Abstract

To identify novel epigenetic signatures that could provide predictive information that is complementary to promoter methylation status of the O-6-methylguanine-DNA methyltransferase (MGMT) gene for predicting temozolomide (TMZ) response, among glioblastomas (GBMs) without glioma-CpGs island methylator phenotype (G-CIMP) Different cohorts of primary non-G-CIMP GBMs with genome-wide DNA methylation microarray data were included for discovery and validation of a multimarker signature, combined using a RISK score model. Different statistical analyses and functional experiments were performed for clinical and biological validation. By employing discovery cohorts with radiotherapy (RT) and TMZ versus RT alone and a strict multistep selection strategy, we identified seven CpGs, each of which was significantly correlated with overall survival (OS) of non-G-CIMP GBMs with RT/TMZ, independent of age, MGMT promoter methylation status, and other identified CpGs. A RISK score signature of the 7 CpGs was developed and validated to distinguish non-G-CIMP GBMs with differential survival outcomes to RT/TMZ, but not to RT alone. The interaction analyses also showed differential outcomes to RT/TMZ versus RT alone within the RISK score-based subgroups. The signature could also improve the risk classification by age and MGMT promoter methylation status. Functional experiments showed that HSBP2 appeared to be epigenetically regulated by one identified CpG and was associated with TMZ resistance, but it was not associated with cell proliferation or apoptosis in GBM cell lines. The predictive value of the single CpG methylation of HSBP2 by pyrosequencing was observed in a local cohort of isocitrate dehydrogenase 1 (IDH1) R132H wild-type GBMs. This novel epigenetic signature might be a promising predictive (but not a general prognostic) biomarker and be helpful for refining the MGMT-based guiding approach to TMZ usage in non-G-CIMP GBMs.