Acetylation of histones in chromatin is one mechanism involved in the regulation of gene transcription and is tightly controlled by the balance of acetyltransferase and deacetylase (HDAC) activities. In cancer, some genes are repressed by the inappropriate recruitment of HDACs, e.g., tumor suppressor genes. To understand the genomic effects of HDAC inhibition on gene transcription we studied the gene expression profiles of T24 bladder and MDA breast carcinoma cells treated with three HDAC inhibitors, suberoylanilide hydroxamic acid, trichostatin A, and MS-27-275. The gene expression profiles of the HDAC inhibitors were generally similar to one another and differed substantially from those produced by structurally related inactive analogues; consequently, the changes in gene expression are mechanism-based. Hierarchical clustering of expression profiles demonstrated a greater similarity between the two hydroxamate-containing inhibitors (suberoylanilide hydroxamic acid and trichostatin A) than with MS-27-275. This difference was also supported by cell phenotypic experiments. As many genes were down-regulated as up-regulated by HDAC inhibitor treatment. Comparison of the data sets defined a common (“core”) set of 13 genes regulated by all of the HDAC inhibitors in three cell lines, 8 up-regulated and 5 down-regulated. Ten of 13 genes were confirmed in dose response studies in T24 cells by quantitative-PCR. The core regulated genes are involved predominantly in cell cycle/apoptosis and DNA synthesis in response to HDAC inhibitors. These data will aide in understanding the complex set of events in cells in response to chromatin remodeling induced by HDAC inhibition, which may be responsible for antitumor effects.