TY - JOUR
T1 - Conjugation of Therapeutic PSD-95 Inhibitors to the Cell-Penetrating Peptide Tat Affects Blood-Brain Barrier Adherence, Uptake, and Permeation
AU - Kristensen, Mie
AU - Kucharz, Krzysztof
AU - Felipe Alves Fernandes, Eduardo
AU - Strømgaard, Kristian
AU - Schallburg Nielsen, Morten
AU - Cederberg Helms, Hans Christian
AU - Bach, Anders
AU - Ulrikkaholm Tofte-Hansen, Malte
AU - Irene Aldana Garcia, Blanca
AU - Lauritzen, Martin
AU - Brodin, Birger
PY - 2020
Y1 - 2020
N2 - 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 N-methyl-D-aspartate (NMDA) receptor has been suggested as a strategy for relieving neuronal damage. The peptides NR2B9c and N-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-N-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-N-dimer may explain this difference, but studies providing a direct comparison of Tat-NR2B9c and Tat-N-dimer are lacking. The aim of the present study was therefore to compare the BBB uptake and permeation of Tat-NR2B9c and Tat-N-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-N-dimer (0.25% ID/g). In conclusion, we demonstrate that attachment of NR2B9c or N-dimer to Tat affects both the chemical stability and the ability of the resulting construct to interact with and permeate the BBB.
AB - 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 N-methyl-D-aspartate (NMDA) receptor has been suggested as a strategy for relieving neuronal damage. The peptides NR2B9c and N-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-N-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-N-dimer may explain this difference, but studies providing a direct comparison of Tat-NR2B9c and Tat-N-dimer are lacking. The aim of the present study was therefore to compare the BBB uptake and permeation of Tat-NR2B9c and Tat-N-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-N-dimer (0.25% ID/g). In conclusion, we demonstrate that attachment of NR2B9c or N-dimer to Tat affects both the chemical stability and the ability of the resulting construct to interact with and permeate the BBB.
U2 - 10.3390/pharmaceutics12070661
DO - 10.3390/pharmaceutics12070661
M3 - Journal article
C2 - 32674358
VL - 12
JO - Pharmaceutics
JF - Pharmaceutics
SN - 1999-4923
IS - 7
M1 - 661
ER -