Deciphering the mechanistic role of mixed lineage leukemia partial tandem duplications (MLLPTD) in MDS and AML Free

The mixed lineage leukemia (MLL) methyltransferase gene is a member of the trithorax group family of factors and plays a significant role in development and hematopoiesis. In a healthy context, MLL is a lysine methyltransferase that catalyzes trimethylation of histone H3 at lysine 4 (H3K4me3), facilitating a transcriptionally permissive environment at promoters. However, defects in MLL are associated with hematopoietic malignancies and leukemias. One class of MLL-related leukemias involves rearrangements of the MLL N-terminal region that manifest as in-frame partial tandem duplications (PTD) of MLL's DNA binding domains, resulting in an elongated protein with aberrant function, with the most common form of MLL^PTD containing duplications of exons 2-6. MLL^PTD appears in up to 10% of patients with myelodysplastic syndrome (MDS) or acute myeloid leukemia (AML) and has been identified as a strong predictor of adverse outcomes, on par with defects in p53. Despite its clinical importance, the MLL^PTD has not been well characterized and its mechanistic functions remain poorly understood. We have characterized the MLL^PTD at the transcript, protein, and genomic level. Moreover, using quantitative mass spectrometry, we show that MLL^PTD cells have a lower overall MLL copy number per nucleus than MLL WT cells. Despite this low copy number, we observe that MLL^PTD exhibits broader binding to chromatin than WT MLL N-term when measured by CUT&Tag. Interestingly, we also observe increased H3K4 trimethylation at genes bound by MLL^PTD. We conclude that the increased number of MLL^PTD DNA binding domains combined with the intrinsic disorder of large regions of the MLL protein create increased opportunities for MLL to interact with distal chromatin regions, resulting in greater methyltransferase activity at a smaller number of loci. We propose that this altered methylation landscape promotes an aberrant gene regulatory program that drives leukemogenesis in MLL^PTD cells. We also hypothesized that MLL^PTD facilitates the development of AML by disrupting normal gene expression through differential protein-protein interactions relative to wild-type MLL. We report the identification of a preferential MLL^PTD interactor, KAT2A, that can be targeted for acute degradation. Pharmacological degradation of KAT2A has a distinctively deleterious effect on cells expressing MLL^PTD. Furthermore, treating MLL^PTD cells with a KAT2A degrader slows engraftment when these cells are transplanted into immunodeficient mice. We conclude that KAT2A plays a crucial role in promoting MLL^PTD-associated leukemogenesis.