Preincubation of Pituitary Tumor Cells With the Epidrugs Zebularine and Trichostatin A Are Permissive for Retinoic Acid-Augmented Expression of the BMP-4 and D2R Genes

Institute of Science and Technology in Medicine, Keele University School of Medicine, Stoke-on-Trent, Staffordshire, ST4 7QB. UK.
Endocrinology (Impact Factor: 4.64). 03/2013; 154(5). DOI: 10.1210/en.2013-1061
Source: PubMed

ABSTRACT Retinoic acid (RA) induced expression of bone morphogenetic protein (BMP-4) inhibits in vitro and in vivo cell proliferation and ACTH synthesis in corticotroph derived tumour cells. Reduced expression of BMP-4 in this adenoma subtype is associated with epigenomic silencing and similar silencing mechanisms are also associated with the RA responsive dopamine D2 receptor (D2R) in somatolactotroph cells. We now show that pre-incubation with the epidrugs, zebularine and TSA is obligate and permissive for RA induced expression of the BMP-4 and the D2R genes in pituitary tumour cells. Combined epidrug challenges are associated with marginal reduction in CpG island methylation. However, significant change to histone tail modifications toward those associated with expression-competent genes is apparent, whereas RA challenge alone or in combined incubations does not impact on these modifications. Epidrug mediated and RA augmented expression of endogenous BMP-4 increased or decreased cell proliferation and CFE in GH3 and AtT-20 pituitary tumour cells respectively, and recapitulating recent reports of challenges of these cells with exogenous ligand. The specificity of the BMP-4 mediated effects was further supported by knock-down experiments of the BMP-4 antagonist noggin (siRNA). Knock-down of noggin, in the absence and the presence of epidrugs, induced and augmented BMP-4 expression respectively. In cell proliferation assays, challenge with either epidrugs or siRNA led to significant increase in cell numbers at the 72hr time point, however, in siRNA treated cells co-incubated with epidrugs a significant increase was apparent at the 48hr time point. These studies show the potential of combined drug challenges as a treatment option, where epidrug renders silenced genes responsive to conventional therapeutic options.

  • [Show abstract] [Hide abstract]
    ABSTRACT: BACKGROUND New therapies are required for castrate-resistant prostate cancer (CRPC), and growth-arrest specific 5 (GAS5) lncRNA, which riborepresses androgen receptor action, may offer novel opportunities in this regard. This lncRNA promotes the apoptosis of prostate cancer cells and its levels decline as prostate cancer cells acquire castrate-resistance, so that enhancing GAS5 expression may improve the effectiveness of chemotherapies. Since GAS5 is a member of the 5' terminal oligopyrimidine gene family, we have examined mTOR inhibition as a strategy to increase GAS5 expression. Furthermore, we have determined if GAS5 itself mediates the action of mTOR inhibitors, as demonstrated for other chemotherapeutic agents in prostate cancer cells.METHODS The effects of mTOR inhibitors on GAS5 lncRNA levels and cell growth were determined in a range of prostate cancer cell lines. Transfection of cells with GAS5 siRNAs and plasmid constructs was performed to determine the involvement of GAS5 lncRNA in mTOR inhibitor action.RESULTSFirst generation mTORC1, combined mTORC1/mTORC2 and dual PI3K/mTOR inhibitors all increased cellular GAS5 levels and inhibited culture growth in androgen-dependent (LNCaP) and androgen-sensitive (22Rv1) cell lines, but not in androgen-independent (PC-3 and DU 145) cell lines. The latter exhibited low endogenous GAS5 expression, and GAS5 silencing in LNCaP and 22Rv1 cells decreased the sensitivity to mTOR inhibitors, whereas transfection of GAS5 lncRNA sensitized PC-3 and DU 145 cells to these agents.CONCLUSIONmTOR inhibition enhances GAS5 transcript levels in certain prostate cancer cell lines. This selectivity is likely to be related to endogenous GAS5 expression levels, since GAS5 lncRNA is itself required for mTOR inhibitor action in prostate cancer cells. Prostate 9999: XX–XX, 2015. © 2015 Wiley Periodicals, Inc.
    The Prostate 02/2015; DOI:10.1002/pros.22952 · 3.57 Impact Factor
  • [Show abstract] [Hide abstract]
    ABSTRACT: Cushing's disease (CD) is a rare condition in which hypercortisolemia is secondary to excessive ACTH release from a pituitary corticotroph adenoma. CD is associated with significant morbidity and mortality, and a safe therapy that effectively targets the pituitary tumor is still lacking. Retinoic acid (RA) and dopamine agonists (DAs) have recently been considered as monotherapy in CD patients, and satisfactory results have been reported, albeit in a limited number of patients. Given the permissive role of RA on the dopamine receptor type-2 (DRD2), the aim of present study was to see whether a combination of 9-cis RA and the DA bromocriptine (Br) might represent a possible treatment for CD. Here we show that 9-cis RA induces a functional DRD2 in the pituitary corticotroph cell line AtT20, and increases cell sensitivity to Br via a mechanism only partially related to corticotroph-to-melanotroph transdifferentiation. In addition, 9-cis RA and Br act synergistically to modulate cell viability, with favorable implications for clinical use. In nearly 45% of corticotropinoma-derived primary cultures, the combined administration of 9-cis RA and Br lowered the steady-state level of the ACTH precursor proopiomelanocortin (POMC) more efficiently than either of the drugs alone. In conclusion, the effects of a combination of 9-cis RA and Br on ACTH synthesis/secretion and cell viability in AtT20, and on POMC transcriptional activity in human corticotropinomas might represent a suitable starting point for assessing the potential of this treatment regimen for ACTH-secreting pituitary adenomas. This study thus has potentially important implications for novel therapeutic approaches to Cushing's disease.
    Endocrinology 06/2014; 155(9):en20131820. DOI:10.1210/en.2013-1820 · 4.64 Impact Factor
  • [Show abstract] [Hide abstract]
    ABSTRACT: To review recent advances in our knowledge and understanding of aberrations that target the epigenome in sporadic pituitary adenomas.
    Current Opinion in Endocrinology Diabetes and Obesity 08/2014; 21(4):299-305. DOI:10.1097/MED.0000000000000078 · 3.77 Impact Factor