Epigenetic Deregulation of Protocadherin PCDHGC3 in Pheochromocytomas/Paragangliomas Associated With SDHB Mutations

Abstract

Context SDHB mutations are found in an increasing number of neoplasms, most notably in paragangliomas and pheochromocytomas (PPGLs). SDHB-PPGLs are slow-growing tumors, but ∼50% of them may develop metastasis. The molecular basis of metastasis in these tumors is a long-standing and unresolved problem. Thus, a better understanding of the biology of metastasis is needed. Objective This study aimed to identify gene methylation changes relevant for metastatic SDHB-PPGLs. Design We performed genome-wide profiling of DNA methylation in diverse clinical and genetic PPGL subtypes, and validated protocadherin γ-C3 (PCDHGC3) gene promoter methylation in metastatic SDHB-PPGLs. Results We define an epigenetic landscape specific for metastatic SDHB-PPGLs. DNA methylation levels were found significantly higher in metastatic SDHB-PPGLs than in SDHB-PPGLs without metastases. One such change included long-range de novo methylation of the PCDHA, PCDHB, and PCDHG gene clusters. High levels of PCDHGC3 promoter methylation were validated in primary metastatic SDHB-PPGLs, it was found amplified in the corresponding metastases, and it was significantly correlated with PCDHGC3 reduced expression. Interestingly, this epigenetic alteration could be detected in primary tumors that developed metastasis several years later. We also show that PCDHGC3 down regulation engages metastasis-initiating capabilities by promoting cell proliferation, migration, and invasion. Conclusions Our data provide a map of the DNA methylome episignature specific to an SDHB-mutated cancer and establish PCDHGC3 as a putative suppressor gene and a potential biomarker to identify patients with SDHB-mutated cancer at high risk of metastasis who might benefit from future targeted therapies.

Metastatic Paragangliomas and Pheochromocytomas: An Epigenetic View

“…More importantly, among the SDHB-mutated PPGLs, those having a metastatic behavior had a significantly higher levels of methylation than tumors that had not developed metastasis at the last follow-up date. Analysis of the RNAseq data confirmed the epigenetic silencing of, not only PCDHGC3 [24], but also PCDHGC4 gene ( Figure 5). The PCDHGC4 mRNA levels were found significantly decreased in SDHx-mutated PPGLs as compared with tumors with other genotypes.…”

“…To identify epigenetic alterations relevant for metastasis, we recently performed a comprehensive analysis of DNA methylation in metastatic PPGLs with and without SDHB mutations. This analysis revealed that over 1000 genes harbored promoter hypermethylation in the metastatic tumors but not in the not metastatic ones thus suggesting that those gene alterations have a role in the pathogenesis of the metastatic disease linked to SDHB mutations [24]. About 15% of these alterations had been also identified in sdhb−/− mouse chromaffin cells and in 41% of SDHx-mutated PPGLs analyzed by Letouzé et al [14].…”

“…Among these hypermethylated genes, we identified CNTN2, SDK1, TENM1, TENM4 encoding neuronal cell adhesion molecules involved in the establishment of connections in the nervous system. More strikingly, the cell-cell adhesion set of hypermethylated genes included a 1 Mb-long chromosomal region that hold clustered protocadherin genes [designated as PCDHA, PCDHB and PCDHG (collectively, PCDHs)] encompassing 50 different genes at the chromosomal locus 5q31.3 [24] (Figure 4). One of the PCDH genes, PCDHGC3, has been further analyzed and found to be of clinical relevance in metastatic SDHB-mutated PPGLs.…”

“…Several recent studies have revealed that epigenetic silencing of clustered PCDHs is present in various human malignant tumors, such as Wilms tumor, neuroblastoma, breast, prostate, colon cancer, gastric and biliary tract cancers, and astrocytoma suggesting that this process plays roles in regulating cancer development and/or progression [34][35][36][37]. By using one of the largest cohorts of epigenetically studied SDHB-mutated PPGLs, we have recently found that the epigenetic silencing of one of the clustered PCDH genes, PCDHGC3, is putatively involved in the metastatic behavior of these tumors [24]. Methylation of PCDHGC3 promoter were found to be null in normal paraganglia, null or low in most SDHB-mutated PPGLs that do not metastasize, high in SDHB-mutated metastatic PPGLs, and much higher in the metastatic tissues derived from these tumors.…”

Identification of Genes Associated with the Metastasis of Pheochromocytoma/Paraganglioma Based on Weighted Gene Coexpression Network Analysis

“…In recent years, genetic changes in PCPG have received increasing attention, and mutations or loss of genes encoding succinate dehydrogenase (SDH) are clinically important genetic changes [4,5], which are associated with the increased metastatic rate of human PCPG. It has been proved that the pathogenesis of nonfamilial PCPG involves SDHB mutation-associated PCPG pro-cadherin c-C3 (PCDHGC3) gene promoter methylation [6] as well as EPAS1 mutations encoding HIF2α [7]. Given that those genetic changes play important roles in the development of PCPG, identification of the genes with a key role in PCPG could be beneficial to the diagnosis, treatment, and prognostic prediction of this neoplasm.…”