Progress on Genetic Basis of Primary Aldosteronism

Primary aldosteronism (PA) is a heterogeneous group of disorders caused by the autonomous overproduction of aldosterone with simultaneous suppression of plasma renin activity (PRA). It is considered to be the most common endocrine cause of secondary arterial hypertension (HT) and is associated with a high rate of cardiovascular complications. PA is most often caused by a bilateral adrenal hyperplasia (BAH) or aldosterone-producing adenoma (APA); rarer causes of PA include genetic disorders of steroidogenesis (familial hyperaldosteronism (FA) type I, II, III and IV), aldosterone-producing adrenocortical carcinoma, and ectopic aldosterone-producing tumors. Over the last few years, significant progress has been made towards understanding the genetic basis of PA, classifying it as a channelopathy. Recently, a growing body of clinical evidence suggests that mutations in ion channels appear to be the major cause of aldosterone-producing adenomas, and several mutations within the ion channel encoding genes have been identified. Somatic mutations in four genes (KCNJ5, ATP1A1, ATP2B3 and CACNA1D) have been identified in nearly 60% of the sporadic APAs, while germline mutations in KCNJ5 and CACNA1H have been reported in different subtypes of familial hyperaldosteronism. These new insights into the molecular mechanisms underlying PA may be associated with potential implications for diagnosis and therapy.

Adrenal venous sampling in the diagnostic workup of primary aldosteronism

Primary aldosteronism (PA) is the primary cause of secondary hypertension. The prevalence of PA has probably been underestimated in the past and recent studies suggest that PA could be present in up to 10% of patients suffering from hypertension. Aldosterone excess in PA can be caused by unilateral adrenal disease, usually adrenal adenoma, or bilateral adrenal hyperplasia. Differentiation between unilateral and bilateral disease is clinically important as the former can effectively be treated by removal of the affected adrenal. CT or MRI cannot reliably distinguish unilateral from bilateral disease. Therefore, adrenal vein sampling (AVS) is an important step of the diagnostic work-up in patients with PA. Current guidelines recommend PA in virtually all patients with biochemically diagnosed PA who would undergo adrenal surgery if unilateral PA was diagnosed. In this narrative review, we give an overview of the current technique used for AVS with a focus on the experience with this technique at the University Hospital Basel, Switzerland.

Bilateral Adrenal Hyperplasia: Pathogenesis and Treatment

Bilateral adrenal hyperplasia is a rare cause of Cushing’s syndrome. Micronodular adrenal hyperplasia, including the primary pigmented micronodular adrenal dysplasia (PPNAD) and the isolated micronodular adrenal hyperplasia (iMAD), can be distinguished from the primary bilateral macronodular adrenal hyperplasia (PBMAH) according to the size of the nodules. They both lead to overt or subclinical CS. In the latter case, PPNAD is usually diagnosed after a systematic screening in patients presenting with Carney complex, while for PBMAH, the diagnosis is often incidental on imaging. Identification of causal genes and genetic counseling also help in the diagnoses. This review discusses the last decades’ findings on genetic and molecular causes of bilateral adrenal hyperplasia, including the several mechanisms altering the PKA pathway, the recent discovery of ARMC5, and the role of the adrenal paracrine regulation. Finally, the treatment of bilateral adrenal hyperplasia will be discussed, focusing on current data on unilateral adrenalectomy.

Novel allosteric ligands of the angiotensin receptor AT1R as autoantibody blockers

While orthosteric ligands of the angiotensin II (AngII) type 1 receptor (AT1R) are available for clinical and research applications, allosteric ligands are not known for this important G protein-coupled receptor (GPCR). Allosteric ligands are useful tools to modulate receptor pharmacology and subtype selectivity. Here, we report AT1R allosteric ligands for a potential application to block autoimmune antibodies. The epitope of autoantibodies for AT1R is outside the orthosteric pocket in the extracellular loop 2. A molecular dynamics simulation study of AT1R structure reveals the presence of a druggable allosteric pocket encompassing the autoantibody epitope. Small molecule binders were then identified for this pocket using structure-based high-throughput virtual screening. The top 18 hits obtained inhibited the binding of antibody to AT1R and modulated agonist-induced calcium response of AT1R. Two compounds out of 18 studied in detail exerted a negative allosteric modulator effect on the functions of the natural agonist AngII. They blocked antibody-enhanced calcium response and reactive oxygen species production in vascular smooth muscle cells as well as AngII-induced constriction of blood vessels, demonstrating their efficacy in vivo. Our study thus demonstrates the feasibility of discovering inhibitors of the disease-causing autoantibodies for GPCRs. Specifically, for AT1R, we anticipate development of more potent allosteric drug candidates for intervention in autoimmune maladies such as preeclampsia, bilateral adrenal hyperplasia, and the rejection of organ transplants.

