Erratum: Aptamers as targeted therapeutics: Current potential and challenges (Nature Reviews Drug Discovery (2017) 16 (181-202) DOI: 10.1038/nrd.2016.199);Aptamers as targeted therapeutics: Current potential and challenges

Nature Reviews Drug Discovery

Volumn 16, Issue 3, 2017, Pages 181-202

  Zhou, Jiehua ^a   Rossi, John ^a  

^a CITY OF HOPE NATIONAL MEDICAL CENTER   (United States)

Author keywords

[No Author keywords available]

Indexed keywords

APTAMER; BLOOD CLOTTING FACTOR; CD134 ANTIGEN; CD28 ANTIGEN; CD40 ANTIGEN; CHOLESTEROL; MICRORNA; NANOMATERIAL; NANOPARTICLE; NUCLEASE; PACLITAXEL; PEGAPTANIB; PROTHROMBIN; SMALL INTERFERING RNA; THROMBIN; TUBERCULIN; NUCLEIC ACID;

DRUG FORMULATION; DRUG TARGETING; GLOMERULUS FILTRATION; HUMAN; MACULAR DEGENERATION; NONHUMAN; ONCOLOGY; PHYSICAL CHEMISTRY; PROTEIN DEGRADATION; REVIEW; SYSTEMATIC EVOLUTION OF LIGANDS BY EXPONENTIAL ENRICHMENT APTAMER TECHNIQUE; TECHNOLOGY; TOXICITY; WHOLE CELL; ANIMAL; DRUG DELIVERY SYSTEM; GENETICS; METABOLISM; PROCEDURES; TRENDS;

ANIMALS; APTAMERS, NUCLEOTIDE; DRUG DELIVERY SYSTEMS; HUMANS; NUCLEIC ACIDS; RNA, SMALL INTERFERING; SELEX APTAMER TECHNIQUE;

EID: 84994134193     PISSN: 14741776     EISSN: 14741784     Source Type: Journal    
DOI: 10.1038/nrd.2017.86     Document Type: Erratum

References (250)

1. Tuerk, C. & Gold, L. Systematic evolution of ligands by exponential enrichment: RNA ligands to bacteriophage T4 DNA polymerase. Science 249, 505-510 (1990).
2. Robertson, D. L. & Joyce, G. F. Selection in vitro of an RNA enzyme that specifically cleaves single-stranded DNA. Nature 344, 467-468 (1990).
3. Ellington, A. D. & Szostak, J. W. In vitro selection of RNA molecules that bind specific ligands. Nature 346, 818-822 (1990).
4. Mayer, G. The chemical biology of aptamers. Angew. Chem. Int. Ed. 48, 2672-2689 (2009).
5. Gelinas, A. D., Davies, D. R. & Janjic, N. Embracing proteins: structural themes in aptamer-protein complexes. Curr. Opin. Struct. Biol. 36, 122-132 (2016).
6. Gold, L. Oligonucleotides as research, diagnostic, and therapeutic agents. J. Biol. Chem. 270, 13581-13584 (1995).
7. Geiger, A., Burgstaller, P., von der Eltz, H., Roeder, A. & Famulok, M. RNA aptamers that bind Larginine with sub-micromolar dissociation constants and high enantioselectivity. Nucleic Acids Res. 24, 1029-1036 (1996).
8. Jenison, R. D., Gill, S. C., Pardi, A. & Polisky, B. High-resolution molecular discrimination by RNA. Science 263, 1425-1429 (1994).
9. Sassanfar, M. & Szostak, J. W. An RNA motif that binds ATP. Nature 364, 550-553 (1993).
10. Chen, L. et al. The isolation of an RNA aptamer targeting to p53 protein with single amino acid mutation. Proc. Natl Acad. Sci. USA 112, 10002-10007 (2015).
11. Keefe, A. D., Pai, S. & Ellington, A. Aptamers as therapeutics. Nat. Rev. Drug Discov. 9, 537-550 (2010).
12. Sundaram, P., Kurniawan, H., Byrne, M. E. & Wower, J. Therapeutic RNA aptamers in clinical trials. Eur. J. Pharm. Sci. 48, 259-271 (2013).
13. Ecker, D. M., Jones, S. D. & Levine, H. L. The therapeutic monoclonal antibody market. MAbs 7, 9-14 (2015).
14. Ng, E. W. et al. Pegaptanib, a targeted anti-VEGF aptamer for ocular vascular disease. Nat. Rev. Drug Discov. 5, 123-132 (2006).
15. Siddiqui, M. A. & Keating, G. M. Pegaptanib: in exudative age-related macular degeneration. Drugs 65, 1571-1577; discussion 1578-1579 (2005).
16. Mousa, S. A. & Mousa, S. S. Current status of vascular endothelial growth factor inhibition in age-related macular degeneration. BioDrugs 24, 183-194 (2010).
17. Ferrara, N. & Adamis, A. P. Ten years of anti-vascular endothelial growth factor therapy. Nat. Rev. Drug Discov. 15, 385-403 (2016).
18. Lincoff, A. M. et al. Effect of the REG1 anticoagulation system versus bivalirudin on outcomes after percutaneous coronary intervention (REGULATE-PCI): A randomised clinical trial. Lancet 387, 349-356 (2016).
19. Verheugt, F. W. An anticoagulant too good to be true for revascularisation. Lancet 387, 314-315 (2016).
20. Bock, L. C., Griffin, L. C., Latham, J. A., Vermaas, E. H. & Toole, J. J. Selection of single-stranded DNA molecules that bind and inhibit human thrombin. Nature 355, 564-566 (1992).
21. Shu, Y. et al. Stable RNA nanoparticles as potential new generation drugs for cancer therapy. Adv. Drug Deliv. Rev. 66, 74-89 (2014).
22. Kulbachinskiy, A. V. Methods for selection of aptamers to protein targets. Biochemistry (Mosc.) 72, 1505-1518 (2007).
23. Ozer, A., Pagano, J. M. & Lis, J. T. New technologies provide quantum changes in the scale, speed, and success of SELEX methods and aptamer characterization. Mol. Ther. Nucleic Acids 3, e183 (2014).
24. Keefe, A. D. & Cload, S. T. SELEX with modified nucleotides. Curr. Opin. Chem. Biol. 12, 448-456 (2008).
25. Pieken, W. A., Olsen, D. B., Benseler, F., Aurup, H. & Eckstein, F. Kinetic characterization of ribonuclease-resistant 2?modified hammerhead ribozymes. Science 253, 314-317 (1991).
