Home>>Signaling Pathways>> TGF-β / Smad Signaling>> ROCK>>K–115 hydrochloride dihydrate

K–115 hydrochloride dihydrate

Catalog No.GC12817

K-115 hydrochloride dihydrate (K-115) is a specific inhibitor of ROCK, with IC50s of 19 and 51 nM for ROCK2 and ROCK1, respectively.

Products are for research use only. Not for human use. We do not sell to patients.

K–115 hydrochloride dihydrate Chemical Structure

Cas No.: 887375-67-9

Size Price Stock Qty
K每115 hydrochloride dihydrate 10mM (in 1mL Water)
$129.00
In stock
2mg
$98.00
In stock
5mg
$148.00
In stock
10mg
$241.00
In stock
25mg
$519.00
In stock
50mg
$918.00
In stock
100mg
$1,483.00
In stock

Tel:(909) 407-4943 Email: sales@glpbio.com

Customer Reviews

Based on customer reviews.

  • GlpBio Citations

    GlpBio Citations
  • Bioactive Compounds Premium Provider

    Bioactive Compounds Premium Provider

Sample solution is provided at 25 µL, 10mM.

Product Documents

Quality Control & SDS

View current batch:

Protocol

Kinase experiment:

ROCK 1 (0.75 ng/mL) and ROCK 2 (0.5 ng/mL) are incubated with various concentrations of Ripasudil, Y-27632, or HA-1077 at 25°C for 90 min in 50 mM Tris-HCl buffer (pH 7.5) containing 100 mM KCl, 10 mM MgCl2, 0.1 mM EGTA, 30 mM Long S6 Kinase Substrate peptide, and 1 mM ATP in a total volume of 40 mL. PKACa, PKC, and CaMKIIa are also incubated with various concentrations of Ripasudil, Y-27632, or HA-1077. PKACa (0.0625 ng/mL) is incubated at 25°C for 30 min in 40 mM Tris-HCl buffer (pH 7.5) containing 20 mM MgCl2, 1 mg/ mL BSA, 5 mM Kemptide peptide substrate, and 1 mM ATP in a total volume of 40 mL. PKC (0.025 ng/mL) is incubated at 25°C for 80 min in 20 mM Tris-HCl buffer (pH 7.5) containing 20 mM MgCl2, 0.4 mM CaCl2, 0.1 mg/mL BSA, 0.25 mM EGTA, 25 ng/mL phosphatidylserine, 2.5 ng/mL diacylglycerol, 0.0075% Triton-X-100, 25 mM DTT, 10 mM Neurogranin (28-43) peptide substrate, and 1 mM ATP in a total volume of 40 mL. CaMKIIa (0.025 ng/mL) is incubated at 25°C for 90 min in 50 mM Tris-HCl buffer (pH 7.5) containing 10 mM MgCl2, 2 mM CaCl2, 0.04 mg/mL BSA, 16 mg/mL purified calmodulin from bovine testis, 500 mM DTT, 50 mM Autocamitide 2, and 1 mM ATP in a total volume of 40 mL. After incubation, 40 mL of KinaseGlo Luminescent Kinase Assay solution is added, and allowed to remain at 25°C for 10 min, and Relative Light Units (RLU) are measured using a luminometer. The RLU without test compound is set as 100% (Control value), and that without enzyme and compound is set as 0% (Normal value). The reaction rate (% of control) is then calculated from the RLU with addition of each concentration of test compounds, and the 50% inhibitory concentrations (IC50) are determined by logistic regression analysis using SAS[1].

Cell experiment:

