Dietary HDAC inhibitors: time to rethink weak ligands in cancer chemoprevention?

Abstract

There is growing interest in the various mechanisms that regulate chromatin remodeling, including modulation of histone deacetylase (HDAC) activities. Competitive HDAC inhibitors disrupt the cell cycle and/or induce apoptosis via de-repression of genes such as P21 and BAX, and cancer cells appear to be more sensitive than non-transformed cells to trichostatin A and related HDAC inhibitory compounds. This apparent selectivity of action in cancer cells makes HDAC inhibitors an attractive avenue for drug development. However, in the search for potent HDAC inhibitors with cancer therapeutic potential there has been a tendency to overlook or dismiss weak ligands that could prove effective in cancer prevention, including agents present in the human diet. Recent reports have described butyrate, diallyl disulfide and sulforaphane as HDAC inhibitors, and many other dietary agents will be probably discovered to attenuate HDAC activity. Here we discuss 'pharmacologic' agents that potently de-repress gene expression (e.g. during therapeutic intervention) versus dietary HDAC inhibitors that, as weak ligands, might subtly regulate the expression of genes involved in cell growth and apoptosis. An important question is the extent to which dietary HDAC inhibitors, and other dietary agents that affect gene expression via chromatin remodeling, modulate the expression of genes such as P21 and BAX so that cells can respond most effectively to external stimuli and toxic insults.

Fig. 1

Indirect and direct mechanisms of HDAC modulation, leading to de-repression of silenced genes. (A) Binding of the retinoic acid receptor (RAR) to the retinoic acid response element (RARE), in conjunction with the promyelocytic leukemia (PML) protein, recruits corepressor-Sin3-HDAC (CoR-SIN3-HDAC) complexes. HDAC removes acetyl groups from histones and causes chromatin condensation, leading to gene silencing. (B) Retinoic acid binds to RAR and induces a conformational change in the protein, leading to the release of CoR-SIN3-HDAC and recruitment of co-activator-histone acetyltransferase (CoA-HAT) complexes. HATs, such as p300 and CREB-binding protein, transfer acetyl groups (blue dots) to the histone amino-terminal tails, leading to nucleosomal repulsion, chromatin relaxation and gene transcription. (C) Resistance to retinoic acid treatment, as in the case of the RAR–PLZF (promyelocytic leukemia zinc finger) fusion protein and its associated CoR/HDAC complexes, can be circumvented through the use of competitive inhibitors of HDAC, such as trichostatin A.

Fig. 2

Structures of dietary HDAC inhibitors. (A) The smallest known HDAC inhibitor, butyrate, contains a short three carbon ‘spacer’ attached to a carboxylic acid functional group. (B) The dietary HDAC inhibitor diallyl disulfide, which is present in garlic, can be converted to the metabolite S-allylmercaptocysteine, containing a spacer and a carboxylic acid functional group. (C) Sulforaphane, an isothiocyanate found in broccoli and broccoli sprouts, can be converted to the metabolite SFN–Cys, which has spacer and carboxylic acid functional that fits within the HDAC active site (Figure 3).

Fig. 3

SFN–Cys/HDAC interaction. Molecular modeling studies revealed a plausible interaction for SFN–Cys within the active site of an HDAC, with the carboxylate group of SFN–Cys positioned as a bidentate ligand with the buried zinc atom. See (30) for further details.

Fig. 4

HDAC inhibitory activity of sulforaphane (SFN) and structurally related isothiocyanates. (A) The mercapturic acid pathway converts SFN sequentially to SFN–GSH (SFN–glutathione), SFN–Cys (SFN–cysteine) and SFN–NAC (SFN–N-acetylcysteine). SFN–Cys formed from SFN–GSH, or after deacetylation of SFN–NAC by HDAC, leads to competitive enzyme inhibition, according to the working hypothesis (see text). Direct addition of SFN or SFN–GSH to isolated nuclear extracts in vitro had no significant effect on HDAC activity, whereas SFN–NAC (blue bar) and SFN–Cys (red bar) attenuated HDAC activity significantly. (B) Incubation of SFN with HCT116 human colon cancer cells followed by testing of the cell lysates revealed significant HDAC inhibition, as seen with several other structurally related isothiocyanates. All concentrations were 15 μM, except butyrate which was 1 mM. Data = mean ± SD, n =3. *P < 0.05, **P <0.01, ***P < 0.001, using Student’s t-test versus the corresponding control (gray bar). For details of the HDAC assay and other conditions, see (30).