Synthesis and in vitro inhibition properties of siRNA conjugates carrying glucose and galactose with different presentations

Abstract

Abstract. Oligoribonucleotides conjugates and the corresponding siRNA duplexes against tumor necrosis factor carrying one, two or four glucose and galactose residues at the 5'-end have been prepared using phosphoramidite chemistry. Carbohydratemodified siRNA duplexes displayed similar or slightly reduced inhibitory properties compared to unmodified RNA duplexes in HeLa cells upon transfection with oligofectamine. When HeLa cells were treated with siRNA carrying one, two or four glucose residues without oligofectamine, no inhibition was observed. The inhibitory properties of siRNA carrying galactose residues without transfecting agent were tested on HuH-7 cells that have abundant asialoglycoprotein receptors. In these cells siRNA carrying galactose residues have slight anti-TNF inhibitory properties (25% in the best case) that are eliminated if the receptors are blocked with a competitor such as a-Dgalactopyranosyl-albumine. These results demonstrate receptor-mediated uptake of siRNA carrying galactose residues although the efficacy of the process is not enough for efficient RNA interference experiments.

Glucose-nucleobase pairs within DNA: impact of hydrophobicity, alternative linking unit and DNA polymerase nucleotide insertion studies

“… 15 We have used standard phosphoramidite chemistry for the preparation of the corresponding carbohydrate oligonucleotide conjugates (COCs). This type of conjugation is quite convenient and straight-forward and it has been reported previously for the synthesis of COCs with different applications such as improving cellular uptake of antisense oligonucleotides 24 , 25 and siRNAs, 26 – 28 preparation of potential anti-HIV drugs, 29 investigation of lectin binding to carbohydrates and glycoarrays 30 and preparation of molecular interaction probes. 11 , 13 , 31 , 32 …”

Covalent Strategies for Targeting Messenger and Non-Coding RNAs: An Updated Review on siRNA, miRNA and antimiR Conjugates

“…Other kind of carbohydrate architectures based on one, two and four Gal residues have been proposed by Aviñó et al with the aim of modifying the 5′-termini of the siRNA passenger strand. The authors carried out the synthesis of a small series of carbohydrate-siRNA conjugates which were designed for inhibiting tumour necrosis factor-α (TNF-α) production and also for enhancing the cellular internalization of siRNAs [ 67 ]. TNF-α plays an important role in many inflammatory diseases and has been proposed to be a therapeutic target for Crohn’s disease, diabetes and obesity among other diseases.…”

The Role of Bridging Water and Hydrogen Bonding as Key Determinants of Noncovalent Protein–Carbohydrate Recognition

“…For example, an enhancement in liver uptake of oligonucleotides was achieved after conjugation. In particular, siRNA sequences were successfully conjugated to glucose and galactose, and subsequently to GalNAc (N‐acetylgalactosamine) . Here, the carbohydrate interacts with the human asialoglycoprotein receptor (ASGPR, also “hepatic lectin”), an integral membrane protein responsible for clearance of desialylated, galactose‐terminal glycoproteins from the circulation by receptor‐mediated endocytosis.…”