BACKGROUND: Primary bilateral macronodular adrenal hyperplasia (PBMAH) is a rare cause of Cushing\'s syndrome. Individuals with PBMAH and glucose-dependent insulinotropic polypeptide (GIP)-dependent Cushing\'s syndrome due to ectopic expression of the GIP receptor (GIPR) typically harbor inactivating KDM1A sequence variants. Primary unilateral macronodular adrenal hyperplasia (PUMAH) with concomitant glucocorticoid and androgen excess has never been encountered or studied.
METHODS: We investigated a woman with a large, heterogeneous adrenal mass and severe adrenocorticotropic hormone-independent glucocorticoid and androgen excess, a biochemical presentation typically suggestive of adrenocortical carcinoma. The patient presented during pregnancy (22nd week of gestation) and reported an 18-month history of oligomenorrhea, hirsutism, and weight gain. We undertook an exploratory study with detailed histopathological and genetic analysis of the resected adrenal mass and leukocyte DNA collected from the patient and her parents.
RESULTS: Histopathology revealed benign macronodular adrenal hyperplasia. Imaging showed a persistently normal contralateral adrenal gland. Whole-exome sequencing of 4 representative nodules detected KDM1A germline variants, benign NM_001009999.3:c.136G > A:p.G46S, and likely pathogenic NM_001009999.3:exon6:c.865_866del:p.R289Dfs*7. Copy number variation analysis demonstrated an additional somatic loss of the KDM1A wild-type allele on chromosome 1p36.12 in all nodules. RNA sequencing of a representative nodule showed low/absent KDM1A expression and increased GIPR expression compared with 52 unilateral sporadic adenomas and 4 normal adrenal glands. Luteinizing hormone/chorionic gonadotropin receptor expression was normal. Sanger sequencing confirmed heterozygous KDM1A variants in both parents (father: p.R289Dfs*7 and mother: p.G46S) who showed no clinical features suggestive of glucocorticoid or androgen excess.
CONCLUSIONS: We investigated the first PUMAH associated with severe Cushing\'s syndrome and concomitant androgen excess, suggesting pathogenic mechanisms involving KDM1A.

UNASSIGNED: Cardiovascular disease is the leading cause of death in Cushingžs syndrome (CS). Primary bilateral macro-nodular adrenal hyperplasia (PBMAH), is a rare cause of CS that is clinically distinct from the other common types of CS, but cardiac characteristics have been poorly studied.
UNASSIGNED: The clinical data, steroid hormones and echocardiographic variables of 17 patients with PBMAH were collected. Twenty-one CS patients with cortisol-producing adenoma (CPA) were collected as controls. The similarities and differences of clinical and cardiac features between the two groups were compared.
UNASSIGNED: Kardiovaskularne bolesti su vodeći uzrok smrti kod Kušingovog sindroma (CS). Primarna bilateralna makro-nodularna hiperplazija nadbubrežne žlezde (PBMA ) je redak uzrok CS koji se klinički razlikuje od drugih uobičajenih tipova CS, ali kardijalne karakteristike su slabo proučavane.
UNASSIGNED: Prikupljeni su klinički podaci, steroidni hormoni i ehokardiografske varijable 17 pacijenata sa PBMA. Dvadeset i jedan pacijent sa CS sa adenomom koji proizvodi kortizol (CPA) je sakupljen kao kontrola. Upoređene su sličnosti i razlike kliničkih i kardijalnih karakteristika između dve grupe.
UNASSIGNED: U grupi PBMA udeo žena je bio manji (35,30 prema 100,00%), starosna dob je bila starija (55,76±2,42 godine prema 39,57±2,72 godine), a tok bolesti je bio duži 13,00 (5,00, 50) godine u odnosu na 1,58 (1,00, 5,00) godina. Procenat pacijenata sa PBMA sa hipertenzijom bio je veći nego kod pacijenata sa CPA (100,00% naspram 61,90%). Kortizol u plazmi i 24-časovni kortizol bez mokraće (24-časovni UFC) bili su niži kod pacijenata sa PBMA nego kod pacijenata sa CPA jutarnji kortizol 19,50 (15,35, 24,48) mg/dL naspram 28,30 (22,88, 29,89) mg/dL, 04h/dL, 04h/dL. 156,58) naspram 506,23 (292,53, 712,18) mg. Grupa PBMA imala je duži prečnik desne komore (24,06±1,23 mm naspram 20,48±0,83 mm), prečnik leve pretkomore (39,41±1,15 mm naspram 32,86±0,76 mm) i veću stopu dijastolne funkcije od 53% (s 76 ± 0,8%). CPA grupa. Univarijantna regresiona analiza je pokazala da kombinacija hipertenzije i trajanja hipertenzije ima značajne efekte na prečnik leve pretkomore (b=6,383, P=0,001; b = 0,276, P<0,001, respektivno) i E/A odnos (b=-0,496, P < 0,001, b=0,022, P<0,001, respektivno). Multivarijantna regresiona analiza je pokazala da nivo kortikosterona ima značajan uticaj na sistolni krvni pritisak (b=6,712, P=0,025).
UNASSIGNED: U poređenju sa CPA, ventrikularna dijastolna disfunkcija je česta kod pacijenata sa PBMA pod relativno nižim nivoom kortizola. PBMA je imao visok nivo kortikosterona koji može igrati ulogu u razvoju hipertenzije i daljih srčanih promena.

