Increased Frequency and Distinct Genotypic Patterns of Somatic Aldosterone-Driver Mutations in Aldosterone-Producing Micronodules from Primary Aldosteronism Patients

Primary aldosteronism (PA) is the leading cause of endocrine hypertension. With the development of highly specific human aldosterone synthase (CYP11B2) antibodies for immunohistochemistry (IHC), the presence of microscopic subcapsular foci of CYP11B2-postive cells – now termed aldosterone-producing micronodules (APM) – has been documented in normal and PA adrenal glands, however, there continues to be debate regarding the role of APM in the pathogenesis of PA. In this study, CYP11B2 IHC-guided targeted next-generation sequencing (NGS) was utilized to characterize the frequency and spectrum of somatic aldosterone-driver mutations in APM identified within adrenal glands from deceased renal donors and patients with primary aldosteronism (PA). 59 subjects were enrolled (31 deceased renal donors and 28 PA patients), 173 APM were collected using CYP11B2-guided IHC (65 APM from 31 deceased renal donors and 108 APM from 28 PA patients), and NGS data was successfully obtained for 131 APM (51 APM from 31 deceased renal donors and 80 APM from 28 PA patients). Importantly, NGS identified aldosterone-driver mutations in a significantly higher proportion of APM from PA patients relative to deceased renal donors (71.3% compared to 45.1%; P-value < 0.05) – with the vast majority in either group being missense mutations in the L-type calcium channel gene CACNA1D. Interestingly, in PA patients, the presence of APA/APN harboring CACNA1D or ATP1A1 mutations was significantly associated with an increased frequency of APM with CACNA1D mutations, while APM from adrenal glands with KCNJ5 mutation-bearing APA/APN were more likely to be mutation-negative (P-values < 0.001 and < 0.05, respectively). Finally, clinical outcomes for PA patients were significantly associated with APM genotype, with post-surgical cure showing higher frequencies of KCNJ5-mutant and mutation-negative APM and a lower frequency of CACNA1D-mutant APM. Overall, our data support a link between the acquisition of somatic aldosterone-driver mutations in APM and PA pathogenesis and suggest the possibility of diverse genetic mechanisms underlying APM development in normal and PA adrenal glands. Furthermore, these results suggest that molecular diagnostic testing of CYP11B2-positive adrenal cortical lesions may help risk stratify patients in clinical practice.