Hydrogel based lipid-oligonucleotides: a new route to self-delivery of therapeutic sequences.

Synthetic OligoNucleotides (ON) provide promising therapeutic tools for controlling specifically genetic expression in a broad range of diseases from cancers to viral infections. Beside their chemical stability and intracellular delivery, the controlled release of therapeutic sequences remains an important challenge for successful clinical applications. In this work, Lipid-OligoNucleotide (LON) conjugates stabilizing hydrogels are reported and characterized by rheology and cryo-scanning electron microscopy (cryo-SEM). These studies revealed that lipid conjugation of antisense oligonucleotides featuring partial self-complementarity resulted in entangled pearl-necklace networks, which were obtained through micelle-micelle interaction driven by duplex formation. Owing to these properties, the Lipid AntiSense Oligonucleotide (LASO) sequences exhibited a prolonged release after subcutaneous administration compared to the non-lipidic antisense (ASO) one. The LASO self-assembly based hydrogels obtained without adjuvant represent an innovative approach for the sustained self-delivery of therapeutic oligonucleotides.

Biochemical study of the comparative inhibition of hepatitis C virus RNA polymerase by VX-222 and filibuvir.

Filibuvir and VX-222 are nonnucleoside inhibitors (NNIs) that bind to the thumb II allosteric pocket of the hepatitis C virus (HCV) RNA-dependent RNA polymerase. Both compounds have shown significant promise in clinical trials and, therefore, it is relevant to better understand their mechanisms of inhibition. In our study, filibuvir and VX-222 inhibited the 1b/Con1 HCV subgenomic replicon, with 50% effective concentrations (EC(50)s) of 70 nM and 5 nM, respectively. Using several RNA templates in biochemical assays, we found that both compounds preferentially inhibited primer-dependent RNA synthesis but had either no or only modest effects on de novo-initiated RNA synthesis. Filibuvir and VX-222 bind to the HCV polymerase with dissociation constants of 29 and 17 nM, respectively. Three potential resistance mutations in the thumb II pocket were analyzed for effects on inhibition by the two compounds. The M423T substitution in the RNA polymerase was at least 100-fold more resistant to filibuvir in the subgenomic replicon and in the enzymatic assays. This resistance was the result of a 250-fold loss in the binding affinity (K(d)) of the mutated enzyme to filibuvir. In contrast, the inhibitory activity of VX-222 was only modestly affected by the M423T substitution but more significantly affected by an I482L substitution.