An Unusual Case of Bilateral Adrenal Hyperplasia

Background: Thrombocytopenia, anasarca, fever, reticulin fibrosis/renal failure, and organomegaly (TAFRO) syndrome is a variant of Castleman Disease, which is a rare lymphoproliferative disease that can be life threatening. Diagnosis is often delayed because of its nonspecific presentation. Bilateral adrenal hyperplasia has been a reported complication, however the majority of cases reported have been in Asian patients. Prior accounts of elevated ACTH in TAFRO have been in the context of adrenal insufficiency. Clinical Case: A 28-year-old Caucasian male with a history of multiple sclerosis was seen in the ED with abdominal pain. On presentation, he was afebrile and normotensive. Physical exam was notable for cervical lymphadenopathy and abdominal tenderness. There was no facial rounding/plethora, bruising, abnormal striae, or proximal muscle weakness. He had normal blood counts, serum chemistry and liver function. An abdominal CT scan showed marked bilateral adrenal hyperplasia with pre-aortic, peri-aortic and retroperitoneal lymphadenopathy. An 8AM serum cortisol was 14.1 mcg/dl (4.8–19.6 mcg/dl) and adrenocorticotrophic hormone (ACTH) was elevated at 152 pg/ml (7.2–63 pg/ml). A repeat serum 8AM cortisol following low dose dexamethasone suppression test (LDDST) was 14.7 mcg/dl, however at that point the patient had developed new fevers and thrombocytopenia. Blood pressure, blood glucose and potassium remained normal. An MRI of the brain showed a normal appearing pituitary gland. An extensive infectious and rheumatologic evaluation was negative, and he underwent an inguinal lymph node biopsy which showed nodal expansion with histiocytes, consistent with TAFRO. High dose methylprednisolone and Siltuximab (an IL-6 inhibitor) were started, and his fever and abdominal pain resolved. He was discharged home on oral prednisone. Conclusion: We describe a case of bilateral adrenal hyperplasia with elevated ACTH and non-suppressed cortisol on LDDST suggestive of ACTH-driven cortisol excess. However, interpretation of his LDDST is made difficult in the context of persistent fevers. Although we cannot definitively exclude pathologic hypercortisolism at this time, given his lack of suggestive features such as proximal muscle weakness, abnormal striae or hypokalemic alkalosis, his over-all presentation was more consistent with hyperplasia secondary to TAFRO rather than an underlying pathologic hypercortisolism. Adrenal hyperplasia has been noted in TAFRO, however its pathogenesis remains poorly understood. TAFRO should be added among the differentials for bilateral adrenal hyperplasia to facilitate early diagnosis and treatment. References: Ducoux G, et al. Thrombocytopenia, Anasarca, Fever, Reticulin Fibrosis/Renal Failure, and Organomegaly (TAFRO) Syndrome with Bilateral Adrenal Hemorrhage in Two Caucasian Patients. Am J Case Rep. 2020;21:e919536.

