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Influence of size and charge of unstructured polypeptides on pharmacokinetics and biodistribution of targeted fusion proteins


Brandl, Fabian; Merten, Hannes; Zimmermann, Martina; Béhé, Martin; Zangemeister-Wittke, Uwe; Plückthun, Andreas (2019). Influence of size and charge of unstructured polypeptides on pharmacokinetics and biodistribution of targeted fusion proteins. Journal of Controlled Release, 307:379-392.

Abstract

Alternative non-IgG binding proteins developed for therapy are small in size and, thus, are rapidly cleared from the circulation by renal filtration. To avoid repeated injection or continuous infusion for the maintenance of therapeutic serum concentrations, extensions of unfolded polypeptides have been developed to prolong serum half-life, but systematic, comparative studies investigating the influence of their size and charge on serum half-life, extravasation, tumor localization and excretion mechanisms have so far been lacking. Here we used a high-affinity Designed Ankyrin Repeat Protein (DARPin) targeting the tumor marker epithelial cell adhesion molecule (EpCAM) in a preclinical tumor xenograft model in mice, and fused it with a series of defined unstructured polypeptides. We used three different sizes of two previously described polypeptides, an uncharged one consisting of only Pro, Ala and Ser (termed PAS) and a charged one consisting of Pro, Ala, Ser, Thr, Gly, Glu (termed XTEN) and performed for the first time a precise comparative localization, distribution and extravasation study. Pharmacokinetic analysis showed a clear linear relationship between hydrodynamic radius and serum half-life across both polypeptides, reaching a half-life of up to 21 h in mice. Tumor uptake was EpCAM-dependent and directly proportional to half-life and size, showing an even tumor penetration for all fusion proteins without unspecific accumulation in non-target tissue. Unexpectedly, charge had no influence on any parameter, neither tumor nor tissue accumulation nor kidney elimination kinetics. Thus, both polypeptide types have a very similar potential for precise half-life modification and tumor targeting.

Abstract

Alternative non-IgG binding proteins developed for therapy are small in size and, thus, are rapidly cleared from the circulation by renal filtration. To avoid repeated injection or continuous infusion for the maintenance of therapeutic serum concentrations, extensions of unfolded polypeptides have been developed to prolong serum half-life, but systematic, comparative studies investigating the influence of their size and charge on serum half-life, extravasation, tumor localization and excretion mechanisms have so far been lacking. Here we used a high-affinity Designed Ankyrin Repeat Protein (DARPin) targeting the tumor marker epithelial cell adhesion molecule (EpCAM) in a preclinical tumor xenograft model in mice, and fused it with a series of defined unstructured polypeptides. We used three different sizes of two previously described polypeptides, an uncharged one consisting of only Pro, Ala and Ser (termed PAS) and a charged one consisting of Pro, Ala, Ser, Thr, Gly, Glu (termed XTEN) and performed for the first time a precise comparative localization, distribution and extravasation study. Pharmacokinetic analysis showed a clear linear relationship between hydrodynamic radius and serum half-life across both polypeptides, reaching a half-life of up to 21 h in mice. Tumor uptake was EpCAM-dependent and directly proportional to half-life and size, showing an even tumor penetration for all fusion proteins without unspecific accumulation in non-target tissue. Unexpectedly, charge had no influence on any parameter, neither tumor nor tissue accumulation nor kidney elimination kinetics. Thus, both polypeptide types have a very similar potential for precise half-life modification and tumor targeting.

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Additional indexing

Item Type:Journal Article, refereed, original work
Communities & Collections:04 Faculty of Medicine > Department of Biochemistry
07 Faculty of Science > Department of Biochemistry
Dewey Decimal Classification:570 Life sciences; biology
610 Medicine & health
Scopus Subject Areas:Life Sciences > Pharmaceutical Science
Language:English
Date:10 August 2019
Deposited On:25 Sep 2019 13:23
Last Modified:29 Jul 2020 11:22
Publisher:Elsevier
ISSN:0168-3659
OA Status:Closed
Publisher DOI:https://doi.org/10.1016/j.jconrel.2019.06.030
PubMed ID:31252038

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