A functional genetic screen identifies retinoic acid signaling as a target of histone deacetylase inhibitors

Abstract

Understanding the pathways that are targeted by cancer drugs is instrumental for their rational use in a clinical setting. Inhibitors of histone deacetylases (HDACI) selectively inhibit proliferation of malignant cells and are used for the treatment of cancer, but their cancer selectivity is understood poorly. We conducted a functional genetic screen to address the mechanism(s) of action of HDACI. We report here that ectopic expression of two genes that act on retinoic acid (RA) signaling can cause resistance to growth arrest and apoptosis induced by HDACI of different chemical classes: the retinoic acid receptor ␣ (RAR␣) and preferentially expressed antigen of melanoma (PRAME), a repressor of RA signaling. Treatment of cells with HDACI induced RA signaling, which was inhibited by RAR␣ or PRAME expression. Conversely, RAR-deficient cells and PRAME-knockdown cells show enhanced sensitivity to HDACI in vitro and in mouse xenograft models. Finally, a combination of RA and HDACI acted synergistically to activate RA signaling and inhibit tumor growth. These experiments identify the RA pathway as a rate-limiting target of HDACI and suggest strategies to enhance the therapeutic efficacy of HDACI.biomarker ͉ chromatin modification ͉ drug resistance ͉ epigenetics ͉ nuclear hormone receptor E pigenetic DNA and histone modifications are appreciated as major determinants in the control of gene activity, and they are extensively deregulated in cancer. Histone acetylation is regulated by the opposing activities of histone acetyltransferases (HATs) and histone deacetylases (HDACs), which catalyze the addition and removal of acetyl groups to histones, respectively, and to a growing list of nonhistone substrates (1). The activities of HATs and HDACs are altered in several human cancers, and modulation of these classes of enzymes provides a potentially attractive therapeutic modality (2, 3). Several classes of HDAC inhibitors (HDACI) have been identified that block enzyme activity, resulting in global histone hyperacetylation. A wide array of literature on HDACI exists, describing their various effects, including G 1 and G 2 /M cell cycle arrests, apoptosis, and differentiation, and several HDACI have entered clinical trials (2-4). Gene expression profiling studies revealed that HDACI treatment induces alterations in transcription of Ͻ5% to Ϸ20% of expressed genes (5, 6) and have not elucidated a consistent picture of the pathway(s) or target(s) that are modulated by HDACI and, consequently, have not provided a comprehensive explanation for their anticancer effects.To identify cellular targets of HDACI action in transformed cells, we used the approach of large-scale functional genetic screening. In this screen we asked which genes or pathways could confer cellular resistance to HDACI. The present work provides evidence that the retinoic acid receptor (RAR) pathway is targeted by HDACI and that the cytotoxic effects of HDACI in solid tumor cells are, at least in part, through derepression of retinoic acid (RA) signaling. ResultsGe...

BCR–ABL-mediated upregulation of PRAME is responsible for knocking down TRAIL in CML patients

“…First, PRAME is a dominant repressor of RA receptor signaling (Epping et al, 2005), whereas TRAIL is a gene inducible by RA treatment. Second, PRAME is capable of inhibiting the induction of apoptosis by histone deacetylase inhibitors in a concentration-dependent manner (Epping et al, 2007), an event shown to be modulated, at least in leukemia, by induction of TRAIL expression (Insinga et al, 2005;Nebbioso et al, 2005). Indeed, our results clearly showed that in the CML line K562, the inhibition of TRAIL expression is mediated by PRAME and this process relies on EZH2 binding to the TRAIL promoter.…”

Thyroid hormone activation of retinoic acid synthesis in hypothalamic tanycytes

“…In some cases, this is believed to be a behavioural control mechanism that, for instance, contributes to imprinting action of the environment on the young animal that results in altered behaviour in the mature animal. Epigenetic control via histone acetylation is particularly active in promoting RA signalling, and derepression of RA signalling may be a major rate‐limiting target of histone deacetylase (HDAC) inhibitors (Epping et al, 2007). To explore whether the RA signalling system in the hypothalamus is acted upon by such an epigenetic mechanism, cultured hypothalamic slices were treated with a low concentration of RA (10 nM) which results in weaker induction of gene expression.…”

“…There is some evidence that the RA signalling pathway is a major target of HDACIs (Epping et al, 2007). Unliganded RARα is known to suppress transcription of RA target genes by recruiting components of the corepressor complex (Hauksdottir et al, 2003), including HDACs, and HDACIs act partly via derepression of RA signalling (Epping et al, 2007).…”

“…There is some evidence that the RA signalling pathway is a major target of HDACIs (Epping et al, 2007). Unliganded RARα is known to suppress transcription of RA target genes by recruiting components of the corepressor complex (Hauksdottir et al, 2003), including HDACs, and HDACIs act partly via derepression of RA signalling (Epping et al, 2007). The data presented in this study, in which induction of Rarb and Ghrh by RA was potentiated in the presence of a class I/II HDACI, suggests that epigenetic regulation of hypothalamic function can, in part, act via the RA signalling pathway.…”

The Drug-Induced Degradation of Oncoproteins: An Unexpected Achilles' Heel of Cancer Cells?

“…These initial alterations are most likely to be the drivers. Very powerful genetic systems using RNA interference approaches have also identified previously unsuspected key pathways (18). However, another issue comes to light: what endpoint to consider?…”