CPPs to the Test: Effects on Binding, Uptake and Biodistribution of a Tumor Targeting Nanobody

Estel Collado Camps, Sanne A. M. van Lith, Cathelijne Frielink, Jordi Lankhof, Ingrid Dijkgraaf, Martin Gotthardt, Roland Brock*

*Corresponding author for this work

Research output: Contribution to journalArticleAcademicpeer-review

3 Citations (Web of Science)

Abstract

Nanobodies are well-established targeting ligands for molecular imaging and therapy. Their short circulation time enables early imaging and reduces systemic radiation exposure. However, shorter circulation time leads to lower tracer accumulation in the target tissue. Cell-penetrating peptides (CPPs) improve cellular uptake of various cargoes, including nanobodies. CPPs could enhance tissue retention without compromising rapid clearance. However, systematic investigations on how the functionalities of nanobody and CPP combine with each other at the level of 2D and 3D cell cultures and in vivo are lacking. Here, we demonstrate that conjugates of the epidermal growth factor receptor (EGFR)-binding nanobody 7D12 with different CPPs (nonaarginine, penetratin, Tat and hLF) differ with respect to cell binding and induction of endocytosis. For nonaarginine and penetratin we compared the competition of EGF binding and performance of L- and D-peptide stereoisomers, and tested the D-peptide conjugates in tumor cell spheroids and in vivo. The D-peptide conjugates showed better penetration into spheroids than the unconjugated 7D12. Both in vivo and in vitro, the behavior of the agent reflects the combination of both functionalities. Although CPPs cause promising increases in in vitro uptake and 3D penetration, the dominant effect of the CPP in the control of biodistribution warrants further investigation.

Original languageEnglish
Article number602
Number of pages22
JournalPharmaceuticals
Volume14
Issue number7
DOIs
Publication statusPublished - Jul 2021

Keywords

  • cell-penetrating peptides
  • molecular imaging
  • nanobody
  • spheroids
  • tumor targeting
  • biodistribution
  • CELL-PENETRATING PEPTIDES
  • 3RD HELIX
  • GROWTH
  • ANTIBODIES
  • ACID
  • INTERNALIZATION
  • INHIBITION
  • RETENTION
  • IMMUNOPET
  • AFFINITY

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