Tuning G-Quadruplex Nanostructures with Lipids. Towards Designing Hybrid Scaffolds for Oligonucleotide Delivery

Santiago Grijalvo,Ramon Eritja,Raimundo Gargallo,Marc Eres,Anna Clua

doi:10.3390/ijms22010121

Abstract

Two G-quadruplex forming oligonucleotides [d(TG4T)4 and d(TG6T)4] were selected as two tetramolecular quadruplex nanostructures because of their demonstrated ability to be modified with hydrophobic molecules. This allowed us to synthesize two series of G-quadruplex conjugates that differed in the number of G-tetrads, as well as in the terminal position of the lipid modification. Both solution and solid-phase syntheses were carried out to yield the corresponding lipid oligonucleotide conjugates modified at their 3′- and 5′-termini, respectively. Biophysical studies confirmed that the presence of saturated alkyl chains with different lengths did not affect the G-quadruplex integrity, but increased the stability. Next, the G-quadruplex domain was added to an 18-mer antisense oligonucleotide. Gene silencing studies confirmed the ability of such G-rich oligonucleotides to facilitate the inhibition of target Renilla luciferase without showing signs of toxicity in tumor cell lines.

Highlights

G-quadruplexes are nucleic acid structures constituted by two or more G-quartet motifs held together by Hoogsteen and π-π stacking interactions
Chemical modification of oligonucleotides with hydrophobic residues represents a lipids [31,32], neutral lipids [33,34] and fatty acids [35] have been attached covalently eiuseful strategy to modulate the intrinsic properties of nucleic acids [30]
G-quadruplex forming oligonucleotides varying the number of guanosines in the oligoour previous findings based on delivering ASOs using G-quadruplex nanostructures modinucleotide chain (TG4T and TG6T) and containing two saturated lipids of different length fied with cationic amino acids [27], we envisaged engineering two series of G-quadruplex (C8 and C14), which were introduced at either the 3′- or 5′-termini of the G-rich oligonuforming oligonucleotides varying the number of guanosines in the oligonucleotide chain cleotide chain

Summary

Introduction

G-quadruplexes are nucleic acid structures constituted by two or more G-quartet motifs held together by Hoogsteen and π-π stacking interactions. These structures are stabilized by monovalent cations (Na+ and K+ , mainly). G-quadruplexes were observed for the first time in the early 1900s and they were characterized by x-ray diffraction in the sixties [2]. In addition G-quadruplex structures have been visualized in human cells [6], being present at the end of the chromosomes (telomeres) as well as in some regions of the genome like 50 -UTR regions and oncogene promoters (e.g., c-kit, bcl-2 and c-myc), emerging as attractive targets for cancer therapy [5]

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Results

Conclusion