KN-92 Hydrochloride, the hydrochloride salt of KN-92, is an inactive derivative of KN-93. KN-92 is intended to be used as a negative control compound in studies designed to elucidate the antagonist activities of KN-93. KN-93 inhibits histamine-induced aminopyrine uptake in parietal cells (IC50 = 300 nM). KN-93 has been used to implicate roles for CaMKII in Ca2+-induced Ca2+ release in cardiac myocytes, constitutive phosphorylation of 5-lipoxygenase in 3T3 cells, and Ca2+-dependent activation of HIF-1α in colon cancer cell.

Physicochemical Properties

Molecular Formula	C24H26CL2N2O3S
Molecular Weight	493.4458
Exact Mass	492.104
Elemental Analysis	C, 58.42; H, 5.31; Cl, 14.37; N, 5.68; O, 9.73; S, 6.50
CAS #	1431698-47-3
Related CAS #	KN-92 phosphate;1135280-28-2;KN-92;176708-42-2
PubChem CID	71576672
Appearance	White to off-white solid powder
Hydrogen Bond Donor Count	2
Hydrogen Bond Acceptor Count	5
Rotatable Bond Count	9
Heavy Atom Count	32
Complexity	650
Defined Atom Stereocenter Count	0
SMILES	CN(C/C=C/C1=CC=C(C=C1)Cl)CC2=CC=CC=C2NS(=O)(=O)C3=CC=C(C=C3)OC.Cl
InChi Key	SHDNWBMPAAXAHM-IPZCTEOASA-N
InChi Code	InChI=1S/C24H25ClN2O3S.ClH/c1-27(17-5-6-19-9-11-21(25)12-10-19)18-20-7-3-4-8-24(20)26-31(28,29)23-15-13-22(30-2)14-16-23;/h3-16,26H,17-18H2,1-2H3;1H/b6-5+;
Chemical Name	(E)-N-(2-(((3-(4-chlorophenyl)allyl)(methyl)amino)methyl)phenyl)-4-methoxybenzenesulfonamide phosphate
Synonyms	KN-92 HCl; KN-92 hydrochloride; 1431698-47-3; KN-92 (hydrochloride); N-[2-[[[(E)-3-(4-chlorophenyl)prop-2-enyl]-methylamino]methyl]phenyl]-4-methoxybenzenesulfonamide;hydrochloride; N-(2-(((3-(4-Chlorophenyl)allyl)(methyl)amino)methyl)phenyl)-4-methoxybenzenesulfonamide hydrochloride; N-[2-({[3-(4-chlorophenyl)prop-2-en-1-yl](methyl)amino}methyl)phenyl]-4-methoxybenzene-1-sulfonamide hydrochloride; KN-92 HCL; KN-92hydrochloride; KN-92; KN 92; KN92
HS Tariff Code	2934.99.9001
Storage	Powder-20°C 3 years 4°C 2 years In solvent -80°C 6 months -20°C 1 month Note: Please store this product in a sealed and protected environment, avoid exposure to moisture.
Shipping Condition	Room temperature (This product is stable at ambient temperature for a few days during ordinary shipping and time spent in Customs)

