ABSTRACT
We report the first well-characterized selective chemical probe for histone deacetylase 10 (HDAC10) with unprecedented selectivity over other HDAC isozymes. HDAC10 deacetylates polyamines and has a distinct substrate specificity, making it unique among the 11 zinc-dependent HDAC hydrolases. Taking inspiration from HDAC10 polyamine substrates, we systematically inserted an amino group ("aza-scan") into the hexyl linker moiety of the approved drug Vorinostat (SAHA). This one-atom replacement (CâN) transformed SAHA from an unselective pan-HDAC inhibitor into a specific HDAC10 inhibitor. Optimization of the aza-SAHA structure yielded the HDAC10 chemical probe DKFZ-748, with potency and selectivity demonstrated by cellular and biochemical target engagement, as well as thermal shift assays. Cocrystal structures of our aza-SAHA derivatives with HDAC10 provide a structural rationale for potency, and chemoproteomic profiling confirmed exquisite cellular HDAC10-selectivity of DKFZ-748 across the target landscape of HDAC drugs. Treatment of cells with DKFZ-748, followed by quantification of selected polyamines, validated for the first time the suspected cellular function of HDAC10 as a polyamine deacetylase. Finally, in a polyamine-limiting in vitro tumor model, DKFZ-748 showed dose-dependent growth inhibition of HeLa cells. We expect DKFZ-748 and related probes to enable further studies on the enigmatic biology of HDAC10 and acetylated polyamines in both physiological and pathological settings.
Subject(s)
Histone Deacetylase Inhibitors , Isoenzymes , Humans , Vorinostat , Histone Deacetylase Inhibitors/pharmacology , Histone Deacetylase Inhibitors/chemistry , HeLa Cells , Histone Deacetylases/chemistry , Polyamines/pharmacology , Zinc , Hydroxamic Acids/pharmacology , Hydroxamic Acids/chemistryABSTRACT
We report the synthesis and evaluation of a class of selective multitarget agents for the inhibition of HDAC6, HDAC8, and HDAC10. The concept for this study grew out of a structural analysis of the two selective inhibitors Tubastatinâ A (HDAC6/10) and PCI-34051 (HDAC8), which we recognized share the same N-benzylindole core. Hybridization of the two inhibitor structures resulted in dihydroxamic acids with benzyl-indole and -indazole core motifs. These substances exhibit potent activity against HDAC6, HDAC8, and HDAC10, while retaining selectivity over HDAC1, HDAC2, and HDAC3. The best substance inhibited the viability of the SK-N-BE(2)C neuroblastoma cell line with an IC50 value similar to a combination treatment with Tubastatinâ A and PCI-34051. This compound class establishes a proof of concept for such hybrid molecules and could serve as a starting point for the further development of enhanced HDAC6/8/10 inhibitors.
Subject(s)
Drug Design , Histone Deacetylase 6/antagonists & inhibitors , Histone Deacetylase Inhibitors/pharmacology , Hydroxamic Acids/pharmacology , Repressor Proteins/antagonists & inhibitors , Cell Proliferation/drug effects , Cell Survival/drug effects , Dose-Response Relationship, Drug , Histone Deacetylase 6/metabolism , Histone Deacetylase Inhibitors/chemical synthesis , Histone Deacetylase Inhibitors/chemistry , Histone Deacetylases/metabolism , Humans , Hydroxamic Acids/chemical synthesis , Hydroxamic Acids/chemistry , Molecular Structure , Repressor Proteins/metabolism , Structure-Activity Relationship , Tumor Cells, CulturedABSTRACT
The discovery of isozyme-selective histone deacetylase (HDAC) inhibitors is critical for understanding the biological functions of individual HDACs and for validating HDACs as drug targets. The isozyme HDAC10 contributes to chemotherapy resistance and has recently been described to be a polyamine deacetylase, but no studies toward selective HDAC10 inhibitors have been published. Using two complementary assays, we found Tubastatin A, an HDAC6 inhibitor, to potently bind HDAC10. We synthesized Tubastatin A derivatives and found that a basic amine in the cap group was required for strong HDAC10 binding. HDAC10 inhibitors mimicked knockdown by causing dose-dependent accumulation of acidic vesicles in a neuroblastoma cell line. Furthermore, docking into human HDAC10 homology models indicated that a hydrogen bond between a cap group nitrogen and the gatekeeper residue Glu272 was responsible for potent HDAC10 binding. Taken together, our data provide an optimal platform for the development of HDAC10-selective inhibitors, as exemplified with the Tubastatin A scaffold.
Subject(s)
Benzamides/metabolism , Glutamic Acid/chemistry , Histone Deacetylase Inhibitors/metabolism , Histone Deacetylases/metabolism , Hydroxamic Acids/metabolism , Animals , Benzamides/chemical synthesis , Benzamides/chemistry , Fluorescence Resonance Energy Transfer , HeLa Cells , Histone Deacetylase 6/chemistry , Histone Deacetylase 6/metabolism , Histone Deacetylase Inhibitors/chemical synthesis , Histone Deacetylase Inhibitors/chemistry , Histone Deacetylases/chemistry , Humans , Hydrogen Bonding , Hydrophobic and Hydrophilic Interactions , Hydroxamic Acids/chemical synthesis , Hydroxamic Acids/chemistry , Ligands , Molecular Docking Simulation , Molecular Structure , Protein Binding , Structure-Activity Relationship , ZebrafishABSTRACT
The first stereoselective total synthesis of the natural product pyrronazolâ B, which contains a chlorinated pyrrole-oxazole-pyrone framework, has been achieved. Genome sequencing of the myxobacterial producer strain Nannocystis pusilla Ari7 led to the identification of the putative biosynthetic gene cluster. The proposed biosynthetic pathway was supported by feeding experiments with stable isotopes of three biosynthetic building blocks, namely l-proline, l-serine, and l-methionine.