TY - JOUR
T1 - Multifunctional peptide-PEG intercalating conjugates
T2 - Programmatic of gene delivery to the blood-brain barrier
AU - Zhang, Hongwei
AU - Gerson, Trevor
AU - Varney, Michelle L.
AU - Singh, Rakesh K.
AU - Vinogradov, Serguei V.
N1 - Funding Information:
This work was supported by NIH grants NS050660 and NS063879 (S.V.V.). Authors are grateful to Mrs. Huai-Yun Han, Arin Zeman, and Galya Warren for excellent technical help in conducting some experiments. The assistance of UNMC Confocal Microscopy and Protein Analysis Core facilities is greatly appreciated.
PY - 2010/12
Y1 - 2010/12
N2 - Purpose To enhance transfection efficacy of pDNA through the application ofmultifunctional peptide-PEG-tris-acridine conjugates ( pPAC) and the formation of biodegradable core-shell polyplexes for gene delivery to the blood-brain barrier (BBB). Methods pPAC-mediated transfection was compositionally optimized in mouse BBB cells (bEnd.3). Cellular uptake and trafficking, and brain accumulation of pDNA was evaluated by fluorescent imaging and histochemistry. We constructed anti- MRP4 siRNA-producing vectors and evaluated the efficacy of MRP4 down-regulation of MRP4 by Western blot and qPCR, and its effect on the uptake of 3H-AZT, an MRP4 substrate. Results A core-shell gene delivery system (GDS) was assembled from pDNA and pPAC, carrying multifunctional peptides with NLS, TAT, and brain-specific BH, or ApoE sequences, and biodegradable pLPEI polyamine. This GDS demonstrated better cellular and nuclear accumulation, and a 25-fold higher transfection efficacy in slow-dividing bEnd.3 cells compared to ExGen500. Inclusion of brain-targeting pPAC enhanced in vivo accumulation of functional pDNA in brain capillaries. Treatment by encapsulated anti-MRP4 siRNAproducing pDNA caused transient down-regulation of MRP4, and, after intravenous injection in Balb/c mice, enhanced AZT uptake in the brain by 230-270%. Conclusions The pPAC represent novel efficient components of GDS that could find various gene therapy applications, including genetic modulation of the BBB.
AB - Purpose To enhance transfection efficacy of pDNA through the application ofmultifunctional peptide-PEG-tris-acridine conjugates ( pPAC) and the formation of biodegradable core-shell polyplexes for gene delivery to the blood-brain barrier (BBB). Methods pPAC-mediated transfection was compositionally optimized in mouse BBB cells (bEnd.3). Cellular uptake and trafficking, and brain accumulation of pDNA was evaluated by fluorescent imaging and histochemistry. We constructed anti- MRP4 siRNA-producing vectors and evaluated the efficacy of MRP4 down-regulation of MRP4 by Western blot and qPCR, and its effect on the uptake of 3H-AZT, an MRP4 substrate. Results A core-shell gene delivery system (GDS) was assembled from pDNA and pPAC, carrying multifunctional peptides with NLS, TAT, and brain-specific BH, or ApoE sequences, and biodegradable pLPEI polyamine. This GDS demonstrated better cellular and nuclear accumulation, and a 25-fold higher transfection efficacy in slow-dividing bEnd.3 cells compared to ExGen500. Inclusion of brain-targeting pPAC enhanced in vivo accumulation of functional pDNA in brain capillaries. Treatment by encapsulated anti-MRP4 siRNAproducing pDNA caused transient down-regulation of MRP4, and, after intravenous injection in Balb/c mice, enhanced AZT uptake in the brain by 230-270%. Conclusions The pPAC represent novel efficient components of GDS that could find various gene therapy applications, including genetic modulation of the BBB.
KW - AZT
KW - ApoE peptide
KW - Blood-brain barrier
KW - Brain capillary endothelial cells
KW - Brain-homing peptide
KW - Intercalating complex
KW - MRP4
KW - NLS peptide
KW - Peptide-PEG conjugate
KW - Plasmid DNA
KW - TAT peptide
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U2 - 10.1007/s11095-010-0256-x
DO - 10.1007/s11095-010-0256-x
M3 - Article
C2 - 20824308
AN - SCOPUS:78651278782
SN - 0724-8741
VL - 27
SP - 2528
EP - 2543
JO - Pharmaceutical Research
JF - Pharmaceutical Research
IS - 12
ER -