to evaluate if optimization of the injected protein dose might improve imaging of EGFR expression in tumours using a novel EGFR-targeting protein, the DOTA-ZEGFR:2377 Affibody molecule.

Methods

An anti-EGFR Affibody molecule, ZEGFR:2377, was labelled with 111In via the DOTA chelator site-specifically conjugated to a C-terminal cysteine. The affinity of DOTA-ZEGFR:2377 for murine and human EGFR was measured by surface plasmon resonance. The cellular processing of 111In-DOTA-ZEGFR:2377 was evaluated in vitro. The biodistribution of radiolabelled Affibody molecules injected in a broad range of injected Affibody protein doses was evaluated in mice bearing EGFR-expressing A431 xenografts.

Results

Site-specific coupling of DOTA provided a uniform conjugate possessing equal affinity for human and murine EGFR. The internalization of 111In-DOTA-ZEGFR:2377 by A431 cells was slow. In vivo, the conjugate accumulated specifically in xenografts and in EGFR-expressing tissues. The curve representing the dependence of tumour uptake on the injected Affibody protein dose was bell-shaped. The highest specific radioactivity (lowest injected protein dose) provided a suboptimal tumour-to-blood ratio. The results of the biodistribution study were confirmed by γ-camera imaging.

Conclusion

The 111In-DOTA-ZEGFR:2377 Affibody molecule is a promising tracer for radionuclide molecular imaging of EGFR expression in malignant tumours. Careful optimization of protein dose is required for high-contrast imaging of EGFR expression in vivo.

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Acknowledgments

This study was supported by grants from the Swedish Cancer Society (Cancerfonden) and the Swedish Research Council (Vetenskapsrådet). We thank Veronika Eriksson and the staff of the animal facility at Rudbeck Laboratory for technical assistance.

Disclosures

The authors, Orlova Anna, Helena Wållberg and Vladimir Tolmachev had earlier, and Daniel Rosik, Anna Sjöberg, Monika Hansson, Anders Wennborg have currently an affiliation (employment) with Affibody AB, Bromma, Sweden, which holds the intellectual property rights and trademarks for Affibody molecules.

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Authors and Affiliations

  1. Unit of Biomedical Radiation Sciences, Rudbeck Laboratory, Uppsala University, 751 81, Uppsala, Sweden

    Vladimir Tolmachev & Anna Orlova

  2. Affibody AB, Bromma, Sweden

    Daniel Rosik, Anna Sjöberg, Monika Hansson & Anders Wennborg

  3. School of Biotechnology, Royal Institute of Technology, Stockholm, Sweden

    Helena Wållberg

  4. Department of Medical Physics, Uppsala University Hospital, Uppsala, Sweden

    Mattias Sandström

  5. Department of Medical Sciences, Nuclear Medicine, Uppsala University, Uppsala, Sweden

    Vladimir Tolmachev

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  1. Vladimir Tolmachev
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Correspondence to Vladimir Tolmachev.

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Tolmachev, V., Rosik, D., Wållberg, H. et al. Imaging of EGFR expression in murine xenografts using site-specifically labelled anti-EGFR 111In-DOTA-ZEGFR:2377 Affibody molecule: aspect of the injected tracer amount. Eur J Nucl Med Mol Imaging 37, 613–622 (2010). https://doi.org/10.1007/s00259-009-1283-x

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