ABSTRACT
Thymic epithelial tumors (TET) consist of thymomas, thymic carcinoma (TC), and neuroendocrine tumors of the thymus (NECTT). Genetic and epigenetic alterations in TET have been the focus of recent research. In the present study, genome-wide screening was performed on aberrantly methylated CpG islands in TET, and this identified neuronal pentraxin 2 (NTPX2) as a significantly hypermethylated CpG island in TC relative to thymomas. NPTX2 is released from pre-synaptic cells in response to neuronal activity/seizure, and plays a role in host immunity and acute inflammation. TET samples were obtained from 38 thymomas, 25 TC, and 6 NECTT. The DNA methylation, mRNA, and protein expression levels of NPTX2 were examined. The DNA methylation rate of the NPTX2 gene was significantly higher in TC than in the normal thymus and thymomas, except B3. The mRNA expression level of NPTX2 was lower in TC than in the normal thymus. An inverse relationship was observed between mRNA expression levels and methylation levels. Relapse-free survival was shorter in patients with high NPTX2 DNA methylation levels than in those with low DNA methylation levels. NECTT showed very high mRNA and protein expression levels and low DNA methylation levels of NPTX2. NPTX2 may function as a tumor suppressor in TC, and have an oncogenic function in NECTT.
ABSTRACT
Our previous study reported that the DNA methylation of growth hormone secretagogue receptor (GHSR) was significantly higher in thymoma or thymic carcinoma (TC) than in normal thymic tissue samples. Thymic epithelial tumors (TETs) with higher GHSR DNA methylation were associated with significantly worse prognosis than those with lower levels of DNA methylation. Diversified components of the ghrelin-GHSR axis may exert opposing effects in cancer progression, depending on the cancer type in question. However, the precise function of the axis remains unclear. In the present study, the mRNA expression of five key components of the ghrelin system [native ligand ghrelin, variant ligand In-1 ghrelin, native receptor GHSR1a, variant receptor GHSR1b and acylation enzyme ghrelin O-acyltransferase (GOAT)] were examined in 58 TET samples by reverse transcription-quantitative PCR, and protein expression of GHSR1a and GHSR1b was assessed in 20 TETs using immunohistochemistry. The results revealed that In-1 ghrelin, GHSR1b (variant forms) and GOAT were more strongly expressed in thymoma compared with thymic-adjacent tissue. By contrast, no significant differences were observed in the expression of ghrelin and GHSR1a (native forms) between thymoma and thymic tissue. The mRNA expression of In-1 ghrelin and GHSR1b (variant forms) was positively associated with GHSR methylation in thymoma tissue samples. However, a relationship was not found between ghrelin, GHSR1a or GOAT expression (native forms) and GHSR methylation in thymoma. Immunohistochemical analysis revealed that mRNA expression of GHSR1a and GHSR1b generally correlated with expression of the corresponding protein, and that the expression of GHSR1b was increased in advanced-stage TETs. These results indicate that the DNA methylation of GHSR is associated with a shift from native expression (ghrelin and GHSR1a) to variant expression (In-1 ghrelin and GHSR1b), which induces the tumorigenesis of thymoma, but not TC.
ABSTRACT
Thymic epithelial tumors (TETs) comprise thymomas and thymic carcinoma (TC). TC has more aggressive features and a poorer prognosis than thymomas. Genetic and epigenetic alterations in thymomas and TC have been investigated in an attempt to identify novel target molecules for TC. In the present study, genome-wide screening was performed on aberrantly methylated CpG islands in thymomas and TC, and the glutamate decarboxylase 1 gene (GAD1) was identified as the 4th significantly hypermethylated CpG island in TC compared with thymomas. GAD1 catalyzes the production of γ-aminobutyric acid from L-glutamic acid. GAD1 expression is abundant in the brain but rare in other tissues, including the thymus. A total of 73 thymomas and 17 TC tissues were obtained from 90 patients who underwent surgery or biopsy at Tokushima University Hospital between 1990 and 2017. DNA methylation was examined by bisulfite pyrosequencing, and the mRNA and protein expression levels of GAD1 were analyzed using reverse transcription-quantitative PCR and immunohistochemistry, respectively. The DNA methylation levels of GAD1 were significantly higher in TC tissues than in the normal thymus and thymoma tissues, and GAD1 methylation exhibited high sensitivity and specificity for discriminating between TC and thymoma. The mRNA and protein expression levels of GAD1 were significantly higher in TC tissues than in thymomas. Patients with TET with high GAD1 DNA hypermethylation and high mRNA and protein expression levels had significantly shorter relapse-free survival rates than those with low levels. In conclusion, significantly more epigenetic alterations were observed in TC tissues compared with in thymomas, which may contribute to the clinical features and prognosis of patients.
