Insights into cell entry and intracellular trafficking of peptide and protein drugs provided by electron microscopy

Advanced Drug Delivery Reviews

Volumn 65, Issue 8, 2013, Pages 1031-1038

  Margus, Helerin ^a   Padari, Kärt ^a   Pooga, Margus ^a  

^a UNIVERSITY OF TARTU   (Estonia)

Author keywords

Cell penetrating peptides; Electron microscopy; Immunolabeling; Intracellular trafficking; Peptide and protein therapeutics

Indexed keywords

CELL-PENETRATING PEPTIDE; IMMUNOFLUORESCENCE MICROSCOPY; IMMUNOLABELING; INTRACELLULAR DELIVERY; INTRACELLULAR TRAFFICKING; MICROSCOPY TECHNIQUE; PEPTIDE THERAPEUTICS; PROTEIN THERAPEUTICS;

CELLS; CYTOLOGY; ELECTRON MICROSCOPES; ELECTRON MICROSCOPY;

PEPTIDES;

DRUG CARRIER; EPIDERMAL GROWTH FACTOR RECEPTOR; PEPTIDE; RECOMBINANT PROTEIN; SUPERPARAMAGNETIC IRON OXIDE NANOPARTICLE; TRANSACTIVATOR PROTEIN;

CELL MIGRATION; CRYOELECTRON MICROSCOPY; DRUG MECHANISM; ELECTRON MICROSCOPY; ELECTRON TOMOGRAPHY; IMMUNOFLUORESCENCE MICROSCOPY; PRIORITY JOURNAL; PROTEIN ANALYSIS; REVIEW; TRANSMISSION ELECTRON MICROSCOPY;

EID: 84881023889     PISSN: 0169409X     EISSN: 18728294     Source Type: Journal    
DOI: 10.1016/j.addr.2013.04.013     Document Type: Review

References (95)

