Mass spectrometry and DigiWest technology emphasize protein acetylation profile from Quisinostat-treated HuT78 CTCL cell line

Histone deacetylases (HDACs) are key enzymes involved in epigenetic modulation and were targeted by HDAC inhibitors (HDACis) for cancer treatment. The action of HDACis is not restricted to histones and also prevents deacetylation of other proteins, supporting their wide biological actions. The HuT78 cell line is recognized as a key tool to support and understand cutaneous T-cell lymphoma (CTCL) biology and was used as a predictive model since HDACi such as Vorinostat and Panobinostat have both demonstrated apoptotic activities in HuT78 cells and in primary blood CTCL cells. In this study, Quisinostat (JNJ-26481585) a novel second-generation HDACi with highest potency for HDAC1, was tested on HuT78 cell line. Quantitative mass spectrometry (MS)-based proteomics after acetylated-lysine peptide enrichment and a targeted antibody-based immunoassay (DigiWest) were used as complementary technologies to assess the modifications of the acetylated proteome. As expected, several acetylated lysines of histones were increased by the HDACi. Additional acetylated non-histone proteins were modulated after treatment with Quisinostat including the nucleolin (a major nucleolar protein), the replication protein A 70 kDa DNA-binding subunit, the phosphoglycerate kinase 1, the stress-70 protein, the proto-oncogene Myc and the serine hydroxymethyltransferase. A better knowledge of histone and non-histone acetylated protein profile after Quisinostat treatment can strongly support the understanding of non-clinical and clinical results of this HDACi. These technological tools can also help in designing new HDACis in a pharmaceutical drug discovery program.

Significance: A better knowledge of histone and non-histone acetylated protein profile after HDAC inhibitors (HDACis) treatment can strongly support the understanding of non-clinical and clinical investigations in a pharmaceutical drug discovery program. Relative quantification using mass spectrometry -based proteomics after acetylated-lysine peptide enrichment and a targeted antibody-based immunoassay (DigiWest) are proposed as complementary technologies to assess the modifications of the acetylated proteome. Quisinostat (JNJ-26481585) a novel second-generation HDACi with highest potency for HDAC1 was better characterized in vitro in HuT78 cells to support and understand cutaneous T-cell lymphoma (CTCL) therapeutic research program.