AS1411 is a G-rich DNA oligonucleotide that functions as an aptamer of the protein nucleolin, found at high levels on the surface of cancer cells but not on the surface of normal cells. Herein, we have studied AS1411 as a supramolecular carrier for the delivery of an acridine-based G-quadruplex ligand, C8, to HeLa cancer cells. Two AS1411 derivatives, LNA-AS1411 and U-AS1411, were also tested, in an attempt to compare AS1411 pharmacological properties. The results showed that AS1411-C8 complexation was made with great binding strength and that it lowered the ligand's cytotoxicity towards non-malignant cells. This effect was suggested to be due to a decreased internalization of the complexed versus free C8 as shown by flow cytometry. The AS1411 derivatives, despite forming a stable complex with C8, lacked the necessary tumour-selective behaviour. The binding of C8 to AS1411 G-quadruplex structure did not negatively affect the recognition of nucleolin by the aptamer. The AS1411-C8 repressed c-MYC expression at the transcriptional level, possibly due to C8 ability to stabilize the c-MYC promoter G-quadruplexes. Overall, this study demonstrates the usefulness of AS1411 as a supramolecular carrier of the G-quadruplex binder C8 and the potential of using its tumour-selective properties for the delivery of ligands for cancer therapy.

Centro de Ciências e Tecnologias Nucleares Instituto Superior Técnico Universidade de Lisboa Estrada Nacional 10 2695 066 Bobadela LRS Portugal

CICS UBI Centro de Investigação em Ciências da Saúde Universidade da Beira Interior Av Infante D Henrique 6200 506 Covilhã Portugal

CIMAGO iCBR CIBB Faculdade de Medicina da Universidade de Coimbra Coimbra Portugal

Institute of Biophysics AS CR v v i Kralovopolska 135 612 65 Brno Czech Republic

Instituto Politécnico de Coimbra ESTESC Coimbra Health School Ciências Biomédicas Laboratoriais Coimbra Portugal

Unidade de Gestão Operacional em Citometria Centro Hospitalar e Universitário de Coimbra Coimbra Portugal

Univ Bordeaux ARNA laboratory INSERM U1212 CNRS UMR 5320 IECB F 33600 Pessac France

Zobrazit více v PubMed

Bates PJ, Laber Da, Miller DM, Thomas SD, Trent JO. Discovery and development of the G-rich oligonucleotide AS1411 as a novel treatment for cancer. Exp. Mol. Pathol. 2009;86:151–164. doi: 10.1016/j.yexmp.2009.01.004. PubMed DOI PMC

Bates PJ, et al. G-quadruplex oligonucleotide AS1411 as a cancer-targeting agent: Uses and mechanisms. Biochim. Biophys. Acta - Gen. Subj. 2017;1861:1414–1428. doi: 10.1016/j.bbagen.2016.12.015. PubMed DOI

Mongelard F, Bouvet P. Nucleolin: a multiFACeTed protein. Trends Cell Biol. 2007;17:80–86. doi: 10.1016/j.tcb.2006.11.010. PubMed DOI

Reyes-Reyes EM, Šalipur FR, Shams M, Forsthoefel MK, Bates PJ. Mechanistic studies of anticancer aptamer AS1411 reveal a novel role for nucleolin in regulating Rac1 activation. Mol. Oncol. 2015;9:1392–1405. doi: 10.1016/j.molonc.2015.03.012. PubMed DOI PMC

Shieh YA, Yang SJ, Wei MF, Shieh MJ. Aptamer-based tumor-targeted drug delivery for photodynamic therapy. ACS Nano. 2010;4:1433–1442. doi: 10.1021/nn901374b. PubMed DOI

Carvalho J, et al. Fluorescent light-up acridine orange derivatives bind and stabilize KRAS-22RT G-quadruplex. Biochimie. 2018;144:144–152. doi: 10.1016/j.biochi.2017.11.004. PubMed DOI

Pereira E, et al. Evaluation of Acridine Orange Derivatives as DNA-Targeted Radiopharmaceuticals for Auger Therapy: Influence of the Radionuclide and Distance to DNA. Sci. Rep. 2017;7:42544. doi: 10.1038/srep42544. PubMed DOI PMC

Mergny J-L, Li J, Lacroix L, Amrane S, Chaires JB. Thermal difference spectra: A specific signature for nucleic acid structures. Nucleic Acids Res. 2005;33:1–6. doi: 10.1093/nar/gki148. PubMed DOI PMC

Kumar, P., Nagarajan, A. & Uchil, P. D. Analysis of Cell Viability by the Lactate Dehydrogenase Assay. Cold Spring Harb. Protoc. 465–469, 10.1101/pdb.prot095497 (2018). PubMed

De Rache A, Mergny J-LL. Assessment of selectivity of G-quadruplex ligands via an optimised FRET melting assay. Biochimie. 2015;115:194–202. doi: 10.1016/j.biochi.2015.06.002. PubMed DOI

Loureiro A, et al. Absence of Albumin Improves in Vitro Cellular Uptake and Disruption of Poloxamer 407-Based Nanoparticles inside Cancer Cells. Mol. Pharm. 2018;15:527–535. doi: 10.1021/acs.molpharmaceut.7b00893. PubMed DOI

Gao, S., Zheng, X., Jiao, B. & Wang, L. Post-SELEX optimization of aptamers. Anal. Bioanal. Chem. 4567–4573, 10.1007/s00216-016-9556-2 (2016). PubMed

Olsen, C. M. & Marky, L. A. Energetic and hydration contributions of the removal of methyl groups from thymine to form uracil in G-quadruplexes. J. Phys. Chem. B, 10.1021/jp808526d (2009). PubMed

