Current Development of siRNA Bioconjugates: From Research to the Clinic

Small interfering RNAs (siRNAs) acting via RNA interference mechanisms are able to recognize a homologous mRNA sequence in the cell and induce its degradation. The main problems in the development of siRNA-based drugs for therapeutic use are the low efficiency of siRNA delivery to target cells and the degradation of siRNAs by nucleases in biological fluids. Various approaches have been proposed to solve the problem of siRNA delivery in vivo (e.g., viruses, cationic lipids, polymers, nanoparticles), but all have limitations for therapeutic use. One of the most promising approaches to solve the problem of siRNA delivery to target cells is bioconjugation; i.e., the covalent connection of siRNAs with biogenic molecules (lipophilic molecules, antibodies, aptamers, ligands, peptides, or polymers). Bioconjugates are “ideal nanoparticles” since they do not need a positive charge to form complexes, are less toxic, and are less effectively recognized by components of the immune system because of their small size. This review is focused on strategies and principles for constructing siRNA bioconjugates for in vivo use

Hydrogen sensors based on In2O3 thin films with bimetallic Pt/Pd catalysts on the surface and tin and dysprosium impuri-ties in the bulk

This paper presents the results of studying the characteristics of hydrogen sensors based on thin In2O3 films modified with tin and dysprosium with dispersed double Pt/Pd catalysts deposited on the surface. To control the content of Sn and Dy in the films, an original technology was developed, and ceramic targets were fabricated from powders of the In–Dy–O, Dy–Sn–O, and In–Dy–Sn–O systems synthesized by the sol–gel method. Films of complex composition were obtained by RF magnetron sputtering of the corresponding targets. Structural features of the obtained thin films were studied by Raman spectroscopy. It is shown that various combinations of tin and dysprosium concentrations, as well as the presence of Pt/Pd catalysts on the surface, have a significant effect on the defectiveness of the films and the density of oxygen adsorption centers. As a result, the resistance of sensors in pure air (R0), the activation energies of the temperature dependences of R0, the bending of the energy bands at the grain boundaries of the semiconductor, and the responses to the action of hydrogen in the concentration range of 20–25,000 ppm change. A unique feature of Pt/Pd/ In2O3: Sn (0.5 at%), Dy (4.95 at%) films is their high sensitivity at 20–100 ppm and the absence of signal saturation in the region of high hydrogen concentrations of 5000–25,000 ppm, allowing them to be used to detect H2 in a wide range of concentrations

The restricted two-body problem in constant curvature spaces

We perform the bifurcation analysis of the Kepler problem on $S^3$ and $L^3$. An analogue of the Delaunay variables is introduced. We investigate the motion of a point mass in the field of the Newtonian center moving along a geodesic on $S^2$ and $L^2$ (the restricted two-body problem). When the curvature is small, the pericenter shift is computed using the perturbation theory. We also present the results of the numerical analysis based on the analogy with the motion of rigid body.Comment: 29 pages, 7 figure

Carrier-free cellular uptake and the gene-silencing activity of the lipophilic siRNAs is strongly affected by the length of the linker between siRNA and lipophilic group

The conjugation of siRNA to molecules, which can be internalized into the cell via natural transport mechanisms, can result in the enhancement of siRNA cellular uptake. Herein, the carrier-free cellular uptake of nuclease-resistant anti-MDR1 siRNA equipped with lipophilic residues (cholesterol, lithocholic acid, oleyl alcohol and litocholic acid oleylamide) attached to the 5′-end of the sense strand via oligomethylene linker of various length was investigated. A convenient combination of H-phosphonate and phosphoramidite methods was developed for the synthesis of 5′-lipophilic conjugates of siRNAs. It was found that lipophilic siRNA are able to effectively penetrate into HEK293, HepG2 and KB-8-5 cancer cells when used in a micromolar concentration range. The efficiency of the uptake is dependent upon the type of lipophilic moiety, the length of the linker between the moiety and the siRNA and cell type. Among all the conjugates tested, the cholesterol-conjugated siRNAs with linkers containing from 6 to 10 carbon atoms demonstrate the optimal uptake and gene silencing properties: the shortening of the linker reduces the efficiency of the cellular uptake of siRNA conjugates, whereas the lengthening of the linker facilitates the uptake but retards the gene silencing effect and decreases the efficiency of the silencing