26. Nimjee, S. M., Rusconi, C. P. & Sullenger, B. A. Aptamers: An emerging class of therapeutics. Annu. Rev. Med. 56, 555-583 (2005).
27. Pestourie, C. et al. Comparison of different strategies to select aptamers against a transmembrane protein target. Oligonucleotides 16, 323-335 (2006).
28. Cerchia, L. & de Franciscis, V. Targeting cancer cells with nucleic acid aptamers. Trends Biotechnol. 28, 517-525 (2010).
29. Cerchia, L. et al. Neutralizing aptamers from whole-cell SELEX inhibit the RET receptor tyrosine kinase. PLoS Biol. 3, e123 (2005).
30. Liu, Y. et al. Aptamers selected against the unglycosylated EGFRvIII ectodomain and delivered intracellularly reduce membrane-bound EGFRvIII and induce apoptosis. Biol. Chem. 390, 137-144 (2009).
31. Guo, K. T., Paul, A., Schichor, C., Ziemer, G. & Wendel, H. P. Cell-SELEX: novel perspectives of aptamer-based therapeutics. Int. J. Mol. Sci. 9, 668-678 (2008).
32. Ohuchi, S. Cell-SELEX technology. Biores Open Access 1, 265-272 (2012).
33. Cerchia, L., Giangrande, P. H., McNamara, J. O. & de Franciscis, V. Cell-specific aptamers for targeted therapies. Methods Mol. Biol. 535, 59-78 (2009).
34. Tang, Z. et al. Selection of aptamers for molecular recognition and characterization of cancer cells. Anal. Chem. 79, 4900-4907 (2007).
35. Xiao, Z., Shangguan, D., Cao, Z., Fang, X. & Tan, W. Cell-specific internalization study of an aptamer from whole cell selection. Chemistry 14, 1769-1775 (2008).
36. Phillips, J. A., Lopez-Colon, D., Zhu, Z., Xu, Y. & Tan, W. Applications of aptamers in cancer cell biology. Anal. Chim. Acta 621, 101-108 (2008).
37. Avci-Adali, M., Metzger, M., Perle, N., Ziemer, G. & Wendel, H. P. Pitfalls of cell-systematic evolution of ligands by exponential enrichment (SELEX): existing dead cells during in vitro selection anticipate the enrichment of specific aptamers. Oligonucleotides 20, 317-323 (2010).
38. Raddatz, M. S. et al. Enrichment of cell-targeting and population-specific aptamers by fluorescence-activated cell sorting. Angew. Chem. Int. Ed. 47, 5190-5193 (2008).
39. Healy, J. M. et al. Pharmacokinetics and biodistribution of novel aptamer compositions. Pharm. Res. 21, 2234-2246 (2004).
40. Mi, J. et al. In vivo selection of tumor-targeting RNA motifs. Nat. Chem. Biol. 6, 22-24 (2010).
41. Cheng, C., Chen, Y. H., Lennox, K. A., Behlke, M. A. & Davidson, B. L. In vivo SELEX for identification of brain-penetrating aptamers. Mol. Ther. Nucleic Acids 2, e67 (2013).
42. Darmostuk, M., Rimpelova, S., Gbelcova, H. & Ruml, T. Current approaches in SELEX: An update to aptamer selection technology. Biotechnol. Adv. 33, 1141-1161 (2015).
43. Ellington, A. D. & Szostak, J. W. Selection in vitro of single-stranded DNA molecules that fold into specific ligand-binding structures. Nature 355, 850-852 (1992).
44. Drabovich, A. P., Berezovski, M., Okhonin, V. & Krylov, S. N. Selection of smart aptamers by methods of kinetic capillary electrophoresis. Anal. Chem. 78, 3171-3178 (2006).
45. Yufa, R. et al. Emulsion PCR significantly improves nonequilibrium capillary electrophoresis of equilibrium mixtures-based aptamer selection: Allowing for efficient and rapid selection of aptamer to unmodified ABH2 protein. Anal. Chem. 87, 1411-1419 (2015).
46. Mosing, R. K., Mendonsa, S. D. & Bowser, M. T. Capillary electrophoresis-SELEX selection of aptamers with affinity for HIV1 reverse transcriptase. Anal. Chem. 77, 6107-6112 (2005).
47. Miyachi, Y., Shimizu, N., Ogino, C. & Kondo, A. Selection of DNA aptamers using atomic force microscopy. Nucleic Acids Res. 38, e21 (2010).
48. Peng, L., Stephens, B. J., Bonin, K., Cubicciotti, R. & Guthold, M. A combined atomic force/fluorescence microscopy technique to select aptamers in a single cycle from a small pool of random oligonucleotides. Microsc. Res. Tech. 70, 372-381 (2007).
49. Mayer, G. et al. Fluorescence-activated cell sorting for aptamer SELEX with cell mixtures. Nat. Protoc. 5, 1993-2004 (2010).
50. Huang, C. J., Lin, H. I., Shiesh, S. C. & Lee, G. B. Integrated microfluidic system for rapid screening of CRP aptamers utilizing systematic evolution of ligands by exponential enrichment (SELEX). Biosens. Bioelectron. 25, 1761-1766 (2010).
51. Cho, M. et al. Quantitative selection of DNA aptamers through microfluidic selection and high-throughput sequencing. Proc. Natl Acad. Sci. USA 107, 15373-15378 (2010).
52. Sayer, N., Ibrahim, J., Turner, K., Tahiri-Alaoui, A. & James, W. Structural characterization of a 2'FRNA aptamer that binds a HIV1 SU glycoprotein, gp120. Biochem. Biophys. Res. Commun. 293, 924-931 (2002).
53. Kanagawa, T. Bias and artifacts in multitemplate polymerase chain reactions (PCR). J. Biosci. Bioeng. 96, 317-323 (2003).
54. Musheev, M. U. & Krylov, S. N. Selection of aptamers by systematic evolution of ligands by exponential enrichment: Addressing the polymerase chain reaction issue. Anal. Chim. Acta 564, 91-96 (2006).
55. Schutze, T. et al. A streamlined protocol for emulsion polymerase chain reaction and subsequent purification. Anal. Biochem. 410, 155-157 (2011).
56. Shao, K. et al. Emulsion PCR: A high efficient way of PCR amplification of random DNA libraries in aptamer selection. PLoS ONE 6, e24910 (2011).