1. Leader B., Baca Q.J., Golan D.E. Protein therapeutics: a summary and pharmacological classification. Nat. Rev. Drug Discov. 2008, 7:21-39.
2. Swain S., Mondal D., Beg S., Patra C.N., Dinda S.C., Sruti J., Rao M.E. Stabilization and delivery approaches for protein and peptide pharmaceuticals: an extensive review of patents. Recent Pat. Biotechnol. 2013, (in press).
3. Agarwal P., Rupenthal I.D. Injectable implants for the sustained release of protein and peptide drugs. Drug Discov. Today 2013, 18:337-349.
4. Schermelleh L., Heintzmann R., Leonhardt H. A guide to super-resolution fluorescence microscopy. J. Cell Biol. 2010, 190:165-175.
5. Murk J.L., Posthuma G., Koster A.J., Geuze H.J., Verkleij A.J., Kleijmeer M.J., Humbel B.M. Influence of aldehyde fixation on the morphology of endosomes and lysosomes: quantitative analysis and electron tomography. J. Microsc. 2003, 212:81-90.
6. Sosinsky G.E., Crum J., Jones Y.Z., Lanman J., Smarr B., Terada M., Martone M.E., Deerinck T.J., Johnson J.E., Ellisman M.H. The combination of chemical fixation procedures with high pressure freezing and freeze substitution preserves highly labile tissue ultrastructure for electron tomography applications. J. Struct. Biol. 2008, 161:359-371.
7. Sosinsky G.E., Deerinck T.J., Greco R., Buitenhuys C.H., Bartol T.M., Ellisman M.H. Development of a model for microphysiological simulations: small nodes of Ranvier from peripheral nerves of mice reconstructed by electron tomography. Neuroinformatics 2005, 3:133-162.
8. Chan A., Holleran W.M., Ferguson T., Crumrine D., Goker-Alpan O., Schiffmann R., Tayebi N., Ginns E.I., Elias P.M., Sidransky E. Skin ultrastructural findings in type 2 Gaucher disease: diagnostic implications. Mol. Genet. Metab. 2011, 104:631-636.
9. Fowler D.J., Weber M.A., Anderson G., Malone M., Sebire N.J., Vellodi A. Ultrastructural features of Gaucher disease treated with enzyme replacement therapy presenting as mesenteric mass lesions. Fetal Pediatr. Pathol. 2006, 25:241-248.
10. Nadaj-Pakleza A., Lusakowska A., Sulek-Piatkowska A., Krysa W., Rajkiewicz M., Kwiecinski H., Kaminska A. Muscle pathology in myotonic dystrophy: light and electron microscopic investigation in eighteen patients. Folia Morphol. (Warsz) 2011, 70:121-129.
11. Hu Y., Qiao C., Lv M., Feng J., Yu M., Shen B., Zhang Q., Li Y. Arg9 facilitates the translocation and downstream signal inhibition of an anti-HER2 single chain antibody. BMC Res. Notes 2012, 5:336.
12. Tokuyasu K.T. A technique for ultracryotomy of cell suspensions and tissues. J. Cell Biol. 1973, 57:551-565.
13. Perera R.M., Zoncu R., Johns T.G., Pypaert M., Lee F.T., Mellman I., Old L.J., Toomre D.K., Scott A.M. Internalization, intracellular trafficking, and biodistribution of monoclonal antibody 806: a novel anti-epidermal growth factor receptor antibody. Neoplasia 2007, 9:1099-1110.
14. Austin C.D., De Maziere A.M., Pisacane P.I., van Dijk S.M., Eigenbrot C., Sliwkowski M.X., Klumperman J., Scheller R.H. Endocytosis and sorting of ErbB2 and the site of action of cancer therapeutics trastuzumab and geldanamycin. Mol. Biol. Cell 2004, 15:5268-5282.
15. Dubochet J., Adrian M., Chang J.J., Homo J.C., Lepault J., McDowall A.W., Schultz P. Cryo-electron microscopy of vitrified specimens. Q. Rev. Biophys. 1988, 21:129-228.
16. Saibil H.R. Conformational changes studied by cryo-electron microscopy. Nat. Struct. Biol. 2000, 7:711-714.
17. Saibil H.R. Macromolecular structure determination by cryo-electron microscopy. Acta Crystallogr. D: Biol. Crystallogr. 2000, 56:1215-1222.
18. Steven A.C., Aebi U. The next ice age: cryo-electron tomography of intact cells. Trends Cell Biol. 2003, 13:107-110.
19. Meryman H.T. Cryopreservation of living cells: principles and practice. Transfusion 2007, 47:935-945.
20. Galway M.E., Heckman J.W., Hyde G.J., Fowke L.C. Advances in high-pressure and plunge-freeze fixation. Methods Cell Biol. 1995, 49:3-19.
