Wnt signaling plays important roles in tumorigenesis, invasiveness and therapeutic resistance of glioblastoma (GBM). We simultaneously investigated six Wnt pathway markers (Wnt5a, Fzd-2, beta-catenin, Wnt3a, Wnt7b, Fzd-10) at epigenetic and protein levels in 21 sequential formalin-fixed paraffin-embedded GBM pairs and controls. Expression levels of Wnt5a, beta-catenin and Wnt3a proteins either moderately or significantly increased, while those of Fzd-2, Wnt7b and Fzd-10 decreased in the primary (GBM-P) and recurrent (GBM-R) tumors compared to the controls. Methylation levels within promoters and genes showed corresponding decreases for Wnt5a, beta-catenin and Wnt3a in tumors vs. controls, while that of Fzd-10 was uniformly high. Comparing the GBM-P and GBM-R pairs, proteins of Fzd-2, beta-catenin and Wnt3a were either moderately or significantly up-, while that of Wnt7b was downregulated in GBM-R, but these patterns were not accompanied by inverse methylation patterns in the corresponding promoters and genes over time. No methylation differences were noted within promoters and genes of the same markers in 112 pairs of primary and recurrent GBMs in a database, suggesting that the observed changes in protein expression levels may not be explained by CpG methylation status alone. The promoter and gene methylation rate was the highest for Fzd-10 in the database cohort too, supporting the noted low Fzd-10 protein expression. These analyses underscore the relevance of Wnt pathway molecules in the context of their methylation profiles in the development and evolution of GBM, and suggest that Wnt pathway regulation as a potential treatment target merits further studies.

Purpose: Glioblastoma is the most aggressive form of brain tumors. A better understanding of the molecular mechanisms leading to its evolution is essential for the development of treatments more effective than the available modalities. Here, we aim to identify molecular drivers of glioblastoma development and recurrence by analyzing DNA CpG methylation patterns in sequential samples. Methods: DNA was isolated from 22 pairs of primary and recurrent formalin-fixed, paraffin-embedded glioblastoma specimens, and subjected to reduced representation bisulfite sequencing. Bioinformatic analyses were conducted to identify differentially methylated sites and pathways, and biostatistics was used to test correlations among clinical and pathological parameters. Results: Differentially methylated pathways likely involved in primary tumor development included those of neuronal differentiation, myelination, metabolic processes, synapse organization and endothelial cell proliferation, while pathways differentially active during glioblastoma recurrence involved those associated with cell processes and differentiation, immune response, Wnt regulation and catecholamine secretion and transport. Conclusion: DNA CpG methylation analyses in sequential clinical specimens revealed hypomethylation in certain pathways such as neuronal tissue development and angiogenesis likely involved in early tumor development and growth, while suggested altered regulation in catecholamine secretion and transport, Wnt expression and immune response contributing to glioblastoma recurrence. These pathways merit further investigations and may represent novel therapeutic targets.

Introduction: Two types of testicular teratomas are distinguished by the current WHO classification. Prepubertal-type teratomas are benign, while postpubertal-type teratomas are considered malignant with metastatic potential, and are associated with germ cell neoplasia in situ. Prepubertal-type cases have been reported in the adult testis potentially causing confusion and overtreatment. Demonstration of the absence of 12p abnormalities with fluorescence in situ hybridization may facilitate diagnosis. Recently, IMP3 has emerged as a potential marker of malignancy in this context. Aims: The aim of this study was to assess histological characteristics, IMP3 expression and the presence of 12p abnormalities of pure testicular teratomas. Results: Thirty-seven cases were studied, 7 patients were children and 30 were adults. Six out of 7 pediatric cases showed no 12p abnormality and were IMP3 positive. Seventy-four percent and 79% of adult cases showed 12p abnormalities and IMP3 expression, respectively. Negative cases were not associated with in situ neoplasia or metastasis, they were smaller (mean, 14 vs 39 mm), showed less histological diversity (2.4 vs 4.0 types of tissues on average) compared to positive cases. Conclusion: Our study provides further evidence that prepubertal-type (type I) teratomas may appear in adult testes, thus teratomas in adults may be either benign (type I) or malignant (type II). IMP3 expression may aid the distinction between type I and type II teratomas of the postpubertal testis even when GCNIS and 12p status cannot be assessed.

Background: Immune evasion in glioblastoma (GBM) shields cancer cells from cytotoxic immune response. Methods: We investigated CpG methylation in promoters, genes, and pathways in 22 pairs of formalin-fixed paraffin-embedded sequential (FFPE) GBM using restricted resolution bisulfite sequencing (RRBS) and bioinformatic analyses. Results: Gene ontology revealed hypermethylation in elements of the innate and adaptive immune system when recurrent GBM samples (GBM^rec) were compared to control (CG) and primary GBM samples (GBM^prim). Higher methylation levels of the IL-7 signaling pathway and response to IL-7 were found in GBM^rec suggesting a progressive blockade of the IL-7 driven T cell response in sequential GBM. Analyses of the Cancer Genome Atlas array-based data confirmed hypermethylation of the IL-7 pathway in recurrent compared with primary GBM. We also quantified DNA CpG methylation in promoter and gene regions of the IL-7 ligand and IL-7 α-receptor subunit in individual samples of a large RRBS-based sequential cohort of GBM in a Viennese database and found significantly higher methylation levels in the IL-7 receptor α-subunit in GBM^rec compared with GBM^prim. Conclusions: This study revealed the progressive suppression of the IL-7 receptor-mediated pathway as a means of immune evasion by GBM and thereby highlighted it as a new treatment target.