597 J. Endocrinol. Invest. 32: 597-600, 2009 DOI: 10.3275/6294 ABSTRACT. Background: Activating mutations of the BRAF oncogene play a central role in the development of various cancer types, but their role in human adrenocortical tumors is unknown. At variance, activating mutations of another onco- gene, CTNNB1, which encodes β β-catenin, have been shown to be common events in both benign and malignant adreno- cortical tumors. Aim: To investigate the prevalence of BRAF and CTNNB1 activating mutations in sporadic adrenocortical tumors. Materials and methods: Tissue samples from 15 adrenocortical carcinomas and 41 adrenocortical adenomas were investigated for the presence of BRAF and CTNNB1 ac- tivating mutations by PCR amplification and direct sequenc- ing. Results: An advanced invasive non-functioning adreno- cortical carcinoma carried a somatic heterozygous BRAF V600E mutation, while 4 functioning and 4 non-functioning adenomas and 3 functioning carcinomas carried different CTNNB1 activating mutations. Conclusions: Activating BRAF somatic mutations may be occasionally found in advanced adrenocortical carcinomas, while CTNNB1 activating muta- tions are early and common events in adrenal tumorigenesis. (J. Endocrinol. Invest. 32: 597-600, 2009) © 2009, Editrice Kurtis INTRODUCTION Sporadic adenomas/hyperplasia of the adrenal cortex are frequent incidental findings in the general population, whereas adrenocortical carcinomas are very rare and as- sociated with a poor prognosis (1). The molecular patho- genesis of adrenocortical cancer is not yet completely understood, even though recent advances have high- lighted the involvement of both tumor suppressor genes, such as TP53, MEN1, PRKAR1A, and oncogenes, such as IGF2, CTNNB1, and RAS (2). Among oncogenes, activating mutations of the CTNNB1 gene and deregulation of the encoded β-catenin protein appear to be early and common events in adrenal tu- morigenesis. In fact, CTNNB1 somatic mutations have been detected in both benign and malignant adreno- cortical tumors, with a prevalence of 27% and 31%, re- spectively (3). The relatively high prevalence of activat- ing CTNNB1 mutations in benign adrenocortical tumors, including primary pigmented nodular adrenocortical dis- eases associated with PRKAR1A mutations, has been con- firmed in other studies (4, 5). Like in other types of tu- mors, activating mutations mostly affect residues in CTNNB1 exon 3 that are crucial for β-catenin targeted degradation, and result in cytoplasmic and nuclear ac- cumulation with subsequent increase in β-catenin tran- scriptional activity (3). Ras proteins are membrane-associated proteins that are frequently mutated in human malignancies, but their role in development of adrenal tumors is still controversial. In fact, somatic mutations of K-ras have been reported in 46% of Conn’s adenomas (6), but other studies have not found any Ras mutation (7, 8). The B-RAF kinase is a downstream effector of the RAS protein and part of the extracellular signal-regulated kinase/mitogen-activated protein kinase (ERK/MAPK) signal transduction pathway. Activating mutations of the BRAF oncogene, generally represented by the amino acid substitution V600E, are key events in the development of numerous cancer types, such as melanoma, thyroid, colon, ovarian, and stomach cancer (9, 10), which are also linked to Wnt/β-catenin pathway activation (11). The involvement of BRAF muta- tions in the development of human adrenocortical tu- mors has not been extensively investigated so far. Davies et al. (9) tested for BRAF mutations the adrenocortical carcinoma cell lines NCI-H295 and SW13 and did not de- tect any genetic abnormalities. Regarding adrenal medullary tumors, two reports investigated the presence of BRAF-activating mutations in a total of 69 pheochro- mocytomas, but did not find any mutation (12, 13). Aim of this study was to investigate a series of function- ing and non-functioning, benign and malignant adreno- cortical tumors for the presence of mutations in BRAF ex- on 15 and in CTNNB1 exon 3, which are the exons where most activating mutations are detected. MATERIALS AND METHODS Patients and tissue samples The presence of somatic BRAF and CTNNB1 mutations was screened in tissue samples from 15 adrenocortical carcinomas, 11 cortisol-producing adenomas, 19 aldosterone-producing adenomas, 1 testosterone-producing adenoma, and 10 non- functioning adrenocortical adenomas. Tissue samples were ob- tained from a series of 56 consecutive patients (Table 1) who underwent adrenalectomy at the Endocrine Surgery Unit of the University of Padua in the period between January 2005 and May 2007. Peripheral whole blood samples were also available *G. Palù and L. Barzon share senior authorship. Key-words: Adrenocortical carcinoma, adrenocortical neoplasm, β-catenin, BRAF, mutation analysis, oncogene. Correspondence: L. Barzon, MD, and G. Palù, MD, Department of Histology, Micro- biology, and Medical Biotechnologies, University of Padova, Via A. Gabelli 63, I-35128 Padova, Italy. E-mail: luisa.barzon@unipd.it; giorgio.palu@unipd.it Accepted February 25, 2009. First published online May 15, 2009. Investigation of BRAF and CTNNB1 activating mutations in adrenocortical tumors G. Masi 1 , E. Lavezzo 1 , M. Iacobone 2 , G. Favia 2 , G. Palù 1 *, and L. Barzon 1 * 1 Department of Histology, Microbiology, and Medical Biotechnologies; 2 Department of Surgical and Gastroenterological Sciences, University of Padua, Padua, Italy