21. Richter T., Biel S.S., Sattler M., Wenck H., Wittern K.P., Wiesendanger R., Wepf R. Pros and cons: cryo-electron microscopic evaluation of block faces versus cryo-sections from frozen-hydrated skin specimens prepared by different techniques. J. Microsc. 2007, 225:201-207.
22. Kellenberger E. The potential of cryofixation and freeze substitution: observations and theoretical considerations. J. Microsc. 1991, 161:183-203.
23. Bohrmann B., Kellenberger E. Cryosubstitution of frozen biological specimens in electron microscopy: use and application as an alternative to chemical fixation. Micron 2001, 32:11-19.
24. Al-Amoudi A., Dubochet J., Studer D. Amorphous solid water produced by cryosectioning of crystalline ice at 113K. J. Microsc. 2002, 207:146-153.
25. Al-Amoudi A., Norlen L.P., Dubochet J. Cryo-electron microscopy of vitreous sections of native biological cells and tissues. J. Struct. Biol. 2004, 148:131-135.
26. Al-Amoudi A., Studer D., Dubochet J. Cutting artefacts and cutting process in vitreous sections for cryo-electron microscopy. J. Struct. Biol. 2005, 150:109-121.
27. Al-Amoudi A., Chang J.J., Leforestier A., McDowall A., Salamin L.M., Norlen L.P., Richter K., Blanc N.S., Studer D., Dubochet J. Cryo-electron microscopy of vitreous sections. EMBO J. 2004, 23:3583-3588.
28. Matias V.R., Beveridge T.J. Cryo-electron microscopy reveals native polymeric cell wall structure in Bacillus subtilis 168 and the existence of a periplasmic space. Mol. Microbiol. 2005, 56:240-251.
29. Sosinsky G.E., Giepmans B.N., Deerinck T.J., Gaietta G.M., Ellisman M.H. Markers for correlated light and electron microscopy. Methods Cell Biol. 2007, 79:575-591.
30. Gaietta G., Deerinck T.J., Adams S.R., Bouwer J., Tour O., Laird D.W., Sosinsky G.E., Tsien R.Y., Ellisman M.H. Multicolor and electron microscopic imaging of connexin trafficking. Science 2002, 296:503-507.
31. Gaietta G.M., Giepmans B.N., Deerinck T.J., Smith W.B., Ngan L., Llopis J., Adams S.R., Tsien R.Y., Ellisman M.H. Golgi twins in late mitosis revealed by genetically encoded tags for live cell imaging and correlated electron microscopy. Proc. Natl. Acad. Sci. U. S. A. 2006, 103:17777-17782.
32. Nair B.G., Fukuda T., Mizuki T., Hanajiri T., Maekawa T. Intracellular trafficking of superparamagnetic iron oxide nanoparticles conjugated with TAT peptide: 3-dimensional electron tomography analysis. Biochem. Biophys. Res. Commun. 2012, 421:763-767.
33. Bennett A., Liu J., Van Ryk D., Bliss D., Arthos J., Henderson R.M., Subramaniam S. Cryoelectron tomographic analysis of an HIV-neutralizing protein and its complex with native viral gp120. J. Biol. Chem. 2007, 282:27754-27759.
34. Sander B., Golas M.M. Visualization of bionanostructures using transmission electron microscopical techniques. Microsc. Res. Tech. 2011, 74:642-663.
35. Le Bihan O., Bonnafous P., Marak L., Bickel T., Trepout S., Mornet S., De Haas F., Talbot H., Taveau J.C., Lambert O. Cryo-electron tomography of nanoparticle transmigration into liposome. J. Struct. Biol. 2009, 168:419-425.
36. Hainfeld J.F., Furuya F.R. A 1.4-nm gold cluster covalently attached to antibodies improves immunolabeling. J. Histochem. Cytochem. 1992, 40:177-184.
37. Hainfeld J.F., Powell R.D. New frontiers in gold labeling. J. Histochem. Cytochem. 2000, 48:471-480.
38. Melan M.A., Sluder G. Redistribution and differential extraction of soluble proteins in permeabilized cultured cells. Implications for immunofluorescence microscopy. J. Cell Sci. 1992, 101(Pt 4):731-743.
39. Faulk W.P., Taylor G.M. An immunocolloid method for the electron microscope. Immunochemistry 1971, 8:1081-1083.
40. Geoghegan W.D., Ackerman G.A. Adsorption of horseradish peroxidase, ovomucoid and anti-immunoglobulin to colloidal gold for the indirect detection of concanavalin A, wheat germ agglutinin and goat anti-human immunoglobulin G on cell surfaces at the electron microscopic level: a new method, theory and application. J. Histochem. Cytochem. 1977, 25:1187-1200.
