Inactive analog of KN-93; negative control

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].

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.

[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.

KN-93 is a Ca²⁺/calmodulin-dependent protein kinase II (CaMKII) inhibitor that inhibits, in a concentration-dependent and reversible manner, intracellular calcium ([Ca²⁺]ᵢ) signaling mediated by the inositol 1,4,5-triphosphate receptor (IP₃R) in mouse oocytes and permeabilized A7r5 smooth muscle cells. Both cell types primarily express type 1 IP₃R (IP₃R-1). KN-92 is an inactive analogue with no inhibitory effect. The inhibitory effect of KN-93 on Ca²⁺ signaling is independent of its effect on IP₃ metabolism and the abundance of Ca²⁺ stores, suggesting that it acts on IP₃R. The inhibitory effect was independent of CaMKII, as other CaMKII inhibitors (KN-62, peptides 281-309, and autologous calmodulin-associated inhibitory peptides) were ineffective under the same conditions, and CaMKII activation was also blocked in permeabilized cells. Furthermore, KN-93 was most effective in the absence of Ca²⁺. Analysis of Ca²⁺ release in A7r5 cells, IP₃R-1 degradation in oocytes, and [³H]IP₃ binding in Sf9 microsomes at different [IP₃] concentrations all indicated that KN-93 did not affect IP₃ binding. The inhibitory effects of KN-93 and calmodulin (CaM), alone or in combination, on Ca²⁺ release and [³H]IP₃ binding were consistent with the specific interaction between KN-93 and the CaM binding site on IP₃R-1. This is consistent with the weak effect of KN-93 in permeabilized 16HBE14o(-) cells, which mainly express type 3 IP(3)R lacking a high-affinity CaM binding site. These findings suggest that KN-93 directly inhibits IP(3)R-1 and may therefore be a useful tool for studying the regulation of IP(3)R function. [1]

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Objective: To investigate the effects of the CaMKII selective inhibitor KN-93 on the proliferation of human hepatic stellate cells and the expression of p53 or p21 proteins. Methods: Human hepatic stellate cells (LX-2) were incubated with different concentrations (0-50 μmol/L) of KN-93 or its inactive derivative KN-92. Cell proliferation was detected by CCK-8 assay, and the expression of two cell cycle regulators, p53 and p21, was detected by SDS-PAGE and Western blotting. Results: KN-93 (5-50 μmol/L) reduced the proliferation of human hepatic stellate cells in a dose-dependent manner. After 24 hours of treatment, the proliferation rate decreased 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). Incubation with 10 μmol/L KN-93 resulted in a time-dependent decrease in cell growth, from 78.27% at 8 hours to 11.48% at 48 hours. However, the inactive derivative of KN-93, KN-92, did not effectively inhibit cell proliferation. Furthermore, cell cycle regulator expression analysis showed that KN-93, but not KN-92, reduced the expression of p53 and p21. Conclusion: KN-93 significantly inhibited the proliferation of LX-2 cells by regulating the expression of two specific cell cycle regulators, p53 and p21. [2]

C24H26CL2N2O3S

493.4458

492.104

C, 58.42; H, 5.31; Cl, 14.37; N, 5.68; O, 9.73; S, 6.50

1431698-47-3

KN-92 phosphate;1135280-28-2;KN-92;176708-42-2

71576672

White to off-white solid powder

2

5

9

32

650

0

CN(C/C=C/C1=CC=C(C=C1)Cl)CC2=CC=CC=C2NS(=O)(=O)C3=CC=C(C=C3)OC.Cl

SHDNWBMPAAXAHM-IPZCTEOASA-N

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+;

2934.99.9001

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.

Room temperature (This product is stable at ambient temperature for a few days during ordinary shipping and time spent in Customs)

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.

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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.

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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.

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 (Please use freshly prepared in vivo formulations for optimal results.)