Nanometer-scale siRNA carriers incorporating peptidomimetic oligomers: physical characterization and biological activity

Utku Konca, Yeliz; Kirshenbaum, Kent; Zuckermann, Ronald N.

doi:10.2147/ijn.s57449

Nanometer-scale siRNA carriers incorporating peptidomimetic oligomers: physical characterization and biological activity

Journal Article · Thu May 01 00:00:00 EDT 2014 · International Journal of Nanomedicine

DOI:https://doi.org/10.2147/ijn.s57449· OSTI ID:1627997

Utku Konca, Yeliz ^[1]; Kirshenbaum, Kent ^[2]; Zuckermann, Ronald N. ^[3]

Koc Univ., Istanbul (Turkey). Dept. of Chemistry; DOE/OSTI
New York Univ. (NYU), NY (United States). Dept. of Chemistry
Lawrence Berkeley National Lab. (LBNL), Berkeley, CA (United States). Molecular Foundry

Synthetic short interfering RNA (siRNA) oligonucleotides can trigger the RNA interference pathway and lead to selective gene silencing. Despite considerable enthusiasm and investment, formidable challenges remain that may deter translating this breakthrough discovery into clinical applications. In particular, the development of efficient, nontoxic, nonimmunogenic methods for delivering siRNA in vivo has proven to be exceptionally challenging. Thorough analysis of the relationship between the structure and function of siRNA carrier systems, both in isolation and in complex with RNA, will facilitate the design of efficient nonviral siRNA delivery vehicles. In this study, we explore the relationship between the physicochemical characteristics and the biological activity of “lipitoid” compounds as potent siRNA delivery vehicles. Lipitoids are cationic peptidomimetic oligomers incorporating a peptoid and a phospholipid moiety. Lipitoids can associate with siRNA oligonucleotides and self-assemble into spherical lipitoid-based nanoparticles (LNPs), with dimensions that are dependent upon the medium and the stoichiometric ratio between the cationic monomers of the lipitoid and anionic siRNA oligonucleotides. The morphology, gene silencing efficiency, and cytotoxicity of the siRNA-loaded LNPs are similarly sensitive to the stoichiometry of the complexes. The medium in which the LNPs are formed affects the assembled cargo particles’ characteristics such as particle size, transfection efficiency, and stability. Formation of the LNPs in the biological, serum-free medium OptiMEM resulted in LNPs an order of magnitude larger than LNPs formed in water, and were twice as efficient in siRNA transfection compared to LNPs formed in water. Inhibitor studies were conducted to elucidate the efficiency of lysosomal escape and the uptake mechanism of the siRNA-loaded LNPs. Our results suggest that these lipitoid-based, siRNA-loaded spherical LNPs are internalized through a lipid raft-dependent and dynaminmediated pathway, circumventing endosomal and lysosomal encapsulation. The lipitoid-siRNA nanospheres proved to be suitable platforms for investigating the critical parameters determining the efficiency of transfection agents, revealing the necessity for conducting characterization studies in biological media. The investigation of the LNP internalization pathway points to an alternative uptake route that bypasses the lysosome, explaining the surprisingly high efficiency of LNPs and suggesting that the uptake mechanism should be probed rather than assumed for the next generation of rationally designed transfection agents.

View Accepted Manuscript (DOE)

Research Organization:: Lawrence Berkeley National Laboratory (LBNL), Berkeley, CA (United States)

Sponsoring Organization:: USDOE Office of Science (SC), Biological and Environmental Research (BER). Biological Systems Science Division

Grant/Contract Number:: AC02-05CH11231

OSTI ID:: 1627997

Journal Information:: International Journal of Nanomedicine, Journal Name: International Journal of Nanomedicine Vol. 9; ISSN 1178-2013

Publisher:: Dove Medical PressCopyright Statement

Country of Publication:: United States

Language:: English

References (43)

