Abstract 883: NUP98-fusion proteins interact with the NSL/MLL1 complexes to drive leukemogenesis

Abstract

Abstract The Nucleoporin 98 gene (NUP98) is fused to a variety of partner genes in an array of hematopoietic malignancies via chromosomal translocations involving 11p15. NUP98-rearranged leukemias show elevated HOXA and HOXB cluster genes and mouse model systems have recapitulated this high-level expression independent of whether the leukemias are derived from mouse or human bone marrow. However, the molecular mechanisms of NUP98-fusion mediated leukemogenesis and elevated HOX gene expression in this leukemia are unclear. Recent studies in Drosophila show that nucleoplasmic Nup98 functions as a potential transcriptional activator, and physically interacts with the non-specific lethal (NSL) and trithorax (Trx)/mixed lineage leukemia (MLL) complex (Kalverda et al., 2010; Pascual-Garcia et al., 2014). To test whether the intranuclear localized NUP98-fusion proteins interact with NSL/MLL1 complexes, coimmunoprecipitation (co-IP) experiments using 293T cells with overexpression of NUP98 fusions and wild type (WT) NUP98 were performed. NUP98-fusion proteins NUP98-HOXA9, NUP98-HOXD13, NUP98-NSD1 and NUP98-PHF23, but not the full-length WT NUP98, show physical interaction with MLL1 and the NSL histone acetyltransferase (HAT) complexes. Genome-wide chromatin immunoprecipitation followed by next generation sequencing (ChIP-seq) illustrated that NUP98-HOXA9 and MLL1 co-localize at Hoxa and Hoxb gene promoters, which correlates with the presence of activating chromatin modifications such as H4K16ac and H3K4me3. In vitro and in vivo functional assays further showed that Mll1 is crucial for the growth of NUP98-HOXA9-transformed cells, and for the initiation and maintenance of NUP98-HOXA9 driven AML. These findings were further supported by transcriptome analyses performed in mouse NUP98-HOXA9 transformed cells lacking Mll1. We found a significant enrichment of co-bound targets of MLL1 and NUP98-HOXA9 and genes downregulated in the absence of Mll1. Gene set enrichment analysis (GSEA) demonstrated strong similarity between Mll1-dependent gene expression signature in our murine NUP98-HOXA9 AML model and the expression profile of human NUP98-fusion AML, indicating that our findings in murine AML models can be extended to human AML. Finally, the overexpression of Hoxa9 and Meis1, direct binding targets of NUP98-HOXA9 and MLL1 that are downregulated upon Mll1 loss, rescues the in vitro transformation defects in NUP98-HOXA9 Mll1-/- cells. In conclusion, our findings support a model where NUP98-fusion proteins recruit the NSL/MLL1 complex as an important step during leukemogenesis. The deregulated HOX gene expression in NUP98 fusion transformed cells remains dependent on MLL1. The discovery of this common leukemogenic pathway for NUP98-fusion proteins opens up new avenues for potential therapeutic opportunities to treat patients with leukemia that harbor various NUP98 rearrangements. Citation Format: Haiming Xu, Daria G. Valerio, Meghan E. Eisold, Amit Sinha, Richard. P. Koche, Wenhuo Hu, Chun-Wei Chen, Scott H. Chu, Gerard L. Brien, Scott A. Armstrong. NUP98-fusion proteins interact with the NSL/MLL1 complexes to drive leukemogenesis. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 883.