41. Humbel B.M., Sibon O.C., Stierhof Y.D., Schwarz H. Ultra-small gold particles and silver enhancement as a detection system in immunolabeling and in situ hybridization experiments. J. Histochem. Cytochem. 1995, 43:735-737.
42. Padari K., Koppel K., Lorents A., Hällbrink M., Mano M., Pedroso de Lima M.C., Pooga M. S4(13)-PV cell-penetrating peptide forms nanoparticle-like structures to gain entry into cells. Bioconjug. Chem. 2010, 21:774-783.
43. Arai R., Geffard M., Calas A. Intensification of labelings of the immunogold silver staining method by gold toning. Brain Res. Bull. 1992, 28:343-345.
44. Zuhorn I.S., Engberts J.B., Hoekstra D. Gene delivery by cationic lipid vectors: overcoming cellular barriers. Eur. Biophys. J. 2007, 36:349-362.
45. Nakamura Y., Kogure K., Futaki S., Harashima H. Octaarginine-modified multifunctional envelope-type nano device for siRNA. J. Control. Release 2007, 119:360-367.
46. Langel Ü. Handbook of Cell-penetrating Peptides 2006, CRC Press, Boca Raton, London, New York, Washington. 2nd ed.
47. Koren E., Apte A., Jani A., Torchilin V.P. Multifunctional PEGylated 2C5-immunoliposomes containing pH-sensitive bonds and TAT peptide for enhanced tumor cell internalization and cytotoxicity. J. Control. Release 2012, 160:264-273.
48. Brooks N.A., Pouniotis D.S., Sheng K.C., Apostolopoulos V., Pietersz G.A. A membrane penetrating multiple antigen peptide (MAP) incorporating ovalbumin CD8 epitope induces potent immune responses in mice. Biochim. Biophys. Acta 2010, 1798:2286-2295.
49. Walker L., Perkins E., Kratz F., Raucher D. Cell penetrating peptides fused to a thermally targeted biopolymer drug carrier improve the delivery and antitumor efficacy of an acid-sensitive doxorubicin derivative. Int. J. Pharm. 2012, 436:825-832.
50. Foged C., Nielsen H.M. Cell-penetrating peptides for drug delivery across membrane barriers. Expert Opin. Drug Deliv. 2008, 5:105-117.
51. Ezzat K., El Andaloussi S., Abdo R., Langel Ü. Peptide-based matrices as drug delivery vehicles. Curr. Pharm. Des. 2010, 16:1167-1178.
52. Johnson R.M., Harrison S.D., Maclean D. Therapeutic applications of cell-penetrating peptides. Methods Mol. Biol. 2011, 683:535-551.
53. Asoh S., Ohsawa I., Mori T., Katsura K.-I., Hiraide T., Katayama Y., Kimura M., Ozaki D., Yamagata K., Ohta S. Protection against ischemic brain injury by protein therapeutics. Proc. Natl. Acad. Sci. U. S. A. 2002, 99:17107-17112.
54. Cao G., Pei W., Ge H., Liang Q., Luo Y., Sharp F.R., Lu A., Ran R., Graham S.H., Chen J. In vivo delivery of a Bcl-xL fusion protein containing the TAT protein transduction domain protects against ischemic brain injury and neuronal apoptosis. J. Neurosci. 2002, 22:5423-5431.
55. Teesalu T., Sugahara K.N., Kotamraju V.R., Ruoslahti E. C-end rule peptides mediate neuropilin-1-dependent cell, vascular, and tissue penetration. Proc. Natl. Acad. Sci. U. S. A. 2009, 106:16157-16162.
56. Sugahara K.N., Teesalu T., Karmali P.P., Kotamraju V.R., Agemy L., Girard O.M., Hanahan D., Mattrey R.F., Ruoslahti E. Tissue-penetrating delivery of compounds and nanoparticles into tumors. Cancer Cell 2009, 16:510-520.
57. Olson E.S., Aguilera T.A., Jiang T., Ellies L.G., Nguyen Q.T., Wong E.H., Gross L.A., Tsien R.Y. In vivo characterization of activatable cell penetrating peptides for targeting protease activity in cancer. Integr. Biol. (Camb) 2009, 1:382-393.
58. Aguilera T.A., Olson E.S., Timmers M.M., Jiang T., Tsien R.Y. Systemic in vivo distribution of activatable cell penetrating peptides is superior to that of cell penetrating peptides. Integr. Biol. (Camb) 2009, 1:371-381.
59. Padari K., Säälik P., Hansen M., Koppel K., Raid R., Langel Ü., Pooga M. Cell transduction pathways of transportans. Bioconjug. Chem. 2005, 16:1399-1410.
60. El-Andaloussi S., Jarver P., Johansson H.J., Langel Ü. Cargo-dependent cytotoxicity and delivery efficacy of cell-penetrating peptides: a comparative study. Biochem. J. 2007, 407:285-292.
