title: Non-degradative intracellular trafficking of highly compacted polymeric DNA nanoparticles.
abstract: Highly compacted DNA nanoparticles (DNPs) composed of polyethylene glycol linked to a 30-mer of poly-l-lysine via a single cysteine residue (CK(30)PEG) have previously been shown to provide efficient gene delivery to the brain, eyes and lungs. In this study, we used a combination of flow cytometry, high-resolution live-cell confocal microscopy, and multiple particle tracking (MPT) to investigate the intracellular trafficking of highly compacted CK(30)PEG DNPs made using two different molecular weights of PEG, CK(30)PEG(10k) and CK(30)PEG(5k). We found that PEG MW did not have a major effect on particle morphology nor nanoparticle intracellular transport. CK(30)PEG(10k) and CK(30)PEG(5k) DNPs both entered human bronchial epithelial (BEAS-2B) cells via a caveolae-mediated pathway, bypassing degradative endolysosomal trafficking. Both nanoparticle formulations were found to rapidly accumulate in the perinuclear region of cells within 2h, 37±19% and 47±8% for CK(30)PEG(10k) and CK(30)PEG(5k), respectively. CK(30)PEG(10k) and CK(30)PEG(5k) DNPs moved within live cells at average velocities of 0.09±0.04μm/s and 0.11±0.04μm/s, respectively, in good agreement with reported values for caveolae. These findings show that highly compacted DNPs employ highly regulated trafficking mechanisms similar to biological pathogens to target specific intracellular compartments.
authors.affiliation: Department of Ophthalmology, The Wilmer Eye Institute, Johns Hopkins University School of Medicine, 400 N. Broadway, Baltimore, MD 21287, USA.
authors.forename: Anthony J
authors.initials: AJ
authors.lastname: Kim
authors.forename: Nicholas J
authors.initials: NJ
authors.lastname: Boylan
authors.forename: Jung Soo
authors.initials: JS
authors.lastname: Suk
authors.forename: Samuel K
authors.initials: SK
authors.lastname: Lai
authors.forename: Justin
authors.initials: J
authors.lastname: Hanes
mesh_terms: D001692:Biological Transport; D002460:Cell Line; D003545:Cysteine / Q000008:administration & dosage* / Q000737:chemistry / Q000493:pharmacokinetics; D004247:DNA / Q000008:administration & dosage* / Q000737:chemistry / Q000493:pharmacokinetics; D018014:Gene Transfer Techniques; D006801:Humans; D046529:Microscopy, Electron, Transmission; D053758:Nanoparticles / Q000008:administration & dosage* / Q000737:chemistry / Q000648:ultrastructure; D010957:Plasmids / Q000235:genetics; D011092:Polyethylene Glycols / Q000008:administration & dosage* / Q000737:chemistry / Q000493:pharmacokinetics
chemical_list: D011092:Polyethylene Glycols; D004247:DNA; D003545:Cysteine
references.citation: J Virol. 2002 Oct;76(20):10455-64
references.pmid: 12239322
references.citation: Bioconjug Chem. 2008 Oct;19(10):2023-9
references.pmid: 18729494
references.citation: Mol Ther. 2008 Feb;16(2):333-42
references.pmid: 18059369
references.citation: J Control Release. 2010 Nov 20;148(1):69-74
references.pmid: 20833214
references.citation: Proc Natl Acad Sci U S A. 2007 Jan 30;104(5):1482-7
references.pmid: 17244708
references.citation: J Cell Sci. 2002 Nov 15;115(Pt 22):4327-39
references.pmid: 12376564
references.citation: Biochim Biophys Acta. 2009 Oct;1788(10):2259-66
references.pmid: 19595666
references.citation: Science. 2004 Apr 9;304(5668):237-42