To perform genetic screening for ARMC5 gene germline pathogenic variants in patients with primary bilateral macronodular adrenal hyperplasia (PBMAH).
In a group of 10 PBMAH patients, we performed complete sequencing of the coding region of the ARMC5 gene and MLPA analysis for large deletion detection. In subjects with the ARMC5 variant, we searched ARMC5 gene somatic variants on tumor samples.
Among 10 PBMAH patients, we identified four ARMC5 germline variants (40%). One variant, c:174dupC p.Glu59Argfs*44, was already known; one variant p.Gly323Asp, was already reported and classified as likely disease-causing VUS (class 3-4); two variants p.Leu596Arg and p.Arg811Pro, were never reported before. For p.Gly323Asp and p.Arg811Pro, we identified second deleterious variants at the somatic level, enforcing the possible pathogenic effect of germline variants.
Our results underscore the importance of performing genetic testing also in sporadic PBMAH patients and broaden the spectrum of molecular variants involved in PBMAH syndrome.

Primary bilateral macronodular adrenal hyperplasia (PBMAH), a rare cause of Cushing syndrome, is often diagnosed as a bilateral adrenal incidentaloma with subclinical cortisol production. Circulating microRNAs (miRNAs) are a characteristic of adrenocortical adenomas, but miRNA expression in PBMAH has not been investigated. We aimed to evaluate the circulating miRNA expression in patients with PBMAH and compare them with those in patients with non-functioning adrenocortical adenoma (NFA) and cortisol-producing adrenocortical adenoma (CPA).
miRNA profiling of plasma samples from four, five, and five patients with NFA, CPA, and PBMAH, respectively, was performed. Selected miRNA expressions were validated using quantitative RT-PCR.
PBMAH samples showed distinct miRNA expression signatures on hierarchical clustering while NFA and CPA samples were separately clustered. PBMAH was distinguished from the adenoma group of NFA and CPA by 135 differentially expressed miRNAs. Hsa-miR-1180-3p, hsa-miR-4732-5p, and hsa-let-7b-5p were differentially expressed between PBMAH and adenoma (P = 0.019, 0.006, and 0.003, respectively). Furthermore, PBMAH could be classified into two subtypes based on miRNA profiling: subtype 1 with a similar profile to those of adenoma and subtype 2 with a distinct profile. Hsa-miR-631, hsa-miR-513b-5p, hsa-miR-6805-5p, and hsa-miR-548av-5p/548k were differentially expressed between PBMAH subtype 2 and adenoma (P = 0.027, 0.027, 0.027, and 1.53E-04, respectively), but not between PBMAH, as a whole, and adenoma.
Circulating miRNA signature was identified specific for PBMAH. The existence of subtype-based miRNA profiles may be associated with the pathophysiological heterogeneity of PBMAH.

