| Description: |
TC-S 7001(Azaindole 1) is a highly potent inhibitor of human ROCK-1 and ROCK-2, with IC50s of 0.6 and 1.1 nM, respectively, and also inhibits murine ROCK-2 or rat ROCK-2 with IC50s of 2.4 and 0.8 nM, respectively. |
| Target: |
ROCK-1:0.6 nM (IC50)
ROCK-2:1.1 nM (IC50)
TRK:252 nM (IC50)
FLT3:303 nM (IC50) |
| In Vivo: |
Azaindole 1 (0.03, 0.1, 0.3 mg/kg, i.v.) results in a dose-dependent and long-lasting decrease in blood pressure in anaesthetized normotensive rats. Azaindole 1 (3 and 10 mg/kg, p.o.) decreases blood pressure dose-dependently and persistently both in normotensive and hypertensive rats, and shows such effects even at 1 mg/kg in hypertensive rats. Azaindole 1 (0.1 and 0.3 mg/kg, i.v. bolus injections) causes decreased mean arterial blood pressure in a dose-related manner and only leads to a moderate and dose-independent increase in heart rate of anaesthetized dogs[1]. |
| In Vitro: |
Azaindole 1 is a highly potent inhibitor of human ROCK-1 and ROCK-2, with IC50s of 0.6 and 1.1 nM, respectively, and also inhibits murine ROCK-2 or rat ROCK-2 with IC50s of 2.4 and 0.8 nM, respectively. Azaindole 1 also inhibits receptor tyrosine kinases TRK and FLT3, with IC50s of 252 and 303 nM, respectively, but shows slight inhibition of MLCK and ZIP-kinase with IC50s of 7.4 μM and 4.1 μM, respectively. Azaindole 1 induces vasorelaxation in vitro, and suppresses the phenylephrine-induced contraction of rabbit saphenous artery in a concentration dependent manner with an IC50 value of 65 nM[1]. |
| Kinase Assay: |
Human ROCK-1 and ROCK-2, murine ROCK-2, human ZIP-kinase and human MLC kinase (MLCK) (aa1425-1771) are preincubated for 10 min with test compounds in 0.05 mL of 50 mM Tris/HCl, pH 7.5, 1 mM EDTA, 5 mM MgCl2 and 0.06% CHAPS or 50 mM HEPES, pH 7.4, 10 mM Mg acetate, 1 mM dithiothreitol, 0.3 mM CaCl2, 1 μM calmodulin (MLCK). As substrate following peptides are used: RRLSSLRA (ROCK; S6 peptide), KEAKEKRQEQIAKRRRLSSLRASTSKSGGSQK (ZIP-kinase; long S6 peptide) and KKRAARATSNVFA (MLCK; MLC peptide). After 10 min preincubation with dimethyl sulphoxide (DMSO) (0.1% final concentration) or with increasing amounts of Azaindole 1 (final concentration 0.1 nM-3 μM), the assays are initiated with [33P]ATP (3000 mCi/mmol) (10 μM). After 20 min incubation, the reaction is terminated by incubation at 95°C for 10 min. After centrifugation at 10 000 g for 1 min, an aliquot of each incubation is spotted on paper matts. The papers are dried and subsequently washed twice in distilled water. After being dried, the papers are coated with scintillator and counted for radioactivity. Inhibition curves are analysed by nonlinear regression using GraphPad Prism[1]. |
| Animal Administration: |
Male Wistar rats weighing 300-350 g are anaesthetized with thiopental 100 mg/kg intraperitoneally (i.p.). A tracheotomy is performed and catheters are inserted into the femoral artery for blood pressure and heart rate measurements and into the femoral vein for test drug administration. The animals are ventilated with room air and their body temperature is controlled. Azaindole 1 is administered intravenously (i.v.) in doses of 0.03-0.1 mg/kg in a solution of Transcutol/Cremophor EL/physiological saline (19/10/80 = v/v/v). The vehicle Transcutol/Cremophor EL/physiological saline (19/10/80 = v/v/v) without test drug is used as control. The volume administered is 1 mL/kg. Six animals are treated per group[1]. |
| References: |
[1]. Kast R, et al. Cardiovascular effects of a novel potent and highly selective azaindole-based inhibitor of Rho-kinase. Br J Pharmacol. 2007 Dec;152(7):1070-80. Epub 2007 Oct 15. |