Trabecular meshwork (TM) cells are plated on 6 well plates at a density of 1 × 104 cells per well in DMEM containing 10% FBS. Following overnight culture, when cells have reached semiconfluence, 1 or 10 μM of Ripasudil, 10 μM of Y-27632, or 10 μM of fasudil are added to culture wells. PBS is used as a control vehicle. After 60 min, drug solutions are removed and replaced with DMEM containing 10% FBS. Cells are observed by phase-contrast microscopy and photographed 60 min after drug application and 2 h after drug removal. For immunohistochemistry, TM cells are plated on gelatin-coated 8 well chamber slides at a density of 1 × 104 cells per well in DMEM containing 10% FBS. After overnight culture, when cells reach semiconfluence, cell are incubated in Ripasudil at 1 or 10 μM, Y-27632 at 10 μM, or fasudil at 10 μM for 60 min. PBS is used as a control vehicle. Drug solutions are removed and replaced with DMEM containing 10% FBS after 2 h. Cells are fixed with 4% paraformaldehyde in PBS for 15 min then washed with cytoskeletal buffer (10 mM MES, 150 mM NaCl, 5 mM EGTA, 5 mM MgCl2, 5 mM glucose, pH 6.1) and serum buffer (10% FBS in PBS). Cells are permeabilized with 0.5% Triton X-100 in PBS for 12 min at room temperature and blocked with serum buffer for at least 2 h at 4°C. Filamentous actin (F-actin) is labeled with 0.05 mg/mL Phalloidin-TRITC for 1 h at room temperature. After washing with PBS, cells are mounted with commercial mounting medium containing DAPI and observed using a fluorescence microscope. The exposure to take images for F-actin and DAPI are 0.1 and 0.05 sec, respectively[2].

Animal experiment:

Rabbits[1]In the rabbit experiments, 50 mL of vehicle or Ripasudil at concentrations of 0.0625%, 0.125%, 0.25, or 0.5% is instilled into one eye. Intraocular pressure (IOP) is measured in both eyes before and 0.5, 1, 2, 3, 4, and 5 h after instillation. The contralateral eye is not treated. Animals are administered all concentrations of Ripasudil assigned using the Latin square method with intervals of at least 2 d.Monkeys[1]In the monkey experiments, 20 mL of Ripasudil at concentrations of 0.1%, 0.2%, or 0.4%, and latanoprost at a concentration of 0.005% are instilled into one eye. IOP is measured in both eyes before and 1, 2, 4, 6, and 8 h after instillation. The contralateral eye is not treated. Animals are arranged to receive all formulations with intervals of at least 1 week using the Latin square method. The IOPs are compared with the results for the instillation side at pre-dose and at each time point after instillation of Ripasudil, and are compared with both eyes at each time point.

References:

[1]. Isobe T, et al. Effects of K-115, a rho-kinase inhibitor, on aqueous humor dynamics in rabbits. Curr Eye Res. 2014 Aug;39(8):813-22.
[2]. Kaneko Y, et al. Effects of K-115 (Ripasudil), a novel ROCK inhibitor, on trabecular meshwork and Schlemm's canal endothelial cells. Sci Rep. 2016 Jan 19;6:19640.
[3]. Yamamoto K, et al. The novel Rho kinase (ROCK) inhibitor K-115: a new candidate drug for neuroprotective treatment in glaucoma. Invest Ophthalmol Vis Sci. 2014 Oct 2;55(11):7126-36.

Background

Ripasudil (K-115) is a novel and potent Rho kinase inhibitor with intraocular pressure-lowering effect. Rho-kinase can be activated by the small GTP-binding protein Rho. Growing evidence has demonstrated that Rho/Rho-kinase pathway has been involved in a variety of cellular functions, such as actin cytoskeleton organization, vascular smooth muscle cell (VSMC) contraction, cell adhesion and motility, cytokinesis, and gene expressions. Dysfunctions of these may result in the pathogenesis of cardiovascular disease such as coronary artery spasm [1].

In vivo: In optic nerve crush (NC) C57BL/6 mice model, oral administration of K-115 (1 mg/kg/d) increased 34 ± 3% survival of RGCs after NC [2].

Clinical Trials: In the phase 1 clinical trials, 0.05%, 0.1%, 0.2%, 0.4%, and 0.8% concentrations of K-115 increased the intraocular pressure (IOP) of healthy male adult volunteers from -1.6 mm Hg for placebo to -3.4, -2.2, -2.6, -4.0, and -4.3 mm Hg when tested in 2 hours after instillation [3]. In the phase 2 randomized clinical study, in patients with primary open-angle glaucoma or ocular hypertension, K-115 treatment (twice daily for 8 weeks) dose-dependently lowered the IOP level [4].

