👤 Lionel Groussin

This study aimed to investigate the genetic cause of food-dependent Cushing syndrome (FDCS) observed in patients with primary bilateral macronodular adrenal hyperplasia (PBMAH) and adrenal ectopic expression of the glucose-dependent insulinotropic polypeptide receptor. Germline ARMC5 alterations have been reported in about 25% of PBMAH index cases but are absent in patients with FDCS. A multiomics analysis of PBMAH tissues from 36 patients treated by adrenalectomy was performed (RNA sequencing, single-nucleotide variant array, methylome, miRNome, exome sequencing). The integrative analysis revealed 3 molecular groups with different clinical features, namely G1, comprising 16 patients with ARMC5 inactivating variants; G2, comprising 6 patients with FDCS with glucose-dependent insulinotropic polypeptide receptor ectopic expression; and G3, comprising 14 patients with a less severe phenotype. Exome sequencing revealed germline truncating variants of KDM1A in 5 G2 patients, constantly associated with a somatic loss of the KDM1A wild-type allele on 1p, leading to a loss of KDM1A expression both at messenger RNA and protein levels (P = 1.2 × 10 KDM1A inactivation explains about 90% of FDCS PBMAH. Genetic screening for ARMC5 and KDM1A can now be offered for most PBMAH operated patients and their families, opening the way to earlier diagnosis and improved management. Show less

Molecular alterations of the MAPK pathway are frequently observed in papillary thyroid carcinomas (PTCs). It leads to a constitutive activation of the signalling pathway through an increase in MEK and ERK phosphorylation. ERK is negatively feedback-regulated by Dual Specificity Phosphatases (DUSPs), especially two ERK-specific DUSPs, DUSP5 (nuclear) and DUSP6 (cytosolic). These negative MAPK regulators may play a role in thyroid carcinogenesis. MAPK pathway activation was analyzed in 11 human thyroid cancer cell lines. Both phosphatases were studied in three PCCL3 rat thyroid cell lines that express doxycycline inducible PTC oncogenes (RET/PTC3, H-RASV12 or BRAFV600E). Expression levels of DUSP5 and DUSP6 were quantified in 39 human PTCs. The functional role of DUSP5 and DUSP6 was investigated through their silencing in two human BRAFV600E carcinoma cell lines. BRAFV600E human thyroid cancer cell lines expressed higher phospho-MEK levels but not higher phospho-ERK levels. DUSP5 and DUSP6 are specifically induced by the MEK-ERK pathway in the three PTC oncogenes inducible thyroid cell lines. This negative feedback loop explains the tight regulation of p-ERK levels. DUSP5 and DUSP6 mRNA are overexpressed in human PTCs, especially in BRAFV600E mutated PTCs. DUSP5 and/or DUSP6 siRNA inactivation did not affect proliferation in two BRAFV600E mutated cell lines, which may be explained by a compensatory increase in other phosphatases. In the light of this, we observed a marked DUSP6 upregulation upon DUSP5 inactivation. Despite this, DUSP5 and DUSP6 positively control cell migration and invasion. Our results are in favor of a stronger activation of the MAPK pathway in BRAFV600E PTCs. DUSP5 and DUSP6 have pro-tumorigenic properties in two BRAFV600E PTC cell line models. Show less

The cyclic AMP (cAMP) pathway plays a major role in the development of endocrine tissues and various molecular defects of key components of this pathway (G protein, receptors, PKA, ...) have been observed in endocrine tumors. Hypersecretion of adrenocorticotropin hormone (ACTH), the key activator of the cAMP pathway in adrenal cortex, is associated with adrenocortical hyperplasia and cortisol oversecretion (Cushing's syndrome). The best example of "illegitimate" membrane receptors expression reported is the abnormal expression of the adenylyl cyclase activating gastric inhibitory peptide receptor (GIP-R) in ACTH-independent Cushing's syndrome (ACS). We have observed that ectopic expression of the GIP-R is frequent in ACTH-Independent Macronodular Adrenal Hyperplasia (AIMAH), rare in benign adrenal adenoma (AA), but seems absent in Adrenal Cancer (AC). In vivo systematic screening of AIMAH shows at least one abnormal response of cortisol (suggesting "illegitimate" membrane receptor expression) in almost all patients. Somatic and germ line inactivating mutations of PRKAR1 (regulatory subunit R1A of PKA) can be observed in patient with isolated primary pigmented nodular adrenocortical disease (PPNAD) and AA responsible for ACS. At the nuclear level, the cAMP pathway regulates transcription mainly by PKA-dependent phosphorylation of the cyclic AMP response element binding (CREB) family of transcription factors (CREB, CREM, and ATF-1). Cyclic AMP response element binding protein (CREB) is expressed in normal adrenal cortex. Alterations of CRE binding proteins with loss of CREB expression and compensatory overexpression of CREMtau is observed in the human adrenocortical cancer cell line H295R. Similar alterations are found at the protein level in human malignant adrenocortical tumors. In conclusion, various alterations leading to activation or inactivation of key components of the cAMP signaling pathway can be observed in adrenocortical tumorigenesis. Show less

Control of cortisol secretion by the abnormal expression of the gastric inhibitory polypeptide receptor (GIP-R) have been observed in some rare cases of ACTH-independent, food-dependent Cushing's syndrome (FD-ACS) due to adrenal adenoma (AA) or bilateral macronodular hyperplasia (AIMAH). This study was performed to determine the prevalence of GIP-R ectopic expression in ACS and its correlation with fasting cortisol levels. GIP-R expression was studied by RT-PCR in 30 unilateral adrenal tumors [16 AA and 14 adrenocortical cancer (AC)] and 8 AIMAH tissues. Fasting and postprandial cortisol levels were assayed, respectively, at 0800 and 1200 h in AA, AC, and AIMAH, and 1 h after a morning standard meal in 6 AIMAH patients. Similar expression of 2 GIP-R isoforms was observed in 1 of 16 AA, 0 of 14 AC, and 4 of 8 AIMAH as well as in the 4 insulinomas used as positive controls. In vitro study of the GIP-R-expressing AA showed stimulation of cortisol secretion and cAMP production by GIP. The fasting 0800-h plasma cortisol level was above 276 nmol/liter in all patients except 1 AA case and 1 AIMAH case, both of whom expressed GIP-R. In the 3 additional AIMAH cases that expressed the GIP-R, fasting plasma cortisol levels were above 276 nmol/liter. This study demonstrates that ectopic expression of GIP-R is rare in AA and is usually associated with the low fasting plasma cortisol levels that characterize FD-ACS. In contrast, GIP-R expression is frequent in AIMAH and might not always be associated with a low fasting plasma cortisol level. This suggests that maintenance of hypercortisolemia in GIP-R- expressing AIMAH does not always depend solely on GIP-R, and that simultaneous abnormal expression of other membrane receptors might be present. The expression of GIP-R could not be observed during malignant transformation of the adrenal cortex. This study highlighted the major role of cAMP alterations secondary to GIP-R ectopic expression in the pathophysiology of AIMAH and in some rare cases of well differentiated benign adrenocortical tumors. Show less