57. Nakano, M. et al. Single-molecule PCR using waterinoil emulsion. J. Biotechnol. 102, 117-124 (2003).
58. Ouellet, E., Foley, J. H., Conway, E. M. & Haynes, C. Hifi SELEX: A high-fidelity digital-PCR based therapeutic aptamer discovery platform. Biotechnol. Bioeng. 112, 1506-1522 (2015).
59. Levay, A. et al. Identifying high-affinity aptamer ligands with defined cross-reactivity using high-throughput guided systematic evolution of ligands by exponential enrichment. Nucleic Acids Res. 43, e82 (2015).
60. Hoon, S., Zhou, B., Janda, K. D., Brenner, S. & Scolnick, J. Aptamer selection by high-throughput sequencing and informatic analysis. BioTechniques 51, 413-416 (2011).
61. Hoinka, J. et al. Large scale analysis of the mutational landscape in HTSELEX improves aptamer discovery. Nucleic Acids Res. 43, 5699-5707 (2015).
62. Thiel, W. H. et al. Nucleotide bias observed with a short SELEX RNA aptamer library. Nucleic Acid. Ther. 21, 253-263 (2011).
63. Thiel, W. H. et al. Rapid identification of cell-specific, internalizing RNA aptamers with bioinformatics analyses of a cell-based aptamer selection. PLoS ONE 7, e43836 (2012).
64. Kuwahara, M. & Sugimoto, N. Molecular evolution of functional nucleic acids with chemical modifications. Molecules 15, 5423-5444 (2010).
65. Jager, S. et al. A versatile toolbox for variable DNA functionalization at high density. J. Am. Chem. Soc. 127, 15071-15082 (2005).
66. Hirao, I. et al. An unnatural hydrophobic base pair system: site-specific incorporation of nucleotide analogs into DNA and RNA. Nat. Methods 3, 729-735 (2006).
67. Lin, Y., Qiu, Q., Gill, S. C. & Jayasena, S. D. Modified RNA sequence pools for in vitro selection. Nucleic Acids Res. 22, 5229-5234 (1994).
68. Ruckman, J. et al. 2?fluoropyrimidine RNA-based aptamers to the 165amino acid form of vascular endothelial growth factor (VEGF165). Inhibition of receptor binding and VEGF-induced vascular permeability through interactions requiring the exon 7encoded domain. J. Biol. Chem. 273, 20556-20567 (1998).
69. Burmeister, P. E. et al. Direct in vitro selection of a 2?Omethyl aptamer to VEGF. Chem. Biol. 12, 25-33 (2005).
70. Kuwahara, M. & Obika, S. In vitro selection of BNA (LNA) aptamers. Artif. DNA PNA XNA 4, 39-48 (2013).
71. Veedu, R. N. & Wengel, J. Locked nucleic acid nucleoside triphosphates and polymerases: on the way towards evolution of LNA aptamers. Mol. Biosyst. 5, 787-792 (2009).
72. Gold, L. et al. Aptamer-based multiplexed proteomic technology for biomarker discovery. PLoS ONE 5, e15004 (2010).
73. Vaught, J. D. et al. Expanding the chemistry of DNA for in vitro selection. J. Am. Chem. Soc. 132, 4141-4151 (2010).
74. Maasch, C., Buchner, K., Eulberg, D., Vonhoff, S. & Klussmann, S. Physicochemical stability of NOXE36, a 40mer lRNA (Spiegelmer) for therapeutic applications. Nucleic Acids Symp. Ser. (Oxf.) 52, 61-62 (2008).
75. Vater, A. & Klussmann, S. Toward third-generation aptamers: Spiegelmers and their therapeutic prospects. Curr. Opin. Drug Discov. Devel. 6, 253-261 (2003).
76. Lee, Y., Urban, J. H., Xu, L., Sullenger, B. A. & Lee, J. 2?fluoro modification differentially modulates the ability of RNAs to activate pattern recognition receptors. Nucleic Acid Ther. 26, 173-182 (2016).
77. Aaldering, L. J. et al. Smart functional nucleic acid chimeras: enabling tissue specific RNA targeting therapy. RNA Biol. 12, 412-425 (2015).
78. Guo, P. The emerging field of RNA nanotechnology. Nat. Nanotechnol. 5, 833-842 (2010).
79. Rusconi, C. P. et al. Antidote-mediated control of an anticoagulant aptamer in vivo. Nat. Biotechnol. 22, 1423-1428 (2004).
80. Lee, C. H. et al. Pharmacokinetics of a cholesterol-conjugated aptamer against the hepatitis C virus (HCV) NS5B protein. Mol. Ther. Nucleic Acids 4, e254 (2015).
81. Dougan, H. et al. Extending the lifetime of anticoagulant oligodeoxynucleotide aptamers in blood. Nucl. Med. Biol. 27, 289-297 (2000).
82. Heo, K. et al. An aptamer-antibody complex (oligobody) as a novel delivery platform for targeted cancer therapies. J. Control. Release 229, 1-9 (2016).
83. Willis, M. C. et al. Liposome-anchored vascular endothelial growth factor aptamers. Bioconjug. Chem. 9, 573-582 (1998).
84. Zhou, J. et al. A hybrid DNA aptamer-dendrimer nanomaterial for targeted cell labeling. Macromol. Biosci. 9, 831-835 (2009).
85. Musumeci, D. & Montesarchio, D. Polyvalent nucleic acid aptamers and modulation of their activity: A focus on the thrombin binding aptamer. Pharmacol. Ther. 136, 202-215 (2012).
86. Soule, E. E., Bompiani, K. M., Woodruff, R. S. & Sullenger, B. A. Targeting two coagulation cascade proteases with a bivalent aptamer yields a potent and antidote-controllable anticoagulant. Nucleic Acid Ther. 26, 1-9 (2016).
87. Kim, Y., Dennis, D. M., Morey, T., Yang, L. & Tan, W. Engineering dendritic aptamer assemblies as superior inhibitors of protein function. Chem. Asian J. 5, 56-59 (2010).
88. Drolet, D. W. et al. Pharmacokinetics and safety of an anti-vascular endothelial growth factor aptamer (NX1838) following injection into the vitreous humor of rhesus monkeys. Pharm. Res. 17, 1503-1510 (2000).
89. Tucker, C. E. et al. Detection and plasma pharmacokinetics of an anti-vascular endothelial growth factor oligonucleotide-aptamer (NX1838) in rhesus monkeys. J. Chromatogr. B Biomed. Sci. Appl. 732, 203-212 (1999).
