Conjugation of Therapeutic PSD-95 Inhibitors to the Cell-Penetrating Peptide Tat Affects Blood-Brain Barrier Adherence, Uptake, and Permeation

• Main Authors: Mie Kristensen (Author), Krzysztof Kucharz (Author), Eduardo Felipe Alves Fernandes (Author), Kristian Strømgaard (Author), Morten Schallburg Nielsen (Author), Hans Christian Cederberg Helms (Author), Anders Bach (Author), Malte Ulrikkaholm Tofte-Hansen (Author), Blanca Irene Aldana Garcia (Author), Martin Lauritzen (Author), Birger Brodin (Author)
• Format: Book
• Published: MDPI AG, 2020-07-01T00:00:00Z.
• Subjects: article
• Online Access: Connect to this object online.

Summary

Novel stroke therapies are needed. Inhibition of the interaction between the postsynaptic density-95 (PSD-95)/disc large/ZO-1 (PDZ) domains of PSD-95 and the <i>N</i>-methyl-D-aspartate (NMDA) receptor has been suggested as a strategy for relieving neuronal damage. The peptides NR2B9c and <i>N</i>-dimer have been designed to hinder this interaction; they are conjugated to the cell-penetrating peptide Tat to facilitate blood-brain barrier (BBB) permeation and neuronal uptake. Tat-<i>N</i>-dimer exhibits 1000-fold better target affinity than Tat-NR2B9c, but the same magnitude of improvement is not observed in terms of therapeutic effect. Differences in BBB permeation by Tat-NR2B9c and Tat-<i>N</i>-dimer may explain this difference, but studies providing a direct comparison of Tat-NR2B9c and Tat-<i>N</i>-dimer are lacking. The aim of the present study was therefore to compare the BBB uptake and permeation of Tat-NR2B9c and Tat-<i>N</i>-dimer. The peptides were conjugated to the fluorophore TAMRA and their chemical stability assessed. Endothelial membrane association and cell uptake, and transendothelial permeation were estimated using co-cultures of primary bovine brain capillary endothelial cells and rat astrocytes. In vivo BBB permeation was demonstrated in mice using two-photon microscopy imaging. Tissue distribution was evaluated in mice demonstrating brain accumulation of TAMRA-Tat (0.4% ID/g), TAMRA-Tat-NR2B9c (0.3% ID/g), and TAMRA-Tat-<i>N</i>-dimer (0.25% ID/g). In conclusion, we demonstrate that attachment of NR2B9c or <i>N</i>-dimer to Tat affects both the chemical stability and the ability of the resulting construct to interact with and permeate the BBB.

Item Description

10.3390/pharmaceutics12070661
1999-4923