Transport Oligonucleotides—A Novel System for Intracellular Delivery of Antisense Therapeutics

Abstract

Biological activity of antisense oligonucleotides (asON), especially those with a neutral backbone, is often attenuated by poor cellular accumulation. In the present proof-of-concept study, we propose a novel delivery system for asONs which implies the delivery of modified antisense oligonucleotides by so-called transport oligonucleotides (tON), which are oligodeoxyribonucleotides complementary to asON conjugated with hydrophobic dodecyl moieties. Two types of tONs, bearing at the 5′-end up to three dodecyl residues attached through non-nucleotide inserts (TD series) or anchored directly to internucleotidic phosphate (TP series), were synthesized. tONs with three dodecyl residues efficiently delivered asON to cells without any signs of cytotoxicity and provided a transfection efficacy comparable to that achieved using Lipofectamine 2000. We found that, in the case of tON with three dodecyl residues, some tON/asON duplexes were excreted from the cells within extracellular vesicles at late stages of transfection. We confirmed the high efficacy of the novel and demonstrated that MDR1 mRNA targeted asON delivered by tON with three dodecyl residues significantly reduced the level of P-glycoprotein and increased the sensitivity of KB-8-5 human carcinoma cells to vinblastine. The obtained results demonstrate the efficacy of lipophilic oligonucleotide carriers and shows they are potentially capable of intracellular delivery of any kind of antisense oligonucleotides.

Amphiphilic “Like-A-Brush” Oligonucleotide Conjugates with Three Dodecyl Chains: Self-Assembly Features of Novel Scaffold Compounds for Nucleic Acids Delivery

“…In recent years, several approaches have been proposed to employ "like-a-brush" LOCs as useful tools in the field of NA-based formulations [10,14,18,22,33,35,[53][54][55][56]. Most of them rely on the conjugates bearing the diacyl lipid group.…”

“…The hydrophobic micellar core of LOC particles also serves as a carrier for extremely poor soluble pharmaceuticals [14,23,29] and fluorescent dyes [13,19]. Substantial characteristics of lipid-oligonucleotide conjugates include good biocompatibility and low toxicity [10,11,33]. LOCs improve pharmacokinetics and biodistribution of antisense oligomers (ASO) compared with native unconjugated oligonucleotides [4].…”

“…Earlier, we designed a non-nucleoside monomer to incorporate dodecyl groups into the oligonucleotides and synthesized dodecyl-containing conjugates with hydrophobicity comparable to that of corresponding cholesterol derivatives [36]. It was recently demonstrated that three dodecyl oligonucleotide conjugates (DOCs) are efficient and non-toxic transport molecules for ASO delivery into the A549 and HEK293 cells, which provide the transfection efficacy comparable to that of Lipofectamine 2000 [33].…”

Effect of Fluorescent Labels on DNA Affinity for Gold Nanoparticles

“…(D)—saturated dodecyl residue based on a phosphodiester of an N-substituted diethanolamine fragment, which replaces the corresponding amount of thymidine residues in T26 ( Figure S5 ). The structure of these compounds was designed to keep the number of phosphate residue charges in ON, so, the number of D-residues (k) corresponded to 26 mer—Tm [ 43 , 44 , 45 ].…”

Modified Oligonucleotides: New Structures, New Properties, and New Spheres of Application

“…This can be demonstrated using the exam-ple of 2',4'-bridged oligonucleotides reviewed by Wan et al [13], which contains information about 30-bridge analogs. Until now, such structural changes that lead to a decrease in the total charge of the modified oligonucleotide and an increase in its hydrophobicity remain in demand [19][20][21][22][23].…”