90. Borbas, K. E., Ferreira, C. S., Perkins, A., Bruce, J. I. & Missailidis, S. Design and synthesis of mono-and multimeric targeted radiopharmaceuticals based on novel cyclen ligands coupled to antiMUC1 aptamers for the diagnostic imaging and targeted radiotherapy of cancer. Bioconjug. Chem. 18, 1205-1212 (2007).
91. Choi, D. Y. et al. Sustained elevated intraocular pressures after intravitreal injection of bevacizumab, ranibizumab, and pegaptanib. Retina 31, 1028-1035 (2011).
92. Steffensmeier, A. C., Azar, A. E., Fuller, J. J., Muller, B. A. & Russell, S. R. Vitreous injections of pegaptanib sodium triggering allergic reactions. Am. J. Ophthalmol. 143, 512-513 (2007).
93. Agrawal, S., Joshi, M. & Christoforidis, J. B. Vitreous inflammation associated with intravitreal anti-VEGF pharmacotherapy. Mediators Inflamm. 2013, 943409 (2013).
94. Falavarjani, K. G. & Nguyen, Q. D. Adverse events and complications associated with intravitreal injection of anti-VEGF agents: A review of literature. Eye (Lond.) 27, 787-794 (2013).
95. Boyer, D. S., Goldbaum, M., Leys, A. M., Starita, C. & V. I. S. I. O. N. Study Group. Effect of pegaptanib sodium 0. 3 mg intravitreal injections (Macugen) in intraocular pressure: posthoc analysis from V. I. S. I. O. N. study. Br. J. Ophthalmol. 98, 1543-1546 (2014).
96. Henry, S. P. et al. Activation of the alternative pathway of complement by a phosphorothioate oligonucleotide: potential mechanism of action. J. Pharmacol. Exp. Ther. 281, 810-816 (1997).
97. Farman, C. A. & Kornbrust, D. J. Oligodeoxynucleotide studies in primates: Antisense and immune stimulatory indications. Toxicol. Pathol. 31, S119-S122 (2003).
98. Swayze, E. E. et al. Antisense oligonucleotides containing locked nucleic acid improve potency but cause significant hepatotoxicity in animals. Nucleic Acids Res. 35, 687-700 (2007).
99. Ganson, N. J. et al. Pre-existing anti-polyethylene glycol antibody linked to first-exposure allergic reactions to pegnivacogin, a PEGylated RNA aptamer. J. Allergy Clin. Immunol. 137, 1610-1613. e7 (2016).
100. Waring, M. J. Lipophilicity in drug discovery. Expert Opin. Drug Discov. 5, 235-248 (2010).
101. Zhou, J. & Rossi, J. J. Cell-type-specific, aptamer-functionalized agents for targeted disease therapy. Mol. Ther. Nucleic Acids 3, e169 (2014).
102. Shum, K. T., Zhou, J. & Rossi, J. J. Aptamer-based therapeutics: new approaches to combat human viral diseases. Pharmaceuticals (Basel) 6, 1507-1542 (2013).
103. Kohn, D. B. et al. A clinical trial of retroviral-mediated transfer of a rev-responsive element decoy gene into CD34+ cells from the bone marrow of human immunodeficiency virus1infected children. Blood 94, 368-371 (1999).
104. DiGiusto, D. L. et al. RNA-based gene therapy for HIV with lentiviral vector-modified CD34+ cells in patients undergoing transplantation for AIDS-related lymphoma. Sci. Transl Med. 2, 36ra43 (2010).
105. Michienzi, A., Li, S., Zaia, J. A. & Rossi, J. J. A nucleolar TAR decoy inhibitor of HIV1 replication. Proc. Natl Acad. Sci. USA 99, 14047-14052 (2002).
106. Unwalla, H. J. & Rossi, J. J. A dual function TAR decoy serves as an anti-HIV siRNA delivery vehicle. Virol. J. 7, 33 (2010).
107. Drolet, D. W., Green, L. S., Gold, L. & Janjic, N. Fit for the eye: Aptamers in ocular disorders. Nucleic Acid Ther. 26, 127-146 (2016).
108. Eyetech Study Group. Preclinical and phase 1A clinical evaluation of an anti-VEGF pegylated aptamer (EYE001) for the treatment of exudative age-related macular degeneration. Retina 22, 143-152 (2002).
109. Bullock, A. N. & Fersht, A. R. Rescuing the function of mutant p53. Nat. Rev. Cancer 1, 68-76 (2001).
110. Axel, D. I. et al. Paclitaxel inhibits arterial smooth muscle cell proliferation and migration in vitro and in vivo using local drug delivery. Circulation 96, 636-645 (1997).
111. Thiel, W. H. et al. Smooth muscle cell-targeted RNA aptamer inhibits neointimal formation. Mol. Ther. 24, 779-787 (2016).
112. Nimjee, S. M., Rusconi, C. P., Harrington, R. A. & Sullenger, B. A. The potential of aptamers as anticoagulants. Trends Cardiovasc. Med. 15, 41-45 (2005).
113. Bompiani, K. M. et al. Probing the coagulation pathway with aptamers identifies combinations that synergistically inhibit blood clot formation. Chem. Biol. 21, 935-944 (2014).
114. Nimjee, S. M. et al. Synergistic effect of aptamers that inhibit exosites 1 and 2 on thrombin. RNA 15, 2105-2111 (2009).
115. Dyke, C. K. et al. Firstinhuman experience of an antidote-controlled anticoagulant using RNA aptamer technology: A phase 1a pharmacodynamic evaluation of a drug-antidote pair for the controlled regulation of factor IXa activity. Circulation 114, 2490-2497 (2006).
116. Chen, C. H., Chernis, G. A., Hoang, V. Q. & Landgraf, R. Inhibition of heregulin signaling by an aptamer that preferentially binds to the oligomeric form of human epidermal growth factor receptor3. Proc. Natl Acad. Sci. USA 100, 9226-9231 (2003).
117. Dollins, C. M. et al. Assembling OX40 aptamers on a molecular scaffold to create a receptor-activating aptamer. Chem. Biol. 15, 675-682 (2008).
118. Pratico, E. D., Sullenger, B. A. & Nair, S. K. Identification and characterization of an agonistic aptamer against the T cell costimulatory receptor, OX40. Nucleic Acid Ther. 23, 35-43 (2013).
119. McNamara, J. O. et al. Multivalent 41BB binding aptamers costimulate CD8+ T cells and inhibit tumor growth in mice. J. Clin. Invest. 118, 376-386 (2008).
