Growing attention is being paid to association of adrenocortical carcinoma (ACC), a rare endocrine malignancy, to cancer predisposition syndromes caused by germline mutations in genes involved in the control of genome stability. Tumour cells with a defective DNA mismatch repair pathway have a high mutation burden, which results in the production of tumour-associated specific neoantigens and in an increase of the sensitivity to therapies that loosen the constraints of tumour attack by the immune system. The study by Landwehr et al. published in the current issue of the European Journal of Endocrinology describes a patient with an aggressive ACC bearing a germline MUTYH mutation with loss of heterozygosity in the tumour and accumulation of 8-hydroxyguanine in its genomic DNA. The authors managed to establish a novel differentiated cell line from that tumour which bears the stigma of the defective DNA repair mechanism in its genome. The availability of this new cell model inside the expanding toolbox of the ACC cell lines will allow for novel experimental possibilities, in particular for the study of the tumour microenvironment and the response to immunotherapy.
Oliver Zwermann, Felix Beuschlein, Enzo Lalli, Albrecht Klink, Paolo Sassone-Corsi, and Martin Reincke
Background: The ACTH receptor (ACTH-R) is a member of the seven transmembrane domain receptor super-family. In non-functional adrenal adenomas and adrenocortical carcinomas, ACTH-R expression is low. However, no inhibitory factor for ACTH-R expression has been defined to date. DAX-1 (dosage-sensitive sex reversal, adrenal hypoplasia congenita, critical region on the X chromosome, gene-1) is a general repressor of steroid production, inhibiting steroidogenic factor-1 (SF-1)-dependent expression of multiple steroidogenic enzymes. The aim of this study was to investigate whether ACTH-R gene transcription is affected by DAX-1 and whether this mechanism is involved in down-regulation of ACTH-R expression in adrenocortical tumors.
Methods: We screened 22 adrenocortical tumors for ACTH-R and DAX-1 mRNA expression by Northern blot. For in vitro analyses we co-transfected mouse Y1 adrenocortical carcinoma cells with the luciferase reporter gene vector pGL3 containing full-length constructs of human (h) or mouse (m) ACTH-R promoter together with a DAX-1 expression plasmid. These experiments were also performed using ACTH-R promoter 5′-deletion constructs and constructs mutated at the SF-1-binding sites.
Results: We found a negative correlation between DAX-1 and ACTH-R mRNA expression (R = −0.47, P < 0.02). Accordingly, in vitro expression of DAX-1 significantly reduced hACTH-R and mACTH-R promoter activity by 89 and 55% respectively. DAX-1 inhibition was also present in the shortest construct of a series of 5′-deletion constructs of the human promoter extending from −64 to +40 bp relative to the transcription start site. Mutation of the SF-1-binding sites within the hACTH-R promoter resulted in reduced or abolished DAX-1 inhibition, arguing for a mechanism that involves SF-1 for DAX-1 inhibition.
Conclusions: These data support the concept that DAX-1 is a major repressor of ACTH-R gene expression in vitro and in vivo.