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First Fluorescent Acetylspermidine Deacetylation Assay for HDAC10 Identifies Inhibitors of Neuroblastoma Cell Colony Growth That Increase Lysosome Accumulation

preprint
revised on 09.07.2020 and posted on 10.07.2020 by Daniel Herp, Johannes Ridinger, Dina Robaa, Stephen A. Shinsky, Karin Schmidtkunz, Talha Z. Yesiloglu, Theresa Bayer, Peter Sehr, Nikolas Gunkel, Aubry K. Miller, David W. Christianson, Ina Oehme, Wolfgang Sippl, Manfred Jung
Histone deacetylases (HDACs) are important epigenetic regulators involved in many diseases, esp. cancer. First HDAC inhibitors have been approved for anticancer therapy and many are in clinical trials. Among the 11 zinc-dependent HDACs, HDAC10 has received relatively little attention by drug discovery campaigns, despite its involvement e.g. in the pathogenesis of neuroblastoma. This is due in part to a lack of robust enzymatic conversion assays. In contrast to the protein lysine deacetylase and deacylase activity of the other HDAC subtypes, it has recently been shown that HDAC10 has strong preferences for deacetylation of oligoamine substrates like spermine or spermidine. Hence, it also termed a polyamine deacetylase (PDAC). Here, we present the first fluorescent enzymatic conversion assay for HDAC10 using an aminocoumarin labelled acetyl spermidine derivative to measure its PDAC activity, which is suitable for high-throughput screening. Using this assay, we identified potent inhibitors of HDAC10 mediated spermidine deacetylation in-vitro. Among those are potent inhibitors of neuroblastoma colony growth in culture that show accumulation of lysosomes, implicating disturbance of autophagic flux.

Funding

Deutsche Forschungsgemeinschaft Ju295/13-1

Deutsche Forschungsgemeinschaft Si868/13-1

Deutsche Forschungsgemeinschaft Oe542/2-1

Deutsche Forschungsgemeinschaft INST 29/931-1

Hector Foundation M91

NIH GM49758

NIH F32GM125141

History

Email Address of Submitting Author

manfred.jung@pharmazie.uni-freiburg.de

Institution

University of Freiburg

Country

Germany

ORCID For Submitting Author

0000-0002-6361-7716

Declaration of Conflict of Interest

No confilict of interest

Version Notes

1.0 Original version 1.1 Minor corrections (esp. Figure 2) 1.2 Minor corrections

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