120. Soldevilla, M. M. et al. 2Fluoro-RNA oligonucleotide CD40 targeted aptamers for the control of B lymphoma and bone-marrow aplasia. Biomaterials 67, 274-285 (2015).
121. Pastor, F. et al. CD28 aptamers as powerful immune response modulators. Mol. Ther. Nucleic Acids 2, e98 (2013).
122. Ramaswamy, V. et al. DNA aptamer assembly as a vascular endothelial growth factor receptor agonist. Nucleic Acid. Ther. 25, 227-234 (2015).
123. Yunn, N. O. et al. Agonistic aptamer to the insulin receptor leads to biased signaling and functional selectivity through allosteric modulation. Nucleic Acids Res. 43, 7688-7701 (2015).
124. Gilboa, E., McNamara, J. & Pastor, F. Use of oligonucleotide aptamer ligands to modulate the function of immune receptors. Clin. Cancer Res. 19, 1054-1062 (2013).
125. Khedri, M., Rafatpanah, H., Abnous, K., Ramezani, P. & Ramezani, M. Cancer immunotherapy via nucleic acid aptamers. Int. Immunopharmacol. 29, 926-936 (2015).
126. Compaan, D. M. & Hymowitz, S. G. The crystal structure of the costimulatory OX40OX40L complex. Structure 14, 1321-1330 (2006).
127. Tasch, J., Gong, M., Sadelain, M. & Heston, W. D. A unique folate hydrolase, prostate-specific membrane antigen (PSMA): A target for immunotherapy? Crit. Rev. Immunol. 21, 249-261 (2001).
128. McNamara, J. O. et al. Cell type-specific delivery of siRNAs with aptamer-siRNA chimeras. Nat. Biotechnol. 24, 1005-1015 (2006).
129. Dassie, J. P. et al. Systemic administration of optimized aptamer-siRNA chimeras promotes regression of PSMA-expressing tumors. Nat. Biotechnol. 27, 839-849 (2009).
130. Pastor, F., Kolonias, D., Giangrande, P. H. & Gilboa, E. Induction of tumour immunity by targeted inhibition of nonsense-mediated mRNA decay. Nature 465, 227-230 (2010).
131. Ni, X. et al. Prostate-targeted radiosensitization via aptamer-shRNA chimeras in human tumor xenografts. J. Clin. Invest. 121, 2383-2390 (2011).
132. Wullner, U. et al. Cell-specific induction of apoptosis by rationally designed bivalent aptamer-siRNA transcripts silencing eukaryotic elongation factor 2. Curr. Cancer Drug Targets 8, 554-565 (2008).
133. Chu, T. C., Twu, K. Y., Ellington, A. D. & Levy, M. Aptamer mediated siRNA delivery. Nucleic Acids Res. 34, e73 (2006).
134. Pastor, F., Kolonias, D., McNamara, J. O. II & Gilboa, E. Targeting 41BB costimulation to disseminated tumor lesions with bispecific oligonucleotide aptamers. Mol. Ther. 19, 1878-1886 (2011).
135. Zhou, J. et al. Selection, characterization and application of new RNA HIV gp 120 aptamers for facile delivery of Dicer substrate siRNAs into HIV infected cells. Nucleic Acids Res. 37, 3094-3109 (2009).
136. Zhou, J., Li, H., Li, S., Zaia, J. & Rossi, J. J. Novel dual inhibitory function aptamer-siRNA delivery system for HIV1 therapy. Mol. Ther. 16, 1481-1489 (2008).
137. Zhou, J. et al. Functional in vivo delivery of multiplexed anti-HIV1 siRNAs via a chemically synthesized aptamer with a sticky bridge. Mol. Ther. 21, 192-200 (2013).
138. Neff, C. P. et al. An aptamer-siRNA chimera suppresses HIV1 viral loads and protects from helper CD4+ T cell decline in humanized mice. Sci. Transl Med. 3, 66ra6 (2011).
139. Catuogno, S., Rienzo, A., Di Vito, A., Esposito, C. L. & de Franciscis, V. Selective delivery of therapeutic single strand antimiRs by aptamer-based conjugates. J. Control. Release 210, 147-159 (2015).
140. Shu, D. et al. Systemic delivery of anti-miRNA for suppression of triple negative breast cancer utilizing RNA nanotechnology. ACS Nano 9, 9731-9740 (2015).
141. Chu, T. C. et al. Aptamer:toxin conjugates that specifically target prostate tumor cells. Cancer Res. 66, 5989-5992 (2006).
142. Hernandez, L. I. et al. Methods for evaluating cell-specific, cell-internalizing RNA aptamers. Pharmaceuticals (Basel) 6, 295-319 (2013).
143. Boyacioglu, O., Stuart, C. H., Kulik, G. & Gmeiner, W. H. Dimeric DNA aptamer complexes for high-capacity-targeted drug delivery using pHsensitive covalent linkages. Mol. Ther. Nucleic Acids 2, e107 (2013).
144. Bagalkot, V., Farokhzad, O. C., Langer, R. & Jon, S. An aptamer-doxorubicin physical conjugate as a novel targeted drug-delivery platform. Angew. Chem. Int. Ed. 45, 8149-8152 (2006).
145. Gijs, M., Aerts, A., Impens, N., Baatout, S. & Luxen, A. Aptamers as radiopharmaceuticals for nuclear imaging and therapy. Nucl. Med. Biol. 43, 253-271 (2016).
146. Younes, C. K., Boisgard, R. & Tavitian, B. Labelled oligonucleotides as radiopharmaceuticals: pitfalls, problems and perspectives. Curr. Pharm. Des. 8, 1451-1466 (2002).
147. Sugiura, G., Kuhn, H., Sauter, M., Haberkorn, U. & Mier, W. Radiolabeling strategies for tumor-targeting proteinaceous drugs. Molecules 19, 2135-2165 (2014).
148. Hicke, B. J. et al. TenascinC aptamers are generated using tumor cells and purified protein. J. Biol. Chem. 276, 48644-48654 (2001).
149. Hicke, B. J. et al. Tumor targeting by an aptamer. J. Nucl. Med. 47, 668-678 (2006).
150. Lao, Y. H., Phua, K. K. & Leong, K. W. Aptamer nanomedicine for cancer therapeutics: barriers and potential for translation. ACS Nano 9, 2235-2254 (2015).
