The bromodomain inhibitor JQ1+ reduces calcium-sensing receptor activity in pituitary cell lines.

Kate E Lines,Anna K Gluck,Caroline M Gorvin,Chas Bountra,Rajesh V Thakker,Supat Thongjuea

doi:10.1530/jme-21-0030

Abstract

Corticotrophinomas represent 10% of all surgically removed pituitary adenomas, however, current treatment options are often not effective, and there is a need for improved pharmacological treatments. Recently, JQ1+, a bromodomain inhibitor that promotes gene transcription by binding acetylated histone residues and recruiting transcriptional machinery, has been shown to reduce proliferation in a murine corticotroph cell line, AtT20. RNA-Seq analysis of AtT20 cells following treatment with JQ1+ identified the calcium-sensing receptor (CaSR) gene as significantly downregulated, which was subsequently confirmed using real-time PCR and Western blot analysis. CaSR is a G protein-coupled receptor that plays a central role in calcium homeostasis but can elicit non-calcitropic effects in multiple tissues, including the anterior pituitary where it helps regulate hormone secretion. However, in AtT20 cells, CaSR activates a tumour-specific cAMP pathway that promotes ACTH and PTHrP hypersecretion. We hypothesised that the Casr promoter may harbour binding sites for BET proteins, and using chromatin immunoprecipitation (ChIP)-sequencing demonstrated that the BET protein Brd3 binds to the promoter of the Casr gene. Assessment of CaSR signalling showed that JQ1+ significantly reduced Ca2+e-mediated increases in intracellular calcium (Ca2+i) mobilisation and cAMP signalling. However, the CaSR-negative allosteric modulator, NPS-2143, was unable to reduce AtT20 cell proliferation, indicating that reducing CaSR expression rather than activity is likely required to reduce pituitary cell proliferation. Thus, these studies demonstrate that reducing CaSR expression may be a viable option in the treatment of pituitary tumours. Moreover, current strategies to reduce CaSR activity, rather than protein expression for cancer treatments, may be ineffective.

Highlights

Pituitary tumours are common neoplasms, which account for 10–15% of primary intracranial tumours and are identified in > 25% of unselected autopsies and approximately 20% of the population undergoing intracranial imaging (Daly et al 2009, Di Ieva et al 2014)
Our studies showed that JQ1+ treatment of corticotroph pituitary cells impairs calcium-sensing receptor (CaSR) mRNA and protein expression (Fig. 1), which may contribute to the reduced cell proliferation observed in earlier studies (Lines et al 2020)
This indicates that targeting CaSR expression could be beneficial in reducing proliferation in neuroendocrine tumours, such as corticotrophinomas that represent up to 10% of all surgically removed pituitary adenomas (Daly et al 2009), this remains to be tested in detail

Summary

Introduction

Pituitary tumours are common neoplasms, which account for 10–15% of primary intracranial tumours and are identified in > 25% of unselected autopsies and approximately 20% of the population undergoing intracranial imaging (Daly et al 2009, Di Ieva et al 2014). Pharmacological treatments, including inhibitors of steroidogenesis, glucocorticoid antagonists, dopamine agonists, and somatostatin analogues, may provide an alternative for patients in whom surgery is contraindicated or has been unsuccessful (Cuevas-Ramos et al 2016). These current medical treatments are often not effective in corticotrophinomas, and, there is a clinically unmet need for improved pharmacological treatments for these patients

Methods

Results

Conclusion