NK-2 Homeobox Transcription Factor NKX2-3 Confers Differentiation Block in NPM1-Mutated in Acute Myeloid Leukaemia

Dysregulation of HOX homeobox family transcription factor (TF) gene expression is important in the pathogenesis of AML in particular in NPM1-mutated and MLL rearranged molecular subtypes. However, little is known about the role of NK2-homeobox (NKX2) family TF genes. NKX2 genes have critical roles i...

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Published in:Blood 2023-11, Vol.142 (Supplement 1), p.2759-2759
Main Authors: Chadwick, John A, Wingelhofer, Bettina, Somervaille, Tim CP
Format: Article
Language:English
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Summary:Dysregulation of HOX homeobox family transcription factor (TF) gene expression is important in the pathogenesis of AML in particular in NPM1-mutated and MLL rearranged molecular subtypes. However, little is known about the role of NK2-homeobox (NKX2) family TF genes. NKX2 genes have critical roles in normal development of the central nervous system, thyroid, lung and pancreas. We observed high expression of NKX2-3 in patient AML samples and hypothesised the transcription factor might have a functional role in the disease. By qPCR, we found that NKX2-3 is highly expressed in normal HSCs but is rapidly down regulated as cells differentiate. In primary AML samples, it is highly expressed in 21-37% cases, including within the immunophenotypic LSC compartment, in particular in cases with NPM1 and/or FLT3-ITD mutations. Forced expression of NKX2-3 in normal murine KIT + bone marrow (BM) HSPCs transiently enhanced their clonogenic activity and impaired differentiation: NKX2-3-expressing HSPCs expressed higher levels of E2F or MYC target genes, and leukemia stem cell (LSC) maintenance genes. Congenic transplant experiments revealed that forced expression of NKX2-3 in normal BM HSPCs was sufficient to enhance multilineage engraftment across four months of follow up without inducing leukemia. In keeping with this, transplantation of Nkx2-3-/- knockout BM cells led to significantly inferior engraftment compared with BM cells from litter mate control mice. Thus, while increased expression of Nkx2-3 enhances engraftment of normal stem cells in transplantation assays, loss of Nkx2-3 impairs it. Analysis of NKX2-3high primary human AML cases demonstrated that HOXA9 was the most highly upregulated transcription factor gene compared with NKX2-3low cases. We expressed both factors singly and together in murine KIT+ BM HSPCs: Hoxa9/NKX2-3 cells exhibited significantly higher clonogenic activity and reduced morphologic and immunophenotypic differentiation compared with Hoxa9/MTV cells. Congenic BM transplant experiments demonstrated that Hoxa9/NKX2-3 recipients developed leukemias significantly earlier than Hoxa9/MTV recipients (median 82 vs 155 days). BM morphology and immunophenotyping confirmed that Hoxa9/NKX2-3AMLs exhibited a greater degree of differentiation block than Hoxa9/MTV AMLs. Leukemia cells from Hoxa9/NKX2-3 recipients coordinately expressed higher levels of E2F or MYC target genes, and LSC maintenance genes. NKX2-3 KD to ~30% of control values reduced prolifer
ISSN:0006-4971
1528-0020
DOI:10.1182/blood-2023-182990