Analytical characterization of NOTA-modified somatropins
dc.contributor.author | Bracke, Nathalie | |
dc.contributor.author | Wynendaele, Evelien | |
dc.contributor.author | D’Hondt, Matthias | |
dc.contributor.author | Haselberg, Rob | |
dc.contributor.author | Somsen, Govert W. | |
dc.contributor.author | Pauwels, Ewald | |
dc.contributor.author | Van de Wiele, Christophe | |
dc.contributor.author | De Spiegeleer, Bart | |
dc.date.accessioned | 2017-03-03T06:07:30Z | |
dc.date.available | 2017-03-03T06:07:30Z | |
dc.date.issued | 2014-08 | |
dc.description.abstract | Chemical modification of biomolecules like the introduction of metal-chelators into proteins can lead to heterogeneous product formation. The nature and extend of the modification is important in interpreting the biological properties of the bioconjugate, given their possible influence on the pharmacokinetics as well as on the binding affinity to the target. The present study describes the synthesis and analytical characterization of somatropin modified on its lysine’s ε-amino groups with the acylating chelator S-2-(4-isothiocyanatobenzyl)-1,4,7-triazacyclononane-1,4,7-triacetic acid (p- SCN-Bn-NOTA). Direct separation and identification techniques (i.e. RP-MS and CE-MS) and peptide mapping after trypsin and chymotrypsin digestion demonstrated that the use of higher amounts of p-SCN-Bn-NOTA during synthesis leads to a complex product composition with higher order substitution degrees (i.e. multiple NOTA-moieties per somatropin molecule), as well as the presence of different position isomers. From the nine lysine (Lys) residues in somatropin, Lys-70 was experimentally found to be the modification hotspot under our synthesis conditions (pH=9.0). This was supported by the in silico calculated lowest pKa value of 8.3 for Lys-70. Based on the crystal structure of somatropin in complex with the extracellular parts of the growth hormone receptor, the Lys-70 residue is positioned outside the binding pockets and will therefore not directly interfere with receptor binding. Gallium chelation by NOTA-somatropin resulted in a 100% complexation. The synthesis of NOTA-somatropin using p-SCN-Bn-NOTA and somatropin under our operational conditions is therefore a suitable synthesis procedure for the production of a target-specific radiopharmaceutical for further investigation towards treatment and visualization of growth hormonespecific cancers. | en_ZA |
dc.description.department | Nuclear Medicine | en_ZA |
dc.description.librarian | hb2017 | en_ZA |
dc.description.sponsorship | This research project was supported by grants from Ghent University (BOF special research fund 01J22510) to BDS and EW and the Institute for the Promotion of Innovation through Science and Technology in Flanders (IWT-Vlaanderen) to MD (101529). | en_ZA |
dc.description.uri | http://www.elsevier.com/locate/jpba | en_ZA |
dc.identifier.citation | Bracke, N, Wynendaele, E, D’Hondt, M, Haselberg, R, Somsen, GW, Pauwels, E, Van de Wiele, C & De Spiegeleer, B 2014, 'Analytical characterization of NOTA-modified somatropins', Journal of Pharmaceutical and Biomedical Analysis, vol. 96, pp. 1-9. | en_ZA |
dc.identifier.issn | 0731-7085 (print) | |
dc.identifier.issn | 1873-264X (online) | |
dc.identifier.other | 10.1016/j.jpba.2014.03.014 | |
dc.identifier.uri | http://hdl.handle.net/2263/59249 | |
dc.language.iso | en | en_ZA |
dc.publisher | Elsevier | en_ZA |
dc.rights | © 2014 Elsevier B.V. All rights reserved. Notice : this is the author’s version of a work that was accepted for publication in Journal of Pharmaceutical and Biomedical Analysis. Changes resulting from the publishing process, such as peer review, editing, corrections, structural formatting, and other quality control mechanisms may not be reflected in this document. A definitive version was subsequently published in Journal of Pharmaceutical and Biomedical Analysis, vol. 96, pp. 1-9, 2014. doi : 10.1016/j.jpba.2014.03.014. | en_ZA |
dc.subject | NOTA modification | en_ZA |
dc.subject | Somatropin | en_ZA |
dc.subject | Peptide mapping | en_ZA |
dc.subject | Liquid chromatography–mass spectrometry (LC-MS) | en_ZA |
dc.subject | Capillary electrophoresis–mass spectrometry (CE-MS) | en_ZA |
dc.title | Analytical characterization of NOTA-modified somatropins | en_ZA |
dc.type | Postprint Article | en_ZA |