Detection of somatic beta-catenin mutations in primary pigmented nodular adrenocortical disease (PPNAD)

Primary pigmented nodular adrenocortical disease (PPNAD) leads to Cushing syndrome (CS) and is often associated with Carney complex (CNC). Genetic alterations of the type 1-alpha regulatory subunit of cAMP-dependent protein kinase A (PRKAR1A) and phosphodiesterase 11A4 (PDE11A) genes have been found in PPNAD. Recent studies have demonstrated that beta-catenin mutations are frequent in adrenocortical adenomas and carcinomas and that the Wnt-signalling pathway is involved in PPNAD tumorigenesis. We hypothesized that adrenocortical adenomas that form in the context of PPNAD may harbour beta-catenin mutations.

We studied 18 patients with CS secondary to PPNAD who were screened for germline PRKAR1A and PDE11A mutations. Tumor DNA was extracted from pigmented adrenocortical adenoma and nodular adrenal hyperplasia. Mutation analysis of exons 3 and 5 of beta-catenin was performed using polymerase chain reaction and direct sequencing. Sections from formalin-fixed, paraffin-embedded tumour samples were studied by immunohistochemistry with an antibody against beta-catenin.

Nine patients were carrying germline PRKAR1A mutations and one patient had a PDE11A mutation. We found somatic beta-catenin mutations in 2 of 18 patients (11%). In both cases, the mutations occurred in relatively large adenomas that had formed in the background of PPNAD. Tumor DNA analysis revealed a heterozygous ACC-to-GCC missense mutation in codon 41 (T41A) and a TCT-to-CCT missense mutation in codon 45 (S45P) of exon 3 of the beta-catenin gene that was confirmed at the cDNA level. There were no alterations in the DNA of PPNAD-adjacent tissues and lymphocytes from the patients, indicating somatic events. Immunohistochemistry showed nuclear accumulation of beta-catenin in more than 90% of cells in adenomatous tissue whereas no nuclear immunoreactivity was detected in adjacent PPNAD nodular cells. Nuclear translocation of beta-catenin protein in the PPNAD adenoma suggests activation of the Wnt-beta-catenin pathway in PPNAD.

We report, for the first time, beta-catenin mutations in adenomas associated with PPNAD, further implicating Wnt-beta-catenin signalling in tumorigenesis linked to bilateral adrenal hyperplasias.