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Title:
Investigating the role of MLL protein in the epigenetic regulation of paediatric diffuse midline glioma
Supervisors:
David Michod and Juan Pedro Martinez-Barbera
Project Description:
Background
The goals of this project are to unravel the role of the protein mixed lineage leukaemia (MLL) in the context of paediatric diffuse midline glioma (pDMG) and to evaluate preclinically the therapeutic potential of targeting the MLL complex against pDMG. pDMG has a dismal prognosis, with no curative treatments, making it one of the most devastating paediatric malignancies. pDMG is characterized by a specific mutation, H3K27M, resulting in the expression of a mutant histone 3 protein in which lysine 27 is replaced by a methionine. H3K27M functions as a dominant negative mutation that severely reduces global levels of H3 lysine 27 tri-methylation (H3K27me3) with a concomitant increase in H3 lysine 27 acetylation (H3K27ac), which result in the transcriptional activation of the oncogenic programme leading to pDMG development.
Our preliminary data have revealed that the oncoprotein MLL is recruited to chromatin regions that are enriched in H3K27M. We further show that upon MLL degradation the H3K27 acetyl transferase p300 is displaced from these regions, which is accompanied by a marked decrease of H3K27ac. This epigenetic normalisation leads to reduced expression of pDMG oncogenic genes. Building on these preliminary data, we seek to explore the following hypotheses: (1) MLL is a key player in activating the oncogenic programme of pDMG; (2) Inhibition of the MLL complex will result in decreased expression of oncogenic drivers and reduced viability of pDMG cancer cells.
Hypothesis, aims and methods.
We hypothesise that in pDMG, MLL is strongly associated with H3K27M mutant histones which stall or relocates MLL complex to genes contributing to the oncogenic properties of the specific histone mutation. Therefore, preventing chromatin occupancy of the MLL complex will reduce the expression of key oncogenic factors in pDMG, which may offer a new therapeutic avenue. The objectives of this proposal are:
1. To identify the MLL-chromatin interactions in pDMG cells. We will use Cut&Tag and Cut&Run assays to reveal the specific chromatin interactions of MLL, its cofactors and the associated histone modifications using a range well characterised patient-derived pDMG cell lines. (year1&2)
2. To reveal the effects of MLL ablation in pDMG cells. We will combine siRNA knockdown and protein degradation approaches to reveal the function of all components of the MLL complex and the associated histone modifications in the specific context of H3K27M expression. (year1&2)
3. To assess the therapeutic efficacy of targeting MLL in vitro and in vivo. We will test the efficacy of a range of inhibitors, some clinically approved, against pDMG in cultured cells and, in both immunosuppressed and immunocompetent murine models to provide data to support a clinical trial. (year3)
References:
1. Hargrave D, Bartels U, Bouffet E. Diffuse brainstem glioma in children: critical review of clinical trials. Lancet Oncol. 2006;7(3):241-8.
2. Schwartzentruber J, Korshunov A, Liu XY, Jones DT, Pfaff E, Jacob K, et al. Driver mutations in histone H3.3 and chromatin remodelling genes in paediatric glioblastoma. Nature. 2012;482(7384):226-31.
3. Bender S, Tang Y, Lindroth AM, Hovestadt V, Jones DT, Kool M, et al. Reduced H3K27me3 and DNA hypomethylation are major drivers of gene expression in K27M mutant pediatric high-grade gliomas. Cancer Cell. 2013;24(5):660-72.
5. Meier S, Cantilena S, Chirou MVN, Anderson J, Hargrave D, Salomoni P, et al. Alcohol-abuse drug disulfiram targets pediatric glioma via MLL degradation. Cell Death Dis. 2021;12(8).
Contact Information:
David Michod