Dailey MM, Clarke Miller M, Bates PJ, Lane AN, Trent JO. Resolution and characterization of the structural polymorphism of a single quadruplex-forming sequence. Nucleic Acids Res. 2010;38:4877–4888. doi: 10.1093/nar/gkq166. PubMed DOI PMC

Karsisiotis AI, et al. Angew. Chemie - Int. Ed. 2011. Topological characterization of nucleic acid G-quadruplexes by UV absorption and circular dichroism; pp. 10645–10648. PubMed

Pedersen EB, Nielsen JT, Nielsen C, Filichev VV. Enhanced anti-HIV-1 activity of G-quadruplexes comprising locked nucleic acids and intercalating nucleic acids. Nucleic Acids Res. 2011;39:2470–2481. doi: 10.1093/nar/gkq1133. PubMed DOI PMC

Bing, T. et al. Triplex-quadruplex structural scaffold: A new binding structure of aptamer. Sci. Rep., 10.1038/s41598-017-15797-5 (2017). PubMed PMC

Lecarme, L. et al. Interaction of Polycationic Ni (II)-Salophen Complexes with G-Quadruplex DNA. (2014). PubMed

Hasegawa, H., Savory, N., Abe, K. & Ikebukuro, K. Methods for improving aptamer binding affinity. Molecules21 (2016). PubMed PMC

D’Onofrio J, et al. 5′-Modified G-quadruplex forming oligonucleotides endowed with anti-HIV activity: Synthesis and biophysical properties. Bioconjug. Chem. 2007;18:1194–1204. doi: 10.1021/bc070062f. PubMed DOI

Bates PJ, Kahlon JB, Thomas SD, Trent JO, Miller DM. Antiproliferative activity of G-rich oligonucleotides correlates with protein binding. J. Biol. Chem. 1999;274:26369–26377. doi: 10.1074/jbc.274.37.26369. PubMed DOI

Girvan AC, et al. AGRO100 inhibits activation of nuclear factor-Œ∫B (NF-Œ∫B) by forming a complex with NF-Œ∫B essential modulator (NEMO) and nucleolin. Mol. Cancer Ther. 2006;5:1790–1799. doi: 10.1158/1535-7163.MCT-05-0361. PubMed DOI

Arzumanov, A. et al. Inhibition of HIV-1 Tat-dependent trans activation by steric block chimeric 2′-O-methyl/LNA oligoribonucleotides. Biochemistry 40, 14645–54 (2001). PubMed

Rai Y, et al. Mitochondrial biogenesis and metabolic hyperactivation limits the application of MTT assay in the estimation of radiation induced growth inhibition. Sci. Rep. 2018;8:1–15. doi: 10.1038/s41598-017-17765-5. PubMed DOI PMC

Yoshimasu T, et al. A theoretical model for the hormetic dose-response curve for anticancer agents. Anticancer Res. 2015;35:5851–5855. PubMed

Ursini CL, et al. Study of Cytotoxic and Genotoxic Effects of Hydroxyl-Functionalized Multiwalled Carbon Nanotubes on Human Pulmonary Cells. J. Nanomater. 2012;2012:1–9. doi: 10.1155/2012/815979. DOI

Kornienko A, Rastogi SK, Lefranc F, Kiss R. Therapeutic Agents Triggering Nonapoptotic Cancer Cell Death. J. Med. Chem. 2013;56:4823–4839. doi: 10.1021/jm400136m. PubMed DOI

Fan X, Sun L, Wu Y, Zhang L, Yang Z. Bioactivity of 2′-deoxyinosine-incorporated aptamer AS1411. Sci. Rep. 2016;6:25799. doi: 10.1038/srep25799. PubMed DOI PMC

Fan X, et al. The Bioactivity of D-/L-Isonucleoside- and 2′-Deoxyinosine-Incorporated Aptamer AS1411s Including DNA Replication/MicroRNA Expression. Mol. Ther. - Nucleic Acids. 2017;9:218–229. doi: 10.1016/j.omtn.2017.09.010. PubMed DOI PMC

Dapić V, et al. Biophysical and biological properties of quadruplex oligodeoxyribonucleotides. Nucleic Acids Res. 2003;31:2097–2107. doi: 10.1093/nar/gkg316. PubMed DOI PMC

Hurley LH, Von Hoff DD, Siddiqui-Jain A, Yang D. Drug Targeting of the c-MYC Promoter to Repress Gene Expression via a G-Quadruplex Silencer Element. Semin. Oncol. 2006;33:498–512. doi: 10.1053/j.seminoncol.2006.04.012. PubMed DOI

Chauhan A, et al. Synthesis of Fluorescent Binaphthyl Amines That Bind c-MYC G-Quadruplex DNA and Repress c-MYC Expression. J. Med. Chem. 2016;59:7275–7281. doi: 10.1021/acs.jmedchem.6b00328. PubMed DOI

Sharma VR, et al. Nucleolin Overexpression Confers Increased Sensitivity to the Anti-Nucleolin Aptamer, AS1411. Cancer Invest. 2018;36:475–491. doi: 10.1080/07357907.2018.1527930. PubMed DOI PMC

Reyes-Reyes EM, Teng Y, Bates PJ. A new paradigm for aptamer therapeutic AS1411 action: Uptake by macropinocytosis and its stimulation by a nucleolin-dependent mechanism. Cancer Res. 2010;70:8617–8629. doi: 10.1158/0008-5472.CAN-10-0920. PubMed DOI PMC

G-Quadruplex Aptamer-Ligand Characterization