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On the Selection of a Tracer for PET Imaging of HER2-Expressing Tumors: Direct Comparison of a (124)I-Labeled Affibody Molecule and Trastuzumab in a Murine Xenograft Model
2009 (English)In: Journal of Nuclear Medicine, ISSN 0161-5505, E-ISSN 1535-5667, Vol. 50, no 3, 417-425 p.Article in journal (Refereed) Published
Abstract [en]

Human epidermal growth factor receptor type 2 (HER2) is a tyrosine kinase, which is often overexpressed in many carcinomas. Imaging HER2 expression in malignant tumors can provide important prognostic and predictive diagnostic information. The use of anti-HER2 tracers labeled with positron-emitting radionuclides may increase the sensitivity of HER2 imaging. The goal of this study was to compare directly 2 approaches for developing anti-HER2 PET tracers: a (124)I-labeled monoclonal antibody and a small (7-kDa) scaffold protein, the Affibody molecule, Methods: The anti-HER2 Affibody Z(HER2:342) and humanized monoclonal antibody trastuzumab were labeled with (124/125)I using p-iodobenzoate (PIB) as a linker. Cellular processing of both tracers by HER2-expressing cells was investigated. The biodistributions of (124)I-PIB-Z(HER2:342) and (125)I-PIB-trastuzumab were compared in BALB/C nu/nu mice bearing HER2-expressing NCI-N87 xenografts using paired labels. Small-animal PET of (124)I-PIB-Z(HER2:342) and (124)I-PIB-trastuzumab in tumor-bearing mice was performed at 6, 24, and 72 h after injection. Results: Both radioiodinated Z(HER2:342) and trastuzumab bound specifically to HER2-expressing cells in vitro and specifically targeted HER2-expressing xenografts in vivo. Radioiodinated trastuzumab was more rapidly internalized and degraded, which resulted in better retention of radioactivity delivered by Z(HER2:342). Total uptake of trastuzumab in tumors was higher than that of (124)I-PIB-Z(HER2:342). However, tumor-to-organ ratios were appreciably higher for (124)I-PIB-Z(HER2:342) due to the more rapid clearance of radioactivity from blood and normal organs. The ex vivo results were confirmed by small-animal PET. Conclusion: The use of the small scaffold targeting Affibody provides better contrast in HER2 imaging than does the monoclonal antibody.

Place, publisher, year, edition, pages
2009. Vol. 50, no 3, 417-425 p.
Keyword [en]
Affibody molecules, imaging, targeting, xenografts, HER2
National Category
Medical and Health Sciences Natural Sciences
URN: urn:nbn:se:kth:diva-40873DOI: 10.2967/jnumed.108.057919ISI: 000264084500021PubMedID: 19223403OAI: diva2:442590
QC 20110922Available from: 2011-09-22 Created: 2011-09-22 Last updated: 2011-09-22Bibliographically approved
In thesis
1. Design and Evaluation of Radiolabeled Affibody Tracers for Imaging of HER2-expressing Tumors
Open this publication in new window or tab >>Design and Evaluation of Radiolabeled Affibody Tracers for Imaging of HER2-expressing Tumors
2011 (English)Doctoral thesis, comprehensive summary (Other academic)
Abstract [en]

The growing understanding of tumor biology and the identification of tumor specificgenetic and molecular alterations, such as the overexpression of human epidermal growthfactor receptor 2 (HER2), opens up for personalization of patient management using targeted therapies. However, this puts stringent demands on the diagnostic tools usedto identify patients that are likely to respond to a particular treatment. Radionuclide molecular imaging is a promising non-invasive method to visualize and characterize the expression of such targets. This thesis, based on five papers, is focused on the development of radiolabeled Affibody molecules for imaging of HER2-expression in malignant tumors. Affibody molecules, which represent a rather novel class of affinity proteins developed by combinatorial protein engineering of the protein A derived Z-domain, display manyfeatures that make them promising tracers for molecular imaging applications. The aim of the work presented here was to further develop the tracer format for improved in vivo properties and flexibility in the choice of radionuclide.

In paper I, the development of an assay that enables quantitative studies of the internalization rate and cellular processing of high affinity Affibody molecules is described. The assay was applied to a HER2-binding Affibody variant that was efficiently retained by HER2-expressing cells, although characterized by a slow internalization rate. This may have implications for the choice of label for Affibody molecules since high affinity to the target may be equally, or more, important for good imaging quality than residualizing properties of the radiolabel. In paper II, a HER2-binding Affibody molecule and the monoclonal antibody trastuzumab were labeled with positron emitting 124I, for a head-to-head in vivocomparison of the two tracer formats. The effects of tracer size and presence of an Fc region on the biodistribution profile were investigated. In paper III, a HER2-binding Affibody molecule was site-specifically labeled with radiocobalt and evaluated in vitro and in vivo.A head-to-head in vivo comparison with the well-studied 111In-labeled counterpart was performed, revealing promising potential for the cobalt-labeled molecule as a PET-tracerfor visualization of HER2. Paper IV describes the in vitro and in vivo evaluation of a panel of Affibody molecules with different C-terminal peptide-based chelators for the coordination of 99mTc. Even small changes in the C-terminal sequence had appreciable impact on the biodistribution of the Affibody molecules and by optimizing the design of the chelator, the kidney uptake of 99mTc could be significantly reduced. Finally, in paper V we describe the development of a HER2-targeting Affibody variant equipped with a Sel-tag for site-specific labeling with the short-lived positron emitter 11C. This novel Affibody tracer could be used to image HER2-expressing tumors in vivo within one hour after injection.

Taken together, Affibody molecules show great promise as targeting tracers for radionuclide molecular imaging of HER2. Careful design and optimization of the tracer protein is important and can be used to improve the biodistribution and targeting properties of Affibody molecules.

Place, publisher, year, edition, pages
Stockholm: KTH Royal Institute of Technology, 2011. xi, 79 p.
Trita-BIO-Report, ISSN 1654-2312 ; 2011:21
Affibody molecule, radionuclide molecular imaging, HER2, radiotracer, SPECT, PET, biodistribution, protein engineering, radiolabeling
National Category
Medical Biotechnology Biochemistry and Molecular Biology
urn:nbn:se:kth:diva-40890 (URN)978-91-7501-092-2 (ISBN)
Public defence
2011-10-21, FD5, AlbaNova University center, Stockholm, 10:00 (English)
QC 20110922Available from: 2011-09-22 Created: 2011-09-22 Last updated: 2011-09-22Bibliographically approved

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