Structural Modifications of siRNA Improve Its Performance In Vivo

The use of small interfering RNA (siRNA) in the clinic gives a wide range of possibilities for the treatment of previously incurable diseases. However, the main limitation for biomedical applications is their delivery to target cells and organs. Currently, delivery of siRNA to liver cells is a solved problem due to the bioconjugation of siRNA with N-acetylgalactosamine; other organs remain challenging for siRNA delivery to them. Despite the important role of the ligand in the composition of the bioconjugate, the structure and molecular weight of siRNA also play an important role in the delivery of siRNA. The basic principle is that siRNAs with smaller molecular weights are more efficient at entering cells, whereas siRNAs with larger molecular weights have advantages at the organism level. Here we review the relationships between siRNA structure and its biodistribution and activity to find new strategies for improving siRNA performance

Structural Modifications of siRNA Improve Its Performance In Vivo

The use of small interfering RNA (siRNA) in the clinic gives a wide range of possibilities for the treatment of previously incurable diseases. However, the main limitation for biomedical applications is their delivery to target cells and organs. Currently, delivery of siRNA to liver cells is a solved problem due to the bioconjugation of siRNA with N-acetylgalactosamine; other organs remain challenging for siRNA delivery to them. Despite the important role of the ligand in the composition of the bioconjugate, the structure and molecular weight of siRNA also play an important role in the delivery of siRNA. The basic principle is that siRNAs with smaller molecular weights are more efficient at entering cells, whereas siRNAs with larger molecular weights have advantages at the organism level. Here we review the relationships between siRNA structure and its biodistribution and activity to find new strategies for improving siRNA performance

Trimeric Small Interfering RNAs and Their Cholesterol-Containing Conjugates Exhibit Improved Accumulation in Tumors, but Dramatically Reduced Silencing Activity

Cholesterol derivatives of nuclease-resistant, anti-MDR1 small-interfering RNAs were designed to contain a 2’-OMe-modified 21-bp siRNA and a 63-bp TsiRNA in order to investigate their accumulation and silencing activity in vitro and in vivo. The results showed that increasing the length of the RNA duplex in such a conjugate increases its biological activity when delivered using a transfection agent. However, the efficiency of accumulation in human drug-resistant KB-8-5 cells during delivery in vitro in a carrier-free mode was reduced as well as efficiency of target gene silencing. TsiRNAs demonstrated a similar biodistribution in KB-8-5 xenograft tumor-bearing SCID mice with more efficient accumulation in organs and tumors than cholesterol-conjugated canonical siRNAs; however, this accumulation did not provide a silencing effect. The lack of correlation between the accumulation in the organ and the silencing activity of cholesterol conjugates of siRNAs of different lengths can be attributed to the fact that trimeric Ch-TsiRNA lags mainly in the intercellular space and does not penetrate sufficiently into the cytoplasm of the cell. Increased accumulation in the organs and in the tumor, by itself, shows that using siRNA with increased molecular weight is an effective approach to control biodistribution and delivery to the target organ

Cholesterol-Containing Nuclease-Resistant siRNA Accumulates in Tumors in a Carrier-free Mode and Silences MDR1 Gene

Chemical modifications are an effective way to improve the therapeutic properties of small interfering RNAs (siRNAs), making them more resistant to degradation in serum and ensuring their delivery to target cells and tissues. Here, we studied the carrier-free biodistribution and biological activity of a nuclease-resistant anti-MDR1 cholesterol-siRNA conjugate in healthy and tumor-bearing severe combined immune deficiency (SCID) mice. The attachment of cholesterol to siRNA provided its efficient accumulation in the liver and in tumors, and reduced its retention in the kidneys after intravenous and intraperitoneal injection. The major part of cholesterol-siRNA after intramuscular and subcutaneous injections remained in the injection place. Confocal microscopy data demonstrated that cholesterol-siRNA spread deep in the tissue and was present in the cytoplasm of almost all the liver and tumor cells. The reduction of P-glycoprotein level in human KB-8-5 xenograft overexpressing the MDR1 gene by 60% was observed at days 5–6 after injection. Then, its initial level recovered by the eighth day. The data showed that, regardless of the mode of administration (intravenous, intraperitoneal, or peritumoral), cholesterol-siMDR efficiently reduced the P-glycoprotein level in tumors. The designed anti-MDR1 conjugate has potential as an adjuvant therapeutic for the reversal of multiple drug resistance of cancer cells. Keywords: cholesterol-containing siRNA, MDR1, 2’-O-methyl modification, tumor xenograft in SCID mice, multiple drug resistance, biodistribution of siRN