Computational Analysis of siRNA Recognition by the Ago2 PAZ Domain and Identification of the Determinants of RNA-Induced Gene Silencing.

Computational Analysis of siRNA Recognition by the Ago2 PAZ Domain and Identification of the Determinants of RNA-Induced Gene Silencing.

PLoS One. 2013; 8(2): e57140
Kandeel M, Kitade Y

RNA interference (RNAi) is a highly specialized process of protein-siRNA interaction that results in the regulation of gene expression and cleavage of target mRNA. The PAZ domain of the Argonaute proteins binds to the 3′ end of siRNA, and during RNAi the attaching end of the siRNA switches between binding and release from its binding pocket. This biphasic interaction of the 3′ end of siRNA with the PAZ domain is essential for RNAi activity; however, it remains unclear whether stronger or weaker binding with PAZ domain will facilitate or hinder the overall RNAi process. Here we report the correlation between the binding of modified siRNA 3′ overhang analogues and their in vivo RNAi efficacy. We found that higher RNAi efficacy was associated with the parameters of lower Ki value, lower total intermolecular energy, lower free energy, higher hydrogen bonding, smaller total surface of interaction and fewer van der Waals interactions. Electrostatic interaction was a minor contributor to compounds recognition, underscoring the presence of phosphate groups in the modified analogues. Thus, compounds with lower binding affinity are associated with better gene silencing. Lower binding strength along with the smaller interaction surface, higher hydrogen bonding and fewer van der Waals interactions were among the markers for favorable RNAi activity. Within the measured parameters, the interaction surface, van der Waals interactions and inhibition constant showed a statistically significant correlation with measured RNAi efficacy. The considerations provided in this report will be helpful in the design of new compounds with better gene silencing ability. HubMed – drug


Pharmacological characterization of memoquin, a multi-target compound for the treatment of Alzheimer’s disease.

PLoS One. 2013; 8(2): e56870
Capurro V, Busquet P, Lopes JP, Bertorelli R, Tarozzo G, Bolognesi ML, Piomelli D, Reggiani A, Cavalli A

Alzheimer’s disease (AD) is characterized by progressive loss of cognitive function, dementia and altered behavior. Over 30 million people worldwide suffer from AD and available therapies are still palliative rather than curative. Recently, Memoquin (MQ), a quinone-bearing polyamine compound, has emerged as a promising anti-AD lead candidate, mainly thanks to its multi-target profile. MQ acts as an acetylcholinesterase and ?-secretase-1 inhibitor, and also possesses anti-amyloid and anti-oxidant properties. Despite this potential interest, in vivo behavioral studies with MQ have been limited. Here, we report on in vivo studies with MQ (acute and sub-chronic treatments; 7-15 mg/kg per os) carried out using two different mouse models: i) scopolamine- and ii) beta-amyloid peptide- (A?-) induced amnesia. Several aspects related to memory were examined using the T-maze, the Morris water maze, the novel object recognition, and the passive avoidance tasks. At the dose of 15 mg/kg, MQ was able to rescue all tested aspects of cognitive impairment including spatial, episodic, aversive, short and long-term memory in both scopolamine- and A?-induced amnesia models. Furthermore, when tested in primary cortical neurons, MQ was able to fully prevent the A?-induced neurotoxicity mediated by oxidative stress. The results support the effectiveness of MQ as a cognitive enhancer, and highlight the value of a multi-target strategy to address the complex nature of cognitive dysfunction in AD. HubMed – drug


Autophagy Inhibition Promotes 5-Fluorouraci-Induced Apoptosis by Stimulating ROS Formation in Human Non-Small Cell Lung Cancer A549 Cells.

PLoS One. 2013; 8(2): e56679
Pan X, Zhang X, Sun H, Zhang J, Yan M, Zhang H

Chemotherapy is an important option for the treatment of various cancers including lung cancer. However, tumor resistance towards cytotoxic chemotherapy has become more common. It has been reported that autophagy is one of the processes contributing to this resistance. In the present study, we found that the anti-cancer drug 5-fluorouraci(5-FU) could induce autophagy in A549 cells. 5-FU treatment could lead to the conversion of LC3 I/II, the up-regulation of Beclin-1, the down-regulation of p62 and the formation of acidic vesicular organelles (AVOs) in A549 cells. Pre-treatment of cancer cells with 3-MA or siAtg7 could enhance 5-FU-induced apoptosis through the activation of caspases, and the caspase inhibitor z-VAD-fmk rescued the cell viability reduction. Furthermore, the inhibition of autophagy also stimulated ROS formation and scavenging of ROS by antioxidant NAC inhibited caspase-3 activity, prevented the release of cyt-c from mitochondria and eventually rescued cancer cells from 5-FU-mediated apoptosis. These results suggest that 5-FU-elicited autophagic response plays a protective role against cell apoptosis and the inhibition of autophagy could sensitize them to 5-FU-induced caspase-dependent apoptosis through the stimulation of ROS formation. HubMed – drug


Enhanced Effectivity of an ALK5-Inhibitor after Cell-Specific Delivery to Hepatic Stellate Cells in Mice with Liver Injury.

PLoS One. 2013; 8(2): e56442
van Beuge MM, Prakash J, Lacombe M, Post E, Reker-Smit C, Beljaars L, Poelstra K

Transforming growth factor-? (TGF-?) is a major pro-fibrotic cytokine, causing the overproduction of extracellular matrix molecules in many fibrotic diseases. Inhibition of its type-I receptor (ALK5) has been shown to effectively inhibit fibrosis in animal models. However, apart from its pro-fibrotic effects, TGF-? also has a regulatory role in the immune system and influences tumorigenesis, which limits the use of inhibitors. We therefore explored the cell-specific delivery of an ALK5-inhibitor to hepatic stellate cells, a key cell in the development of liver fibrosis. We synthesized a conjugate of the ALK5-inhibitor LY-364947 coupled to mannose-6-phosphate human serum albumin (M6PHSA), which binds to the insulin-like growth factor II receptor on activated HSC. The effectivity of the conjugate was evaluated in primary HSC and in an acute liver injury model in mice. In vitro, the free drug and the conjugate significantly inhibited fibrotic markers in HSC. In hepatocytes, TGF-?-dependent signaling was inhibited by free drug, but not by the conjugate, thus showing its cell-specificity. In vivo, the conjugate localized in desmin-positive cells in the liver and not in hepatocytes or immune cells. In the acute liver injury model in mice, the conjugate reduced fibrogenic markers and collagen deposition more effectively than free drug. We conclude that we can specifically deliver an ALK5-inhibitor to HSC using the M6PHSA carrier and that this targeted drug reduces fibrogenic parameters in vivo, without affecting other cell-types. HubMed – drug