title: Structure-property relationship for in vitro siRNA delivery performance of cationic 2-hydroxypropyl-β-cyclodextrin: PEG-PPG-PEG polyrotaxane vectors.
abstract: Nanoparticle-mediated siRNA delivery is a promising therapeutic approach, however, the processes required for transport of these materials across the numerous extracellular and intracellular barriers are poorly understood. Efficient delivery of siRNA-containing nanoparticles would ultimately benefit from an improved understanding of how parameters associated with these barriers relate to the physicochemical properties of the nanoparticle vectors. We report the synthesis of three Pluronic(®)-based, cholesterol end-capped cationic polyrotaxanes (PR(+)) threaded with 2-hydroxypropyl-β-cyclodextrin (HPβCD) for siRNA delivery. The biological data showed that PR(+):siRNA complexes were well tolerated (∼90% cell viability) and produced efficient silencing (>80%) in HeLa-GFP and NIH 3T3-GFP cell lines. We further used a multi-parametric approach to identify relationships between the PR(+) structure, PR(+):siRNA complex physical properties, and biological activity. Small angle X-ray scattering and cryoelectron microscopy studies reveal periodicity and lamellar architectures for PR(+):siRNA complexes, whereas the biological assays, ζ potential measurements, and imaging studies suggest that silencing efficiency is influenced by the effective charge ratio (ρeff), polypropylene oxide (PO) block length, and central PO block coverage (i.e., rigidity) of the PR(+) core. We infer from our findings that more compact PR(+):siRNA nanostructures arising from lower molecular weight, rigid rod-like PR(+) polymer cores produce improved silencing efficiency relative to higher molecular weight, more flexible PR(+) vectors of similar effective charge. This study demonstrates that PR(+):siRNA complex formulations can be produced having higher performance than Lipofectamine(®) 2000, while maintaining good cell viability and siRNA sequence protection in cell culture.
authors.affiliation: Department of Chemistry, Multi-disciplinary Cancer Research Facility, Bindley Bioscience Center, 1203 W. State Street, West Lafayette, IN 47907, USA.
authors.forename: Vivek D
authors.initials: VD
authors.lastname: Badwaik
authors.affiliation: Department of Chemistry, Multi-disciplinary Cancer Research Facility, Bindley Bioscience Center, 1203 W. State Street, West Lafayette, IN 47907, USA.
authors.forename: Emilio
authors.initials: E
authors.lastname: Aicart
authors.affiliation: Department of Chemistry, Multi-disciplinary Cancer Research Facility, Bindley Bioscience Center, 1203 W. State Street, West Lafayette, IN 47907, USA.
authors.forename: Yawo A
authors.initials: YA
authors.lastname: Mondjinou
authors.affiliation: Department of Physics, Indiana University-Purdue University Indianapolis, IN 46202, USA.
authors.forename: Merrell A
authors.initials: MA
authors.lastname: Johnson
authors.affiliation: Discovery Park, Hockmeyer Hall of Structural Biology, Purdue University, West Lafayette, IN 47907, USA.
authors.forename: Valorie D
authors.initials: VD
authors.lastname: Bowman
authors.affiliation: Department of Chemistry, Multi-disciplinary Cancer Research Facility, Bindley Bioscience Center, 1203 W. State Street, West Lafayette, IN 47907, USA. Electronic address: davethom@purdue.edu.
authors.forename: David H
authors.initials: DH
authors.lastname: Thompson
mesh_terms: D000073738:2-Hydroxypropyl-beta-cyclodextrin; D000818:Animals; D016923:Cell Death; D002470:Cell Survival; D003505:Cyclodextrins / Q000737:chemistry*; D004247:DNA / Q000378:metabolism; D005434:Flow Cytometry; D055785:Gene Knockdown Techniques; D005822:Genetic Vectors / Q000378:metabolism*; D049452:Green Fluorescent Proteins / Q000378:metabolism; D006367:HeLa Cells; D006801:Humans; D051379:Mice; D041681:NIH 3T3 Cells; D010957:Plasmids / Q000378:metabolism; D020442:Poloxamer / Q000737:chemistry*; D011092:Polyethylene Glycols / Q000138:chemical synthesis / Q000737:chemistry*; D011409:Propylene Glycols / Q000138:chemical synthesis / Q000737:chemistry*; D034622:RNA Interference; D034741:RNA, Small Interfering / Q000378:metabolism*; D043862:Rotaxanes / Q000737:chemistry*; D055672:Static Electricity; D013329:Structure-Activity Relationship; D047392:beta-Cyclodextrins / Q000138:chemical synthesis / Q000737:chemistry*
chemical_list: D003505:Cyclodextrins; D011409:Propylene Glycols; D034741:RNA, Small Interfering; D043862:Rotaxanes; D047392:beta-Cyclodextrins; C528875:poly(ethylene glycol)-block-poly(propylene glycol)-block-poly(ethylene glycol) copolymer; C475673:polyrotaxane; D020442:Poloxamer; D049452:Green Fluorescent Proteins; D000073738:2-Hydroxypropyl-beta-cyclodextrin; D011092:Polyethylene Glycols; D004247:DNA