61. Mueller J., Kretzschmar I., Volkmer R., Boisguerin P. Comparison of cellular uptake using 22 CPPs in 4 different cell lines. Bioconjug. Chem. 2008, 19:2363-2374.
62. Nakase I., Niwa M., Takeuchi T., Sonomura K., Kawabata N., Koike Y., Takehashi M., Tanaka S., Ueda K., Simpson J.C., Jones A.T., Sugiura Y., Futaki S. Cellular uptake of arginine-rich peptides: roles for macropinocytosis and actin rearrangement. Mol. Ther. 2004, 10:1011-1022.
63. Richard J.P., Melikov K., Brooks H., Prevot P., Lebleu B., Chernomordik L.V. Cellular uptake of unconjugated TAT peptide involves clathrin-dependent endocytosis and heparan sulfate receptors. J. Biol. Chem. 2005, 280:15300-15306.
64. Jones A.T. Macropinocytosis: searching for an endocytic identity and role in the uptake of cell penetrating peptides. J. Cell. Mol. Med. 2007, 11:670-684.
65. Lundin P., Johansson H., Guterstam P., Holm T., Hansen M., Langel Ü., E.L.A.S Distinct uptake routes of cell-penetrating peptide conjugates. Bioconjug. Chem. 2008, 19:2535-2542.
66. El Andaloussi S., Said Hassane F., Boisguerin P., Sillard R., Langel Ü., Lebleu B. Cell-penetrating peptides-based strategies for the delivery of splice redirecting antisense oligonucleotides. Methods Mol. Biol. 2011, 764:75-89.
67. Morris M.C., Deshayes S., Heitz F., Divita G. Cell-penetrating peptides: from molecular mechanisms to therapeutics. Biol. Cell 2008, 100:201-217.
68. Räägel H., Säälik P., Pooga M. Peptide-mediated protein delivery - which pathways are penetrable?. Biochim. Biophys. Acta 2010, 1798:2240-2248.
69. Nakase I., Tadokoro A., Kawabata N., Takeuchi T., Katoh H., Hiramoto K., Negishi M., Nomizu M., Sugiura Y., Futaki S. Interaction of arginine-rich peptides with membrane-associated proteoglycans is crucial for induction of actin organization and macropinocytosis. Biochemistry 2007, 46:492-501.
70. Tyagi M., Rusnati M., Presta M., Giacca M. Internalization of HIV-1 Tat requires cell surface heparan sulfate proteoglycans. J. Biol. Chem. 2001, 276:3254-3261. (Epub 2000 Oct 3256).
71. Poon G.M., Gariepy J. Cell-surface proteoglycans as molecular portals for cationic peptide and polymer entry into cells. Biochem. Soc. Trans. 2007, 35:788-793.
72. Ziegler A., Nervi P., Dürrenberger M., Seelig J. The cationic cell-penetrating peptide CPP(TAT) derived from the HIV-1 protein TAT is rapidly transported into living fibroblasts: optical, biophysical, and metabolic evidence. Biochemistry 2005, 44:138-148.
73. Pujals S., Fernandez-Carneado J., Lopez-Iglesias C., Kogan M.J., Giralt E. Mechanistic aspects of CPP-mediated intracellular drug delivery: relevance of CPP self-assembly. Biochim. Biophys. Acta 2006, 1758:264-279.
74. Munoz-Morris M.A., Heitz F., Divita G., Morris M.C. The peptide carrier Pep-1 forms biologically efficient nanoparticle complexes. Biochem. Biophys. Res. Commun. 2007, 355:877-882.
75. Henriques S.T., Costa J., Castanho M.A. Re-evaluating the role of strongly charged sequences in amphipathic cell-penetrating peptides: a fluorescence study using Pep-1. FEBS Lett. 2005, 579:4498-4502.
76. Crombez L., Morris M.C., Deshayes S., Heitz F., Divita G. Peptide-based nanoparticle for ex vivo and in vivo drug delivery. Curr. Pharm. Des. 2008, 14:3656-3665.
77. Palm-Apergi C., Lorents A., Padari K., Pooga M., Hällbrink M. The membrane repair response masks membrane disturbances caused by cell-penetrating peptide uptake. FASEB J. 2009, 23:214-223.
78. Rydström A., Deshayes S., Konate K., Crombez L., Padari K., Boukhaddaoui H., Aldrian G., Pooga M., Divita G. Direct translocation as major cellular uptake for CADY self-assembling peptide-based nanoparticles. PLoS One 2011, 6:e25924.
79. Oskolkov N., Arukuusk P., Copolovici D.M., Lindberg S., Margus H., Padari K., Pooga M., Langel Ü. NickFects, phosphorylated derivatives of transportan 10 for cellular delivery of oligonucleotides. Int. J. Pept. Res. Ther. 2011, 17:147-157.