Primary bilateral macronodular adrenocortical hyperplasia (PBMAH) is a rare form of adrenal Cushing\'s syndrome. The slowly progressing expansion of bilateral adrenal tissues usually persists for dozens of years, leading to delayed onset with severe conditions due to chronic mild hypercortisolism. About 20-50% cases were found to be caused by inactivating mutation of armadillo repeat-containing protein 5 (ARMC5) gene.
A 51-year-old man was admitted for severe diabetes mellitus, resistant hypertension, centripedal obesity and edema. PBMAH was diagnosed after determination of adrenocorticotropic hormone and cortisol levels, dexamethasone suppression tests and abdominal contrast-enhanced CT scanning. The metabolic disorders of the patient remarkably improved after sequentially bilateral laparoscopic adrenalectomy combined with hormone replacement. Sanger sequencing showed germline nonsense mutation of ARMC5 c.967C>T (p.Gln323Ter). The second somatic missense mutation of ARMC5 was detected in one out of two resected nodules, reflecting the second-hit model of tumorigenesis. Routine genetic testing in his apparently healthy offspring showed one of two daughters and one son harbored the germline mutation.
In conclusion, our case report highlight the importance of genetic testing in the molecular diagnosis of PBMAH. Genetic screening in related family members will find out asymptomatic variant carriers to guide life-long follow-up.

Primary bilateral macronodular adrenal hyperplasia (PBMAH), characterized by bilateral benign adrenal macronodules (>1 cm) potentially responsible for variable levels of cortisol excess, is a rare and heterogeneous disease. However, its frequency increases due to incidentally diagnosed cases on abdominal imaging carried out for reasons other than suspected adrenal disease. Mostly isolated, it can also be associated with dominantly inherited genetic conditions in rare cases. Considering the bilateral nature of adrenal involvement and the description of familial cases, the search of a genetic predisposition has led to the identification of germline heterozygous inactivating mutations of the putative tumor suppressor gene ARMC5, causing around 25% of the apparent sporadic cases. Rigorous biochemical and imaging assessment are key elements in the management of this challenging disease in terms of diagnosis. Treatment is reserved for symptomatic patients with overt or subclinical Cushing syndrome, and was historically based on bilateral adrenalectomy, which nowadays tends to be replaced by unilateral adrenalectomy or lifelong treatment with cortisol synthesis inhibitors.

BACKGROUND: Radiological characterization of adrenal size in primary bilateral macronodular adrenocortical hyperplasia (PBMAH) has not been previously investigated.
OBJECTIVE: We hypothesized that volumetric modeling of adrenal gland size may correlate with biochemical disease severity in patients with PBMAH. Secondary analysis of patients with concurrent primary aldosteronism (PA) was performed.
METHODS: A retrospective cross-sectional analysis of 44 patients with PBMAH was conducted from 2000 to 2019.
METHODS: Tertiary care clinical research center.
METHODS: Patients were diagnosed with PBMAH based upon clinical, genetic, radiographic and biochemical characteristics.
METHODS: Clinical, biochemical, and genetic data were obtained. Computed tomography scans were used to create volumetric models by manually contouring both adrenal glands in each slice using Vitrea Core Fx v6.3 software (Vital Images, Minnetonka, Minnesota).
METHODS: 17-hydroxycorticosteroids (17-OHS), ARMC5 genetics, and aldosterone-to-renin ratio (ARR) were retrospectively obtained. Pearson test was used for correlation analysis of biochemical data with adrenal volume.
RESULTS: A cohort of 44 patients with PBMAH was evaluated, with a mean age (±SD) of 53 ± 11.53. Eight patients met the diagnostic criteria for PA, of whom 6 (75%) were Black. In the Black cohort, total adrenal volumes positively correlated with midnight cortisol (R = 0.76, P = 0.028), urinary free cortisol (R = 0.70, P = 0.035), and 17-OHS (R = 0.87, P = 0.0045), with a more pronounced correlation with left adrenal volume alone. 17-OHS concentration positively correlated with total, left, and right adrenal volume in patients harboring pathogenic variants in ARMC5 (R = 0.72, P = 0.018; R = 0.65, P = 0.042; and R = 0.73, P = 0.016, respectively).
CONCLUSIONS: Volumetric modeling of adrenal gland size may associate with biochemical severity in patients with PBMAH, with particular utility in Black patients.

