Mitotane activates ATF4/ATF3 axis triggering endoplasmic reticulum stress in adrenocortical carcinoma cells

Biomed Pharmacother. 2025 Mar:184:117917. doi: 10.1016/j.biopha.2025.117917.

Aurora Schiavon ¹, Laura Saba ², Carlotta Evaristo ³, Jessica Petiti ⁴, Ymera Pignochino ⁵, Giulio Ferrero ⁶, Giorgia Giordano ⁷, Cristina Tucciarello ⁸, Soraya Puglisi ⁹, Giuseppe Reimondo ¹⁰, Massimo Terzolo ¹¹, Marco Lo Iacono ¹²

Affiliations

1 Department of Clinical and Biological Sciences, University of Turin, Turin, Italy. Electronic address: aurora.schiavon@unito.it.
2 Department of Clinical and Biological Sciences, University of Turin, Turin, Italy. Electronic address: laura.saba@unito.it.
3 Department of Clinical and Biological Sciences, University of Turin, Turin, Italy. Electronic address: carlotta.evaristo@unito.it.
4 Division of Advanced Materials Metrology and Life Sciences, Istituto Nazionale di Ricerca Metrologica (INRiM), Turin, Italy. Electronic address: j.petiti@inrim.it.
5 Department of Clinical and Biological Sciences, University of Turin, Turin, Italy; Candiolo Cancer Institute, FPO-IRCCS, Candiolo, Turin, Italy. Electronic address: ymera.pignochino@unito.it.
6 Department of Clinical and Biological Sciences, University of Turin, Turin, Italy. Electronic address: giulio.ferrero@unito.it.
7 Candiolo Cancer Institute, FPO-IRCCS, Candiolo, Turin, Italy; Department of Oncology, University of Turin, Turin, Italy. Electronic address: giorgia.giordano@unito.it.
8 Department of Clinical and Biological Sciences, University of Turin, Turin, Italy; Candiolo Cancer Institute, FPO-IRCCS, Candiolo, Turin, Italy. Electronic address: cristina.tucciarello@unito.it.
9 Department of Clinical and Biological Sciences, University of Turin, Turin, Italy. Electronic address: soraya.puglisi@unito.it.
10 Department of Clinical and Biological Sciences, University of Turin, Turin, Italy. Electronic address: giuseppe.reimondo@unito.it.
11 Department of Clinical and Biological Sciences, University of Turin, Turin, Italy. Electronic address: massimo.terzolo@unito.it.
12 Department of Clinical and Biological Sciences, University of Turin, Turin, Italy. Electronic address: marco.loiacono@unito.it.

PMID: 39965324 DOI: 10.1016/j.biopha.2025.117917

Abstract

Adrenocortical Carcinoma is a rare and aggressive endocrine malignancy, that arises from cells of one of the three cortical layers of the adrenal gland. Radical surgery is the only curative treatment, even if recurrence rates are high. Therapeutic options are limited, with mitotane as the cornerstone of medical therapy. Despite 50 years of clinical use, the mechanism of action of mitotane has not yet been fully established, possibly due to the drug's susceptibility to interaction with confounding factors that reduce its biological activity. In the present study, we evaluated by RNAseq the effect of mitotane on gene expression in the H295R cell line, in an environment free of known confounding factors. Our approach allowed us to identify transcriptional deregulation of the ATF4/ATF3 axis, often involved in ER stress. These results were also validated by ddPCR in independent experiments. Mitotane-mediated ATF4 overexpression was also confirmed at the protein level. We observed how an incremental concentration of mitotane could deregulate main biological pathways. Further, we confirmed, both at RNAseq and ddPCR level, the mitotane-mediated downmodulation of genes such as STAR, CYP11A1, CYP21A2, and HSD3B2, highlighting its effect on steroid Hormones biosynthesis. Through our approach, we identified biological pathways altered by mitotane in early response stages and with low drug concentrations. Some of these pathways could potentially be investigated in the future as functional biomarkers to monitor adrenocortical carcinoma treatment or as new pharmacological targets for this rare disease.

Keywords

ATF3; ATF4; Adrenocortical carcinoma; ER stress; Mitotane; RNAseq.

Products

Cat. No.

Product Name

Description

Target

Research Area
HY-13690

Mitotane

99.94%, Antineoplastic Agent

Apoptosis

Cancer

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