Supravital exposure to propidium iodide identifies apoptosis on adherent cells Zamai, Loris; Canonico, Barbara; Luchetti, Francesca Cytometry, Vol. 44, Issue 1 https://doi.org/10.1002/1097-0320(20010501)44:1<57::aid-cyto1082>3.0.co;2-o	journal	January 2001
Current progress of siRNA/shRNA therapeutics in clinical trials Burnett, John C.; Rossi, John J.; Tiemann, Katrin Biotechnology Journal, Vol. 6, Issue 9 https://doi.org/10.1002/biot.201100054	journal	July 2011
Structure/Function Analysis of Peptoid/Lipitoid:DNA Complexes Lobo, Brian A.; Vetro, Joseph A.; Suich, Daniel M. Journal of Pharmaceutical Sciences, Vol. 92, Issue 9 https://doi.org/10.1002/jps.10450	journal	September 2003
Optimization of a lipitoid-based plasmid DNA transfection protocol for bovine trophectoderm CT-1 cells Schiffmacher, Andrew T.; Keefer, Carol L. In Vitro Cellular & Developmental Biology - Animal, Vol. 48, Issue 7 https://doi.org/10.1007/s11626-012-9525-9	journal	July 2012
Delivery of siRNA from lyophilized polymeric surfaces Andersen, Morten Ø.; Howard, Kenneth A.; Paludan, Søren R. Biomaterials, Vol. 29, Issue 4 https://doi.org/10.1016/j.biomaterials.2007.10.003	journal	February 2008
The role of adenosine receptor and caveolae-mediated endocytosis in oligonucleotide-mediated gene transfer Chung, Yi-Chen; Cheng, Ting-Yun; Young, Tai-Horng Biomaterials, Vol. 32, Issue 19 https://doi.org/10.1016/j.biomaterials.2011.02.060	journal	July 2011
A review of the current status of siRNA nanomedicines in the treatment of cancer Resnier, Pauline; Montier, Tristan; Mathieu, Véronique Biomaterials, Vol. 34, Issue 27 https://doi.org/10.1016/j.biomaterials.2013.04.060	journal	September 2013
RNA-Based Therapeutics: Current Progress and Future Prospects Burnett, John C.; Rossi, John J. Chemistry & Biology, Vol. 19, Issue 1 https://doi.org/10.1016/j.chembiol.2011.12.008	journal	January 2012
Comparison of different commercially available cationic liposome–DNA lipoplexes: Parameters influencing toxicity and transfection efficiency Masotti, A.; Mossa, G.; Cametti, C. Colloids and Surfaces B: Biointerfaces, Vol. 68, Issue 2 https://doi.org/10.1016/j.colsurfb.2008.09.017	journal	February 2009
Mechanistic investigation of poly(ethylene imine)-based siRNA delivery: Disulfide bonds boost intracellular release of the cargo Breunig, Miriam; Hozsa, Constantin; Lungwitz, Uta Journal of Controlled Release, Vol. 130, Issue 1 https://doi.org/10.1016/j.jconrel.2008.05.016	journal	August 2008
Lipitoids — novel cationic lipids for cellular delivery of plasmid DNA in vitro Huang, Chin-Yi; Uno, Tetsuo; Murphy, John E. Chemistry & Biology, Vol. 5, Issue 6 https://doi.org/10.1016/s1074-5521(98)90173-9	journal	June 1998
A Combinatorial Polymer Library Approach Yields Insight into Nonviral Gene Delivery Green, Jordan J.; Langer, Robert; Anderson, Daniel G. Accounts of Chemical Research, Vol. 41, Issue 6 https://doi.org/10.1021/ar7002336	journal	June 2008
Internally Cationic Polyamidoamine PAMAM-OH Dendrimers for siRNA Delivery: Effect of the Degree of Quaternization and Cancer Targeting Patil, Mahesh L.; Zhang, Min; Taratula, Oleh Biomacromolecules, Vol. 10, Issue 2 https://doi.org/10.1021/bm8009973	journal	January 2009
Efficient Delivery of Bcl-2-Targeted siRNA Using Cationic Polymer Nanoparticles: Downregulating mRNA Expression Level and Sensitizing Cancer Cells to Anticancer Drug Beh, Cyrus W.; Seow, Wei Yang; Wang, Yong Biomacromolecules, Vol. 10, Issue 1 https://doi.org/10.1021/bm801109g	journal	December 2008
Rapid Discovery of Potent siRNA-Containing Lipid Nanoparticles Enabled by Controlled Microfluidic Formulation Chen, Delai; Love, Kevin T.; Chen, Yi Journal of the American Chemical Society, Vol. 134, Issue 16 https://doi.org/10.1021/ja301621z	journal	April 2012
Functional Core/Shell Nanoparticles via Layer-by-Layer Assembly. Investigation of the Experimental Parameters for Controlling Particle Aggregation and for Enhancing Dispersion Stability Schneider, Grégory; Decher, Gero Langmuir, Vol. 24, Issue 5 https://doi.org/10.1021/la7021837	journal	January 2008
Nanotechnology in Drug Delivery and Tissue Engineering: From Discovery to Applications Shi, Jinjun; Votruba, Alexander R.; Farokhzad, Omid C. Nano Letters, Vol. 10, Issue 9 https://doi.org/10.1021/nl102184c	journal	September 2010
Molecular Parameters of siRNA–Cell Penetrating Peptide Nanocomplexes for Efficient Cellular Delivery van Asbeck, Alexander H.; Beyerle, Andrea; McNeill, Hesta ACS Nano, Vol. 7, Issue 5 https://doi.org/10.1021/nn305754c	journal	April 2013
Peptoid Polymers: A Highly Designable Bioinspired Material Sun, Jing; Zuckermann, Ronald N. ACS Nano, Vol. 7, Issue 6 https://doi.org/10.1021/nn4015714	journal	May 2013
A New Potent Secondary Amphipathic Cell–penetrating Peptide for siRNA Delivery Into Mammalian Cells Crombez, Laurence; Aldrian-Herrada, Gudrun; Konate, Karidia Molecular Therapy, Vol. 17, Issue 1 https://doi.org/10.1038/mt.2008.215	journal	January 2009
