IC50: 0.4 mM
Valproic acid (VPA) is a histone deacetylase inhibitor.
Histone deacetylases (HDACs), which have been recognized for their roles in controlling chromatin remodeling through histone acetylation/deacetylation, are currently known to modify a large number of non-histone proteins to regulate diverse cell processes.
In vitro: VPA showed to have cellular neuroprotective properties. In cultured neurons, VPA protected from thapsigargin-induced endoplasmic reticulum stress, glutamate-induced excitotoxicity, as well as lipopolysaccharide (LPS)-induced dopaminergic neuronal death. In midbrain neuron-glia cultures, VPA was also shown to inhibit LPS-induced, microglia-mediated inflammation [1].
In vivo: Post-pMCAO injections with VPA could decrease the brain infarct volume. Postinsult treatment with VPA also reduced the number of microglia, suppressed microglial activation, and inhibited other inflammatory markers in the ischemic brain. The reduction in acetylated histone H3 was prevented by treatment with VPA. Moreover, VPA superinduced heat-shock protein 70 and blocked pMCAO-induced down-regulation of cyclooxygenase-2. The sensory, motor, and reflex performance of pMCAO rats was improved by VPA treatment [1].
Clinical trial: In CRPC patients, VPA levels and VPA exposure duration were independent predictors for declining prostate-specific antigen (PSA). Oral VPA was not well tolerated in this patient population due to grade 1 and 2 neurologic symptoms and fatigue. Total and free VPA levels were useful for preventing severe toxicities from oral VPA. It was unlikely that PSA responses from oral VPA were related to HDAC inhibition [2].
参考文献:
[1] Kim HJ,Rowe M,Ren M,Hong JS,Chen PS,Chuang DM.? Histone deacetylase inhibitors exhibit anti-inflammatory and neuroprotective effects in a rat permanent ischemic model of stroke: multiple mechanisms of action. J Pharmacol Exp Ther.2007 Jun;321(3):892-901. Epub 2007 Mar 19.
[2] Sharma S,Symanowski J,Wong B,Dino P,Manno P,Vogelzang N.? A Phase II Clinical Trial of Oral Valproic Acid in Patients with Castration-Resistant Prostate Cancers Using an Intensive Biomarker Sampling Strategy. Transl Oncol.2008 Sep;1(3):141-7.
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Kinase experiment: |
The activity of caspase-3, -8 and -9 is assessed using the caspase-3, -8 and -9 colorimetric assay kits, respectively. In brief, 1×106?cells in a 60-mm culture dish are incubated with 10 mM Valproic acid for 24 h. The cells are then washed in PBS and suspended in 5 volumes of lysis buffer provided with the kit. Protein concentrations are determined using the Bradford method. Supernatants containing 50 μg total protein are used to determine caspase-3, -8 and -9 activities. The supernatants are added to each well in 96-well microtiter plates with DEVD-pNA, IETD-pNA or LEHD-pNA as caspase-3, -8 and -9 substrates and the plates are incubated at 37°C for 1 h. The optical density of each well is measured at 405 nm using a microplate reader. The activity of caspase-3, -8 and -9 is expressed in arbitrary absorbance units. |
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Cell experiment: |
In brief, 5×105?cells are seeded in 96-well microtiter plates for MTT assays. After exposure to the designated doses of Valproic acid for the indicated times, MTT solution [20 mL: 2 mg/mL in phosphate-buffered saline (PBS)] is added to each well of the 96-well plates. The plates are additionally incubated for 3 h at 37°C. Medium is withdrawn from the plates by pipetting and 200 mL DMSO is added to each well to solubilize the formazan crystals. The optical density is measured at 570 nm using a microplate reader. |
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Animal experiment: |
Splenectomies are performed on the BALB/c nude mice. One week after the splenectomies, the mice receiv whole body irradiation with?137Cs at a dose of 4 Gy. At 48-72 h post-irradiation, the mice are subcutaneously implanted with Kasumi-1 cells (2×107?cells/mouse with 0.15-0.2 mL) in the right axillary region. The mice are randomLy assigned to two groups, the Valproic acid (n=6) and control (n=6) groups. When the tumors are appr 200 mm3?in size at appr 10 days post-implantation, 0.2 mL Valproic acid (500 mg/kg body weight) or 0.2 mL saline is injected intraperitoneally every day. Valproic acid is dissolved in saline at a concentration of 25 mg/mL. The longest diameter (a) and the shortest diameter (b) of the tumor are measured every three days, and the tumor volume (TV) is calculated according to the following formula: TV=1/2×a×b2. Following two weeks of injections, the mice are sacrificed by cervical dislocation and the tumor masses are removed for the following experiments. |
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参考文献: [1]. Han BR, et al. Valproic acid inhibits the growth of HeLa cervical cancer cells via caspase-dependent apoptosis. Oncol Rep. 2013 Dec;30(6):2999-3005. |
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