keywords: Cryoelectron microscopy; Gene silencing; Polyrotaxane; Small angle X-ray scattering; siRNA; β-Cyclodextrin
references.citation: Biomacromolecules. 2007 Nov;8(11):3365-74
references.pmid: 17929967
references.citation: Macromol Biosci. 2016 Jan;16(1):63-73
references.pmid: 26257319
references.citation: J Am Chem Soc. 2006 Mar 29;128(12):3852-3
references.pmid: 16551060
references.citation: Biochemistry. 2007 Apr 24;46(16):4785-92
references.pmid: 17391006
references.citation: Integr Biol (Camb). 2013 Jan;5(1):115-21
references.pmid: 23042106
references.citation: Biomacromolecules. 2010 Dec 13;11(12):3571-7
references.pmid: 21067145
references.citation: Bioconjug Chem. 2006 Jul-Aug;17(4):877-88
references.pmid: 16848393
references.citation: Small. 2011 Feb 7;7(3):364-70
references.pmid: 21294265
references.citation: Colloids Surf B Biointerfaces. 2011 Jan 1;82(1):141-6
references.pmid: 20843667
references.citation: Macromol Biosci. 2008 Jul 7;8(7):665-9
references.pmid: 18366149
references.citation: Langmuir. 2009 Apr 21;25(8):4402-11
references.pmid: 19366220
references.citation: J Mater Chem B. 2014 Aug 7;2(29):4640-4652
references.pmid: 32262276
references.citation: Mol Pharm. 2012 May 7;9(5):1067-76
references.pmid: 22482910
references.citation: Beilstein J Org Chem. 2012;8:1318-32
references.pmid: 23019466
references.citation: J Am Chem Soc. 2011 Nov 16;133(45):18014-7
references.pmid: 21985329
references.citation: Drug Deliv Transl Res. 2014 Feb;4(1):74-83
references.pmid: 25786618
references.citation: Biomacromolecules. 2013 Nov 11;14(11):3951-63
references.pmid: 24083552
references.citation: Theranostics. 2014 Jun 11;4(9):872-92
references.pmid: 25057313
references.citation: Methods Enzymol. 1999;304:191-213
references.pmid: 10372361
references.citation: Biomacromolecules. 2014 Mar 10;15(3):997-1001
references.pmid: 24547884
references.citation: Biomater Sci. 2013 Feb 3;1(2):152-170
references.pmid: 32481795
references.citation: Angew Chem Int Ed Engl. 2011 Jul 25;50(31):7027-31
references.pmid: 21698724
references.citation: Q Rev Biophys. 1988 May;21(2):129-228
references.pmid: 3043536
references.citation: Biomacromolecules. 2012 Dec 10;13(12):3926-37
references.pmid: 23130552
references.citation: Biomacromolecules. 2013 Oct 14;14(10):3643-52
references.pmid: 24040909
references.citation: Proc Natl Acad Sci U S A. 2007 Oct 16;104(42):16534-9
references.pmid: 17923669
references.citation: Biomaterials. 2013 Mar;34(10):2480-91
references.pmid: 23332177
references.citation: Macromol Biosci. 2014 Mar;14(3):359-68
references.pmid: 24634263
references.citation: J Am Chem Soc. 2010 Dec 1;132(47):16841-7
references.pmid: 21028803
references.citation: Drug Deliv Transl Res. 2014 Feb;4(1):96-103
references.pmid: 25786620
references.citation: Proc Natl Acad Sci U S A. 2007 Sep 4;104(36):14454-9
references.pmid: 17726101
references.citation: Mol Pharm. 2013 Apr 1;10(4):1299-305
references.pmid: 23398604
references.citation: Expert Opin Drug Deliv. 2011 Nov;8(11):1455-68
references.pmid: 21867463
references.citation: Biomaterials. 2013 Sep;34(27):6429-43
references.pmid: 23727262
references.citation: Mol Ther. 2012 Mar;20(3):513-24
references.pmid: 22252451
references.citation: J Control Release. 2008 Oct 21;131(2):137-44
references.pmid: 18700157
references.citation: ACS Cent Sci. 2015 Sep 23;1(6):303-312
references.pmid: 26436138
references.citation: J Phys Chem B. 2005 Dec 22;109(50):23816-22
references.pmid: 16375366
references.citation: J Phys Chem B. 2010 Feb 11;114(5):2028-32
references.pmid: 20078061
references.citation: Biomaterials. 2010 May;31(14):4186-93
references.pmid: 20149427
references.citation: J Control Release. 2012 Dec 28;164(3):387-93
references.pmid: 22789529
references.citation: Food Chem Toxicol. 2005 Oct;43(10):1451-9
references.pmid: 16018907
references.citation: Biophys J. 1999 Aug;77(2):915-24
references.pmid: 10423436
references.citation: Biomaterials. 2012 May;33(15):3952-8
references.pmid: 22386920
references.citation: Nat Mater. 2013 Nov;12(11):967-77
references.pmid: 24150415
references.citation: J Colloid Interface Sci. 2012 Dec 1;387(1):84-94
references.pmid: 22980740
references.citation: Biomacromolecules. 2013 Dec 9;14(12):4189-97
references.pmid: 24180231
references.citation: Biomaterials. 2012 Oct;33(29):7138-50
references.pmid: 22796160
references.citation: Expert Opin Biol Ther. 2015 Feb;15(2):269-85
references.pmid: 25399911
references.citation: J Control Release. 2010 Mar 19;142(3):416-21
references.pmid: 19919845
references.citation: Biomacromolecules. 2013 Dec 9;14(12):4289-300
references.pmid: 24144000
references.citation: J Mater Chem B. 2015 Feb 28;3(8):1495-1506
references.pmid: 32262422
references.citation: Biomaterials. 2012 Mar;33(8):2546-69
references.pmid: 22209641