Development of Lipidoid–siRNA Formulations for Systemic Delivery to the Liver Akinc, Akin; Goldberg, Michael; Qin, June Molecular Therapy, Vol. 17, Issue 5 https://doi.org/10.1038/mt.2009.36	journal	May 2009
Rational design of cationic lipids for siRNA delivery Semple, Sean C.; Akinc, Akin; Chen, Jianxin Nature Biotechnology, Vol. 28, Issue 2 https://doi.org/10.1038/nbt.1602	journal	January 2010
Efficiency of siRNA delivery by lipid nanoparticles is limited by endocytic recycling Sahay, Gaurav; Querbes, William; Alabi, Christopher Nature Biotechnology, Vol. 31, Issue 7 https://doi.org/10.1038/nbt.2614	journal	June 2013
A combinatorial library of lipid-like materials for delivery of RNAi therapeutics Akinc, Akin; Zumbuehl, Andreas; Goldberg, Michael Nature Biotechnology, Vol. 26, Issue 5 https://doi.org/10.1038/nbt1402	journal	April 2008
Efficient polyethylenimine-mediated gene delivery proceeds via a caveolar pathway in HeLa cells. Gabrielson, Nathan; Pack, Daniel Nature Precedings https://doi.org/10.1038/npre.2008.2262.1	journal	September 2008
Caveolae: mining little caves for new cancer targets Carver, Lucy A.; Schnitzer, Jan E. Nature Reviews Cancer, Vol. 3, Issue 8 https://doi.org/10.1038/nrc1146	journal	August 2003
Knocking down barriers: advances in siRNA delivery Whitehead, Kathryn A.; Langer, Robert; Anderson, Daniel G. Nature Reviews Drug Discovery, Vol. 8, Issue 2 https://doi.org/10.1038/nrd2742	journal	February 2009
Bio-inspired, bioengineered and biomimetic drug delivery carriers Yoo, Jin-Wook; Irvine, Darrell J.; Discher, Dennis E. Nature Reviews Drug Discovery, Vol. 10, Issue 7 https://doi.org/10.1038/nrd3499	journal	July 2011
Clathrin- and non-clathrin-mediated endocytic regulation of cell signalling Le Roy, Christine; Wrana, Jeffrey L. Nature Reviews Molecular Cell Biology, Vol. 6, Issue 2 https://doi.org/10.1038/nrm1571	journal	February 2005
A peptidomimetic siRNA transfection reagent for highly effective gene silencing Utku, Yeliz; Dehan, Elinor; Ouerfelli, Ouathek Molecular BioSystems, Vol. 2, Issue 6-7 https://doi.org/10.1039/b603229j	journal	January 2006
Size-dependent internalization of particles via the pathways of clathrin- and caveolae-mediated endocytosis Rejman, Joanna; Oberle, Volker; Zuhorn, Inge S. Biochemical Journal, Vol. 377, Issue 1 https://doi.org/10.1042/bj20031253	journal	January 2004
The effect of particle design on cellular internalization pathways Gratton, S. E. A.; Ropp, P. A.; Pohlhaus, P. D. Proceedings of the National Academy of Sciences, Vol. 105, Issue 33, p. 11613-11618 https://doi.org/10.1073/pnas.0801763105	journal	August 2008
Lipid-like materials for low-dose, in vivo gene silencing Love, K. T.; Mahon, K. P.; Levins, C. G. Proceedings of the National Academy of Sciences, Vol. 107, Issue 5 https://doi.org/10.1073/pnas.0910603106	journal	January 2010
A versatile vector for gene and oligonucleotide transfer into cells in culture and in vivo: polyethylenimine. Boussif, O.; Lezoualc'h, F.; Zanta, M. A. Proceedings of the National Academy of Sciences, Vol. 92, Issue 16 https://doi.org/10.1073/pnas.92.16.7297	journal	August 1995
A combinatorial approach to the discovery of efficient cationic peptoid reagents for gene delivery Murphy, J. E.; Uno, T.; Hamer, J. D. Proceedings of the National Academy of Sciences, Vol. 95, Issue 4 https://doi.org/10.1073/pnas.95.4.1517	journal	February 1998
Caveolae/raft-dependent endocytosis Nabi, Ivan R.; Le, Phuong U. The Journal of Cell Biology, Vol. 161, Issue 4 https://doi.org/10.1083/jcb.200302028	journal	May 2003
The Business of RNAi Therapeutics Haussecker, Dirk Human Gene Therapy, Vol. 19, Issue 5 https://doi.org/10.1089/hum.2008.007	journal	May 2008
Caveolae Are Highly Immobile Plasma Membrane Microdomains, Which Are not Involved in Constitutive Endocytic Trafficking Thomsen, Peter; Roepstorff, Kirstine; Stahlhut, Martin Molecular Biology of the Cell, Vol. 13, Issue 1 https://doi.org/10.1091/mbc.01-06-0317	journal	January 2002
RNA interference is mediated by 21- and 22-nucleotide RNAs Elbashir, S. M. Genes & Development, Vol. 15, Issue 2 https://doi.org/10.1101/gad.862301	journal	January 2001
The Caveolae Membrane System Anderson, Richard G. W. Annual Review of Biochemistry, Vol. 67, Issue 1 https://doi.org/10.1146/annurev.biochem.67.1.199	journal	June 1998
Uptake and intracellular traffic of siRNA dendriplexes in glioblastoma cells and macrophages Morilla, Maria International Journal of Nanomedicine https://doi.org/10.2147/ijn.s25235	journal	November 2011
Cellular Delivery In Vivo of siRNA-Based Therapeutics Aigner, A. Current Pharmaceutical Design, Vol. 14, Issue 34 https://doi.org/10.2174/138161208786898815	journal	December 2008
Improving Conventional Enhanced Permeability and Retention (EPR) Effects; What Is the Appropriate Target? Kobayashi, Hisataka; Watanabe, Rira; Choyke, Peter L. Theranostics, Vol. 4, Issue 1 https://doi.org/10.7150/thno.7193	journal	January 2014

