Copy number variations alter methylation and parallel IGF2 overexpression in adrenal tumors.

Helene Myrtue Nielsen,Frederic Castinetti,Vessela N Kristensen,Lise Lotte Hansen,Jörg Tost,Anne Barlier,Celine Besse,Hans Kristian Moen Vollan,Frederic Sebag,Catherine De Micco,Antoine Daunay,Peter Van Loo,David Taieb,Carole Guerin,Alexandre How-Kit

doi:10.1530/erc-15-0086

Abstract

Overexpression of insulin growth factor 2 (IGF2) is a hallmark of adrenocortical carcinomas and pheochromocytomas. Previous studies investigating the IGF2/H19 locus have mainly focused on a single molecular level such as genomic alterations or altered DNA methylation levels and the causal changes underlying IGF2 overexpression are still not fully established. In the current study, we analyzed 62 tumors of the adrenal gland from patients with Conn's adenoma (CA, n=12), pheochromocytomas (PCC, n=10), adrenocortical benign tumors (ACBT, n=20), and adrenocortical carcinomas (ACC, n=20). Gene expression, somatic copy number variation of chr11p15.5, and DNA methylation status of three differential methylated regions of the IGF2/H19 locus including the H19 imprinting control region were integratively analyzed. IGF2 overexpression was found in 85% of the ACCs and 100% of the PCCs compared to 23% observed in CAs and ACBTs. Copy number aberrations of chr11p15.5 were abundant in both PCCs and ACCs but while PCCs retained a diploid state, ACCs were frequently tetraploid (7/19). Loss of either a single allele or loss of two alleles of the same parental origin in tetraploid samples resulted in a uniparental disomy-like genotype. These copy number changes correlated with hypermethylation of the H19 ICR suggesting that the lost alleles were the unmethylated maternal alleles. Our data provide conclusive evidence that loss of the maternal allele correlates with IGF2 overexpression in adrenal tumors and that hypermethylation of the H19 ICR is a consequence thereof.

Highlights

Tumors developing in the adrenal gland include a heterogeneous group of adrenocortical and adrenomedullary malignancies (Brunt & Moley 2001, Grumbach et al 2003)
To provide a comprehensive analysis of the mechanisms underlying insulin growth factor 2 (IGF2) overexpression in adrenal tumors, gene expression levels were integrated with copy number and ploidy analysis, and the DNA methylation status of three differentially methylated regions (DMRs) including the imprinting control region (ICR) upstream of H19 (H19 DMR) and two secondary DMRs (DMR0 and DMR2) within IGF2 (Supplementary Fig. 1, see section on supplementary data given at the end of this article)
Studies investigating the underlying mechanisms of IGF2 overexpression in adrenal tumors at more than a single molecular level are limited (Gicquel et al 1994, 1997, Gao et al 2002) and no study has so far integrated epigenetic and copy number variation obtained with quantitative high-resolution technologies with gene expression

Summary

Introduction

Tumors developing in the adrenal gland include a heterogeneous group of adrenocortical and adrenomedullary malignancies (Brunt & Moley 2001, Grumbach et al 2003). Adrenocortical masses require attention as hypersecretion of steroid hormones and metabolic changes may be present, which might lead to hypertension, abdominal pain, gender disorders, weight gain, or increased blood sugar levels depending on which adrenal hormone is dysregulated (Low & Sahi 2012, Hodin et al 2014). Adrenal tumors with increased aldosterone secretion lead to Conn’s syndrome (CA), whereas pheochromocytomas (PCCs) secrete excess amounts of catecholamines (Low & Sahi 2012, Hodin et al 2014). Adrenocortical carcinomas (ACCs) can present as either non-functional or functional tumors and represent the most aggressive group of adrenal tumors. ACCs have often metastasized at the time of diagnosis and are associated with poor prognosis (Volante et al 2008)

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Conclusion