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Multifunctional Peptide-PEG Intercalating Conjugates: Programmatic of Gene Delivery to the Blood-Brain Barrier

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ABSTRACT

Purpose

To enhance transfection efficacy of pDNA through the application of multifunctional 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 siRNA-producing 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.

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Abbreviations

Acr:

acridine

ApoE:

Apolipoprotein E-derived peptide

BBB:

blood-brain barrier

BCEC:

brain capillary endothelial cells

BH:

brain-homing peptide

BSA:

bovine serum albumin

DAB:

Diaminobenzidine

DET:

drug efflux transporters

DLS:

dynamic light scattering

ECL:

chemoluminescence detection kit

GDS:

gene delivery system

GFP:

green fluorescent protein

HAART:

highly active antiretroviral therapy

HPLC:

high performance liquid chromatography

LPEI:

linear polyethylenimine

NLS:

SV40 nuclear localization signal peptide

NRTI:

Nucleoside reverse transcriptase inhibitors

PAGE:

Polyacrylamide gel electrophoresis

PEG:

Poly(ethylene glycol)

TAT:

HIV-1 trans-activator of transcription peptide

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ACKNOWLEDGMENTS

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.

Author information

Author notes

  1. Hongwei Zhang

    Present address: Gene Therapy Center, University of Massachusetts Medical School, Worcester, Massachusetts, 01605, USA

Authors and Affiliations

  1. Center for Drug Delivery and Nanomedicine and Department of Pharmaceutical Sciences, College of Pharmacy, University of Nebraska Medical Center, Omaha, Nebraska, 68198, USA

    Hongwei Zhang, Trevor Gerson & Serguei V. Vinogradov

  2. Department of Pathology and Microbiology, College of Medicine, University of Nebraska Medical Center, Omaha, Nebraska, 68198, USA

    Michelle L. Varney & Rakesh K. Singh

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  1. Hongwei Zhang
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  2. Trevor Gerson
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  5. Serguei V. Vinogradov
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Corresponding author

Correspondence to Serguei V. Vinogradov.

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Zhang, H., Gerson, T., Varney, M.L. et al. Multifunctional Peptide-PEG Intercalating Conjugates: Programmatic of Gene Delivery to the Blood-Brain Barrier. Pharm Res 27, 2528–2543 (2010). https://doi.org/10.1007/s11095-010-0256-x

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  • DOI: https://doi.org/10.1007/s11095-010-0256-x

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