Eun-Kyoung Bang

Modified Oligonucleotides Containing Lithocholic Acid in Their Backbones: Their Enhanced Cellular Uptake and Their Mimicking of Hairpin Structures

Their enhanced cell permeability and their ability to mimic DNA structures make modified oligodeoxyribonucleotides (ODNs) very important substances for increasing our understanding of cell biology and for therapeutic applications. Lithocholic acid is a hydrophobic secondary bile acid that is a substrate of nuclear Pregnane X receptor (PXR). We designed and synthesized novel lithocholic acid‐based ODNs (L‐ODNs) by using a new phosphoramidite derived from lithocholic acid. By comparing data obtained from circular‐dichroism, melting‐point, and theoretical studies, we believe that these L‐ODNs adopt DNA hairpin structures. Furthermore, L‐ODNs have enhanced cellular uptake properties with respect to regular ODNs. To demonstrate their enhanced cell permeabilities, we carried out cellular uptake experiments of L‐ODNs in HeLa cells. By attaching fluorescein as a fluorescence label and using confocal microscopy, we observed that the permeability of L‐ODNs is much higher than that of natural ODNs.

Synthesis and efficient siRNA delivery of polyamine -conjugated cationic nucleoside lipids

To advance their use as nonviral gene delivery agents, we have synthesized cationic nucleoside lipids featuring naturally occurring polyamines conjugated at the 5′ position and oleyl groups at the 2′ and 3′ positions of uridine, through chemically stable, but biodegradable, carbamate linkers. The corresponding lipoplexes are efficient and nontoxic vectors for the delivery of siRNA into cells in vitro.

A Self-Complementary Nucleoside: Synthesis, Solid-State Structure, and Fluorescence Behavior

A novel self-complementary nucleoside ((A)T), featuring two complementary nucleobases linked through an ethynyl group has been synthesized. The rigid aromatic nucleobases provided (A)T with a pale-blue fluorescence. Unlike most fluorescent organic molecules, nucleoside (A)T gives enhanced fluorescence in solid state. It exhibits considerably enhanced fluorescence intensity and a remarkable redshift (ca. 70 nm) in its emission maximum upon an increase in concentration or decrease in temperature as a result of the formation of aggregates stabilized through hydrogen bonding and π-π stacking of well-organized (A) T assemblies; these interactions are also evident in the solid-state structure, determined by X-ray crystallography. DFT calculations supported the preference of such aggregation processes.

Coordinative Amphiphiles as Tunable siRNA Transporters

In this study, we developed coordinative amphiphiles for use as novel siRNA transporters. As a modification of a conventional cationic lipid structure, we replaced the cationic head with zinc(II)-dipicolylamine complex (Zn/DPA) as a phosphate-directing group, and used various membrane-directing groups in the place of the hydrophobic tails. These simple amphiphiles are readily synthesized and easy to modify. The Zn/DPA head groups bind to the phosphate backbones of siRNAs, and to our surprise, they prevented the enzymatic degradation of siRNAs by RNase A. Interestingly, the Zn/DPA head itself exhibited moderate transfection efficiency, and its combination with a membrane-directing group-oleoyl (CA1), pyrenebutyryl (CA2), or biotin (CA3)-enhanced the delivery efficiency without imparting significant cytotoxicity. Notably, the uptake pathway was tunable depending on the nature of the membrane-directing group. CA1 delivered siRNAs mainly through caveolae-mediated endocytosis, and CA2 through clathrin- and caveolin-independent endocytosis; CA3 recruited siRNAs specifically into biotin receptor-positive HepG2 cells through receptor-mediated endocytosis. Thus, it appears possible to develop tunable siRNA transporters simply by changing the membrane-directing parts. These are the first examples of amphiphilic siRNA transporters accompanying coordinative interactions between the amphiphiles and siRNAs.