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Imaging of HER2-expressing tumours using a synthetic Affibody molecule containing the 99mTc-chelating mercaptoacetyl-glycyl-glycyl-glycyl (MAG3) sequence

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Abstract

Purpose

Expression of human epidermal growth factor receptor type 2 (HER2) in malignant tumours possesses well-documented prognostic and predictive value. Non-invasive imaging of expression can provide valuable diagnostic information, thereby influencing patient management. Previously, we reported a phage display selection of a small (about 7 kDa) protein, the Affibody molecule ZHER2:342, which binds HER2 with subnanomolar affinity, and demonstrated the feasibility of targeting of HER2-expressing xenografts using radioiodinated ZHER2:342. The goal of this study was to develop a method for 99mTc labelling of ZHER2:342 using the MAG3 chelator, which was incorporated into ZHER2:342 using peptide synthesis, and evaluate the targeting properties of the labelled conjugate.

Methods

MAG3-ZHER2:342 was assembled using Fmoc/tBu solid phase peptide synthesis. Biochemical characterisation of the agent was performed using RP-HPLC, ESI-MS, biosensor studies and circular dichroism. A procedure for 99mTc labelling in the presence of sodium/potassium tartrate was established. Tumour targeting was evaluated by biodistribution study and gamma camera imaging in xenograft-bearing mice. Biodistribution of 99mTc-MAG3-ZHER2:342 and 125I-para-iodobenzoate -ZHER2:342 was compared 6 h p.i.

Results

Synthetic MAG3-ZHER2:342 possessed an affinity of 0.2 nM for HER2 receptors. The peptide was labelled with 99mTc with an efficiency of about 75–80%. Labelled 99mTc-MAG3-ZHER2:342 retained capacity to bind specifically HER2-expressing SKOV-3 cells in vitro. 99mTc-MAG3-ZHER2:342 showed specific tumour targeting with a contrast similar to a radioiodinated analogue in mice bearing LS174T xenografts. Gamma camera imaging demonstrated clear and specific visualisation of HER2 expression.

Conclusion

Incorporation of a mercaptoacetyl-containing chelating sequence during chemical synthesis enabled site-specific 99mTc labelling of the ZHER2:342 Affibody molecule with preserved targeting capacity.

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Acknowledgements

This study was financially supported by Affibody AB and grants from the Swedish Cancer Society and Swedish Research Council (VR). We thank Veronika Eriksson at BMS and the animal facility staff of Rudbeck laboratory for technical assistance, and Gustav Sundqvist at KTH for running the ESI-MS analyses. We thank Dr. Joachim Feldwisch, Dr. Anders Wennborg and Dr. Lars Abrahmsén (Affibody AB) for interesting and inspiring discussions on the use of Affibody molecules for imaging and for comments on the manuscript.

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Authors and Affiliations

  1. School of Biotechnology, Division of Molecular Biotechnology, Royal Institute of Technology, Stockholm, Sweden

    Torun Engfeldt & Amelie Eriksson Karlström

  2. Biomedical Radiation Sciences, Rudbeck Laboratory, Uppsala University, Uppsala, Sweden

    Anna Orlova, Thuy Tran, Alexander Bruskin & Vladimir Tolmachev

  3. Affibody AB, Bromma, Sweden

    Anna Orlova & Vladimir Tolmachev

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

    Charles Widström

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  1. Torun Engfeldt
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  2. Anna Orlova
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  3. Thuy Tran
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  4. Alexander Bruskin
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  5. Charles Widström
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  6. Amelie Eriksson Karlström
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  7. Vladimir Tolmachev
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Correspondence to Vladimir Tolmachev.

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Engfeldt, T., Orlova, A., Tran, T. et al. Imaging of HER2-expressing tumours using a synthetic Affibody molecule containing the 99mTc-chelating mercaptoacetyl-glycyl-glycyl-glycyl (MAG3) sequence. Eur J Nucl Med Mol Imaging 34, 722–733 (2007). https://doi.org/10.1007/s00259-006-0266-4

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  • DOI: https://doi.org/10.1007/s00259-006-0266-4

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