80. Ezzat K., Helmfors H., Tudoran O., Juks C., Lindberg S., Padari K., El-Andaloussi S., Pooga M., Langel Ü. Scavenger receptor-mediated uptake of cell-penetrating peptide nanocomplexes with oligonucleotides. FASEB J. 2012, 26:1172-1180.
81. Cardoso A.M., Trabulo S., Cardoso A.L., Lorents A., Morais C.M., Gomes P., Nunes C., Lucio M., Reis S., Padari K., Pooga M., Pedroso de Lima M.C., Jurado A.S. S4(13)-PV cell-penetrating peptide induces physical and morphological changes in membrane-mimetic lipid systems and cell membranes: implications for cell internalization. Biochim. Biophys. Acta 2012, 1818:877-888.
82. Säälik P., Padari K., Niinep A., Lorents A., Hansen M., Jokitalo E., Langel Ü., Pooga M. Protein delivery with transportans is mediated by caveolae rather than flotillin-dependent pathways. Bioconjug. Chem. 2009, 20:877-887.
83. Wadia J.S., Stan R.V., Dowdy S.F. Transducible TAT-HA fusogenic peptide enhances escape of TAT-fusion proteins after lipid raft macropinocytosis. Nat. Med. 2004, 10:310-315.
84. Khalil I.A., Kogure K., Futaki S., Harashima H. High density of octaarginine stimulates macropinocytosis leading to efficient intracellular trafficking for gene expression. J. Biol. Chem. 2006, 281:3544-3551.
85. Al Soraj M., He L., Peynshaert K., Cousaert J., Vercauteren D., Braeckmans K., De Smedt S.C., Jones A.T. siRNA and pharmacological inhibition of endocytic pathways to characterize the differential role of macropinocytosis and the actin cytoskeleton on cellular uptake of dextran and cationic cell penetrating peptides octaarginine (R8) and HIV-Tat. J. Control. Release 2012, 161:132-141.
86. Gerbal-Chaloin S., Gondeau C., Aldrian-Herrada G., Heitz F., Gauthier-Rouviere C., Divita G. First step of the cell-penetrating peptide mechanism involves Rac1 GTPase-dependent actin-network remodelling. Biol. Cell 2007, 99:223-238.
87. Magzoub M., Sandgren S., Lundberg P., Oglecka K., Lilja J., Wittrup A., Goran Eriksson L.E., Langel Ü., Belting M., Graslund A. N-terminal peptides from unprocessed prion proteins enter cells by macropinocytosis. Biochem. Biophys. Res. Commun. 2006, 348:379-385.
88. Löfgren K., Wöhlstrom A., Lundberg P., Langel Ü., Gräslund A., Bedecs K. Antiprion properties of prion protein-derived cell-penetrating peptides. FASEB J. 2008, 22:2177-2184.
89. Glebov O.O., Bright N.A., Nichols B.J. Flotillin-1 defines a clathrin-independent endocytic pathway in mammalian cells. Nat. Cell Biol. 2006, 8:46-54.
90. El-Andaloussi S., Johansson H., Lundberg P., Langel Ü. Induction of splice correction by cell-penetrating peptide nucleic acids. J. Gene Med. 2006, 8:1262-1273.
91. Krpetic Z., Saleemi S., Prior I.A., See V., Qureshi R., Brust M. Negotiation of intracellular membrane barriers by TAT-modified gold nanoparticles. ACS Nano. 2011, 5:5195-5201.
92. Mäe M., El Andaloussi S., Lundin P., Oskolkov N., Johansson H.J., Guterstam P., Langel Ü. A stearylated CPP for delivery of splice correcting oligonucleotides using a non-covalent co-incubation strategy. J. Control. Release 2009, 134:221-227.
93. Lehto T., Abes R., Oskolkov N., Suhorutsenko J., Copolovici D.M., Mäger I., Viola J.R., Simonson O.E., Ezzat K., Guterstam P., Eriste E., Smith C.I., Lebleu B., Samir El A., Langel Ü. Delivery of nucleic acids with a stearylated (RxR)4 peptide using a non-covalent co-incubation strategy. J. Control. Release 2010, 141:42-51.
94. El-Andaloussi S., Lehto T., Mäger I., Rosenthal-Aizman K., Oprea, Simonson O.E., Sork H., Ezzat K., Copolovici D.M., Kurrikoff K., Viola J.R., Zaghloul E.M., Sillard R., Johansson H.J., Said Hassane F., Guterstam P., Suhorutsenko J., Moreno P.M., Oskolkov N., Halldin J., Tedebark U., Metspalu A., Lebleu B., Lehtio J., Smith C.I., Langel Ü. Design of a peptide-based vector, PepFect6, for efficient delivery of siRNA in cell culture and systemically in vivo. Nucleic Acids Res. 2011, 39:3972-3987.
95. Lehto T., Kurrikoff K., Langel Ü. Cell-penetrating peptides for the delivery of nucleic acids. Expert Opin. Drug Deliv. 2012, 9:823-836.