Effects of the Lipophilic Core of Polymer Nanoassemblies on Intracellular Delivery and Transfection of siRNA

Steven Rheiner; Piotr Rychahou; Younsoo Bae; Steven Rheiner; Piotr Rychahou; Younsoo Bae

doi:10.3934/biophy.2015.3.284

AIMS Biophysics

2015, Volume 2, Issue 3: 284-302. doi: 10.3934/biophy.2015.3.284

Previous Article Next Article

Research article Special Issues

Effects of the Lipophilic Core of Polymer Nanoassemblies on Intracellular Delivery and Transfection of siRNA

1.
Department of Pharmaceutical Sciences, College of Pharmacy, University of Kentucky, 789 South Limestone, Lexington, KY 40536, USA;
2.
Markey Cancer Center, University of Kentucky, 800 Rose Street, CC140, Lexington, KY 40536;
3.
Department of Surgery, College of Medicine, University of Kentucky, 741 South Limestone, Lexington, KY 40536, USA

Received: 07 May 2015 Accepted: 02 July 2015 Published: 30 July 2015

Despite effective gene silencing in vitro, in vivo delivery and transfection of siRNA remain challenging due to the lack of carriers that protect siRNA stably in the body. This study is focused to elucidate the correlation between complex stability and transfection efficiency of siRNA carriers. The carriers were prepared by using polymer nanoassemblies made of a cationic branched polymer [poly(ethylene imine): bPEI] to which hydrophilic poly(ethylene glycol) polymers were tethered covalently. These polymer tethered nanoassemblies (TNAs) were further modified with lipophilic chains (palmitate: PAL) in the core to stabilize siRNA TNAs complexes through ionic and hydrophobic interactions in combination. The effects of PAL in the core of TNAs were investigated with respect to in vitro transfection, intracellular gene delivery, and toxicity of the complexes, using a human colon cancer HT29 cell line stably expressing a luciferase reporter gene. A commercial transfection agent (RNAiMax) was used as a control. TNAs entrapping siRNA showed the greatest complex stability in the absence of PAL although TNAs with a greater PAL content induced effective intracellular siRNA delivery, while luciferase expression decreased as the amount of PAL increased in the core of TNAs. These results demonstrate that lipophilic components in carriers affect not only complex stability but also intracellular distribution and transfection of siRNA in cancer cells.
- polymer nanoassemblies,
- gene therapy,
- siRNA delivery,
- non-viral vectors,
- cancer,
- nanotechnology
Citation: Steven Rheiner, Piotr Rychahou, Younsoo Bae. Effects of the Lipophilic Core of Polymer Nanoassemblies on Intracellular Delivery and Transfection of siRNA[J]. AIMS Biophysics, 2015, 2(3): 284-302. doi: 10.3934/biophy.2015.3.284

Related Papers:

Abstract

Despite effective gene silencing in vitro, in vivo delivery and transfection of siRNA remain challenging due to the lack of carriers that protect siRNA stably in the body. This study is focused to elucidate the correlation between complex stability and transfection efficiency of siRNA carriers. The carriers were prepared by using polymer nanoassemblies made of a cationic branched polymer [poly(ethylene imine): bPEI] to which hydrophilic poly(ethylene glycol) polymers were tethered covalently. These polymer tethered nanoassemblies (TNAs) were further modified with lipophilic chains (palmitate: PAL) in the core to stabilize siRNA TNAs complexes through ionic and hydrophobic interactions in combination. The effects of PAL in the core of TNAs were investigated with respect to in vitro transfection, intracellular gene delivery, and toxicity of the complexes, using a human colon cancer HT29 cell line stably expressing a luciferase reporter gene. A commercial transfection agent (RNAiMax) was used as a control. TNAs entrapping siRNA showed the greatest complex stability in the absence of PAL although TNAs with a greater PAL content induced effective intracellular siRNA delivery, while luciferase expression decreased as the amount of PAL increased in the core of TNAs. These results demonstrate that lipophilic components in carriers affect not only complex stability but also intracellular distribution and transfection of siRNA in cancer cells.

