title: Methylation of IGF2 regulatory regions to diagnose adrenocortical carcinomas.
abstract: Adrenocortical carcinoma (ACC) is a rare malignancy with a poor prognosis. Discrimination of ACCs from adrenocortical adenomas (ACAs) is challenging on both imaging and histopathological grounds. High IGF2 expression is associated with malignancy, but shows large variability. In this study, we investigate whether specific methylation patterns of IGF2 regulatory regions could serve as a valuable biomarker in distinguishing ACCs from ACAs. Pyrosequencing was used to analyse methylation percentages in DMR0, DMR2, imprinting control region (ICR) (consisting of CTCF3 and CTCF6) and the H19 promoter. Expression of IGF2 and H19 mRNA was assessed by real-time quantitative PCR. Analyses were performed in 24 ACCs, 14 ACAs and 11 normal adrenals. Using receiver operating characteristic (ROC) analysis, we evaluated which regions showed the best predictive value for diagnosis of ACC and determined the diagnostic accuracy of these regions. In ACCs, the DMR0, CTCF3, CTCF6 and the H19 promoter were positively correlated with IGF2 mRNA expression (P<0.05). Methylation in the most discriminating regions distinguished ACCs from ACAs with a sensitivity of 96%, specificity of 100% and an area under the curve (AUC) of 0.997±0.005. Our findings were validated in an independent cohort of 9 ACCs and 13 ACAs, resulting in a sensitivity of 89% and a specificity of 92%. Thus, methylation patterns of IGF2 regulatory regions can discriminate ACCs from ACAs with high diagnostic accuracy. This proposed test may become the first objective diagnostic tool to assess malignancy in adrenal tumours and facilitate the choice of therapeutic strategies in this group of patients.
authors.affiliation: Department of Internal MedicineDivision of Endocrinology, Erasmus MC, University Medical Center Rotterdam, Rotterdam, The Netherlands.
authors.forename: S G
authors.initials: SG
authors.lastname: Creemers
authors.affiliation: Department of Internal MedicineDivision of Endocrinology, Erasmus MC, University Medical Center Rotterdam, Rotterdam, The Netherlands.
authors.forename: P M
authors.initials: PM
authors.lastname: van Koetsveld
authors.affiliation: Department of PathologyErasmus MC, University Medical Center Rotterdam, Rotterdam, The Netherlands.
authors.forename: F J
authors.initials: FJ
authors.lastname: van Kemenade
authors.affiliation: Department of PathologyErasmus MC, University Medical Center Rotterdam, Rotterdam, The Netherlands Department of HistopathologyKing's College Hospital, Denmark Hill, London, UK.
authors.forename: T G
authors.initials: TG
authors.lastname: Papathomas
authors.affiliation: Department of SurgeryErasmus MC, University Medical Center Rotterdam, Rotterdam, The Netherlands.
authors.forename: G J H
authors.initials: GJ
authors.lastname: Franssen
authors.affiliation: Department of Internal MedicineDivision of Endocrinology, Erasmus MC, University Medical Center Rotterdam, Rotterdam, The Netherlands.
authors.forename: F
authors.initials: F
authors.lastname: Dogan
authors.affiliation: Department of PathologyVU University Medical Center, Amsterdam, The Netherlands.
authors.forename: E M W
authors.initials: EM
authors.lastname: Eekhoff
authors.affiliation: Department of PathologyVU University Medical Center, Amsterdam, The Netherlands.
authors.forename: P
authors.initials: P
authors.lastname: van der Valk
authors.affiliation: Department of Internal MedicineDivision of Endocrinology, Erasmus MC, University Medical Center Rotterdam, Rotterdam, The Netherlands.
authors.forename: W W
authors.initials: WW
authors.lastname: de Herder
authors.affiliation: Department of Internal MedicineDivision of Endocrinology, Erasmus MC, University Medical Center Rotterdam, Rotterdam, The Netherlands.
authors.forename: J A M J L
authors.initials: JA
authors.lastname: Janssen
authors.affiliation: Department of Internal MedicineDivision of Endocrinology, Erasmus MC, University Medical Center Rotterdam, Rotterdam, The Netherlands.
authors.forename: R A
authors.initials: RA
authors.lastname: Feelders
authors.affiliation: Department of Internal MedicineDivision of Endocrinology, Erasmus MC, University Medical Center Rotterdam, Rotterdam, The Netherlands l.hofland@erasmusmc.nl.
authors.forename: L J
authors.initials: LJ
authors.lastname: Hofland
mesh_terms: D000293:Adolescent; D000306:Adrenal Cortex Neoplasms / Q000175:diagnosis / Q000235:genetics*; D018246:Adrenocortical Adenoma / Q000175:diagnosis / Q000235:genetics*; D018268:Adrenocortical Carcinoma / Q000175:diagnosis / Q000235:genetics*; D000328:Adult; D000368:Aged; D000964:Antimetabolites, Antineoplastic / Q000494:pharmacology; D001374:Azacitidine / Q000031:analogs & derivatives / Q000494:pharmacology; D045744:Cell Line, Tumor; D002648:Child; D019175:DNA Methylation; D000077209:Decitabine; D005260:Female; D006801:Humans; D007335:Insulin-Like Growth Factor II / Q000235:genetics*; D008297:Male; D008875:Middle Aged; D012333:RNA, Messenger / Q000378:metabolism; D012045:Regulatory Sequences, Nucleic Acid; D055815:Young Adult
chemical_list: D000964:Antimetabolites, Antineoplastic; C035170:IGF2 protein, human; D012333:RNA, Messenger; D007335:Insulin-Like Growth Factor II; D000077209:Decitabine; D001374:Azacitidine
keywords: DNA methylation; IGF2; adrenocortical carcinoma; biomarker; epigenetics