Pediatric brainstem glioma is among the most devastating childhood cancers. Since these tumors occur in the brainstem, there are no surgical options for providing relief to patients, and chemotherapy as well as radiation therapy provide palliative relief at best. In contrast to most types of human cancer, there has been no significant improvement in treatment outcomes for brainstem glioma patients. Until recently, the lack of improvement for treating brainstem glioma was attributable, in part, to infrequent brainstem tumor tissue acquisition for analyzing tumor molecular characteristics: the underlying genetic basis for brainstem glioma occurrence remained largely unknown, until as recently as 2012. Fortunately, two groups were able to address this information deficiency by acquiring sufficient numbers of brainstem tumors to conduct meaningful mutation analysis. Exome sequencing identified recurrent mutation of the H3F3A gene, resulting in replacement of lysine 27 by methionine (K27M) in the encoded histone H3.3 protein, in as many as 60% of these tumors. To date, this mutation remains unique and specific to brainstem tumors. The K27M mutation causes substantial reduction in histone H3 K27 methylation in cellular chromatin, and we have recently shown that inhibition of the histone H3 K27 demethylase JMJD3 acts to restore K27 methylation in brainstem glioma cells, while demonstrating potent anti-tumor activity, both in cell culture and in xenograft models of brainstem glioma. These results, as well as results published by others, support JMJD3 as having pro-tumorigenic activity in at least some types of cancer. However, it is likely that K27 methylation status in brainstem tumors is influenced by activities i addition to JMJD3: there are several enzymes that regulate K27 methylation, and whether these activities act in concert with JMJD3, to promote brainstem tumor development, or, alternatively, oppose tumor development, is largely unknown. This application will examine how different H3 K27 methyltransferase and demethylase activities influence brainstem tumor growth, and in so doing will determine whether JMJD3's role in tumor maintenance is dependent upon other proteins that regulate K27 methylation. This project will, in addition, determine how brainstem tumors adapt to extended JMJD3 inhibition, which is important for identifying secondary therapeutics to use in treating brainstem tumors that may acquire resistance to JMJD3 inhibition, or that could potentially be used in combination with JMJD3 inhibitors. Finally, this proposal will examine the effects of JMJD3 inhibition when used in combination with radiation, as one of the cellular properties affected by JMJD3 inhibition is DNA repair. JMJD3 inhibition, therefore, may enhance the cytotoxic effects of radiation.
Diffuse intrinsic pontine gliomas (DIPGs) carry a dismal prognosis despite the use of aggressive multi-modality treatment. Recent genetic analysis revealed the presence of a novel gene mutation in these tumors that alters a DNA binding protein, histone H3.3, and our recently published work was the first to demonstrate that pharmacologic inhibition of the histone modifying enzyme, JMJD3, increases cellular histone methylation in mutant DIPG tumor cells, while demonstrating potent anti-tumor activity. As such, JMJD3 appears to have oncogenic activity in contributing to the development or maintenance of DIPG, the major goals of this project are to determine whether JMJD3 oncogenic activity is influenced by other histone-modifying enzymes, how other histone modifying activities respond to sustained JMJD3 inhibition, and if JMJD3 inhibition can be used to advantage, in treating DIPG, when combined with radiotherapy.
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