Functional delivery of DNAzyme with iron oxide nanoparticles for hepatitis C virus gene knockdown.

DNAzyme is an attractive therapeutic oligonucleotide which enables cleavage of mRNA in a sequence-specific manner and thus, silencing target gene. A particularly important challenge in achieving the successful down-regulation of gene expression is to efficiently deliver DNAzymes to disease sites and cells. Here, we report the nanoparticle-assisted functional delivery of therapeutic DNAzyme for the treatment of hepatitis C by inducing knockdown of hepatitis C virus (HCV) gene, NS3.

3-O-arylmethylgalangin, a novel isostere for anti-HCV 1,3-diketoacids (DKAs).

Through chelation of the metal ions at the enzyme active site, 1,3-diketoacids (DKAs) show potent inhibition of viral enzymes such as HIV integrase and HCV NS5B. In order to optimize the antiviral activity of the DKAs, structural modification of their metal-binding units, keto-enol acids or monoketo acids, have been actively performed. In this study, we proposed 3-O-arylmethylgalangin 3 as an alternative to ortho-substituted aromatic DKA, a potent inhibitor of HCV NS5B.

Suppression of hepatitis C virus genome replication in cells with RNA-cleaving DNA enzymes and short-hairpin RNA.

A class of antisense oligodeoxyribozymes, known as the 10-23 DNA enzymes (DNAzyme), has been shown to efficiently cleave target RNA at purine-pyrimidine junctions in vitro. Herein we have utilized a strategy to identify accessible cleavage sites for DNAzyme in the target RNA, the hepatitis C virus nonstructural gene 3 (HCV NS3) RNA that encodes viral helicase and protease, from a pool of randomized DNAzyme library. The screening procedure identified 18 potential cleavage sites in the target RNA.

7-O-Arylmethylgalangin as a novel scaffold for anti-HCV agents.

In spite of potent antiviral activity, suboptimal physicochemical properties of aryl diketo acids (ADKs) necessitates modification of the core 1,3-diketo acid functionality into a novel scaffold. As the metal-binding affinity of the diketo acid is the key to the antiviral activity of ADKs, we anticipated 3,5-dihydroxy-4-oxo arrangement of galangin scaffold would serve as an excellent mimic for the diketo acid functionality. In this study, through synthesis and biological evaluation of various galangin derivatives, we have shown that the diketo acid functionality can be successfully replaced with the galangin scaffold by specific combination of the substituents to result in identification of a novel galangin derivative (3s) with anti-HCV activity (EC(50)=0.

Facilitation of polymerase chain reaction with thermostable inorganic pyrophosphatase from hyperthermophilic archaeon Pyrococcus horikoshii.

An inorganic pyrophosphatase (PPases) was cloned from the hyperthermophilic archaeon Pyrococcus horikoshii and was expressed in and purified from Escherichia coli. The recombinant inorganic pyrophosphatase (PhPPase) exhibited robust catalytic activity of the hydrolysis of pyrophosphate into two orthophosphates at high temperatures (70 degrees C to 95 degrees C). Thermostable pyrophosphatase activity was applied into polymerase chain reaction (PCR) due to its ability to push chemical equilibrium toward the synthesis of DNA by removing pyrophosphate from the reaction.