Biochemical characterization of primary bilateral macronodular adrenocortical hyperplasia (PBMAH) by distinct plasma steroid profiles and its putative correlation to disease has not been previously studied. LC-MS/MS-based steroid profiling of 16 plasma steroids was applied to 36 subjects (22 females, 14 males) with PBMAH, 19 subjects (16 females, 3 males) with other forms of adrenal Cushing\'s syndrome (ACS), and an age and sex-matched control group. Germline ARMC5 sequencing was performed in all PBMAH cases. Compared to controls, PBMAH showed increased plasma 11-deoxycortisol, corticosterone, 11-deoxycorticosterone, 18-hydroxycortisol, and aldosterone, but lower progesterone, DHEA, and DHEA-S with distinct differences in subjects with and without pathogenic variants in ARMC5. Steroids that showed isolated differences included cortisol and 18-oxocortisol with higher (P < 0.05) concentrations in ACS than in controls and aldosterone with higher concentrations in PBMAH when compared to controls. Larger differences in PBMAH than with ACS were most clear for corticosterone, but there were also trends in this direction for 18-hydroxycortisol and aldosterone. Logistic regression analysis indicated four steroids - DHEA, 11-deoxycortisol, 18-oxocortisol, and corticosterone - with the most power for distinguishing the groups. Discriminant analyses with step-wise variable selection indicated correct classification of 95.2% of all subjects of the four groups using a panel of nine steroids; correct classification of subjects with and without germline variants in ARMC5 was achieved in 91.7% of subjects with PBMAH. Subjects with PBMAH show distinctive plasma steroid profiles that may offer a supplementary single-test alternative for screening purposes.

Primary bilateral macronodular adrenocortical hyperplasia (PBMAH) is a rare cause of ACTH-independent Cushing syndrome (CS), which has been associated with ectopic G-protein coupled receptors (GPCRs) in the adrenal cortex. We recently studied a 51-year-old male with PBMAH who presented with severe CS and hyperestronemia, manifesting clinically with a Cushingoid appearance, gynecomastia, and telangiectasias. Analysis of adrenal tissues following bilateral adrenalectomy showed high expression of P450 aromatase (CYP19A1). The patient carried a germline non-sense pathogenic variant in ARMC5 (p.R173*), with two independent somatic pathogenic variants identified in the right (p.S571*) and left (p.Q235*) adrenal tissues, respectively. The expression of ARMC5 was drastically decreased in the hyperplastic regions when compared to either the adjacent non-hyperplastic regions and samples from PBMAH without pathogenic variants in ARMC5. We found expression of CYP19A1 in other cases of PBMAH, although there were no differences in aromatase expression between ARMC5-mutant and ARMC5-non-mutant cases. We conclude that in select cases, PBMAH can be associated with aromatase expression resulting in elevated estrogens, irrespective of sex. Additionally, CYP19A1 expression does not appear to depend on the ARMC5 variant status.

BACKGROUND: Primary bilateral macronodular adrenal hyperplasia is a rare cause of Cushing\'s syndrome characterized by the presence of bilateral secretory adrenal nodules. Recent studies have shown that primary bilateral macronodular adrenal hyperplasia is caused by combined germline and somatic mutations of the ARMC5 gene. Exophthalmos is an underappreciated sign of Cushing\'s syndrome.
METHODS: A 52-year-old Chinese woman with progressively worsening bilateral proptosis presented to our hospital. Subsequently she was diagnosed as having primary bilateral macronodular adrenal hyperplasia and underwent bilateral laparoscopic adrenalectomy. Genomic deoxyribonucleic acid was isolated from lymphocytes as well as seven different adrenal nodules and the ARMC5 sequence was determined by Sanger sequencing. We identified one heterozygous ARMC5 germline mutation c.682C>T (p. Gln228*) and five heterozygous somatic mutations (c.310delG, c.347_357del11, c.267delC, c.283_289del7, and c.205-322del118) in five different adrenal nodules. All mutations are novel and were not found in any of the available online databases. To test whether the ARMC5 mutation induced messenger ribonucleic acid decay, real-time reverse transcriptase polymerase chain reaction was performed on patient and control adrenal tissue. We found that the adrenal cortex of our patient showed a low ARMC5 messenger ribonucleic acid expression compared with normal adrenal cortex, possibly as a result of nonsense-mediated messenger ribonucleic acid decay CONCLUSIONS: We demonstrated extensive genetic diversity of ARMC5 in a patient with primary bilateral macronodular adrenal hyperplasia that started with exophthalmos, which contributes to further understanding of the pathogenesis of this disease. Early recognition of atypical symptoms and screening for ARMC5 mutation in patients with primary bilateral macronodular adrenal hyperplasia has important clinical implications for the diagnosis and genetic counseling.