All 12 glioblastoma samples were classified as isocitrate dehydrogenase (IDH) wild type or mutated by methylation profiling, but subclass differences were noted in 5 tumors.
Intratumor heterogeneity was identified as a characteristic of glioblastoma.
Some samples showed greater variations in DNA methylation within the same tumor compared to differences between tumors.
Approximately 17,000 cytosine-phosphate-guanine (CpG) sites exhibited different methylation levels within the tumors.
MGMT methylation status was found to vary in IDH mutated patients, with 1 out of 1 showing differences.
The heterogeneity observed may affect the reliability of methylation-based biomarkers for glioblastoma diagnosis and patient stratification.
Simplified
BACKGROUND: A feature of glioblastoma (GBM) is cellular and molecular heterogeneity, both within and between tumors. This variability causes a risk for sampling bias and potential tumor escape from future targeted therapy. Heterogeneous intratumor gene expression in GBM is well documented, but little is known regarding the epigenetic heterogeneity. Variability in within tumors would have implications for diagnostics, as methylation can be used for tumor classification, subtyping, and determination of the clinically used biomarker O6-methylguanine-DNA methyltransferase (MGMT) promoter methylation. We therefore aimed to profile the intratumor DNA methylation heterogeneity in GBM and its effect on diagnostic properties.
METHODS: Three to 4 spatially separated biopsies per tumor were collected from 12 GBM patients. We performed genome-wide DNA methylation analysis and investigated intratumor variation.
RESULTS: All samples were classified as GBM isocitrate dehydrogenase (IDH) wild type (wt)/mutated by methylation profiling, but the subclass differed within 5 tumors. Some GBM samples exhibited higher DNA methylation differences within tumors than between, and many cytosine-phosphate-guanine (CpG) sites (mean: 17 000) had different methylation levels within the tumors. MGMT methylation status differed in IDH mutated patients (1/1).
CONCLUSIONS: We demonstrated that intratumor DNA methylation heterogeneity is a feature of GBM. Although all biopsies were classified as GBM IDH wt/mutated by methylation analysis, the assigned subclass differed in samples from the same patient. The observed heterogeneity within tumors is important to consider for methylation-based biomarkers and future improvements in stratification of GBM patients.
Key numbers
5 of 12
Subclass Variation
Subclass assignments varied within 5 tumors out of 12.
17,000
Differentially Methylated Probes
Mean number of DMP within GBM tumors.
7 years
Methylation Age Difference
Mean difference in methylation age within GBM tumors.
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