Chemical phylogenetics of histone deacetylases - PubMed (original) (raw)

Chemical phylogenetics of histone deacetylases

James E Bradner et al. Nat Chem Biol. 2010 Mar.

Abstract

The broad study of histone deacetylases in chemistry, biology and medicine relies on tool compounds to derive mechanistic insights. A phylogenetic analysis of class I and II histone deacetylases (HDACs) as targets of a comprehensive, structurally diverse panel of inhibitors revealed unexpected isoform selectivity even among compounds widely perceived as nonselective. The synthesis and study of a focused library of cinnamic hydroxamates allowed the identification of, to our knowledge, the first nonselective HDAC inhibitor. These data will guide a more informed use of HDAC inhibitors as chemical probes and therapeutic agents.

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Figures

Figure 1

Figure 1

Development of a platform for biochemical profiling of human deacetylases. (a) Chemical structure of substrates 3 - 6. (b) Comparative enzymatic activity of HDAC1-9 with tripeptide substrate 3 and trifluoro acetyl-lysine tripeptide substrate 6, studied at equivalent substrate concentration (10 μM). Data represent mean values of three measurements ± s.d. Substrate 6 allows miniaturized, kinetic study of HDAC4, 5, 7, 8 and 9.

Figure 2

Figure 2

Chemical phylogenetic analysis of HDACs identifies unexpected selectivity of HDAC inhibitors. (a) Hierarchical clustering of HDACs and a representative panel of structurally-diverse HDAC inhibitor tool and investigational compounds 1, 2, 7-20 weighted by inhibitory potency (Ki). Complete quantitative data are shown in Supplementary Table 1. Data based on mean values of triplicate measurements. (b,c) Chemical structure and enzymatic selectivity profile of (b) MS-275 19 and (c) SAHA 1, overlaying molecular phylogeny. HDAC dendrograms are adapted from Supplementary Figure 4. Circles are proportionate in size to Ki on a logarithmic scale, as shown.

Figure 3

Figure 3

Inhibition of Class IIa HDACs by acetylated lysine based substrates. Inhibition of trifluoroacetyl-lysine substrate 6 processing by (a) acetyl-lysine substrate 3 (b) acetyl-lysine substrate 21 is presented in dose-response format for Class IIa HDACs. Data represent mean values of triplicate measurements ± s.d. IC50 curves were fit by logistic regression.

Figure 4

Figure 4

Synthesis and testing of an HDAC-biased chemical library and identification of a non-selective HDAC inhibitor. (a) Library design of meta- and para-substituted hydroxamic acid HDAC inhibitors, utilizing parallel condensation of aldehydes, efficiently samples chemical diversity at the capping feature. (b) Biochemical profiling data for the para-substituted sub-library (n=160 compounds), presented in dose-response format for inhibition of HDAC5. Structural variation in the capping feature was observed to confer a broad range of potency, as illustrated with the most (IC50= 18 nM) and least (IC50 = 55 μM) potent small molecules tested. (c) Comparative biochemical profiling of meta- (red) and para-substituted (blue) sub-libraries for relative inhibition of HDAC2 and HDAC3. The complete library was studied and is displayed at a range of concentrations (0.03, 0.3, 3.0 and 30.0 μM). Compounds of this structural class do not discriminate between HDAC2 and HDAC3. (d) Comparative biochemical profiling of meta- (red) and para-substituted (blue) sub-libraries for relative inhibition of HDAC5 and HDAC7. The complete library was studied and is displayed at a range of concentrations (0.03, 0.3, 3.0 and 30.0 μM). Para-substituted cinnamic hydroxamic acids exhibit increased potency for HDAC5, relative to meta-substituted regioisomers. (e) Specificity profile of pandacostat 22 overlaying molecular phylogeny. HDAC dendrograms are adapted from Supplementary Figure 4. Circles are proportionate in size to Ki on a logarithmic scale, as shown. (f) Immunoblot of Jurkat cells treated with pandacostat for 24 hours and stained for acetylated histones (AcH3K18), acetylated alpha-tubulin (AcTub) or GAPDH. (g) Chemical structure of pandacostat 22.

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