Identifying the mechanisms responsible for low- to high-grade transformation in IDH mutant astrocytoma

Background: IDH mutant (IDHmut) astrocytoma is an incurable brain cancer that typically presents as a low-grade lesion but high-grade transformation occurs uniformly, after which average survival is 2-years. There is a lack of understanding as to how this transformation occurs. The low-grade state provides a novel (and prolonged) therapeutic window that remains unexploited, currently.

IDH mutation causes 2-hydroxyglutarate production, inhibiting 2-oxoglutarate-dependent dioxygenases (2-OGDD) leading to hypermethylation (via TET inhibition), aberrant chromatin configuration, and oncogene activation. Little is known about how IDHmut affects other 2-OGDDs and subsequent transcription factor (TF) binding.

Hypothesis: Aberrant metabolism and 2-OGDD dysfunction dysregulate epigenomic/transcriptional networks causing high-grade transformation in IDHmut astrocytoma.

Aims: Aim 1: Map transforming IDHmut astrocytoma across human resection specimens High-grade IDHmut glioma contain significant co-existent areas of low-grade disease.

I reason that by looking ‘back in time’ from the high-grade regions, and sampling regions sequentially from this area, in multiple directions, we will map the molecular landscape of transformation ‘in reverse’ by layering different spatial and single cell -omic technologies.

My approach eliminates inter-patient variation (e.g. genomic/epigenomic differences), sampling bias (published literature explore primary/recurrent disease, with spatially incongruent regions compared) and treatment bias (intervening surgery, chemotherapy, radiotherapy).

We will perform: a) Multi-region mass spectrometry imaging (MSI) of sequential high to low-grade areas using ultra-rapid intraoperative freezing (<​2s). b) Corresponding multi-region spatial transcriptomic analysis using Visium and proteomic analysis using Imaging Mass Cytometry. c) Nanopore, histone modification and TF analysis of FACS/FANS from corresponding regions Aim 2: Identify how epigenomic factors orchestrate gene expression programmes that drive low to high-grade transformation in IDHmut astrocytoma Modelling IDHmut glioma has been challenging for the field of glioma biology.

We have developed a novel canonical low-grade IDHmut mouse model that produces low grade features in vivo and can be cultured in vitro. We have also recently derived IDHmut cell lines from primary human material. Untargeted and metabolically targeted CRISPR screens will be performed in human and mouse IDHmut cell lines in vitro.

Moreover, we will CRISPR ‘hits’ from Aim 1 (patient material) and Aim 2 in vitro and in vivo.

Outcomes: These data will be used to identify potential therapeutic targets that block transformation, potentially improving patient outcomes.