Surgically Treated Resistant Hypertension Due to Bilateral Adrenal Hyperplasia

Background: Treatment resistant hypertension (TRH) may affect about 15% of patients with hypertension. While primary hyperaldosteronism is a known etiology of TRH, clinicians must consider the possibility of elevations of other adrenal hormones. Early identification of an etiology may prevent or delay the onset of complications. Case Report: Endocrinology was consulted on a 46 years-old man for evaluation of TRH. His past medical history is significant for TRH, nonischemic cardiomyopathy, congestive heart failure (Ejection Fraction 45–50%), and chronic kidney disease. Physical examination was unremarkable except for a blood pressure of 193/124 mmHg while on furosemide, isosorbide mononitrate, hydralazine, carvedilol, spironolactone, and clonidine. Chart review revealed mild hypokalemia. Computed tomography (CT) of the abdomen without contrast showed bilateral enlarged nodular adrenal glands with an increase in size over the last three years (left adrenal gland: 6.3 cm, right adrenal gland: 5.6 cm). Initial workup showed normal free plasma metanephrine, normetanephrine, aldosterone renin ratio, 17-hydroxyprogesterone, and undetectable random adrenocorticotropic hormone (ACTH) with random cortisol of 29 mcg/dl. Subsequent evaluation revealed elevated deoxycorticosterone (3030 ng/dL), 11-deoxycorticosterone (42 ng/d) and 18-Hydroxycorticosterone (640 ng/dL). He subsequently developed Cushing’s syndrome and diabetes mellitus. The patient underwent laparoscopic left adrenalectomy and subtotal right adrenalectomy. Pathology showed macro-nodular adrenal cortical hyperplasia. He was started on hydrocortisone for postoperative adrenal insufficiency. On his most recent follow-up, his blood pressure was well controlled on bumetanide, carvedilol, metolazone, and nifedipine. (Hydralazine, isosorbide mononitrate, spironolactone, and clonidine were stopped). After surgery, Corticosterone (92.10 ng/dL), 11-Deoxycorticosterone (<5.00 ng/dL) and ACTH(9 pg/mL) normalized. Conclusion: Determining the etiology of TRH should not be stopped after ruling out the “usual suspects” since malignant hypertension with end-organ dysfunction can develop, if not appropriately treated. In our patient, TRH was due to elevated 18-Hydroxycorticosterone (precursor of aldosterone), which improved after adrenalectomy.

Resistant Hypertension Due to Familial Hyperaldosteronism Type III: First Report From Indian Sub-Continent

Background: Familial hyperaldosteronism type III (FH-III) is caused by germline mutations in KCNJ5 gene. FH-III presents with phenotypic variability from spironolactone-responsive hypertension to massive adrenal hyperplasia requiring bilateral adrenalectomy. Till date, seven different pathogenic mutations in KCNJ5 gene have been identified. Here we describe a sporadic FH-III case, due to a Gly151Arg mutation, first from Indian subcontinent, presenting with extremely high plasma aldosterone concentration (PAC) values, further expanding our knowledge of this rare condition. Clinical Case: A 24-year-old female, symptomatic since age of 5 years with periodic limb weakness and gradual increase in frequency of episodes over the years. Her blood pressure (BP) was recorded for first time at 9 years of age, and it was 170/110 mm Hg. On evaluation at this time, PAC was highly elevated at 1007 ng/dL and plasma renin activity (PRA) was suppressed at 0.04 ng/ml/h with aldosterone renin ratio (ARR) of 25,175 ng/dL per ng/mL/h (>20 suggestive of primary aldosteronism). CT scan demonstrated mild enlargement of bilateral adrenal glands. A presumptive diagnosis of Glucocorticoid-remediable aldosteronism was made. She was started on dexamethasone, spironolactone and nifedipine but was not improved. Dexamethasone was stopped after 1 year of initiation. Before presenting to our referral center in 2018, she was having uncontrolled hypertension with recurrent episodes of hypokalemic paralysis. She was on maximal doses on four antihypertensive agents, further increased to six agents (including spironolactone 100 mg BD), and potassium chloride supplementation (120 mEq/day). Despite this, she had a serum potassium of 2.6 mEq/L. Her biochemical investigations demonstrated that PAC was 2070 ng/dL, direct renin concentration (DRC) was 2.35 µIU/mL (PRA 0.2 ng/ml/h) and ARR was 880.9 ng/dL per µIU/mL. CT scan revealed massive bilateral adrenal hyperplasia. Genetic analysis by whole exome sequencing detected KCNJ5 (p.Gly151Arg) mutation, confirming the diagnosis of FH-III. She was subjected to bilateral adrenalectomy and she became normokalemic. Dramatic reduction in antihypertensives with BP control achieved only on amlodipine post-operatively. Genetic testing of family members was not done but they were normotensive and normokalemic. Histopathological examination revealed bilateral adrenal hyperplasia. PAC levels up to 297 ng/dL have been described previously in FH-III but our patient had exceedingly high-level of 2070 ng/dL. Conclusion: This case demonstrates florid clinical and biochemical manifestations of FH-III and gradual worsening of symptoms, consistent with progression of disease with age. It illustrates that proper investigations and treatment can lead to remission of symptoms. Further studies are warranted to elucidate the full clinical and biochemical spectrum of FH-III.

