WP5 – Identification and validation of novel therapeutic targets
The treatment failure of GBM has been attributed to the persistence of GSCs within the GBM tumours, which are thought to drive tumour growth and progression.
This suggests that long-term disease control is likely to require elimination of this stem-like cell population, in addition to the more differentiated tumour bulk. Thus, there is an immediate need to develop novel therapies against GBM that target the stem-like cell population in GBM.
In this WP, we will identify proteins as being required for the maintenance of the stem-like cell population in GBM and qualify them as therapeutic targets for the development of precision medicine in GBM.
- identification of genes required for GBM maintenance through CRISPR-based functional genetic screens
- validation and molecular characterization of hits
Several research groups have isolated a subpopulation of GSCs, GBMs expressing immature neural stem cell markers that are thought to drive tumour growth.
These GSCs and their normal neural stem cell counterparts can be readily expanded in culture providing a disease-relevant model system for fundamental and translational studies.
We will use such human GSCs from WP4 and additionally from external collaborators (Steve Pollard, University of Edinburgh and Lene Uhrbom, Uppsala University) to identify proteins that are required for their growth in vitro and in vivo (Figure 6).
Specifically, we will screen these cell lines for genetic vulnerabilities (“drop-out screens”) using CRISPR-Cas9 technology to identify genes required for their proliferation in vitro. To do this, we have already generated focused guide RNA (gRNA) libraries targeting 1,218 human genes, coding for putative chromatin-associated proteins.
Moreover, we have generated agRNA library targeting 520 human kinase genes.
As a third subgroup of potentially druggable proteins, we will generate gRNAs to a smaller subset of genes that are found in areas of genomic-amplification and found to be over-expressed in GBMs (WP1 and publically available data).
The requirement for each of these genes, as well as hits identified using the two libraries, will be tested in PDX GBM models (from WP4). The validated proteins will be functionally characterised as an essential step for the development of precision medicine.
In addition to the in vitro approach, we are also currently testing the feasibility to identify genes required for GBM maintenance by performing screens in PDX GBM models in vivo using custom made gRNA libraries.
As for the in vitro screens, we will validate the generated hits.
Once the hits from the in vitro and in vivo screens have been validated and characterised, we will test already existing inhibitors or newly developed inhibitors (in collaboration with biotech/pharma, WP14 and 15) in the PDX models and, subsequently, in clinical trials for GBM patients.
Novel drug-targets that can be utilised in prioritisation of drug-targets and inclusion in dedicated clinical trials.