Novel somatic mutations in primary hyperaldosteronism are related to the clinical, radiological and pathological phenotype

Abstract

Aldosterone-producing adenomas (APAs) and bilateral adrenal hyperplasia are important causes of secondary hypertension. Somatic mutations in KCNJ5, CACNA1D, ATP1A1, ATP2B3 and CTNNB1 have been described in APAs.

Objective: To characterize clinical-pathological features in APAs and unilateral adrenal hyperplasia, and correlate them with genotypes.

Design: Retrospective study.

Subjects and measurements: Clinical and pathological characteristics of 90 APAs and seven diffusely or focally hyperplastic adrenal glands were reviewed, and samples were examined for mutations in known disease genes by Sanger or exome sequencing.

Results: Mutation frequencies were as follows: KCNJ5, 37·1%; CACNA1D, 10·3%; ATP1A1, 8·2%; ATP2B3, 3·1%; and CTNNB1, 2·1%. Previously unidentified mutations included I157K, F154C and two insertions (I150_G151insM and I144_E145insAI) in KCNJ5, all close to the selectivity filter, V426G_V427Q_A428_L433del in ATP2B3 and A39Efs*3 in CTNNB1. Mutations in KCNJ5 were associated with female and other mutations with male gender (P = 0·007). On computed tomography, KCNJ5-mutant tumours displayed significantly greater diameter (P = 0·023), calculated area (P = 0·002) and lower precontrast Hounsfield units (P = 0·0002) vs tumours with mutations in other genes. Accordingly, KCNJ5-mutant tumours were predominantly comprised of lipid-rich fasciculata-like clear cells, whereas other tumours were heterogeneous (P = 5 × 10(-6) vs non-KCNJ5 mutant and P = 0·0003 vs wild-type tumours, respectively). CACNA1D mutations were present in two samples with hyperplasia without adenoma.

Conclusions: KCNJ5-mutant tumours appear to be associated with fasciculata-like clear cell predominant histology and tend to be larger with a characteristic imaging phenotype. Novel somatic KCNJ5 variants likely cause adenomas by loss of potassium selectivity, similar to previously described mutations.

Figure 1. Gross pathology and hematoxylin-eosin staining of representative cases of uninodular and multinodular aldosterone producing adrenal lesions

A) APA55. Solitary uninodular adrenocortical 1.9 cm adenoma (arrow) with unremarkable cortex and surrounding fat tissue; tumor has the KCNJ5-G151R mutation. B) APA60. Macronodular adrenocortical hyperplasia with at least 3 distinct macronodules (arrows); no mutations were identified. C) APA50. Uninodular tumor with surrounding adrenocortical hyperplasia; tumor has an ATP2B3 V426_V427del mutation. D) APA62. Diffuse adrenocortical hyperplasia. Specimen carries the CACNA1D F767V mutation. E) APA57. H&E staining of APA displays cells with large clear microvesicular cytoplasm with numerous lipid droplets; tumor is KCNJ5 G151R positive (x40), F) APA59. H&E staining of APA showing cells with compact lipid sparse eosinophilic cytoplasm arranged in trabeculae. Occasional cells in the center show intracytoplasmic brown pigment consistent with lipofuscin; tumor is CACNA1D G403R positive. Scale bars: 100 μm.

Figure 2. Previously undescribed variants and analysis of multinodular gland

A) Sanger sequences of tumor (and, if available, normal) areas from aldosterone-producing adenomas, demonstrating previously undescribed variants. Fwd, forward; Rev, reverse; WT, wild type. Reverse sequence shown reverse complement for comparison. Cf. table 2 for genomic positions of variants. B) H&E stains of three blocks of APA81 are shown above. Sampling of four tumor nodules (B/T1, C/T1, C/T2, D/T1), one area of nodular hyperplasia (B/T2) and three areas with normal adrenal tissue (B/N, C/N, D/N) demonstrate the presence of an L168R KCNJ5 mutation exclusively in the largest nodule (B/T1), whereas all other samples are wild type for this mutation. Corresponding Sanger sequences are depicted below.

Figure 3. Representative CT scans demonstrating findings typical of different genotypes of aldosterone-producing adenomas

A) KCNJ5-mutant left aldosteronoma displaying larger tumor size and lower average H.U. reading. Left aldosterone-producing tumors with CACNA1D (B) and ATP1A1 (C) mutations, respectively, demonstrating smaller tumor size and higher H.U. D) Wild type left aldosteronoma.