Biological Activity

Targets	Inactive analog of KN-93; negative control
ln Vitro	LX-2 cell growth is inhibited by KN-93 (5-50μM; 24 hours), while KN-92 (5-50μM; 24 hours) is ineffective in blocking cell growth[2]. KN-93, not KN-92, decreased the expression of p53 and p21, according to an analysis of cell cycle regulator expression[2].
ln Vivo
Cell Assay	Cell Viability Assay[2] Cell Types: Human hepatic stellate cells (LX-2) Tested Concentrations: 5-50 μM Incubation Duration: 24 hrs (hours) Experimental Results: Ineffective in blocking cell growth.
Animal Protocol
References	[1]. Inhibition of the inositol trisphosphate receptor of mouse eggs and A7r5 cells by KN-93 via a mechanism unrelated to Ca2+/calmodulin-dependent protein kinase II antagonism. J Biol Chem. 2002;277(38):35061-35070. [2]. KN-93, a specific inhibitor of CaMKII inhibits human hepatic stellate cell proliferation in vitro. World J Gastroenterol. 2007;13(9):1445-1448.
Additional Infomation	KN-93, a Ca(2+)/calmodulin-dependent protein kinase II (CaMKII) inhibitor, concentration-dependently and reversibly inhibited inositol 1,4,5-trisphosphate receptor (IP(3)R)-mediated [Ca(2+)](i) signaling in mouse eggs and permeabilized A7r5 smooth muscle cells, two cell types predominantly expressing type-1 IP(3)R (IP(3)R-1). KN-92, an inactive analog, was ineffective. The inhibitory action of KN-93 on Ca(2+) signaling depended neither on effects on IP(3) metabolism nor on the filling grade of Ca(2+) stores, suggesting a direct action on the IP(3)R. Inhibition was independent of CaMKII, since in identical conditions other CaMKII inhibitors (KN-62, peptide 281-309, and autocamtide-related inhibitory peptide) were ineffective and since CaMKII activation was precluded in permeabilized cells. Moreover, KN-93 was most effective in the absence of Ca(2+). Analysis of Ca(2+) release in A7r5 cells at varying [IP(3)], of IP(3)R-1 degradation in eggs, and of [(3)H]IP(3) binding in Sf9 microsomes all indicated that KN-93 did not affect IP(3) binding. Comparison of the inhibition of Ca(2+) release and of [(3)H]IP(3) binding by KN-93 and calmodulin (CaM), either separately or combined, was compatible with a specific interaction of KN-93 with a CaM-binding site on IP(3)R-1. This was also consistent with the much smaller effect of KN-93 in permeabilized 16HBE14o(-) cells that predominantly express type 3 IP(3)R, which lacks the high affinity CaM-binding site. These findings indicate that KN-93 inhibits IP(3)R-1 directly and may therefore be a useful tool in the study of IP(3)R functional regulation. [1] Aim: To investigate the effects of KN-93, a CaMKII selective inhibitor on cell proliferation and the expression of p53 or p21 protein in human hepatic stellate cells. Methods: Human hepatic stellate cells (LX-2) were incubated with various concentrations (0-50 micromol/L) of KN-93 or its inactive derivative, KN-92. Cell proliferation was measured by CCK-8 assay, and the expression of two cell cycle regulators, p53 and p21, was determined by SDS-PAGE and Western blotting. Results: KN-93 (5-50 micromol/L) decreased the proliferation of human hepatic stellate cells in a dose-dependent manner from 81.76% (81.76% +/- 2.58% vs 96.63% +/- 2.69%, P < 0.05) to 27.15% (27.15% +/- 2.86% vs 96.59% +/- 2.44%, P < 0.01) after 24 h treatment. Incubation of 10 micromol/L KN-93 induced the cell growth reduction in a time-dependent manner from 78.27% at 8 h to 11.48% at 48 h. However, KN-92, an inactive derivative of KN-93, did not inhibit cell proliferation effectively. Moreover, analysis of cell cycle regulator expression revealed that KN-93 rather than KN-92 reduced the expression of p53 and p21. Conclusion: KN-93 has potent inhibitory effect on proliferation of LX-2 cells by modulating the expression of two special cell cycle regulators, p53 and p21. [2]

Solubility Data

Solubility (In Vitro)

DMSO: ~25mg/mL Water: N/A Ethanol:N/A

Solubility (In Vivo)

Solubility in Formulation 1: ≥ 2.75 mg/mL (5.57 mM) (saturation unknown) in 10% DMSO + 40% PEG300 + 5% Tween80 + 45% Saline (add these co-solvents sequentially from left to right, and one by one), clear solution. For example, if 1 mL of working solution is to be prepared, you can add 100 μL of 27.5 mg/mL clear DMSO stock solution to 400 μL PEG300 and mix evenly; then add 50 μL Tween-80 to the above solution and mix evenly; then add 450 μL normal saline to adjust the volume to 1 mL. Preparation of saline: Dissolve 0.9 g of sodium chloride in 100 mL ddH₂ O to obtain a clear solution. Solubility in Formulation 2: ≥ 2.75 mg/mL (5.57 mM) (saturation unknown) in 10% DMSO + 90% (20% SBE-β-CD in Saline) (add these co-solvents sequentially from left to right, and one by one), clear solution. For example, if 1 mL of working solution is to be prepared, you can add 100 μL of 27.5 mg/mL clear DMSO stock solution to 900 μL of 20% SBE-β-CD physiological saline solution and mix evenly. Preparation of 20% SBE-β-CD in Saline (4°C，1 week): Dissolve 2 g SBE-β-CD in 10 mL saline to obtain a clear solution. Solubility in Formulation 3: ≥ 2.75 mg/mL (5.57 mM) (saturation unknown) in 10% DMSO + 90% Corn Oil (add these co-solvents sequentially from left to right, and one by one), clear solution. For example, if 1 mL of working solution is to be prepared, you can add 100 μL of 27.5 mg/mL clear DMSO stock solution to 900 μL of corn oil and mix evenly.  (Please use freshly prepared in vivo formulations for optimal results.)

Preparing Stock Solutions		1 mg	5 mg	10 mg
	1 mM	2.0265 mL	10.1327 mL	20.2655 mL
	5 mM	0.4053 mL	2.0265 mL	4.0531 mL
	10 mM	0.2027 mL	1.0133 mL	2.0265 mL

*Note: Please select an appropriate solvent for the preparation of stock solution based on your experiment needs. For most products, DMSO can be used for preparing stock solutions (e.g. 5 mM, 10 mM, or 20 mM concentration); some products with high aqueous solubility may be dissolved in water directly. Solubility information is available at the above Solubility Data section. Once the stock solution is prepared, aliquot it to routine usage volumes and store at -20°C or -80°C. Avoid repeated freeze and thaw cycles.