3 research outputs found
Discovery of first-in-class inhibitors of ASH1L histone methyltransferase with anti-leukemic activity
ASH1L histone methyltransferase plays a crucial role in the pathogenesis of different diseases, including acute leukemia. While ASH1L represents an attractive drug target, developing ASH1L inhibitors is challenging, as the catalytic SET domain adapts an inactive conformation with autoinhibitory loop blocking the access to the active site. Here, by applying fragment-based screening followed by medicinal chemistry and a structure-based design, we developed first-in-class small molecule inhibitors of the ASH1L SET domain. The crystal structures of ASH1L-inhibitor complexes reveal compound binding to the autoinhibitory loop region in the SET domain. When tested in MLL leukemia models, our lead compound, AS-99, blocks cell proliferation, induces apoptosis and differentiation, downregulates MLL fusion target genes, and reduces the leukemia burden in vivo. This work validates the ASH1L SET domain as a druggable target and provides a chemical probe to further study the biological functions of ASH1L as well as to develop therapeutic agents
Two Loops Undergoing Concerted Dynamics Regulate the Activity of the ASH1L Histone Methyltransferase
ASH1L
(absent, small, or homeotic-like 1) is a histone methyltransferase
(HMTase) involved in gene activation that is overexpressed in multiple
forms of cancer. Previous studies of ASH1L’s catalytic SET
domain identified an autoinhibitory loop that blocks access of histone
substrate to the enzyme active site. Here, we used both nuclear magnetic
resonance and X-ray crystallography to identify conformational dynamics
in the ASH1L autoinhibitory loop. Using site-directed mutagenesis,
we found that point mutations in the autoinhibitory loop that perturb
the structure of the SET domain result in decreased enzyme activity,
indicating that the autoinhibitory loop is not a simple gate to the
active site but is rather a key feature critical to ASH1L function.
We also identified a second loop in the SET-I subdomain of ASH1L that
experiences conformational dynamics, and we trapped two different
conformations of this loop using crystallographic studies. Mutation
of the SET-I loop led to a large decrease in ASH1L enzymatic activity
in addition to a significant conformational change in the SET-I loop,
demonstrating the importance of the structure and dynamics of the
SET-I loop to ASH1L function. Furthermore, we found that three C-terminal
chromatin-interacting domains greatly enhance ASH1L enzymatic activity
and that ASH1L requires native nucleosome substrate for robust activity.
Our study illuminates the role of concerted conformational dynamics
in ASH1L function and identifies structural features important for
ASH1L enzymatic activity