Primary aldosteronism (PA) is a common cause of secondary hypertension. Recent technological advances in genetic analysis have provided a better understanding of the molecular pathogenesis of this disease. The application of next-generation sequencing has resulted in the identification of somatic mutations in aldosterone-producing adenoma (APA), a major subtype of PA. Based on the recent findings using a sequencing method that selectively targets the tumor region where aldosterone synthase (CYP11B2) is expressed, the vast majority of APAs appear to harbor a somatic mutation in one of the aldosterone-driver genes, including KCNJ5, ATP1A1, ATP2B3, CACNA1D, CACNA1H, and CLCN2. Mutations in these genes alter intracellular ion homeostasis and enhance aldosterone production. In a small subset of APAs, somatic activating mutations in the CTNNB1 gene, which encodes β-catenin, have also been detected. Accumulating evidence suggests that race and sex impact the somatic mutation spectrum of APA. Specifically, somatic mutations in the KCNJ5 gene, encoding an inwardly rectifying K+ channel, are common in APAs from Asian populations as well as women regardless of race. Associations between APA histology, genotype, and patient clinical characteristics have also been proposed, suggesting a potential need to consider race and sex for the management of PA patients. Herein, we review recent findings regarding somatic mutations in APA and discuss potential roles of race and sex on the pathophysiology of APA as well as possible clinical implications.
You are looking at 1 - 6 of 6 items for
- Author: William E Rainey x
- Refine by Access: Content accessible to me x
Kazutaka Nanba and William E Rainey
Tao Wang, Fumitoshi Satoh, Ryo Morimoto, Yasuhiro Nakamura, Hironobu Sasano, Richard J Auchus, Michael A Edwards, and William E Rainey
Primary aldosteronism (PA) is the most common form of endocrine hypertension affecting ∼8–10% of hypertensive subjects. Aldosterone production in PA occurs under low-renin conditions, and the mechanisms that maintain the production of aldosterone in PA remain unknown.
This study was designed to compare the transcript profiles between aldosterone-producing adenoma (APA) and their adjacent adrenal gland (AAG) from the same adrenal.
Total RNA was extracted from ten APA and ten AAG; and subsequently analyzed by microarray and real-time quantitative RT-PCR (qPCR). The microarray data were paired for each APA–AAG, and analyzed by GeneSpring GX 11 with paired t-test and fold change calculations for each transcript. Changes identified by microarray analysis were confirmed by qPCR.
Microarray analysis indicated that 14 genes had significantly up-regulated expression in APA compared to AAG. Among the elevated genes were aldosterone synthase (CYP11B2) as well as novel transcription factors, calmodulin-binding proteins, and other genes that have not been previously studied in APA. Selective analysis of 11 steroidogenic enzymes using microarray demonstrated that only CYP11B2 showed a significantly higher transcript level in APA compared to AAG (P<0.001). In contrast, AKR1C3 (17β-hydroxysteroid dehydrogenase type 5), CYP17 (17α-hydroxylase/17,20 lyase), and CYB5 (cytochrome b5) showed significantly lower transcript level in APA (P<0.05).
The transcriptome analysis of APA compared with AAG showed several novel genes that are associated with APA phenotype. This gene list provides new candidates for the elucidation of the molecular mechanisms leading to PA.
Adina F Turcu, Lili Zhao, Xuan Chen, Rebecca Yang, Juilee Rege, William E Rainey, Johannes D Veldhuis, and Richard J Auchus
Many hormones display distinct circadian rhythms, driven by central regulators, hormonal bioavailability, and half-life. A set of 11-oxygenated C19 steroids (11-oxyandrogens) and pregnenolone sulfate (PregS) are elevated in congenital adrenal hyperplasia and other disorders, but their circadian patterns have not been characterized.
Participants and methods
Peripheral blood was collected every 2 h over 24 h from healthy volunteer men (10 young, 18–30 years, and 10 older, 60–80 years). We used mass spectrometry to quantify 15 steroids, including androstenedione (A4), testosterone (T), 11β-hydroxy- and 11-ketotestosterone (11OHT, 11KT),11β-hydroxy- and 11-ketoandrostenedione (11OHA4, 11KA4), and 4 ∆5-steroid sulfates. Diurnal models including mesor (rhythm adjusted median), peak, and nadir concentrations, acrophase, and amplitude were computed.
11OHA4 followed a rhythm similar to cortisol: acrophase 8:00 h, nadir 21:00 h and were similar in young and old men. 11KT had similar diurnal patterns, but the peak was lower in older than in young men, as was the case for A4. All four steroid sulfates were higher in young vs older men. PregS and 17-hydroxypregnenolone sulfate (17OHPregS) showed sustained elevations between 8:00 and 18:00 h, and nadirs around midnight, while DHEAS and AdiolS displayed minimal diurnal variations. All 4 11-oxyandrogens correlated tightly with cortisol (r from 0.54 for 11OHT to 0.81 for 11OHA4, P < 0.0001 for all), but very weakly with T, supporting their adrenal origin and ACTH governance.
11-Oxyandrogens, PregS, and 17OHPregS display distinct circadian and age variations, which should be accounted for when used as clinical biomarkers.
Kazutaka Nanba, Kei Omata, Scott A Tomlins, Thomas J Giordano, Gary D Hammer, William E Rainey, and Tobias Else
Co-secretion of cortisol and aldosterone can be observed in adrenal adenomas. The aim of this study was to investigate the molecular characteristics of a co-existing aldosterone- and a cortisol-producing adenoma (CPA) in the same patient.
