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

Abstract

<jats:p>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.</jats:p>

Aza-SAHA Derivatives are Selective Histone Deacetylase 10 Chemical Probes That Inhibit Polyamine Deacetylation and Phenocopy HDAC10 Knockout

“…comparatively little attention by the medicinal chemistry community (7)(8)(9)(10)(11), resulting in a lack of appropriate pharmacological tools. Indeed, it took 15 years after its isolation before HDAC10 was shown to be a poor lysine deacetylase, which instead recognizes acetylated polyamines as substrates.…”

Identification of histone deacetylase 10 (HDAC10) inhibitors that modulate autophagy in transformed cells

“…In case of drHDAC10 we recently developed an enzymatic in vitro assay that was used in the current study. 24 Compound 6a, bearing an N-piperidinomethylene capping group, showed only weak HDAC10 inhibition and also very low activity against other HDACs. Meanwhile, replacement of the piperidine ring of 6a with an N-methylpiperazine moiety yielded compound 6b which was found to a potent inhibitor against drHDAC10 (IC50 43 ± 7 nM).…”

Identification of HDAC10 Inhibitors that Modulate Autophagy-Related Proteins in Transformed Cells

“…In case of drHDAC10 we recently developed an enzymatic in vitro assay that was used in the current study. 24 Compound 6a, bearing an N-piperidinomethylene capping group, showed only weak HDAC10 inhibition and also very low activity against other HDACs. Meanwhile, replacement of the piperidine ring of 6a with an N-methylpiperazine moiety yielded compound 6b which was found to a potent inhibitor against drHDAC10 (IC 50 43 ± 7 nM).…”