Novel Assay Identifies Potential Ewing Sarcoma Treatments

A novel high-throughput screening assay was used to identify a number of compounds that could potentially offer therapeutic benefit in Ewing sarcoma.

A novel high-throughput screening assay was used to identify a number of compounds that could potentially offer therapeutic benefit in the bone and soft tissue tumor known as Ewing sarcoma. The assay specifically targeted genomic regions where the Ewing sarcoma-specific transcription factor EWSR1-FLI1 mediated regions of aberrant nucleosome depletion.

More and more cancers have been associated in recent years with genetic mutations relating to chromatin regulation; the chimeric transcription factor EWSR1-FLI1 is one such example. “There are characteristic regions where this oncogene goes in Ewing sarcoma to open up regions of DNA,” said study author Ian J. Davis, MD, PhD, of the University of North Carolina at Chapel Hill, in a press release. “So this offers us a unique signature for this cancer, one that is not observed in other cancers, creating an opportunity to use it in our new screen.”

The researchers adapted a formaldehyde-assisted isolation of regulatory elements (FAIRE) method into a high-throughput automated assay (HT-FAIRE). They used this to screen a targeted small molecule library by specifically evaluating the regions of nucleosome depletion caused by the EWSR1-FLI1 protein in cells from Ewing sarcoma patients. The results were published online ahead of print in Proceedings of the National Academy of Sciences.

The screen included a total of 640 small molecules. Researchers calculated a “relative chromatin inhibition” score measuring the ability to reduce chromatin accessibility at targeted sites, and those compounds with a score more than two standard deviations from the mean were deemed “hits.” There were 58 hits; a secondary screen method with a more stringent threshold identified 15 promising compounds.

Surprisingly, histone deacetylase (HDAC) inhibitors were by far the most promising of compounds. Of 21 HDAC inhibitors in the screen library, 14 scored as hits on both primary and secondary screens. Several other uncharacterized compounds also were found to be promising.

Though the HDAC inhibitors surprised the authors based on the mechanisms of the drugs, there have been earlier studies showing activity of these agents in Ewing sarcoma. Specifically, romidepsin, vorinostat, and others have been shown to have anti-proliferative effects on Ewing sarcoma cells. All HDAC inhibitors that previously showed promise in this malignancy were identified by the HT-FAIRE assay.

The authors noted that the development of the assay is broadly useful across other malignancies as well. “HT-FAIRE is applicable to any cellular model associated with a specific chromatin accessibility signature and offers a general strategy to disrupt the function of proteins lacking structure amenable to small molecular targeting,” they wrote.