References

[1]	Resnier P, Montier T, Mathieu V, et al. (2013) A review of the current status of siRNA nanomedicines in the treatment of cancer. Biomaterials 34: 6429-6443. doi: 10.1016/j.biomaterials.2013.04.060
[2]	Oh Y-K, Park TG (2009) siRNA delivery systems for cancer treatment. Adv Drug Deliver Rev 61: 850-862. doi: 10.1016/j.addr.2009.04.018
[3]	Fire A (1999) RNA-triggered gene silencing. Trends Genet 15: 358-363. doi: 10.1016/S0168-9525(99)01818-1
[4]	Fire A, Xu SQ, Montgomery MK, et al. (1998) Potent and specific genetic interference by double-stranded RNA in Caenorhabditis elegans. Nature 391: 806-811. doi: 10.1038/35888
[5]	Aliabadi HM, Landry B, Sun C, et al. (2012) Supramolecular assemblies in functional siRNA delivery: where do we stand? Biomaterials 33: 2546-2569. doi: 10.1016/j.biomaterials.2011.11.079
[6]	Guo P, Coban O, Snead NM, et al. (2010) Engineering RNA for targeted siRNA delivery and medical application. Adv Drug Deliver Rev 62: 650-666. doi: 10.1016/j.addr.2010.03.008
[7]	Sheikhi Mehrabadi F, Fischer W, Haag R (2012) Dendritic and lipid-based carriers for gene/siRNA delivery (a review). Curr Opin Solid St M 16: 310-322. doi: 10.1016/j.cossms.2013.01.003
[8]	Gonzalez H, Hwang SJ, Davis ME (1999) New class of polymers for the delivery of macromolecular therapeutics. Bioconjugate Chem 10: 1068-1074. doi: 10.1021/bc990072j
[9]	Zhang S, Zhao B, Jiang H, et al. (2007) Cationic lipids and polymers mediated vectors for delivery of siRNA. J Control Release 123: 1-10. doi: 10.1016/j.jconrel.2007.07.016
[10]	Ballarín-González B, Ebbesen MF, Howard KA (2013) Polycation-based nanoparticles for RNAi-mediated cancer treatment. Cancer Lett.
[11]	Fischer D, Bieber T, Li Y, et al. (1999) A Novel Non-Viral Vector for DNA Delivery Based on Low Molecular Weight, Branched Polyethylenimine: Effect of Molecular Weight on Transfection Efficiency and Cytotoxicity. Pharm Res 16: 1273-1279. doi: 10.1023/A:1014861900478
[12]	Richards Grayson A, Doody A, Putnam D (2006) Biophysical and Structural Characterization of Polyethylenimine-Mediated siRNA Delivery in Vitro. Pharm Res 23: 1868-1876. doi: 10.1007/s11095-006-9009-2
[13]	Wightman L, Kircheis R, Rössler V, et al. (2001) Different behavior of branched and linear polyethylenimine for gene delivery in vitro and in vivo. J Gene Med 3: 362-372. doi: 10.1002/jgm.187
[14]	Merkel OM, Beyerle A, Beckmann BM, et al. (2011) Polymer-related off-target effects in non-viral siRNA delivery. Biomaterials 32: 2388-2398. doi: 10.1016/j.biomaterials.2010.11.081
[15]	Akhtar S, Benter I (2007) Toxicogenomics of non-viral drug delivery systems for RNAi: potential impact on siRNA-mediated gene silencing activity and specificity. Adv Drug Deliver Rev 59: 164-182. doi: 10.1016/j.addr.2007.03.010
[16]	Zuckerman JE, Choi CHJ, Han H, et al. (2012) Polycation-siRNA nanoparticles can disassemble at the kidney glomerular basement membrane. PNAS 109: 3137-3142. doi: 10.1073/pnas.1200718109
[17]	Nel AE, Madler L, Velegol D, et al. (2009) Understanding biophysicochemical interactions at the nano-bio interface. Nat Mater 8: 543-557. doi: 10.1038/nmat2442
[18]	Albanese A, Tang PS, Chan WCW (2012) The effect of nanoparticle size, shape, and surface chemistry on biological systems. Annu Rev Biomed Eng 14: 1-16. doi: 10.1146/annurev-bioeng-071811-150124
[19]	Lv H, Zhang S, Wang B, et al. (2006) Toxicity of cationic lipids and cationic polymers in gene delivery. J Control Release 114: 100-109. doi: 10.1016/j.jconrel.2006.04.014
[20]	He C, Hu Y, Yin L, et al. (2010) Effects of particle size and surface charge on cellular uptake and biodistribution of polymeric nanoparticles. Biomaterials 31: 3657-3666. doi: 10.1016/j.biomaterials.2010.01.065
[21]	Zintchenko A, Philipp A, Dehshahri A, et al. (2008) Simple modifications of branched PEI lead to highly efficient siRNA carriers with low toxicity. Bioconjugate Chem 19: 1448-1455. doi: 10.1021/bc800065f
[22]	Pasche S, Vörös J, Griesser HJ, et al. (2005) Effects of Ionic Strength and Surface Charge on Protein Adsorption at PEGylated Surfaces. J Phys Chem B 109: 17545-17552.
[23]	Miteva M, Kirkbride KC, Kilchrist KV, et al. (2014) Tuning PEGylation of mixed micelles to overcome intracellular and systemic siRNA delivery barriers. Biomaterials.
