Artikel
Molecular and functional characterization of SMOC1 in IDH1 wild type and IDH1 mutant gliomas
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Autoren
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Marc Maximilian Weber
- Heinrich-Heine Universität Düsseldorf, Institute for Neuropathology Düsseldorf, Düsseldorf, Germany
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Wolfgang Kaisers
- Heinrich-Heine University Düsseldorf, Mathematics, Düsseldorf, Germany
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Guido Reifenberger
- Heinrich-Heine Universität Düsseldorf, Institute for Neuropathology Düsseldorf, Düsseldorf, Germany
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Christiane B. Knobbe-Thomsen
- Heinrich-Heine Universität Düsseldorf, Institute for Neuropathology Düsseldorf, Düsseldorf, Germany
| Veröffentlicht: | 14. September 2016 |
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Gliederung
Text
Questions: In 2010 Noushmehr and coworkers showed that IDH1 mutations are associated with a CpG island methylator phenotype (CIMP) in glioma [1]. This phenotype is accompanied with longer overall survival, but its molecular role and function remains to be elucidated.
Methods: Using data from the The Cancer Genome Atlas we generated a list of differentially methylated candidate genes. We performed mRNA expression analyses of 15 candidate genes in a group of 79 glioma samples via qRT-PCR. IDH1 status of the 79 tumor samples was determined by pyrosequencing. To assess the promotor methylation status we performed Sangersequencing. We transfected U87 MG glioblastoma cells with a vector containing the coding sequence of SMOC1 or an empty vector as control. MTT-Assays were performed to investigate effects of cytostatic agents on SMOC1 expressing cells. To analyze the influence of SMOC1 expression on TGF-β signaling we treated SMOC1 expressing glioblastoma cells and controls with BMP-2 or TGF-β1 for different time periods and performed Westernblot afterwards. We used mass spectrometry of the lysate and secretome of SMOC1 expressing cells to analyze the impact of SMOC1 on protein expression.
Results: One of the 15 candidate genes – SPARC related modular calcium binding 1 (SMOC1) – showed a significantly higher mRNA expression in IDH1 mutant samples compared to IDH1 wildtype samples (p < 0.0001, t-test with Welch’s correction).
Analyses of the promotor region of SMOC1 via Sangersequencing revealed that SMOC1 mRNA expression levels however do not significantly correlate with its promotor methylation. MTT-Assays showed that SMOC1 expressing cells are less vulnerable to different cytotoxic agents like etoposide (p < 0.05), gemcitabine (p <0.0001) and hydroxyurea (p < 0.01). Analyses of the MS data and Westernblots are still in progress.
Conclusions: In summary we show that SMOC1 is differentially expressed in IDH1 wild type and IDH1 mutant gliomas and further experimental results will elucidate how SMOC1 is involved in the pathogenesis of diffuse gliomas. Further research will be needed to determine the exact regulation of SMOC1.
References
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