This compound caused inhibition of HDAC 8 activity with no effect on the activity of HDAC 1/2. The protein levels of HDAC 2, 3 and 8 were found to be drastically reduced with no change in HDAC 4 6 upon chrysin treat ment. Chrysin caused histone modifications such selleck chem as acetylation and methylation at p21 promoter particu larly at STAT binding site and resulted in increased p21 promoter activity. More over chrysin as a HDAC inhibitor cause apoptosis by decreasing the levels of NF kB targeted and HDACi related genes such as Bcl xL, survivin and increased the level of caspase 3 proteins. Methods Chemical structure and extraction of natural compounds The dried stem bark of dundilum tree, Oroxylum indicum was grinded and extracted consecutively with hexane in a soxhlet apparatus.

Solid residue in the hexane extract was filtered and subjected to silica gel column chromatography to isolate two major frac tions. Fraction F1 was purified on silica gel column chromatography eluted with 0. 5 % MeoH in Chloroform to isolate methoxy chrysin. Simi larly, Fraction F2 was subjected to repeated column chro matography with the elution of 2 % MeoH in Chloroform to isolate oroxylin A and chrysin. The purifi cation, chemical structure and characterization of all three compounds were determined via extensive spectroscopic NMR, ESI MS, and HPLC methods. The conserved methyl oxide and hydroxyl group are shown in the chemical struc ture of small flavonoid compounds. Cell culture A375, U3A cell lines were maintained in DMEM. Whereas K562 cell line was maintained in RPMI media.

All three cell lines were supplemented with 10 % FCS, 1 % pencillin/ streptomycin 5 % glutamine. These cell lines were grown at 370 C in a humidified chamber containing 5 % CO2. MTT assay Cell viability was assessed by the MTT assay, a mitochon drial function assay. It is based on the ability of viable cells to reduce the MTT to insoluble formazan crystals by mitochondrial dehydrogenase. A375 cells were seeded in a 96 well plate at a density 10,000 cells/well. After overnight incubation, cells were treated with compounds chrysin, methoxy chrysin, oroxylin A at a final concentration of 40 uM and Trichostatin A at a final concentration of 4 uM and incubated for 24 h. Medium was then discarded and replaced with 10 uL MTT dye. Plates were incubated at 37 C for 2 h.

The resulting formazan crystals were solu bilized in 100 uL extraction buffer. The optical density was read at 570 using micro plate reader. Cell Cycle Analysis 5 X 105 A375 cells were seeded in 60 mm dish and were allowed to grow for 24 h. Compounds chrysin, oroxylin A, methoxy chrysin at 40 uM final concentration as well as TSA at 4 uM final concentration were added Anacetrapib to the culture media, and the cells were incubated for an additional 24, 48 and 72 h.