Research And Grants

Harnessing Developmental Neurobiology to Reprogram the Cell of Origin of Diffuse Midline Gliomas

Transcription factors can be considered as “molecular switches” that can turn ON or OFF specific sets of genes. The overall objective of this project is to understand how the gene networks up- and downstream of the transcription factor DLX2, and how changes to DLX2 itself, affect brain and DMG development. This will be accomplished in two scientific aims: In Aim 1, we will use advanced genomics and gene-editing technologies to investigate the developing mouse brain (collected from developing mouse embryos). We will determine what other proteins and genes change the behaviour (or function) of DLX2, as well as what proteins and genes are impacted by DLX2. This will help us understand the networks that regulate DLX2, and how normal and abnormal brain development in this region of the brain occurs. In Aim 2, we will manipulate DLX2 in cells that are derived from the DMG tumors of patients, to determine how the cell growth is impacted by changes in DLX2. These manipulated DMG cells will be observed in cell-culture (in vitro) and in live mice where these modified DMG cells have been surgically implanted into newborn mouse brainstem (in vivo).

Despite significant efforts, there has been no improvement in the treatment and outcomes for DIPG (diffuse intrinsic pontine glioma)/DMG (diffuse midline glioma) in the past 50 years. In 2012, the co-discovery that the majority of DMG, including DIPG, have specific mutations of variant histones (known as H3.3 and H3.1) fundamentally changed what we know about these tumors, and as a result, many researchers have redirected their efforts to systemically unravel the underlying biology and to apply these learnings towards the development of new therapies.

Histones help to “wrap” our DNA so that it fits inside the nucleus of the cell. Changes to histones, known as histone modifications, further assist to “unwrap” or “wrap” specific regions of the DNA within the nucleus of the cell so that specific genes or sets of genes can either be expressed or are blocked from being expressed. The more recent application of single cell genomics platforms using DMG samples, including RNA sequencing, support that there is a specific cell of origin for DMGs, an oligodendroglial progenitor cell (OPC). In the adult brain, OPCs eventually become oligodendrocytes that make myelin, a substance that insulates the projections of other brain cells called neurons and coordinates brain function. However, current and proposed therapies for DMG do not target this cell of origin. Our novel work in the emerging field of Cancer Neuroscience addresses the developmental neurobiology of these tumors and through sophisticated and comprehensive analysis of genetic and epigenetic networks and structures, as well as new models (using cell culture and mouse models), we will unlock the underlying biological basis of these tumors towards approaches to novel therapies for DMG.

The Eisenstat laboratory is one of three labs in the world that are expert in understanding the role of the distalless (DLX) class of transcription factors in brain development and the only one focused on applying this knowledge to help solve DMG. The Neuro-oncology laboratory based at the Murdoch Children’s Research Institute (MCRI) and the Brain Cancer Research laboratory based at the Walter and Eliza Hall Institute (WEHI), both affiliated with the University of Melbourne, Australia, have the specialised skillsets and laboratory platforms to perform this work. The MCRI has established and maintained the genetically engineered mouse models lacking Dlx gene function, and has the expertise required for the genetic manipulation of DMG cell lines, including advanced gene editing techniques known as CRISPR. The MCRI also has access to multiple cell-lines derived from patients with DMG which will be used for the these experiments. The WEHI has the specialised equipment required for performing the newborn mouse implantation experiments, as well as the required imaging technology to visualise the tumor development in live mice.

Ultimately, Aim 1 will provide a comprehensive understanding of the role DLX2 plays in normal brain development (and how it interacts with other transcription factors), while Aim 2 will apply this knowledge to how varying levels of DLX2 impacts DMG development. These findings will inform future experiments at the MCRI, including more complex cell culture experiments and sophisticated characterisation of the DLX2 protein itself towards making this transcription factor “druggable”. Our longer-term translational goal is to alter the level of DLX2 in patients with DMG as a novel form of differentiation therapy, so that we may take advantage of the underlying developmental neurobiology of the tumor to change outcomes for DMG patients.