Preparation and Biochemical Analysis of Classical Histone Deacetylases.

Histone deacetylase assays were first developed in the 1970s, and subsequently refined in the 1990s with the cloning of HDAC enzymes. Most of these early assays, relying on traditional in vitro chemical methodologies, are still applicable today. More recently, however, cell-based HDAC assays that measure HDAC activities in physiological conditions are emerging. Also, there is a continuing development of assays that can measure an isolated HDAC in the absence of other HDAC activities. This chapter reviews some of the older established methods for assaying HDAC activities, as well as introduces more recently developed nontraditional assays.

Targeting histone deacetylase 6 mediates a dual anti-melanoma effect: Enhanced antitumor immunity and impaired cell proliferation.

The median survival for metastatic melanoma is in the realm of 8-16 months and there are few therapies that offer significant improvement in overall survival. One of the recent advances in cancer treatment focuses on epigenetic modifiers to alter the survivability and immunogenicity of cancer cells. Our group and others have previously demonstrated that pan-HDAC inhibitors induce apoptosis, cell cycle arrest and changes in the immunogenicity of melanoma cells. Here we interrogated specific HDACs which may be responsible for this effect. We found that both genetic abrogation and pharmacologic inhibition of HDAC6 decreases in vitro proliferation and induces G1 arrest of melanoma cell lines without inducing apoptosis. Moreover, targeting this molecule led to an important upregulation in the expression of tumor associated antigens and MHC class I, suggesting a potential improvement in the immunogenicity of these cells. Of note, this anti-melanoma activity was operative regardless of mutational status of the cells. These effects translated into a pronounced delay of in vivo melanoma tumor growth which was, at least in part, dependent on intact immunity as evidenced by the restoration of tumor growth after CD4+ and CD8+ depletion. Given our findings, we provide the initial rationale for the further development of selective HDAC6 inhibitors as potential therapeutic anti-melanoma agents.