Pheochromocytoma as the Etiology of Cushing Syndrome Through Paracrine ACTH Signaling

Introduction: Pheochromocytomas are rare catecholamine-secreting tumors of the adrenal medulla that may also secrete ACTH. There are approximately 100 cases of ACTH-producing pheochromocytomas reported in the peer reviewed literature, and they typically cause Cushing syndrome by significantly raising circulating ACTH levels. We report a pheochromocytoma causing apparent ACTH-independent Cushing syndrome by paracrine action of locally produced ACTH on ipsilateral adrenal cortex. Case: A 64-year-old female was referred for an incidentally discovered left adrenal mass. The mass was 4 cm and 37 Hounsfield units on non-contrast computed tomography, and the right adrenal gland was unremarkable. The patient’s history was notable for three years of resistant hypertension, and examination revealed stigmata of Cushing syndrome including moon facies, scattered bruises, and wide, pink striae. Three measurements of plasma metanephrines ranged from 3- to 4.5-fold elevated, and 24 h urine metanephrines were 5.2-fold elevated. Urine 24 h epinephrine was 2.7-fold elevated. Three 8 AM cortisol measurements on overnight dexamethasone suppression tests were 16.3–17.4 mcg/dL (< 1.8), and 8 AM dexamethasone on one test was 494 ng/dL (140–295). Two midnight salivary cortisol measurements were 0.348 and 0.416 mcg/dL (< 0.112), and 8 AM ACTH levels on two occasions were 6.8 and 7.9 pg/mL (7.2–63.3). After adrenergic blockade, the patient underwent left adrenalectomy. Her tumor stained for synaptophysin and chromogranin consistent with pheochromocytoma, and focal staining for ACTH was also observed. Hyperplasia of the adjacent adrenal cortex was present. Hypertension resolved after adrenalectomy, and the patient developed secondary adrenal insufficiency (8 AM cortisol and ACTH 1.0 mcg/dL [> 15] and 2.9 pg/mL, respectively). Discussion: There are five published reports of pheochromocytoma and subclinical hypercortisolemia due to paracrine ACTH or IL-6 activity on ipsilateral adrenal cortex, but this case is distinguished by the occurrence of overt Cushing syndrome. Our patient had ACTH < 10 pg/mL on two measurements, her pheochromocytoma showed focal staining for ACTH, and hyperplasia of adjacent adrenal cortex was observed. In contrast, median ACTH level was 5-6x above the upper limit of normal in a recent review of 95 cases of pheochromocytomas with Cushing syndrome due to ectopic ACTH secretion. If Cushing syndrome was due to bilateral adrenal hyperplasia independent of the patient’s pheochromocytoma, cortisol excess would not have resolved after unilateral adrenalectomy. Communication between cortical and medullary circulations through venous radicles provides a mechanism for ACTH produced by the patient’s pheochromocytoma to affect the function of adjacent cortex. This case demonstrates a novel mechanism by which ACTH from a pheochromocytoma can cause Cushing syndrome.

Phosphodiesterase 2A and 3B variants are associated with primary aldosteronism

Familial primary aldosteronism (PA) is rare and mostly diagnosed in early-onset hypertension (HT). However, ‘sporadic’ bilateral adrenal hyperplasia (BAH) is the most frequent cause of PA and remains without genetic etiology in most cases. Our aim was to investigate new genetic defects associated with BAH and PA. We performed whole-exome sequencing (paired blood and adrenal tissue) in six patients with PA caused by BAH that underwent unilateral adrenalectomy. Additionally, we conducted functional studies in adrenal hyperplastic tissue and transfected cells to confirm the pathogenicity of the identified genetic variants. Rare germline variants in phosphodiesterase 2A (PDE2A) and 3B (PDE3B) genes were identified in three patients. The PDE2A heterozygous variant (p.Ile629Val) was identified in a patient with BAH and early-onset HT at 13 years of age. Two PDE3B heterozygous variants (p.Arg217Gln and p.Gly392Val) were identified in patients with BAH and HT diagnosed at 18 and 33 years of age, respectively. A strong PDE2A staining was found in all cases of BAH in zona glomerulosa and/or micronodules (that were also positive for CYP11B2). PKA activity in frozen tissue was significantly higher in BAH from patients harboring PDE2A and PDE3B variants. PDE2A and PDE3B variants significantly reduced protein expression in mutant transfected cells compared to WT. Interestingly, PDE2A and PDE3B variants increased SGK1 and SCNN1G/ENaCg at mRNA or protein levels. In conclusion, PDE2A and PDE3B variants were associated with PA caused by BAH. These novel genetic findings expand the spectrum of genetic etiologies of PA.