Page 17 - 2020_07-Haematologica-web

Editorials
associated with high HOX gene expression. Similarly to wildtype NUP98, the NUP98-HOXA9 fusion also inter- acts with wildtype KMT2A through the second GLFG repeat of NUP98 and KMT2A is important for the recruit- ment of NUP98-HOXA9 to the HOXA locus and NUP98- HOXA9-dependent HOXA genes expression.17
Fisher et al. performed functional modeling of a peculiar NUP98-KMT2A alteration, resulting from an inv(11)(p15;q23) characterized in two AML patients and leading to the fusion of NUP98 exon 13 to KMT2A exon 2. The predicted NUP98-KMT2A fusion encodes a 4,340 amino acid protein lacking the MEN1-interacting domain but containing most of KMT2A including the H3K4 methylation SET domain, as opposed to KMT2A-X fusions (Figure 1). As the reciprocal KMT2A-NUP98 fusion transcript (between exon 1 of KMT2A and exon 14 of NUP98) was detected in only one of the two original patients,2 the hypothesis was that the NUP98-KMT2A fusion may represent the important disease driver. Fisher et al. achieved this tour de force through the development of a novel inducible NUP98-KMT2A transgenic mouse model.
The authors demonstrate the expansion and increased competitiveness of NUP98-KMT2A-expressing hematopoietic progenitor cells (Lineage-Sca1+Kit+ cells) and concomitant cell cycle abnormalities without signifi- cant changes in the relative distribution between long- term hematopoietic stem cells and multi-potent progeni- tors. Upon long-term NUP98-KMT2A expression, mice succumbed to lethal myelodysplasia or AML. The median latency for development of a hematopoietic malignancy in inducible NUP98-KMT2A mice was rather long (>1 year) and sublethal irradiation to generate DNA damage accelerated disease. Together with the observation of additional mutations in other human NUP98-rearranged15 or KMT2A-PTD,11 this strongly suggests that cooperating mechanisms are required for induction of full-blown AML by NUP98-KMT2A. Interestingly, however, the co- expression of Flt3-ITD did not accelerate the disease in this inducible NUP98-KMT2A model, suggesting differ- ent cooperating networks as compared to the NUP98- NSD1 fusion.18
Inducible models allow elegant and formal testing of whether continuous expression of the driver oncogene is required for leukemia maintenance by removing doxycy- cline treatment in diseased animals. Previously, full dependence of AML on KMT2A-MLLT3 expression was observed using a similar model.19 Here, however, ceasing the doxycycline treatment in recipients of inducible NUP98-KMT2A cells did not abrogate the disease pro- gression in all the mice. The authors suggest that this may result from a “leak” of residual NUP98-KMT2A expres- sion inherent to this inducible system. In this regard, given that KMT2A-X and NUP98-KMT2A fusion tran- script expression is controlled by different endogenous regulatory elements - KMT2A and NUP98 promoters, respectively - significantly different levels of fusion expression could be required for leukemia induction and maintenance. It remains to be formally tested whether NUP98-KMT2A expression is essential for the mainte- nance of already transformed leukemic cells in all cases.
At the molecular level, inducible NUP98-KMT2A murine leukemia cells, similarly to NUP98-KMT2A patient’s leukemic cells,2 do not show significant upregu- lation of HOX genes as compared to control cells or leukemia from two retroviral models of KMT2A fusions (KMT2A-ENL and KMT2A-MLLT3). Gene expression analyses in transgenic mouse embryonic fibroblasts con- firmed cell cycle deregulation and further demonstrated a block in induction of senescence. Notably, a subset of cell cycle- and senescence-associated genes deregulated by transgene induction in mouse embryonic fibroblasts was also found to be deregulated in murine hematopoietic pro- genitor cells (e.g. Sirt1, Rbl2, Tert2). These data suggest that NUP98-KMT2A does not transform hematopoietic pro- genitor cells through aberrant expression of HOX genes and cofactors but through an alternative mechanism asso- ciated with a defective cell cycle checkpoint. Notably, this is further supported by the absence of significant cell cycle perturbation in inducible NUP98-KMT2A cells mediated by small MEN1 or BRD inhibitors, as opposed to their effects on KMT2A-MLLT3 cells.
Three NUP98 fusion partners (NSD1, NSD3, and KMT2A) have a SET domain and another partner is a known interactor of SET-containing proteins (SETBP1) with histone methyltransferase function. Although addi- tional genome-wide chromatin analyses will be required to assess H3K4me3 profiles and NUP98-KMT2A DNA binding sites in NUP98-KMT2A cells, it could be hypoth- esized that aberrant deposition of H3K4me3 at NUP98 target genes enhances or ectopically creates promoter activities. More generally, it also remains to be deter- mined: (i) whether the NUP98 or KMT2A moiety con- trols the identity of the target genes; (ii) whether dimer- ization is required for transformation as for other KMT2A-X fusions;20 and (iii) whether the location of NUP98-KMT2A at the nuclear pore, reported to be in close proximity to the loci of actively transcribed cell cycle regulators, in part controls the identity of the target genes in a cell context-dependent manner.
Together, the identification of transcriptional targets of NUP98-KMT2A represents a first step toward the devel- opment of novel therapeutic strategies. Based on the pro- tein structure, the NUP98-KMT2A transforming proper- ties could depend on cleavage by TASPASE 1 and SET domain catalytic activity. As interference with these activities has been proposed,21 future assessment of the efficacy and specificity of targeted therapies could be of interest in these human leukemias.
Acknowledgments
We thank Olivier A. Bernard and Brian J. Huntly for scientific discussions.
Funding
AF is supported by Fondation pour la Recherche Médicale. TM is supported by Institut National Du Cancer (PLBIO- 2018-169), PAIR-Pédiatrie/CONECT-AML (Collaborative Network for Children and Teenagers with Acute Myeloblastic Leukemia: INCa-ARC-LIGUE_11905 and Association Laurette Fugain), Société Française des Cancers de l'Enfant, INCa-DGOS-INSERM_12551.
haematologica | 2020; 105(7)
1759