Histone deacetylases (HDAC) are a family of enzymes

that have been identified as a promising target to reverse aberrant epigenetic states associated with cancer via the regulation of acetylation levels in histone. Aberrant histone acetylation has been observed in the development of numerous malignancies, and HDAC inhibitors (HDACIs) are a promising new treatment strategy for malignant disease. The hydroxamic acid trichostatin A (TSA) have been known as a differentiation inducer for tumor cells, but its clinical use has been limited by high reactivity and instability. Furthermore, TSA has also been reported to be unstable in vivo following intravenous injection in mice. Suberoylanilide hydroxamic acid (SAHA, vorinostat, Zolinza), is a potent HDACI that has demonstrated antitumor activity in vitro against a variety of cell lines and in vivo against several human tumor xenograft models. Furthermore, SAHA is currently prescribed to treat cutaneous T-cell lymphoma and is also being tested in several phase I to III clinical trials for the treatment of a variety of other cancers, including breast, lung, and colon. Although SAHA is orally administrated because of its hydrophobic property, it is easily metabolized in the liver. Pharmacokinetic studies in patients after administration of SAHA have identified two inactive metabolites, SAHA-glucuronide and 4-anilino-4oxobutanoic acid, which are pharmacologically inactive, and the mean terminal half-life (t1/2) of SAHA was ca. 2.0 h ; therefore, the liposomal SAHA formulation warrants in vivo experiments because it allows intravenous administration and prevents metabolism in the liver. Liposomes have the advantage of an enhanced permeability and retention effect (EPR), increasing their tumor accumulation when they are intravenously injected. Such a type of nanocarrier is supposed to improve the pharmacokinetic and pharmacodistribution of the encapsulated drug. Liposomal LAQ824, hydrophilic hydroxamic acid HDACI, was shown to be both long-circulating and highly stable in vivo; however, in the encapsulation of hydrophobic SAHA into liposome, SAHA must be incorporated into the lipid membrane, which might be limited in its capacity to entrap SAHA. We therefore attempted to synthesize hydroxamatetype HDACI-lipid conjugates as a component of the liposome formulation for efficient incorporation into the liposomal membrane. Previously, we have reported that inclusion of HDACI-lipid conjugates into cationic nanopartilces as a non-viral vector could increase gene expression via hyper acetylation of histone. In this study, taking into consideration the diverse derivatization, we selected SAHA and one of the K-series compounds, K-182, which has two OH groups to tether other groups with a biodegradable ester bond, and synthesized cholesteryl HDACI and HDACI-polyethylene glycol (PEG)-lipid conjugates to prepare liposomal HDACIlipid conjugates (Fig. 1). Furthermore, we evaluated their cytotoxic effect on tumor cells via the acetylation of histone H3 and apoptotic activity.