The condensation of nucleic acids into compact nanoparticles with cationic carriers is a powerful tool for translocating exogenous nucleic acids into cells. To date, most efforts have been focused on the development of novel gene carriers for safe and efficient gene delivery. However, small interfering RNA (siRNA) is generally not strongly associated with cationic carriers due to its stiff structure and low spatial charge density. To overcome this limitation, this work introduces a well‐defined dimeric conjugate of small internally segment interfering RNA (sisiRNA) linked via a disulfide bond for enhanced cellular uptake and gene silencing. Dimeric sisiRNA is synthesized through oxidizing two monomeric sisiRNA molecules, each of which consists of a sense strand carrying a nick and an antisense strand modified with a thiol group at the 3′‐end. The nick in the sense strand enables the dimeric sisiRNA to be more effectively condensed into nanosized complexes due to the increased structural flexibility, which results in a higher gene silencing efficiency compared with the dimeric siRNA containing the intact sense strands. The results indicate that the discontinuity of the sense strands is a simple method of adding more flexibility to various siRNA‐based nanostructures for enhanced gene silencing.
Cell surface integrins, which play important roles in the survival, proliferation, migration, and invasion of cancer cells, are a viable target for treatment of metastatic breast cancer. This line of therapy still remains challenging due to the lack of proper identification and validation of effective targets as well as the lack of suitable therapeutic agents for treatment. The focus is on one such molecular target for this purpose, namely integrin‐β1, and effective lowering of integrin‐β1 levels on a breast cancer model (MDA‐MB‐231 cells) is achieved by delivering a dicer‐substrate short interfering RNA (siRNA) targeting integrin‐β1 with lipid‐modified low molecular weight polyethylenimine polymers. Reduction of integrin‐β1 levels leads to reduced adhesion of MDA‐MB‐231 cells to extracellular matrix component fibronectin as well as to human bone marrow cells. A reduced migration of the breast cancer cells is also observed after integrin‐β1 silencing in “scratch” and “transwell” migration assays. These results highlight the importance of integrin‐β1 for the migration of metastatic breast cancer cells by effectively silencing this target with a practical dose of siRNA.
Plant Dicer-like proteins (DCLs) belong to the Ribonuclease III (RNase III) enzyme family. They are involved in the regulation of gene expression and antiviral defense through RNA interference pathways. A model plant, Arabidopsis thaliana encodes four DCL proteins (AtDCL1-4) that produce different classes of small regulatory RNAs. Our studies focus on AtDCL4 that processes double-stranded RNAs (dsRNAs) into 21 nucleotide trans-acting small interfering RNAs. So far, little is known about the structures of plant DCLs and the complexes they form with dsRNA. In this work, we present models of the catalytic core of AtDCL4 and AtDCL4-dsRNA complex constructed by computational methods.We built a homology model of the catalytic core of AtDCL4 comprising Platform, PAZ, Connector helix and two RNase III domains. To assemble the AtDCL4-dsRNA complex two modeling approaches were used. In the first method, to establish conformations that allow building a consistent model of the complex, we used Normal Mode Analysis for both dsRNA and AtDCL4. The second strategy involved template-based approach for positioning of the PAZ domain and manual arrangement of the Connector helix. Our results suggest that the spatial orientation of the Connector helix, Platform and PAZ relative to the RNase III domains is crucial for measuring dsRNA of defined length. The modeled complexes provide information about interactions that may contribute to the relative orientations of these domains and to dsRNA binding. All these information can be helpful for understanding the mechanism of AtDCL4-mediated dsRNA recognition and binding, to produce small RNA of specific size. 相似文献
Multivalent aptamer–siRNA conjugates containing multiple mucin‐1 aptamers and BCL2‐specific siRNA are synthesized, and doxorubicin, an anthracycline anticancer drug, is loaded into these conjugates through intercalation with nucleic acids. These doxorubicin‐incorporated multivalent aptamer–siRNA conjugates are transfected to mucin‐1 overexpressing MCF‐7 breast cancer cells and their multidrug‐resistant cell lines. Doxorubicin‐incorporated multivalent aptamer–siRNA conjugates exert promising anticancer effects, such as activation of caspase‐3/7 and decrease of cell viability, on multidrug‐resistant cancer cells because of their high intracellular uptake efficiency. Thus, this delivery system is an efficient tool for combination oncotherapy with chemotherapeutics and nucleic acid drugs to overcome multidrug resistance.
Polyethylenimine-poly(L-lysine)(PEI-PLL) copolymer was synthesized via ring-opening polymerization of L-lysine N-carboxyanhydride(Lys(Z)-NCA) initiated by PEI. The complexation of PEI-PLL with si RNA was studied by particle size and zeta potential measurements. The flow cytometric analysis and confocal imaging showed its excellent intracellular trafficking ability. PEI-PLL displayed higher gene silencing efficiency and lower cytotoxicity than commercial PEI-25 k in vitro. In the antitumor study, PEI-PLL was further combined with si VEGF and showed obviously tumor inhibition effect for the treatment of CT26 tumor model. Therefore, PEI-PLL is a promising si RNA carrier candidate for further antitumor treatment in vivo. 相似文献
Programmable molecular self‐assembly of siRNA molecules provides precisely controlled generation of dendrimeric siRNA nanostructures. The second‐generation dendrimers of siRNA can be effectively complexed with a low‐molecular‐weight, cationic polymer (poly(β‐amino ester), PBAE) to generate stable nanostructures about 160 nm in diameter via strong electrostatic interactions. Condensation and gene silencing efficiencies increase with the increased generation of siRNA dendrimers due to a high charge density and structural flexibility. 相似文献
Short interfering RNA (siRNA) functions directly in the cytoplasm, where it is assembled into an RNA-induced silencing complex (RISC). The localized delivery of siRNA to a specific site in vivo is highly challenging. There are many disease states in which a systemic effect of RNAi may be desirable; some examples include non-localized cancers, HIV, neurodegenerative diseases, respiratory viruses, and heart and vascular disease. In this Concept, we will focus on the localized delivery of siRNA to a target site using various delivery modalities. In certain tissues, such as the eye, central nervous system and lung, it has been demonstrated that a simple injection of naked siRNA will silence gene expression specifically in that tissue. To achieve local gene silencing in other tissues, a variety of approaches have been pursued to help stabilize the siRNA and facilitate uptake; they include chemical modification of the siRNA or complexation within liposomes or polymers to form nanoparticles. Recently, the use of macroscopic biomaterial scaffolds for siRNA delivery has been reported, and although there is still significant work to be done in this area to optimize the delivery systems, it is an important area of research that offers the potential for having great impact on the field of siRNA delivery. 相似文献