151. Farokhzad, O. C. et al. Nanoparticle-aptamer bioconjugates: A new approach for targeting prostate cancer cells. Cancer Res. 64, 7668-7672 (2004).
152. Liang, C. et al. Aptamer-functionalized lipid nanoparticles targeting osteoblasts as a novel RNA interference-based bone anabolic strategy. Nat. Med. 21, 288-294 (2015).
153. Barakat, M. R. & Kaiser, P. K. VEGF inhibitors for the treatment of neovascular age-related macular degeneration. Expert Opin. Investig. Drugs 18, 637-646 (2009).
154. VEGF Inhibition Study in Oracular Neovascularization (V. I. S. I. O. N.) Clinical Trial Group et al. Year 2 efficacy results of 2 randomized controlled clinical trials of pegaptanib for neovascular age-related macular degeneration. Ophthalmology 113, 1508. e1-1508. e25 (2006).
155. Jellinek, D., Lynott, C. K., Rifkin, D. B. & Janjic, N. High-affinity RNA ligands to basic fibroblast growth factor inhibit receptor binding. Proc. Natl Acad. Sci. USA 90, 11227-11231 (1993).
156. Shaw, J. P., Kent, K., Bird, J., Fishback, J. & Froehler, B. Modified deoxyoligonucleotides stable to exonuclease degradation in serum. Nucleic Acids Res. 19, 747-750 (1991).
157. Jellinek, D. et al. Potent 2?amino2?deoxypyrimidine RNA inhibitors of basic fibroblast growth factor. Biochemistry 34, 11363-11372 (1995).
158. Jaschke, A. et al. Synthesis and properties of oligodeoxyribonucleotide-polyethylene glycol conjugates. Nucleic Acids Res. 22, 4810-4817 (1994).
159. Kawaguchi, T., Asakawa, H., Tashiro, Y., Juni, K. & Sueishi, T. Stability, specific binding activity, and plasma concentration in mice of an oligodeoxynucleotide modified at 5?terminal with poly(ethylene glycol). Biol. Pharm. Bull. 18, 474-476 (1995).
160. CATT Research Group et al. Ranibizumab and bevacizumab for neovascular age-related macular degeneration. N. Engl. J. Med. 364, 1897-1908 (2011).
161. Brown, D. M. et al. Ranibizumab versus verteporfin for neovascular age-related macular degeneration. N. Engl. J. Med. 355, 1432-1444 (2006).
162. Rosenfeld, P. J. et al. Ranibizumab for neovascular age-related macular degeneration. N. Engl. J. Med. 355, 1419-1431 (2006).
163. Heier, J. S. et al. Intravitreal aflibercept (VEGF trap-eye) in wet age-related macular degeneration. Ophthalmology 119, 2537-2548 (2012).
164. D'Amore, P. A. Vascular endothelial cell growth factora: not just for endothelial cells anymore. Am. J. Pathol. 171, 14-18 (2007).
165. Ratanji, K. D., Derrick, J. P., Dearman, R. J. & Kimber, I. Immunogenicity of therapeutic proteins: influence of aggregation. J. Immunotoxicol. 11, 99-109 (2014).
166. Kawa, M. P., Machalinska, A., Roginska, D. & Machalinski, B. Complement system in pathogenesis of AMD: dual player in degeneration and protection of retinal tissue. J. Immunol. Res. 2014, 483960 (2014).
167. Baas, D. C. et al. The complement component 5 gene and age-related macular degeneration. Ophthalmology 117, 500-511 (2010).
168. Biesecker, G., Dihel, L., Enney, K. & Bendele, R. A. Derivation of RNA aptamer inhibitors of human complement C5. Immunopharmacology 42, 219-230 (1999).
169. US National Library of Medicine. ClinicalTrials. gov https://clinicaltrials. gov/ct2/show/NCT02397954? term=NCT02397954&rank=1 (2016).
170. US National Library of Medicine. Clinicaltrials. gov https://clinicaltrials. gov/ct2/show/NCT02686658? term=NCT02686658&rank=1 (2016).
171. Sadiq, M. A. et al. Platelet derived growth factor inhibitors: A potential therapeutic approach for ocular neovascularization. Saudi J. Ophthalmol. 29, 287-291 (2015).
172. Mabry, R. et al. A dual-targeting PDGFR?/VEGFA molecule assembled from stable antibody fragments demonstrates anti-angiogenic activity in vitro and in vivo. MAbs 2, 20-34 (2010).
173. Jo, N. et al. Inhibition of platelet-derived growth factor B signaling enhances the efficacy of anti-vascular endothelial growth factor therapy in multiple models of ocular neovascularization. Am. J. Pathol. 168, 2036-2053 (2006).
174. Boyer, D. S. Phase 2b study of FovistaTM, a platelet derived growth factor (PDGF) inhibitor in combination with a vascular endothelial growth factor (VEGF) inhibitor for neovascular age-related macular degeneration (AMD). Invest. Ophthalmol. Vis. Sci. 54, abstr. 2175 (2013).
175. Boyer, D. S. Combined inhibition of platelet derived (PDGF) and vascular endothelial (VEGF) growth factors for the treatment of neovascular age-related macular degeration (NVAMD)-results of a Phase I study. Invest. Ophthalmol. Vis. Sci. 50, abstr. 1260 (2009).
176. Green, L. S. et al. Inhibitory DNA ligands to platelet-derived growth factor Bchain. Biochemistry 35, 14413-14424 (1996).
177. Floege, J. et al. Novel approach to specific growth factor inhibition in vivo: Antagonism of platelet-derived growth factor in glomerulonephritis by aptamers. Am. J. Pathol. 154, 169-179 (1999).
178. US National Library of Medicine. Clinicaltrials. gov https://clinicaltrials. gov/ct2/show/NCT01089517? term=NCT01089517&rank=1 (2013).
179. US National Library of Medicine. Clinicaltrials. gov https://clinicaltrials. gov/ct2/show/NCT01944839? term=NCT01944839&rank=1 (2005).
180. US National Library of Medicine. Clinicaltrials. gov https://clinicaltrials. gov/ct2/show/NCT01940900? term=NCT01940900&rank=1 (2015).
181. US National Library of Medicine. Clinicaltrials. gov https://clinicaltrials. gov/ct2/show/NCT01940887? term=NCT01940887&rank=1 (2016).
182. Smiley, D. A. & Becker, R. C. Factor IXa as a target for anticoagulation in thrombotic disorders and conditions. Drug Discov. Today 19, 1445-1453 (2014).