References: 

[1].Shimokawa H, Takeshita A. Rho-kinase is an important therapeutic target in cardiovascular medicine[J]. Arteriosclerosis, thrombosis, and vascular biology, 2005, 25(9): 1767-1775.
[2].Yamamoto K, Maruyama K, Himori N, et al. The Novel Rho Kinase (ROCK) Inhibitor K-115: A New Candidate Drug for Neuroprotective Treatment in GlaucomaNovel Rho Kinase Inhibitor[J]. Investigative ophthalmology & visual science, 2014, 55(11): 7126-7136.
[3].Tanihara H, Inoue T, Yamamoto T, et al. Phase 1 clinical trials of a selective Rho kinase inhibitor, K-115[J]. JAMA ophthalmology, 2013, 131(10): 1288-1295.
[4].Tanihara H, Inoue T, Yamamoto T, et al. Phase 2 randomized clinical study of a Rho kinase inhibitor, K-115, in primary open-angle glaucoma and ocular hypertension[J]. American journal of ophthalmology, 2013, 156(4): 731-736. e2.

Chemical Properties

Cas No. 887375-67-9 SDF
Chemical Name 4-fluoro-5-[[(2S)-2-methyl-1,4-diazepan-1-yl]sulfonyl]isoquinoline;dihydrate;hydrochloride
Canonical SMILES CC1CNCCCN1S(=O)(=O)C2=CC=CC3=CN=CC(=C32)F.O.O.Cl
Formula C15H23ClFN3O4S M.Wt 395.88
Solubility ≥ 123.2mg/mL in DMSO Storage Store at -20°C
General tips Please select the appropriate solvent to prepare the stock solution according to the solubility of the product in different solvents; once the solution is prepared, please store it in separate packages to avoid product failure caused by repeated freezing and thawing.Storage method and period of the stock solution: When stored at -80°C, please use it within 6 months; when stored at -20°C, please use it within 1 month.
To increase solubility, heat the tube to 37°C and then oscillate in an ultrasonic bath for some time.
Shipping Condition Evaluation sample solution: shipped with blue ice. All other sizes available: with RT, or with Blue Ice upon request.

Complete Stock Solution Preparation Table

Prepare stock solution
1 mg 5 mg 10 mg
1 mM 2.526 mL 12.6301 mL 25.2602 mL
5 mM 0.5052 mL 2.526 mL 5.052 mL
10 mM 0.2526 mL 1.263 mL 2.526 mL
  • Molarity Calculator

  • Dilution Calculator

Mass
=
Concentration
x
Volume
x
MW*
 
 
 
**When preparing stock solutions always use the batch-specific molecular weight of the product found on the vial label and MSDS / CoA (available online).

Calculate

In vivo Formulation Calculator (Clear solution)

Step 1: Enter information below (Recommended: An additional animal making an allowance for loss during the experiment)

mg/kg g μL

Step 2: Enter the in vivo formulation (This is only the calculator, not formulation. Please contact us first if there is no in vivo formulation at the solubility Section.)

% DMSO % % Tween 80 % ddH2O
%DMSO %

Calculation results:

Working concentration: mg/ml;

Method for preparing DMSO master liquid: mg drug pre-dissolved in μL DMSO ( Master liquid concentration mg/mL, Please contact us first if the concentration exceeds the DMSO solubility of the batch of drug. )

Method for preparing in vivo formulation: Take μL DMSO master liquid, next addμL PEG300, mix and clarify, next addμL Tween 80, mix and clarify, next add μL ddH2O, mix and clarify.

Method for preparing in vivo formulation: Take μL DMSO master liquid, next add μL Corn oil, mix and clarify.

Note: 1. Please make sure the liquid is clear before adding the next solvent.
2. Be sure to add the solvent(s) in order. You must ensure that the solution obtained, in the previous addition, is a clear solution before proceeding to add the next solvent. Physical methods such as vortex, ultrasound or hot water bath can be used to aid dissolving.
3. All of the above co-solvents are available for purchase on the GlpBio website.

Reviews

Review for K–115 hydrochloride dihydrate

Average Rating: 5 ★★★★★ (Based on Reviews and 33 reference(s) in Google Scholar.)

5 Star
100%
4 Star
0%
3 Star
0%
2 Star
0%
1 Star
0%
Review for K–115 hydrochloride dihydrate

GLPBIO products are for RESEARCH USE ONLY. Please make sure your review or question is research based.

Required fields are marked with *

You may receive emails regarding this submission. Any emails will include the ability to opt-out of future communications.