KN-93 Phosphate

KN-93 inhibits dopamine formation in PC12h cells by modulating the reaction rate of TH to reduce the Ca(2+)-mediated phosphorylation levels of the TH molecule. ^[1] KN-93 inhibits serum-induced fibroblast cell growth with IC50 of 8 μM, and induces apoptosis after prolonged G1 arrest. ^[2] KN-93 inhibits androgen receptor activity and p53-independently induces cell death in PCa cells. ^[3]

KN-93 (5 μg) ameliorates levodopa-induced dyskinesia by lowering the expression of pGluR1S845 in a rat model of Parkinson’s disease. ^[4] In MRL/lpr Foxp3-GFP mice, KN-93 results in a significant induction of Treg cells in the spleen, peripheral lymph nodes and peripheral blood, and decreases skin and kidney damage. ^[5]

• Measurement of activities of autophosphorylated/non-autophosphorylate CaMKII

  CaMKII activity is measured utilizing syntideII as a substrate. Purified CaMKII is pre-incubated in the assay mixture ( 35 mM Hepes-Na ( pH 8.0 ), 10 mM MgC1₂, 0.5 μM CaM, 5 μM ATP, 1 mM CaCl₂ or 1 mM EGTA, total 25 μL) at 30 °C for 2 minutes. After this pre-incubation, the protein substrate/radioactive ATP mixture is added to the same test tube and the preparation is further incubated at 30 °C, for 5 minutes ( final assay condition; 35 mM Hepes-Na (pH 8.0), 10 mM MgCl₂, 0.125 μM CaCl₂, 20 μM syntideII, 11.25 μM [ γ-³²P] ATP, 10 % DMSO and indicated concentrations of KN-93, supplemented with 0.25 mM CaCl₂ and 2 mM EGTA (for autophosphorylated samples) or 0.25 mM EGTA and 2 mM CaCl2 (for nonautophosphorylated samples ), total 100 μL ). The reaction is terminated by adding of 25 μL of 100 % ( w/v ) ice-cold TCA. After centrifugation, 80 μL of the supernatant is applied to phosphocellulose paper. The filters are then washed with 75 mM H3P04 for 15 min with continuous agitation. After 4-cycles of washing, the radioactivity retains on the filter paper is quantified in a liquid scintillation counter.

• Cell lines

  NIH 3T3 fibroblasts

• Concentrations

  ~24 μM

• Incubation Time

  70 hours

• Method

  NIH 3T3 fibroblasts are cultured on polystyrene dishes in DMEM and fetal bovine serum, supplemented with penicillin/streptomycmn in a 5% CO2 humidified chamber at 37 癈. Cell growth is measured by using the MTT dye reduction method.

• Animal Models

  Sprague Dawley female rats

• Dosages

  ~5 μg

• Administration

  Intrastriatal administration