183. Rusconi, C. P. et al. RNA aptamers as reversible antagonists of coagulation factor IXa. Nature 419, 90-94 (2002).
184. Povsic, T. J. et al. Use of the REG1 anticoagulation system in patients with acute coronary syndromes undergoing percutaneous coronary intervention: results from the phase II RADAR-PCI study. EuroIntervention 10, 431-438 (2014).
185. Povsic, T. J. et al. A Phase 2, randomized, partially blinded, active-controlled study assessing the efficacy and safety of variable anticoagulation reversal using the REG1 system in patients with acute coronary syndromes: results of the RADAR trial. Eur. Heart J. 34, 2481-2489 (2013).
186. Povsic, T. J. et al. Pegnivacogin results in near complete FIX inhibition in acute coronary syndrome patients: RADAR pharmacokinetic and pharmacodynamic substudy. Eur. Heart J. 32, 2412-2419 (2011).
187. Cohen, M. G. et al. First clinical application of an actively reversible direct factor IXa inhibitor as an anticoagulation strategy in patients undergoing percutaneous coronary intervention. Circulation 122, 614-622 (2010).
188. US National Library of Medicine. Clinicaltrials. gov https://clinicaltrials. gov/ct2/show/NCT00932100? term=NCT00932100&rank=1 (2012).
189. US National Library of Medicine. Clinicaltrials. gov https://clinicaltrials. gov/ct2/show/NCT00715455? term=NCT00715455&rank=1 (2013).
190. US National Library of Medicine. Clinicaltrials. gov https://clinicaltrials. gov/ct2/show/NCT01872572? term=NCT01872572&rank=1 (2013).
191. US National Library of Medicine. Clinicaltrials. gov https://clinicaltrials. gov/ct2/show/NCT01848106? term=NCT01848106&rank=1 (2014).
192. Diener, J. L. et al. Inhibition of von Willebrand factor-mediated platelet activation and thrombosis by the anti-von Willebrand factor A1domain aptamer ARC1779. J. Thromb. Haemost. 7, 1155-1162 (2009).
193. Jilma, B. et al. A randomised pilot trial of the anti-von Willebrand factor aptamer ARC1779 in patients with type 2b von Willebrand disease. Thromb. Haemost. 104, 563-570 (2010).
194. Gilbert, J. C. et al. Firstinhuman evaluation of anti von Willebrand factor therapeutic aptamer ARC1779 in healthy volunteers. Circulation 116, 2678-2686 (2007).
195. US National Library of Medicine. Clinicaltrials. gov https://clinicaltrials. gov/ct2/show/NCT00432770? term=ARC1779&rank=6 (2007).
196. US National Library of Medicine. Clinicaltrials. gov https://clinicaltrials. gov/ct2/show/NCT00632242? term=ARC1779&rank=1 (2009).
197. US National Library of Medicine. Clinicaltrials. gov https://clinicaltrials. gov/ct2/show/NCT00742612? term=ARC1779&rank=2 (2010).
198. US National Library of Medicine. Clinicaltrials. gov https://clinicaltrials. gov/ct2/show/NCT00507338? term=ARC1779&rank=5 (2009).
199. Waters, E. K., Richardson, J., Schaub, R. G. & Kurz, J. C. Effect of NU172 and bivalirudin on ecarin clotting time in human plasma and whole blood. J. Thromb. Haemost. 7, 683 (2009).
200. US National Library of Medicine. Clinicaltrials. gov https://clinicaltrials. gov/ct2/show/NCT00808964? term=00808964&rank=1 (2011).
201. Waters, E. K. et al. Aptamer ARC19499 mediates a procoagulant hemostatic effect by inhibiting tissue factor pathway inhibitor. Blood 117, 5514-5522 (2011).
202. Gorczyca, M. E. et al. Inhibition of tissue factor pathway inhibitor (TFPI) by ARC19499 improves clotting of hemophiliac blood. BMC Pharmacol. 10, abstr. A44 (2010).
203. US National Library of Medicine. Clinicaltrials. gov https://clinicaltrials. gov/ct2/show/NCT01191372? term=NCT01191372&rank=1 (2015).
204. Soundararajan, S. et al. Plasma membrane nucleolin is a receptor for the anticancer aptamer AS1411 in MV411 leukemia cells. Mol. Pharmacol. 76, 984-991 (2009).
205. Bates, P. J., Kahlon, J. B., Thomas, S. D., Trent, J. O. & Miller, D. M. Antiproliferative activity of Grich oligonucleotides correlates with protein binding. J. Biol. Chem. 274, 26369-26377 (1999).
206. Soundararajan, S., Chen, W., Spicer, E. K., Courtenay-Luck, N. & Fernandes, D. J. The nucleolin targeting aptamer AS1411 destabilizes Bcl2 messenger RNA in human breast cancer cells. Cancer Res. 68, 2358-2365 (2008).
207. Berger, C. M., Gaume, X. & Bouvet, P. The roles of nucleolin subcellular localization in cancer. Biochimie 113, 78-85 (2015).
208. Mongelard, F. & Bouvet, P. AS1411, a guanosine-rich oligonucleotide aptamer targeting nucleolin for the potential treatment of cancer, including acute myeloid leukemia. Curr. Opin. Mol. Ther. 12, 107-114 (2010).
209. Reyes-Reyes, E. M., Teng, Y. & Bates, P. J. A new paradigm for aptamer therapeutic AS1411 action: uptake by macropinocytosis and its stimulation by a nucleolin-dependent mechanism. Cancer Res. 70, 8617-8629 (2010).
210. Bates, P. J., Laber, D. A., Miller, D. M., Thomas, S. D. & Trent, J. O. Discovery and development of the Grich oligonucleotide AS1411 as a novel treatment for cancer. Exp. Mol. Pathol. 86, 151-164 (2009).
211. US National Library of Medicine. Clinicaltrials. gov https://clinicaltrials. gov/ct2/show/NCT00881244? term=NCT00881244&rank=1 (2009).
212. US National Library of Medicine. Clinicaltrials. gov https://clinicaltrials. gov/ct2/show/NCT00512083? term=NCT00512083&rank=1 (2009).
213. US National Library of Medicine. Clinicaltrials. gov https://clinicaltrials. gov/ct2/show/NCT00740441? term=NCT00740441&rank=1 (2009).
214. Hoellenriegel, J. et al. The Spiegelmer NOXA12, a novel CXCL12 inhibitor, interferes with chronic lymphocytic leukemia cell motility and causes chemosensitization. Blood 123, 1032-1039 (2014).