Design and methods
Two different adenomas within the same adrenal gland from a 49-year-old female patient with primary aldosteronism (PA) and Cushing's syndrome (CS) were studied. Multiple formalin-fixed paraffin-embedded tumor blocks were used for the analysis. Immunohistochemistry (IHC) was performed using a specific antibody against aldosterone synthase (CYP11B2). DNA and RNA were isolated separately from CYP11B2-positive and -negative tumor regions based on CYP11B2 IHC results.
CYP11B2 IHC clearly demonstrated that three pieces from one adenoma were positive for CYP11B2 and the remaining three from the other adenoma were negative for CYP11B2. In quantitative real-time RT-PCR, CYP11B2 mRNA was upregulated in CYP11B2-positive tumor specimens (219-fold vs CYP11B2-negative tumor specimens). Targeted next-generation sequencing (NGS) detected novel KCNJ5 gene mutations (p.T148I/T149S, present in the same reads) and a PRKACA gene hotspot mutation (p.L206R) in the CYP11B2-positive and -negative tumors, respectively. Sanger sequencing of DNA from each tumor specimen (CYP11B2-positive tumor, n=3; CYP11B2-negative tumor, n=3) showed concordant results with targeted NGS.
Our findings illustrate the co-existence of two different adrenocortical adenomas causing the concurrent diagnosis of PA and CS in the same patient. Molecular analysis was able to demonstrate that the two diseases resulted from independent somatic mutations seen in double adrenocortical adenomas.
Adina F Turcu, Aya T Nanba, Robert Chomic, Sunil K Upadhyay, Thomas J Giordano, James J Shields, Deborah P Merke, William E Rainey, and Richard J Auchus
To comprehensively characterize androgens and androgen precursors in classic 21-hydroxylase deficiency (21OHD) and to gain insights into the mechanisms of their formation.
Serum samples were obtained from 38 patients (19 men) with classic 21OHD, aged 3–59, and 38 sex- and age-matched controls; 3 patients with 11β-hydroxylase deficiency; 4 patients with adrenal insufficiency; and 16 patients (8 men) undergoing adrenal vein sampling. Paraffin-embedded normal (n = 5) and 21OHD adrenal tissues (n = 3) were used for immunohistochemical studies.
We measured 11 steroids in all sera by liquid chromatography-tandem mass spectrometry. Immunofluroescence localized 3β-hydroxysteroid dehydrogenase type 2 (HSD3B2) and cytochrome b5 (CYB5A) within the normal and 21OHD adrenals.
Four 11-oxygenated 19-carbon (11oxC19) steroids were significantly higher in male and female 21OHD patients than in controls: 11β-hydroxyandrostenedione, 11-ketoandrostenedione 11β-hydroxytestosterone, and 11-ketotestosterone (3–4-fold, P < 0.0001). For 21OHD patients, testosterone and 11-ketotestosterone were positively correlated in females, but inversely correlated in males. All 11oxC19 steroids were higher in the adrenal vein than in the inferior vena cava samples from men and women and rose with cosyntropin stimulation. Only trace amounts of 11oxC19 steroids were found in the sera of patients with 11β-hydroxylase deficiency and adrenal insufficiency, confirming their adrenal origin. HSD3B2 and CYB5A immunoreactivities were sharply segregated in the normal adrenal glands, whereas areas of overlapping expression were identified in the 21OHD adrenals.
All four 11oxC19 steroids are elevated in both men and women with classic 21OHD. Our data suggest that 11oxC19 steroids are specific biomarkers of adrenal-derived androgen excess.
Takuya Kitamura, Amy R Blinder, Kazutaka Nanba, Mika Tsuiki, Mutsuki Mishina, Hiroshi Okuno, Koki Moriyoshi, Yuto Yamazaki, Hironobu Sasano, Keisuke Yoneyama, Aaron M Udager, William E Rainey, Akihiro Yasoda, Noriko Satoh-Asahara, and Tetsuya Tagami
Although excess production of androgens and glucocorticoids has often been observed in adrenocortical carcinomas, adrenocortical adenoma with such hormonal activity is rare. Herein, we report a 41-year-old woman who presented with hyperandrogenemia and mild autonomous cortisol secretion with an undetectable level of adrenocorticotropic hormone. Imaging demonstrated a 6 cm left adrenal tumor. The histologic diagnosis of the resected adrenal tumor was adrenocortical adenoma. Pre- and post-operative serum samples were used for steroid profiling with liquid chromatography-tandem mass spectrometry (LC-MS/MS). LC-MS/MS analysis of pre-operative serum revealed an abnormal buildup of steroid precursors and androgens. Importantly, circulating levels of 11-oxygenated androgens, including 11β-hydroxytestosterone (11OHT) and 11-ketotestosterone (11KT), were highly elevated. Both androgen and glucocorticoid levels significantly decreased post-operatively. Immunohistochemical analysis of steroidogenic enzymes and cofactor protein supported the tumor’s ability to directly produce 11OHT and 11KT. This study is the first to describe and characterize an adrenocortical adenoma that co-secretes glucocorticoids and 11-oxygenated androgens.
Due to its rarity, biochemical and histologic characteristics of androgen and glucocorticoid co-secreting adrenocortical adenomas are largely unknown. Herein, we report a case of adrenocortical adenoma that caused marked hyperandrogenemia and mild autonomous cortisol secretion. In this study, we investigated serum steroid profiles using liquid chromatography-tandem mass spectrometry (LC-MS/MS) and histologic characteristics of the resected tumor. LC-MS/MS revealed highly elevated levels of 11-oxygenated androgens which have not been well studied in adrenal tumors. The expression patterns of steroidogenic enzymes determined by immunohistochemistry supported the results of steroid profiling and suggested the capacity of the tumor cells to produce 11-oxygenated androgens. Measurement of 11-oxygenated steroids should facilitate a better understanding of androgen-producing adrenocortical neoplasms.