piperidines and fasudil

Accumulative indoxyl sulfate (IS) retained in chronic kidney disease (CKD) can potentiate vascular endothelial dysfunction, and herein, we aim at elucidating the underlying mechanisms from the perspective of possible association between reactive oxygen species (ROS) and RhoA/ROCK pathway. IS-treated nephrectomized rats are administered with antioxidants including NADPH oxidase inhibitor apocynin, SOD analog tempol, and mitochondrion-targeted SOD mimetic mito-TEMPO to scavenge ROS, or ROCK inhibitor fasudil to obstruct RhoA/ROCK pathway. First, we find in response to IS stimulation, antioxidants treatments suppress increased aortic ROCK activity and expression levels. Additionally, ROCK blockade prevent IS-induced increased NADPH oxidase expression (mainly p22phox and p47phox), mitochondrial and intracellular ROS (superoxide and hydrogen peroxide) generation, and decreased Cu/Zn-SOD expression in thoracic aortas. Apocynin, mito-TEMPO, and tempol also reverse these markers of oxidative stress. These results suggest that IS induces excessive ROS production and ROCK activation involving a circuitous relationship in which ROS activate ROCK and ROCK promotes ROS overproduction. Finally, ROS and ROCK depletion attenuate IS-induced decrease in nitric oxide (NO) production and eNOS expression levels, and alleviate impaired vasomotor responses including increased vasocontraction to phenylephrine and decreased vasorelaxation to acetylcholine, thereby preventing cardiovascular complications accompanied by CKD. Taken together, excessive ROS derived from NADPH oxidase and mitochondria coordinate with RhoA/ROCK activation in a form of positive reciprocal relationship to induce endothelial dysfunction through disturbing endothelium-dependent NO signaling upon IS stimulation in CKD status.

Topics: 1-(5-Isoquinolinesulfonyl)-2-Methylpiperazine; Acetophenones; Animals; Antioxidants; Cyclic N-Oxides; Endothelium, Vascular; Gene Expression Regulation; Humans; Indican; NADPH Oxidases; Nitric Oxide; Nitric Oxide Synthase Type III; Organophosphorus Compounds; Oxidative Stress; Piperidines; Rats; Reactive Oxygen Species; Renal Insufficiency, Chronic; rho GTP-Binding Proteins; rho-Associated Kinases; Signal Transduction; Spin Labels

The Rho-kinase (ROCK) inhibitor Fasudil has proven beneficial in experimental autoimmune encephalomyelitis (EAE). Given the small safety window of Fasudil, we are looking for novel ROCK inhibitors, which have similar or stronger effect on EAE with greater safety. In this study, we report that WAR-5, a Y-27632 derivative, alleviates the clinical symptoms, attenuates myelin damage and reduces CNS inflammatory responses in EAE C57BL/6 mice at an extent similar to Fasudil, while exhibits less vasodilator and adverse reaction in vivo. WAR-5 inhibits ROCK activity, and selectively suppresses the expression of ROCK II in spleen, brain and spinal cord of EAE mice, especially in spinal cord, accompanied by decreased expression of Nogo. WAR-5 also regulates the imbalance of Th1/Th17 T cells and regulatory T cells, inhibits inflammatory microenvironment induced with NF-κB-IL-1β pathway. Importantly, WAR-5 converts M1 toward M2 microglia/macrophages that are positively correlated with BDNF and NT-3 production. Taken together, WAR-5 exhibits therapeutic potential in EAE by more selectively inhibits ROCK II, with a greater safety than Fasudil, and is worthy of further clinical study to clarify its clinical value.

Topics: 1-(5-Isoquinolinesulfonyl)-2-Methylpiperazine; Amides; Aminopyridines; Animals; Cytokines; Encephalomyelitis, Autoimmune, Experimental; Enzyme-Linked Immunosorbent Assay; Female; Immunoblotting; Immunoenzyme Techniques; Inflammation; Macrophages; Mice; Mice, Inbred C57BL; Microglia; Nerve Growth Factors; Piperidines; Protein Kinase Inhibitors; Pyridines; rho-Associated Kinases; T-Lymphocytes, Regulatory

Rho kinase is an important target implicated in a variety of cardiovascular diseases. Herein, we report the optimisation of the fragment derived ATP-competitive ROCK inhibitors 1 and 2 into lead compound 14A. The initial goal of improving ROCK-I potency relative to 1, whilst maintaining a good PK profile, was achieved through removal of the aminoisoquinoline basic centre. Lead 14A was equipotent against both ROCK-I and ROCK-II, showed good in vivo efficacy in the spontaneous hypertensive rat model, and was further optimised to demonstrate the scope for improving selectivity over PKA versus hydroxy Fasudil 3.

Topics: 1-(5-Isoquinolinesulfonyl)-2-Methylpiperazine; Amines; Animals; Disease Models, Animal; Hypertension; Isoquinolines; Models, Chemical; Models, Molecular; Piperidines; Protein Kinase Inhibitors; Quinolones; Rats; rho-Associated Kinases; Structure-Activity Relationship

A bottleneck in drug discovery is the identification of the molecular targets of a compound (mode of action, MoA) and of its off-target effects. Previous approaches to elucidate drug MoA include analysis of chemical structures, transcriptional responses following treatment, and text mining. Methods based on transcriptional responses require the least amount of information and can be quickly applied to new compounds. Available methods are inefficient and are not able to support network pharmacology. We developed an automatic and robust approach that exploits similarity in gene expression profiles following drug treatment, across multiple cell lines and dosages, to predict similarities in drug effect and MoA. We constructed a "drug network" of 1,302 nodes (drugs) and 41,047 edges (indicating similarities between pair of drugs). We applied network theory, partitioning drugs into groups of densely interconnected nodes (i.e., communities). These communities are significantly enriched for compounds with similar MoA, or acting on the same pathway, and can be used to identify the compound-targeted biological pathways. New compounds can be integrated into the network to predict their therapeutic and off-target effects. Using this network, we correctly predicted the MoA for nine anticancer compounds, and we were able to discover an unreported effect for a well-known drug. We verified an unexpected similarity between cyclin-dependent kinase 2 inhibitors and Topoisomerase inhibitors. We discovered that Fasudil (a Rho-kinase inhibitor) might be "repositioned" as an enhancer of cellular autophagy, potentially applicable to several neurodegenerative disorders. Our approach was implemented in a tool (Mode of Action by NeTwoRk Analysis, MANTRA, http://mantra.tigem.it).

Topics: 1-(5-Isoquinolinesulfonyl)-2-Methylpiperazine; Algorithms; Antineoplastic Agents; Autophagy; Blotting, Western; Camptothecin; Cell Line, Tumor; Doxorubicin; Drug Discovery; Drug Screening Assays, Antitumor; Flavonoids; Fuzzy Logic; Gene Expression Profiling; Gene Expression Regulation, Neoplastic; HeLa Cells; Humans; Irinotecan; Neoplasms; Oligonucleotide Array Sequence Analysis; Phosphorylation; Piperidines; Pyrazoles; Pyrroles; RNA Polymerase II