Drug discovery targeting the interaction of MLL-AF4 with AF9 in pediatric MLL-rearranged leukemia
Date
2013
Authors
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Publisher
University of Delaware
Abstract
Rearrangement of the mixed-lineage leukemia (MLL) in pediatric leukemia is generally a predictor of a very poor prognosis. These chromosomal rearrangements result in fusion of the protein MLL to one of more than 60 protein partners. MLL fusions are potent inducers of leukemia through activation of oncogene expression; therefore, targeting this transcriptional activation function may arrest MLL-rearranged (MLL-R) leukemia. Leukemic cell lines harboring the most common fusion protein, MLL-AF4, require the direct interaction of AF4 with the transcription factor AF9 to survive and self-renew. Disrupting this interaction with a cell-penetrating AF4-derived peptide results in cell death, suggesting that the AF4-AF9 interaction could be a viable target for a novel MLL-R leukemia therapy. To discover chemical compounds that disrupt AF4-AF9 binding, I used a high-throughput screening (HTS) assay using homogeneous proximity-based AlphaScreen technology to detect non-peptidic inhibitors of AF4-AF9 binding. The assay had been previously validated in the HTS lab by screening small compound libraries containing 5,680 compounds. I confirmed the activity of hits from this pilot screen by testing in dose response, and further characterized the most promising active compound. After pilot screening, I used this assay to screen a library from the Lankenau Chemical Genomics Center (LCGC) comprising 96,000 orthogonally pooled compounds. The assay was also transferred to the Broad Institute (Cambridge, MA) and used to screen 350,000 compounds from the NIH collection. I confirmed hits from both screens by testing in dose-response, and identified one hit from the Broad Institute screening with promising activity. All other remaining hits from both the Broad Institute and LCGC screening were found to be artifacts interfering with the assay. In an attempt to improve potency of the two promising hits from the Nemours pilot screen and the Broad Institute HTS, analogs were synthesized in Dr. John Koh’s lab at the University of Delaware (Department of Chemistry and Biochemistry) and tested by me at Nemours. Work will continue to further optimize inhibitors of the interaction of MLL-AF4 and AF9 so that these may be tested in leukemia cells and ultimately as potential therapeutics for MLL-R leukemia.