Experimental glioma with high bHLH expression harbor increased replicative stress and are sensitive toward ATR inhibition

Abstract

Background The overexpression of (basic)helix-loop-helix ((b)HLH) transcription factors is frequent in malignant glioma. We investigated molecular effects upon disruption of the (b)HLH network by a dominant negative variant of the E47 protein (dnE47). Our goal was to identify novel molecular subgroup-specific therapeutic strategies. Methods Glioma cell lines LN229, LNZ308 and GS-2/GS-9 were lentivirally transduced. Functional characterization included immunocytochemistry, immunoblots, cytotoxic and clonogenic survival assays in vitro, and latency until neurological symptoms in vivo. Results of cap analysis gene expression (CAGE) and RNA sequencing (RNA-Seq) were further validated by immunoblot, flow cytometry and functional assays in vitro. Results The induction of dnE47-RFP led to cytoplasmic sequestration of (b)HLH transcription factors and anti-glioma activity in vitro and in vivo. Downstream molecular events, i.e. alterations in transcription start site usage and in the transcriptome revealed enrichment of cancer-relevant pathways, particularly of the DNA damage response (DDR) pathway. Pharmacologic validation of this result using ATR inhibition led to a significantly enhanced early and late apoptotic effect compared to temozolomide alone. Conclusions Gliomas overexpressing (b)HLH transcription factors are sensitive towards inhibition of the ATR kinase. The combination of ATR inhibition plus temozolomide or radiation therapy in this molecular subgroup are warranted.

Multiparametric Longitudinal Profiling of RCAS-tva-Induced PDGFB-Driven Experimental Glioma

“…Intracranially implanted mice were carefully monitored and longitudinally imaged as outlined in Section 2.3 . Tumor-bearing mice were euthanized at the appearance of moderate clinical signs, which were regularly assessed according to a predefined, previously described scoring system shown in detail in Supplementary Table S1 [ 57 , 58 , 59 , 60 ].…”