Abstract 462: The mutated chromatin modifier gene, H3F3AK27M, is a druggable target of DNA hypomethylating agents in pediatric high-grade glioma
Authors: Gujar Hemant , Zhang Lian , Li Hong-Tao , Otani Hitoshi , Weisenberger Daniel J. , Erdreich-Epstein Anat , Liang Gangning 
Published: September 2020

Abstract
Purpose: With the current standard of care for Pediatric high-grade glioma (pHGG) patients the 5-year survival rate remain low at 5-35%. Thus, there is an urgent need to develop improved therapies for pHGG patients. A specific point mutation of histone H3 (H3F3A) has been identified at amino acid 27 (lysine to methionine (K27M)), resulting in inhibition of EZH2 (histone H3K27 tri-methylase), and reduced global levels of H3K27 tri-methylation (H3K27me3). The H3K27M alteration can be found in the majority (~80%) of pHGGs and represents an attractive target for diagnostic and treatment. Since the H3K27me3 mark correlate with gene silencing, mutations resulting in global decrease in H3K27me3 may result in aberrant gene regulation. Majority of demethylated promoters remain silenced after treatment with hypomethylating agent (HMA) due to the presence of repressive H3K27me3. HMAs treatment is also rendered ineffective in case of repression of endogenous retroviruses (ERVs) due to the presence of H3K27me3. Hence, pHGGs harboring the H3K27M mutation with low levels of H3K27me3 represent an attractive therapeutic target for treatment with HMAs. HMAs are approved by FDA for treatment of several forms of human cancers.
Methods: We had introduced mutant H3K27M in human pediatric glioma cell (SF188). Pediatric glioma cell lines (wild type and endogenous or endogenous expressed with H3K27M) were treated with DNA hypomethylating agents (HMAs). Cell doubling time was calculated. DNA methylation and gene expression were monitored at different time points. Additional RNA-seq data were obtained from pediatric glioma patient and patient derived cell lines.
Results: We demonstrate that pHGG cells carrying H3K27M mutation are more aggressive and showed down-regulation of innate immune response genes (immune deficient or immune cold) and up-regulation of cancer stem like genes as compared to the wild-type pHGG cells. These findings are further confirmed with published data from patient-derived pHGG cells. The prolonged antitumor effects in pHGG cells with H3K27M mutation observed after HMA treatment may be due to the failure of DNA remethylation and upregulated innate immune response pathway genes due to ERV reactivation.
Conclusion: Our findings suggest that HMA treatment may be effective against pHGGs with the H3K27M mutation and critical for converting immune deficient cells into immune reactive cells (immune hot). The success of this project will lead to clinical trials for pHGG patients to evaluate HMAs with immune-based therapies in pHGG patients, for whom there are currently no effective treatments.
Citation Format: Hemant Gujar, Lian Zhang, Hong-Tao Li, Hitoshi Otani, Daniel J. Weisenberger, Anat Erdreich-Epstein, Gangning Liang. The mutated chromatin modifier gene, H3F3AK27M, is a druggable target of DNA hypomethylating agents in pediatric high-grade glioma [abstract]. In: Proceedings of the Annual Meeting of the American Association for Cancer Research 2020; 2020 Apr 27-28 and Jun 22-24. Philadelphia (PA): AACR; Cancer Res 2020;80(16 Suppl):Abstract nr 462.