Succinate Dehydrogenase Deficiency Is Associated with Decreased 5-Hydroxymethylcytosine Production in Gastrointestinal Stromal Tumors: Implications for Mechanisms of Tumorigenesis
Emily F Mason, Jason L Hornick. Brigham and Women's Hospital, Harvard Medical School, Boston, MA
Background: Gastrointestinal stromal tumors (GISTs) usually harbor activating mutations in KIT or PDGFRA, which promote tumorigenesis through growth factor receptor signaling pathways. Around 15% of GISTs in adults and >90% in children lack such mutations ("wild-type" GISTs). Most gastric wild-type GISTs show loss of function of the Krebs cycle enzyme complex succinate dehydrogenase (SDH). The mechanism by which SDH deficiency drives tumorigenesis is unclear. Loss of SDH leads to succinate accumulation, which inhibits α-ketoglutarate-dependent dioxygenase enzymes, such as the TET family of DNA hydroxylases. TET proteins catalyze conversion of 5-methylcytosine to 5-hydroxymethylcytosine (5-hmc), required for subsequent DNA demethylation. Thus, TET-mediated 5-hmc production alters global DNA methylation patterns and thereby influences gene expression. In this study, we investigated 5-hmc levels in a cohort of genotyped GISTs to determine whether loss of SDH was associated with inhibition of TET activity.
Design: 30 genotyped GISTs were examined, including 10 SDH-deficient tumors (5 SDHA mutant; 1 SDHB mutant; 1 SDHC mutant; 3 unknown), 14 with KIT mutations (10 exon 11; 3 exon 9; 1 exon 17), and 6 with PDGFRA mutations (all exon 18). Immunohistochemistry was performed following antigen retrieval using a rabbit anti-5-hmc polyclonal antibody (1:10,000; Active Motif). Nuclear staining for 5-hmc in tumor cells was compared to endothelial cells and lymphocytes, which served as positive controls, and was scored as positive, weak (<50% intensity of controls), or negative.
Results: 5-hmc was negative in 9 of 10 (90%) SDH-deficient GISTs, 3 of 14 (21%) KIT-mutant GISTs (2 exon 11, 1 exon 17), and 1 of 6 (17%) PDGFRA-mutant GISTs. One SDH-deficient GIST showed weak staining for 5-hmc. All 3 KIT exon 9-mutant GISTs and 2 PDGFRA-mutant GISTs showed weak 5-hmc staining.
Conclusions: 5-hmc is absent in nearly all SDH-deficient GISTs. These findings suggest that SDH deficiency may promote tumorigenesis through accumulation of succinate and inhibition of dioxygenase enzymes. Inhibition of TET activity may, in turn, alter global DNA methylation in SDH-deficient tumors.
Monday, March 4, 2013 1:00 PM
Poster Session II # 275, Monday Afternoon