Hdac11

The in vitro HDACs inhibitory assay of all compounds was performed by Sundia MediTech Company, Ltd. (Shanghai, China). It was conducted by fluorescent assay. Briefly, all compounds were serially diluted to certain concentrations. Then, the HDAC1 enzymes (Active Motif, 31504), HDAC2 (BPS bioscience, 50002), HDAC5 (BPS bioscience, 50005), HDAC6 (BPS bioscience, 50006), HDAC8 (Active Motif, 31566), HDAC11 (BPS bioscience, 50011), and compounds solution were dissolved in 1×Assay buffer, which was then added into a 384-well microplate. It was mixed briefly with gentle centrifuging and the plate was incubated at r.t. Substrate peptide and Trypsin mixture were added to each well, shaken for 1 minute, and the 384-plate was placed in a BioTek Synergy plate reader for data collection.

Dulbecco's modified Eagle's medium (DMEM) with L-glutamine was purchased from GenDEPOT (Barker, TX), fetal bovine serum was purchased from HyClone, and penicillin and streptomycin were purchased from Gibco BRL (Gaithersburg, MD). Antibodies for α-tubulin, Ac-α-tubulin (Lys40), Histone H3, Ac-Histone H3 (Lys9), HDAC1, HDAC6, and glyceraldehyde 3-phosphate dehydrogenase (GAPDH) were purchased from Cell Signaling Technology (Boston, MA). Goat anti-rabbit IgG horseradish peroxidase conjugate was purchased from Santa Cruz Biotechnology (Santa Cruz, CA). Cell Titer 96 Aqueous One Solution cell proliferation assay kit was purchased from Promega (Madison, WI). Amersham ECL Select western blotting Detection Reagent was purchased from GE Healthcare. HDAC fluorogenic assay kits (HDAC1, #50061; HDAC2, #50062; HDAC3, #50073; HDAC4, #50064; HDAC6, #50076; HDAC11, #50687) were bought from BPS Bioscience (San Diego, CA).