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Effects of lysine-containing mercaptoacetyl-based chelators on the biodistribution of 99mTc-labeled anti-HER2 Affibody molecules
Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Oncology, Radiology and Clinical Immunology, Biomedical Radiation Sciences.
Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Oncology, Radiology and Clinical Immunology, Biomedical Radiation Sciences.ORCID iD: 0000-0001-6120-2683
Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Oncology, Radiology and Clinical Immunology, Oncology.
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2008 (English)In: Bioconjugate chemistry, ISSN 1043-1802, E-ISSN 1520-4812, Vol. 19, no 12, 2568-2576 p.Article in journal (Refereed) Published
Abstract [en]

The effects of polar (mercaptoacetyl-triseryl) and negatively charged (mercaptoacetyl-triglumatyl) chelators on the biodistribution of 99mTc-labeled anti-HER2 Affibody molecules were previously investigated. With glycine, serine, and glutamate, we demonstrated that substitution with a single amino acid in the chelator can significantly influence the biodistribution properties and the excretion pathways. Here, we have taken this investigation further, by analyzing the effects of introduction of a positive amino acid residue on the in vivo properties of the 99mTc-labeled Affibody molecule. The Affibody molecules with mercaptoacetyl-seryl-lysyl-seryl (maSKS) and mercaptoacetyl-trilysyl (maKKK) extensions were produced by peptide synthesis and labeled with 99mTc in alkaline conditions. A comparative biodistribution was performed in normal mice to evaluate the excretion pathway. A shift toward renal excretion was obtained when serine was substituted with lysine in the chelating sequence. The radioactivity in the gastrointestinal tract was reduced 3-fold for the 99mTc-maSKS-Z(HER2:342) and 99mTc-maKKK-Z(HER2: 342) in comparison with the 99mTc-maSSS-Z(HER2:342) conjugate 4 h post injection (p.i.). The radioactivity in the liver was elevated when a triple substitution of positively charged lysine was used. The tumor targeting properties of 99mTc-maSKS-Z(HER2:342) were further investigated in SKOV-3 xenografts. The tumor uptake of 99mTc-maSKS-Z(HER2: 342) was 17+/-7% IA/g 4 h p.i. Tumor xenografts were well-visualized by gamma scintigraphy. In conclusion, the substitution with one single lysine in the chelator results in better tumor imaging properties of the Affibody molecule Z(HER2:342) and is favorable for imaging of tumors and metastases in the abdominal area. Multiple lysine residues in the chelator are, however, undesirable due to elevated uptake both in the liver and kidneys.

Place, publisher, year, edition, pages
2008. Vol. 19, no 12, 2568-2576 p.
Keyword [en]
peptide synthesis, imaging, affibody, HER2, technetium, radionuclides, chelator
National Category
Medical and Health Sciences
Identifiers
URN: urn:nbn:se:uu:diva-98942DOI: 10.1021/bc800244bISI: 000261767800036PubMedID: 19035668OAI: oai:DiVA.org:uu-98942DiVA: diva2:201686
Available from: 2009-03-05 Created: 2009-03-05 Last updated: 2017-12-13Bibliographically approved
In thesis
1. Molecular Imaging of HER2 Expression using Synthetic Affibody Molecules: Design, Synthesis and Biological Evaluation
Open this publication in new window or tab >>Molecular Imaging of HER2 Expression using Synthetic Affibody Molecules: Design, Synthesis and Biological Evaluation
2009 (English)Doctoral thesis, comprehensive summary (Other academic)
Abstract [en]

Molecular imaging is an emerging multidisciplinary field that addresses the visualisation of diseases at the cellular and molecular levels. This thesis focuses on the development of a synthetic Affibody molecule-based imaging tracer for the detection of HER2 expression in malignant tumours.

Papers I-IV report the development of the HER2-specific Affibody molecule, ZHER2:342 by peptide synthesis and the use of different chelators attached to the N-terminus to allow 99mTc-labelling. Paper I described the optimisation of labelling of Affibody molecules using cysteine-based chelator sequences, in which the direct labelling method under alkaline conditions was the most suitable one. Papers II-IV report the development and optimisation of the in vivo properties of the HER2-specific Affibody molecule for high-contrast imaging. By using an array of mercaptoacetyl-based chelators, it was found that the substitution of a single amino acid in a 60 amino acid-long Affibody molecule can dramatically change the pharmacokinetics of the tracer. Strategic approaches that utilised hydrophilic amino acids, such as serine, glutamate and lysine, changed the excretion pathway from hepatobiliary to renal excretion. Problems with the high accumulation of radioactivity in the abdomen area and restricted imaging were resolved by the use of mercaptoacetyl-triglutamyl, maEEE or mercaptoacetyl-seryl-lysyl-seryl, maSKS chelators.

Paper V reports the re-engineering of the HER2-specific Affibody molecule to provide a C-terminal SECG sequence using peptide synthesis. Incorporation of this sequence provided a multifunctional platform for labelling (with technetium or trivalent radiometals) and a flexible production route (recombinant or chemical synthesis). Combination of a serine, a glutamic acid and a thiol-bearing group in the chelating sequence reduced the renal accumulation of Affibody molecules.

Altogether, the in vivo efficiency of Affibody molecules to target tumours and their biodistribution properties can be improved by strategic design and suitable chemistry. Hopefully, these observations will be applicable to other small peptide and protein scaffold-based tracers.

Place, publisher, year, edition, pages
Uppsala: Acta Universitatis Upsaliensis, 2009. 53 p.
Series
Digital Comprehensive Summaries of Uppsala Dissertations from the Faculty of Medicine, ISSN 1651-6206 ; 445
Series
National Category
Medical and Health Sciences
Research subject
Biomedical Radiation Science
Identifiers
urn:nbn:se:uu:diva-99248 (URN)978-91-554-7489-8 (ISBN)
Public defence
2009-05-16, Rudbecksalen, Rudbecklaboratoriet, Uppsala, 09:15 (English)
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Available from: 2009-04-24 Created: 2009-03-10 Last updated: 2009-04-24Bibliographically approved

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Orlova, AnnaTolmachev, Vladimir

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