Discovery, Development, and Evaluations of Human MLL1 Core Complex Inhibitors.

Abstract

In higher eukaryotes, epigenetic modulation of gene expression lies at the heart of cell fate and identity determination. One of the major regulatory epigenetic mark is histone H3 Lysine 4 methylation (H3K4me), the general hallmark of transcription activation. In metazoans, majority of H3K4me is deposited by the Mixed Lineage Leukemia (MLL) family of methyltransferases. The family comprises of three distinct subclasses (a total of six members): MLL1/2 (KMT2A/2B), MLL3/4 (KMT2C/2D), and SET1A/B (KMT2F/2G). Each MLL family subclass has distinct product specificity, complex compositions, and consequently, diverse functions in cells. However, they share identical enzymatic core subunits (MLL SET domain, WDR5, RBBP5, and ASH2L) and recognize the same lysine residue on histone tail. To delineate the unique physiological functions of the MLL family methyltransferases, we resort to develop a series of molecular tools to complement the tranditional genetic knockdown and knockout approaches to probe MLL family functions in normal and malignant physiology. To this end, we employ two major strategies to target the MLL1 core complex: 1) discover small molecule inhibitors that directly target the enzymatic activity of MLL1 via High-Throughput Screening, and 2) rationally design a series of chemical probes specifically disrupting the WDR5-MLL1 SET domain interaction. Through the work done in this thesis, we were able to establish a high- throughput screening assay to screen for MLL1 small molecule inhibitors, as well as setting up secondary assays to further validate and characterize the inhibitors. Through this process, we identified small molecule leads that were specific for inhibiting MLL1, as well as leads that target more than one MLL family enzymes, which are the first-in-class to our knowledge. In addition to discovery of MLL1 small molecule leads, we further improved the in-vitro and in- vivo stability and potency of peptidomimetic inhibitors MM-101 and 102 by 10 and 3-6 folds, respectively. Taken together, we have developed a series of molecular probes that will facilitate future studies of MLL and serve as lead compounds for development of cancer therapeutics.