An overwhelming proportion of drug development in both academia and industry is focused on identifying novel drug targets for the treatment of cancer. The Sussex Drug Discovery Centre is no exception; our oncology portfolio is constantly expanding and changing in order to be at the cutting edge of research for a disease that has an economic cost of £15.8bn a year in the UK alone, and is responsible for almost 15% of human deaths worldwide.
Typically, the drug development process begins with identifying inhibitors of a therapeutic target, which involves screening small molecules against a biological component involved in the disease pathway – often a specific enzyme or receptor. Because cancers are caused by mutations to genes that regulate cell growth and differentiation, resulting in unregulated cell division, most targets in cancer research are transcription factors or their corresponding gene products. While many drugs have entered clinical development this way – such as those targeting the bromodomain reader BRD4, histone deacetylase (HDAC) enzymes, and DNA methylation – challenges remain in identifying drugs that do more than simply indiscriminately kill cells, and that are able to target key transcription factors with molecular features that are considered ‘undruggable’.
In a revolutionary new study, researchers at the University of North Carolina developed and tested a new high-throughput screening technique that makes it possible to test potential drug compounds against an altered transcription factor previously thought to be ‘undruggable’ that is present in most Ewing sarcomas – a highly malignant bone and soft tissue tumour that affects children and young adults. Ewing sarcoma is characterized by a chromosomal translocation that produces the chimeric transcription factor EWSR1-FLI1; its corresponding gene product then localizes to specific regions of DNA, causing the chromatin to unwind. This creates a unique, disease-specific chromatin signature resulting from nucleosome depletion, making EWSR1-FLI1 an attractive drug target despite the fact that the mechanism for its action remains unknown. In this study, the researchers adapted formaldehyde-assisted isolation of regulatory elements (FAIRE), a biochemical assay for the identification of nucleosome-depleted regions of the genome, to a miniaturized and automated form that allowed high-throughput testing of the effects of compounds on proteins that regulate chromatin.
FAIRE in its standard format is dependent on organic extraction with a mixture of phenol and chloroform; in high-throughput (HT)-FAIRE this phase was substituted with a column-based approach to allow for the switch to automation, and performed similarly when compared side-by-side with standard FAIRE. Then, using a custom library of 640 small molecules, a screen was performed against a cell model of Ewing sarcoma to see if any of the molecules could restore normal chromatin structure. This was defined by a reduction in the FAIRE signal, indicative of a decrease in chromatin accessibility as the EWSR1-FLI1-mediated effects are reversed and the chromatin re-folds. Through the screen it was found that HDAC inhibitors were particularly effective at restoring chromatin structure, and while this class of compounds had been previously identified at a potential treatment for Ewing sarcoma, the screen also elicited novel compounds that were active in the cell model.
These results are undoubtedly remarkable, and highlight the advantages of a technique such as HT-FAIRE over the target-based screening approach that is the staple of modern drug discovery. By targeting a specific characteristic of Ewing sarcoma, HT-FAIRE removes the need for lengthy enzyme assay optimization or a comprehensive understanding of the molecular mechanisms underpinning the disease. If this method can identify potential drugs in one specific type of cancer, it may be possible to apply this technique to other cancers, and potentially reform drug discovery in the process.
Blog written by Chloe Koulouris