references.pmid: 15073366
references.citation: Pharmacol Rev. 2006 Mar;58(1):32-45
references.pmid: 16507881
references.citation: J Control Release. 2008 Jan 22;125(2):107-11
references.pmid: 18053606
references.citation: Mol Ther. 2003 Dec;8(6):936-47
references.pmid: 14664796
references.citation: Mol Pharm. 2010 Jun 7;7(3):738-50
references.pmid: 20349982
references.citation: Proc Natl Acad Sci U S A. 2003 Apr 1;100(7):3878-82
references.pmid: 12644705
references.citation: Adv Drug Deliv Rev. 2000 Mar 30;41(2):201-8
references.pmid: 10699315
references.citation: PLoS One. 2009 Oct 12;4(10):e7410
references.pmid: 19823583
references.citation: Adv Drug Deliv Rev. 2005 Jan 2;57(1):63-78
references.pmid: 15518921
references.citation: J Control Release. 2003 Feb 21;87(1-3):89-105
references.pmid: 12618026
references.citation: Science. 2002 Apr 19;296(5567):535-9
references.pmid: 11964480
references.citation: J Aerosol Med. 2005 Summer;18(2):137-82
references.pmid: 15966771
references.citation: Proc Natl Acad Sci U S A. 2010 Jan 12;107(2):598-603
references.pmid: 20018745
references.citation: J Control Release. 2009 Oct 15;139(2):88-93
references.pmid: 19580830
references.citation: Adv Drug Deliv Rev. 2009 Feb 27;61(2):158-71
references.pmid: 19133304
references.citation: J Biol Chem. 2003 Aug 29;278(35):32578-86
references.pmid: 12807905
references.citation: Biomaterials. 2009 May;30(13):2591-7
references.pmid: 19176245
references.citation: Angew Chem Int Ed Engl. 2006 Feb 27;45(10):1568-72
references.pmid: 16470762
references.citation: Mol Ther. 2009 Apr;17(4):641-50
references.pmid: 19223866
references.citation: Adv Drug Deliv Rev. 2007 Aug 10;59(8):798-809
references.pmid: 17707545
references.citation: Mol Ther. 2011 Jan;19(1):93-102
references.pmid: 20959809
references.citation: J Control Release. 2009 May 21;136(1):54-61
references.pmid: 19217921
references.citation: Int Rev Cytol. 2000;192:189-221
references.pmid: 10553280
references.citation: Biochem J. 2004 Jan 1;377(Pt 1):159-69
references.pmid: 14505488
references.citation: Nat Cell Biol. 2001 May;3(5):473-83
references.pmid: 11331875
references.citation: J Biol Chem. 2000 Jan 21;275(3):1625-9
references.pmid: 10636854
references.citation: Mol Pharm. 2006 Nov-Dec;3(6):644-53
references.pmid: 17140252
references.citation: Biomaterials. 2007 Jun;28(18):2876-84
references.pmid: 17363053
references.citation: J Gen Virol. 2002 Jul;83(Pt 7):1535-1545
references.pmid: 12075072
references.citation: Angew Chem Int Ed Engl. 2008;47(50):9726-9
references.pmid: 18979480
references.citation: J Control Release. 2010 Aug 3;145(3):182-95
references.pmid: 20226220
references.citation: Methods Mol Biol. 2008;434:81-97
references.pmid: 18470640
references.citation: Pharm Res. 2000 Sep;17(9):1035-48
references.pmid: 11087034
references.citation: J Liposome Res. 2006;16(3):237-47
references.pmid: 16952878
references.citation: J Control Release. 2006 Dec 1;116(3):314-21
references.pmid: 17112618
references.citation: Biomaterials. 2010 Mar;31(7):1757-64
references.pmid: 19963266
references.citation: J Cell Biol. 2001 Oct 1;155(1):77-88
references.pmid: 11581287
references.citation: J Phys Chem B. 2006 Mar 16;110(10):4548-54
references.pmid: 16526683
references.citation: Gene Ther. 1999 Apr;6(4):595-605
references.pmid: 10476219