215. Marasca, R. & Maffei, R. NOXA12: mobilizing CLL away from home. Blood 123, 952-953 (2014).
216. Liu, S. C. et al. Blockade of SDF1 after irradiation inhibits tumor recurrences of autochthonous brain tumors in rats. Neuro Oncol. 16, 21-28 (2014).
217. Sayyed, S. G. et al. Podocytes produce homeostatic chemokine stromal cell-derived factor1/CXCL12, which contributes to glomerulosclerosis, podocyte loss and albuminuria in a mouse model of type 2 diabetes. Diabetologia 52, 2445-2454 (2009).
218. Burns, J. M. et al. A novel chemokine receptor for SDF1 and ITAC involved in cell survival, cell adhesion, and tumor development. J. Exp. Med. 203, 2201-2213 (2006).
219. US National Library of Medicine. Clinicaltrials. gov https://clinicaltrials. gov/ct2/show/NCT00976378? term=NCT00976378&rank=1 (2014).
220. US National Library of Medicine. Clinicaltrials. gov https://clinicaltrials. gov/ct2/show/NCT01194934? term=NCT01194934&rank=1 (2014).
221. US National Library of Medicine. Clinicaltrials. gov https://clinicaltrials. gov/ct2/show/NCT01486797? term=NCT01486797&rank=1 (2016).
222. US National Library of Medicine. Clinicaltrials. gov https://clinicaltrials. gov/ct2/show/NCT01521533? term=NCT01521533&rank=1 (2015).
223. Oberthur, D. et al. Crystal structure of a mirror-image lRNA aptamer (Spiegelmer) in complex with the natural lprotein target CCL2. Nat. Commun. 6, 6923 (2015).
224. Ninichuk, V. et al. Late onset of Ccl2 blockade with the Spiegelmer mNOXE363'PEG prevents glomerulosclerosis and improves glomerular filtration rate in db/ db mice. Am. J. Pathol. 172, 628-637 (2008).
225. Kulkarni, O. et al. Spiegelmer inhibition of CCL2/ MCP1 ameliorates lupus nephritis in MRL-(Fas)lpr mice. J. Am. Soc. Nephrol. 18, 2350-2358 (2007).
226. US National Library of Medicine. Clinicaltrials. gov https://clinicaltrials. gov/ct2/show/NCT01085292? term=NCT01085292&rank=1 (2013).
227. US National Library of Medicine. Clinicaltrials. gov https://clinicaltrials. gov/ct2/show/NCT01547897? term=NCT01547897&rank=1 (2014).
228. Schwoebel, F. et al. The effects of the anti-hepcidin Spiegelmer NOXH94 on inflammation-induced anemia in cynomolgus monkeys. Blood 121, 2311-2315 (2013).
229. Ganz, T. Hepcidin, a key regulator of iron metabolism and mediator of anemia of inflammation. Blood 102, 783-788 (2003).
230. US National Library of Medicine. Clinicaltrials. gov https://clinicaltrials. gov/ct2/show/NCT01372137? term=NCT01372137&rank=1 (2016).
231. US National Library of Medicine. Clinicaltrials. gov https://clinicaltrials. gov/ct2/show/NCT01691040? term=NCT01691040&rank=1 (2014).
232. US National Library of Medicine. Clinicaltrials. gov https://clinicaltrials. gov/ct2/show/NCT02079896? term=NCT02079896&rank=1 (2015).
233. US National Library of Medicine. Clinicaltrials. gov https://clinicaltrials. gov/ct2/show/NCT00021736? term=NCT00021736&rank=1 (2005).
234. US National Library of Medicine. Clinicaltrials. gov https://clinicaltrials. gov/ct2/show/NCT00040313? term=00040313&rank=1 (2006).
235. US National Library of Medicine. Clinicaltrials. gov https://clinicaltrials. gov/ct2/show/NCT00056199? term=00056199&rank=1 (2008).
236. US National Library of Medicine. Clinicaltrials. gov https://clinicaltrials. gov/ct2/show/NCT00312351? term=00312351&rank=1 (2007).
237. US National Library of Medicine. Clinicaltrials. gov https://clinicaltrials. gov/ct2/show/NCT00321997? term=NCT00321997&rank=1 (2006).
238. US National Library of Medicine. Clinicaltrials. gov https://clinicaltrials. gov/ct2/show/NCT01487070? term=NCT01487070&rank=1 (2011).
239. US National Library of Medicine. Clinicaltrials. gov https://clinicaltrials. gov/ct2/show/NCT00709527? term=NCT00709527&rank=1 (2012).
240. US National Library of Medicine. Clinicaltrials. gov https://clinicaltrials. gov/ct2/show/NCT00950638? term=NCT00950638&rank=1 (2013).
241. US National Library of Medicine. Clinicaltrials. gov https://clinicaltrials. gov/ct2/show/NCT00569140? term=NCT00569140&rank=1 (2010).
242. US National Library of Medicine. Clinicaltrials. gov https://clinicaltrials. gov/ct2/show/ NCT02387957?term=NCT02387957&rank=1 (2016).
243. US National Library of Medicine. Clinicaltrials. gov https://clinicaltrials. gov/ct2/show/NCT02591914? term=NCT02591914&rank=1 (2005).
244. US National Library of Medicine. Clinicaltrials. gov https://clinicaltrials. gov/ct2/show/NCT00113997? term=NCT00113997&rank=1 (2008).
245. US National Library of Medicine. Clinicaltrials. gov https://clinicaltrials. gov/ct2/show/NCT00694785? term=ARC1779&rank=3 (2009).
246. Jilma-Stohlawetz, P. et al. Inhibition of von Willebrand factor by ARC1779 in patients with acute thrombotic thrombocytopenic purpura. Thromb. Haemost. 105, 545-552 (2011).
247. US National Library of Medicine. Clinicaltrials. gov https://clinicaltrials. gov/ct2/show/NCT01034410? term=NCT01034410&rank=1 (2011).
248. US National Library of Medicine. Clinicaltrials. gov https://clinicaltrials. gov/ct2/show/NCT00976729? term=NCT00976729&rank=1 (2013).
249. US National Library of Medicine. Clinicaltrials. gov https://clinicaltrials. gov/ct2/show/NCT01372124? term=NCT01372124&rank=1 (2012).
250. US National Library of Medicine. Clinicaltrials. gov https://clinicaltrials. gov/ct2/show/ NCT01522794?term=NCT01522794&rank=1 (2014).