Cited By (4)

Sequence Programmable Peptoid Polymers for Diverse Materials Applications Knight, Abigail S.; Zhou, Effie Y.; Francis, Matthew B. Advanced Materials, Vol. 27, Issue 38 https://doi.org/10.1002/adma.201500275	journal	April 2015
Peptoids as tools and sensors Culf, Adrian S. Biopolymers, Vol. 110, Issue 6 https://doi.org/10.1002/bip.23285	journal	April 2019
Implementing Efficient Peptoid‐Mediated Delivery of RNA‐Based Therapeutics to the Vocal Folds Mukudai, Shigeyuki; Kraja, Iv; Bing, Renjie Laryngoscope Investigative Otolaryngology, Vol. 4, Issue 6 https://doi.org/10.1002/lio2.310	journal	October 2019
Tunable biomaterials from synthetic, sequence-controlled polymers Austin, Mariah J.; Rosales, Adrianne M. Biomaterials Science, Vol. 7, Issue 2 https://doi.org/10.1039/c8bm01215f	journal	January 2019

Similar Records

A peptidomimetic siRNA transfection reagent forhighly effectivegene silencing

Journal Article · Wed May 17 00:00:00 EDT 2006 · Molecular BioSystems · OSTI ID:918632

Structure And Gene Silencing Activities of Monovalent And Pentavalent Cationic Lipid Vectors Complexed With Sirna

Journal Article · Tue Jul 03 00:00:00 EDT 2007 · Biochem.46:4785-4792,2007 · OSTI ID:909557

Cationic Lipid-Nucleic Acid Complexes for Gene Delivery And Silencing: Pathways And Mechanisms for Plasmid Dna And Sirna

Journal Article · Tue Jul 17 00:00:00 EDT 2012 · Top.Curr.Chem. 296:191-226,2010 · OSTI ID:1046374

Related Subjects

59 BASIC BIOLOGICAL SCIENCES
Pharmacology & Pharmacy
Science & Technology - Other Topics
lipitoid
peptoid
siRNA delivery
therapeutic oligonucleotides

Nanometer-scale siRNA carriers incorporating peptidomimetic oligomers: physical characterization and biological activity

Citation Formats

References (43)

Cited By (4)

Similar Records

Related Subjects