[24]	Zhu C, Jung S, Luo S, et al. (2010) Biomaterials Co-delivery of siRNA and paclitaxel into cancer cells by biodegradable cationic micelles based on PDMAEMA - PCL - PDMAEMA triblock copolymers. Biomaterials 31: 2408-2416. doi: 10.1016/j.biomaterials.2009.11.077
[25]	Kim D, Lee D, Jang YL, et al. (2012) Facial amphipathic deoxycholic acid-modified polyethyleneimine for efficient MMP-2 siRNA delivery in vascular smooth muscle cells. Eur J Pharm Biopharm 81: 14-23. doi: 10.1016/j.ejpb.2012.01.013
[26]	Hong J, Ku SH, Lee MS, et al. (2014) Cardiac RNAi therapy using RAGE siRNA/deoxycholic acid-modified polyethylenimine complexes for myocardial infarction. Biomaterials 35: 7562-7573. doi: 10.1016/j.biomaterials.2014.05.025
[27]	Patel MM, Anchordoquy TJ (2005) Contribution of Hydrophobicity to Thermodynamics of Ligand-DNA Binding and DNA Collapse. Biophys J 88: 2089-2103. doi: 10.1529/biophysj.104.052100
[28]	Kim HJ, Miyata K, Nomoto T, et al. (2014) siRNA delivery from triblock copolymer micelles with spatially-ordered compartments of PEG shell, siRNA-loaded intermediate layer, and hydrophobic core. Biomaterials 35: 4548-4556. doi: 10.1016/j.biomaterials.2014.02.016
[29]	Bartlett DW, Davis ME (2006) Insights into the kinetics of siRNA-mediated gene silencing from live-cell and live-animal bioluminescent imaging. Nucleic Acids Res 34: 322-333. doi: 10.1093/nar/gkj439
[30]	Matsumoto S, Christie RJ, Nishiyama N, et al. (2009) Environment-Responsive Block Copolymer Micelles with a Disulfide Cross-Linked Core for Enhanced siRNA Delivery. Biomacromolecules 10: 119-127. doi: 10.1021/bm800985e
[31]	Xia W, Wang P, Lin C, et al. (2012) Bioreducible polyethylenimine-delivered siRNA targeting human telomerase reverse transcriptase inhibits HepG2 cell growth in vitro and in vivo. J Control Release 157: 427-436. doi: 10.1016/j.jconrel.2011.10.011
[32]	Nelson CE, Kintzing JR, Hanna A, et al. (2013) Balancing cationic and hydrophobic content of PEGylated siRNA polyplexes enhances endosome escape, stability, blood circulation time, and bioactivity in vivo. ACS nano 7: 8870-8880. doi: 10.1021/nn403325f
[33]	Petros Ra, DeSimone JM (2010) Strategies in the design of nanoparticles for therapeutic applications. Nat Rev Drug Discov 9: 615-627. doi: 10.1038/nrd2591
[34]	Maeda H, Wu J, Sawa T, et al. (2000) Tumor vascular permeability and the EPR effect in macromolecular therapeutics: a review. J Control Release 65: 271-284. doi: 10.1016/S0168-3659(99)00248-5
[35]	Holzerny P, Ajdini B, Heusermann W, et al. (2012) Biophysical properties of chitosan/siRNA polyplexes: profiling the polymer/siRNA interactions and bioactivity. J Control Release 157: 297-304. doi: 10.1016/j.jconrel.2011.08.023
[36]	Guo G, Zhou L, Chen Z, et al. (2013) Alkane-modified low-molecular-weight polyethylenimine with enhanced gene silencing for siRNA delivery. Int J Pharm 450: 44-52. doi: 10.1016/j.ijpharm.2013.04.024
[37]	Canton I, Battaglia G (2012) Endocytosis at the nanoscale. Chem Soc Rev.
[38]	Noguchi A, Furuno T, Kawaura C, et al. (1998) Membrane fusion plays an important role in gene transfection mediated by cationic liposomes. FEBS Lett 433: 169-173. doi: 10.1016/S0014-5793(98)00837-0
[39]	Cevc G, Richardsen H (1999) Lipid vesicles and membrane fusion. Adv Drug Deliver Rev 38: 207-232. doi: 10.1016/S0169-409X(99)00030-7
[40]	Akinc A, Thomas M, Klibanov AM, et al. (2005) Exploring polyethylenimine-mediated DNA transfection and the proton sponge hypothesis. J Gene Med 7: 657-663.
[41]	Pack DW, Hoffman AS, Pun S, et al. (2005) Design and development of polymers for gene delivery. Nat Rev Drug Discov 4: 581-593. doi: 10.1038/nrd1775
[42]	Kurtulus I, Yilmaz G, Ucuncu M, et al. (2014) A new proton sponge polymer synthesized by RAFT polymerization for intracellular delivery of biotherapeutics. Polymer Chem 5: 1593-1604. doi: 10.1039/C3PY01244A
[43]	Richard I, Thibault M, De Crescenzo G, et al. (2013) Ionization Behavior of Chitosan and Chitosan-DNA Polyplexes Indicate That Chitosan Has a Similar Capability to Induce a Proton-Sponge Effect as PEI. Biomacromolecules 14: 1732-1740. doi: 10.1021/bm4000713

Reader Comments

Your name:*

Email:*
© 2015 the Author(s), licensee AIMS Press. This is an open access article distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/4.0)